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Editor’s Choice Articles

Editor’s Choice articles are based on recommendations by the scientific editors of MDPI journals from around the world. Editors select a small number of articles recently published in the journal that they believe will be particularly interesting to readers, or important in the respective research area. The aim is to provide a snapshot of some of the most exciting work published in the various research areas of the journal.

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17 pages, 4849 KiB  
Article
Solid Lipid Nanoparticles Loaded with Dexamethasone Palmitate for Pulmonary Inflammation Treatment by Nebulization Approach
by Hsin-Hung Chen, Chen-Hsiang Sang, Chang-Wei Chou, Yi-Ting Lin, Yi-Shou Chang and Hsin-Cheng Chiu
Pharmaceutics 2024, 16(7), 878; https://doi.org/10.3390/pharmaceutics16070878 - 29 Jun 2024
Viewed by 1605
Abstract
Pneumonia stands as the leading infectious cause of childhood mortality annually, underscoring its significant impact on pediatric health. Although dexamethasone (DXMS) is effective for treating pulmonary inflammation, its therapeutic potential is compromised by systemic side effects and suboptimal carrier systems. To address this [...] Read more.
Pneumonia stands as the leading infectious cause of childhood mortality annually, underscoring its significant impact on pediatric health. Although dexamethasone (DXMS) is effective for treating pulmonary inflammation, its therapeutic potential is compromised by systemic side effects and suboptimal carrier systems. To address this issue, the current study introduces solid lipid nanoparticles encapsulating hydrophobic dexamethasone palmitate (DXMS-Pal-SLNs) as an anti-inflammatory nanoplatform to treat pneumonia. The specialized nanoparticle formulation is characterized by high drug loading efficiency, low drug leakage and excellent colloidal stability in particular during nebulization and is proficiently designed to target alveolar macrophages in deep lung regions via local delivery with the nebulization administration. In vitro analyses revealed substantial reductions in the secretions of tumor necrosis factor-α and interleukin-6 from alveolar macrophages, highlighting the potential efficacy of DXMS-Pal-SLNs in alleviating pneumonia-related inflammation. Similarly, in vivo experiments showed a significant reduction in the levels of these cytokines in the lungs of mice experiencing lipopolysaccharide-induced pulmonary inflammation after the administration of DXMS-Pal-SLNs via nebulization. Furthermore, the study demonstrated that DXMS-Pal-SLNs effectively control acute infections without causing pulmonary infiltration or excessive recruitment of immunocytes in lung tissues. These findings highlight the potential of nebulized DXMS-Pal-SLNs as a promising therapeutic strategy for mitigating pneumonia-related inflammations. Full article
(This article belongs to the Special Issue Lipid-Based Nanoparticulate Drug Delivery Systems: Preparation, Biomedical Applications, and Evaluation)
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Figure 1

Figure 1
<p>TEM images of (<b>a</b>) DXMS-Pal-SLNs-1 and (<b>b</b>) DXMS-Pal-SLNs-2.</p> Full article ">Figure 2
<p>(<b>a</b>) Fluorescence images of MH-S cells co-incubated with DiI-DXMS-Pal-SLNs for 4 h at 37 °C. Scale bars are 20 μm. DiI was employed as a fluorescence probe for monitoring the cellular uptake of the SLNs by the alveolar macrophages. The nuclei were stained with Hoechst 33342 in blue. (<b>b</b>) Intracellular DiI fluorescence intensity of MH-S cells after co-incubation with DiI-DXMS-Pal-SLNs for 2 h and 4 h, respectively, followed by the cell lysis with DMSO (<span class="html-italic">n</span> = 3). The DiI fluorescence intensity was determined by fluorescence spectroscopy. NS <span class="html-italic">p</span> &gt; 0.05 and ** <span class="html-italic">p</span> &lt; 0.01.</p> Full article ">Figure 3
<p>(<b>a</b>) Cell viability of MH-S cells treated with DSP, DXMS-Pal-SLNs-1 and DXMS-Pal-SLNs-2 by MTT assay (<span class="html-italic">n</span> = 3). The secretion levels of (<b>b</b>) TNF-α and (<b>c</b>) IL-6 in LPS-induced inflammatory MH-S cells after the treatment with DSP and DXMS-Pal-SLNs at various DXMS concentrations (<span class="html-italic">n</span> = 3). The cytokine levels were determined by ELISA kits. NS <span class="html-italic">p</span> &gt; 0.05 and ** <span class="html-italic">p</span> &lt; 0.01.</p> Full article ">Figure 4
<p>Determination of the levels of (<b>a</b>) TNF-α and (<b>b</b>) IL-6 in the BALF of mice experiencing LPS-induced lung inflammation (LPS 1.0 mg/kg in 100 μL of PBS) at 22 h after the treatments with DXMS-Pal-SLNs and DSP (DXMS 5.0 mg/kg in 100 μL of PBS), respectively, via intratracheal injection (<span class="html-italic">n</span> = 6). The cytokine levels were determined by ELISA kits. NS <span class="html-italic">p</span> &gt; 0.05 and ** <span class="html-italic">p</span> &lt; 0.01.</p> Full article ">Figure 5
<p>Determination of the levels of TNF-α and IL-6 in the BALF of mice experiencing the LPS-induced lung inflammation (LPS 1.0 mg/kg) at 22 h after the treatments with DXMS-Pal-SLNs and DSP, respectively, via the nebulization administration (<span class="html-italic">n</span> = 6) at the DXMS doses of (<b>a</b>,<b>b</b>) 5.0 mg/kg, (<b>c</b>,<b>d</b>) 10.0 mg/kg and (<b>e</b>,<b>f</b>) 15.0 mg/kg in 6 mL of PBS. The cytokine levels were evaluated by ELISA kits. NS <span class="html-italic">p</span> &gt; 0.05, * <span class="html-italic">p</span> &lt; 0.05 and ** <span class="html-italic">p</span> &lt; 0.01.</p> Full article ">Figure 5 Cont.
<p>Determination of the levels of TNF-α and IL-6 in the BALF of mice experiencing the LPS-induced lung inflammation (LPS 1.0 mg/kg) at 22 h after the treatments with DXMS-Pal-SLNs and DSP, respectively, via the nebulization administration (<span class="html-italic">n</span> = 6) at the DXMS doses of (<b>a</b>,<b>b</b>) 5.0 mg/kg, (<b>c</b>,<b>d</b>) 10.0 mg/kg and (<b>e</b>,<b>f</b>) 15.0 mg/kg in 6 mL of PBS. The cytokine levels were evaluated by ELISA kits. NS <span class="html-italic">p</span> &gt; 0.05, * <span class="html-italic">p</span> &lt; 0.05 and ** <span class="html-italic">p</span> &lt; 0.01.</p> Full article ">Figure 6
<p>Determination of the levels of (<b>a</b>) TNF-α and (<b>b</b>) IL-6 in the BALF of mice experiencing the LPS-induced lung inflammation (with an LPS dose of 2.0 mg/kg) at 22 h after the treatments with DXMS-Pal-SLNs and DSP (DXMS 15 mg/kg in 6.0 mL PBS), respectively, via the nebulization administration (<span class="html-italic">n</span> = 6). NS <span class="html-italic">p</span> &gt; 0.05 and ** <span class="html-italic">p</span> &lt; 0.01.</p> Full article ">Figure 7
<p>H and E-stained lung tissues of mice experiencing LPS-induced lung inflammation after the treatment with DSP, DXMS-Pal-SLNs-1 and DXMS-Pal-SLNs-2 via the nebulizer administration. The recruitment of alveolar macrophages and immunocytes in the alveolar sacs and bronchioles is highlighted by red and green arrows, respectively. The scale bars represent 50 μm.</p> Full article ">Scheme 1
<p>Schematic representation of the working mechanism of nebulized DXMS-Pal-SLNs for the treatment of pneumonia-related inflammation.</p> Full article ">
29 pages, 1354 KiB  
Review
Gene Therapy with Chitosan Nanoparticles: Modern Formulation Strategies for Enhancing Cancer Cell Transfection
by Varvara Antoniou, Elena A. Mourelatou, Eleftheria Galatou, Konstantinos Avgoustakis and Sophia Hatziantoniou
Pharmaceutics 2024, 16(7), 868; https://doi.org/10.3390/pharmaceutics16070868 - 27 Jun 2024
Cited by 3 | Viewed by 1251
Abstract
Gene therapy involves the introduction of exogenous genetic material into host tissues to modify gene expression or cellular properties for therapeutic purposes. Initially developed to address genetic disorders, gene therapy has expanded to encompass a wide range of conditions, notably cancer. Effective delivery [...] Read more.
Gene therapy involves the introduction of exogenous genetic material into host tissues to modify gene expression or cellular properties for therapeutic purposes. Initially developed to address genetic disorders, gene therapy has expanded to encompass a wide range of conditions, notably cancer. Effective delivery of nucleic acids into target cells relies on carriers, with non-viral systems gaining prominence due to their enhanced safety profile compared to viral vectors. Chitosan, a biopolymer, is frequently utilized to fabricate nanoparticles for various biomedical applications, particularly nucleic acid delivery, with recent emphasis on targeting cancer cells. Chitosan’s positively charged amino groups enable the formation of stable nanocomplexes with nucleic acids and facilitate interaction with cell membranes, thereby promoting cellular uptake. Despite these advantages, chitosan-based nanoparticles face challenges such as poor solubility at physiological pH, non-specificity for cancer cells, and inefficient endosomal escape, limiting their transfection efficiency. To address these limitations, researchers have focused on enhancing the functionality of chitosan nanoparticles. Strategies include improving stability, enhancing targeting specificity, increasing cellular uptake efficiency, and promoting endosomal escape. This review critically evaluates recent formulation approaches within these categories, aiming to provide insights into advancing chitosan-based gene delivery systems for improved efficacy, particularly in cancer therapy. Full article
(This article belongs to the Special Issue Cancer Gene Therapy with Non-Viral Nanocarriers, 2nd Edition)
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Figure 1
<p>Chitosan production from chitin. * indicate the points at which the polymer chain continues, denoting the repetition of the enclosed unit.</p> Full article ">Figure 2
<p>Modern strategies for improving transfection efficiency of ChNPs in cancer cells.</p> Full article ">Figure 3
<p>Chemical structure of Ch and Ch derivatives. * indicate the points at which the polymer chain continues, denoting the repetition of the enclosed unit.</p> Full article ">
10 pages, 2992 KiB  
Article
Histological Assessment of Respiratory Tract and Liver of BALB/c Mice Nebulized with Tocilizumab
by Paloma Jimena de Andres, Sergio Ferreiro, Angela Flores, Almudena Garcia and Cesar Henriquez-Camacho
Pharmaceutics 2024, 16(7), 862; https://doi.org/10.3390/pharmaceutics16070862 - 27 Jun 2024
Viewed by 893
Abstract
Pulmonary drug delivery offers a minimally invasive and efficient method for treating lung conditions, leveraging the lungs’ extensive surface area and blood flow for rapid drug absorption. Nebulized therapies aim to deliver drugs directly to the lung tissue. This study investigates the histological [...] Read more.
Pulmonary drug delivery offers a minimally invasive and efficient method for treating lung conditions, leveraging the lungs’ extensive surface area and blood flow for rapid drug absorption. Nebulized therapies aim to deliver drugs directly to the lung tissue. This study investigates the histological impact of nebulized tocilizumab—a monoclonal antibody targeting IL-6, traditionally administered intravenously for rheumatoid arthritis and severe COVID-19—on a murine model. Thirty BALB/c mice were nebulized with tocilizumab (10 mg, 5 mg, and 2.5 mg) and six controls were nebulized with saline solution. They were euthanized 48 h later, and their organs (lungs, nasal mucosa, and liver) were analyzed by a microscopic histological evaluation. The results indicate that all the mice survived the 48 h post-nebulization period without systemic compromise. The macroscopic examination showed no abnormalities, and the histopathological analysis revealed greater lung vascular changes in the control group than in the nebulized animals, which is attributable to the euthanasia with carbon dioxide. Additionally, increased alveolar macrophages were observed in the nebulized groups compared to controls. No significant histological changes were observed in the liver, indicating the safety of nebulized tocilizumab. In conclusion, these findings suggest the potential of nebulized tocilizumab for treating pulmonary inflammation, warranting further research to establish its efficacy and safety in clinical settings. Full article
(This article belongs to the Section Physical Pharmacy and Formulation)
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Figure 1
<p>Nebulization of tocilizumab (200 mg/mL) using a polymethyl methacrylate chamber (Emka technologies SAS).</p> Full article ">Figure 2
<p>Normal histological architecture in organs from mice nebulized with saline (control group). (<b>A</b>) Lung, original magnification (OM) ×10. (<b>B</b>) Liver, OM ×20.</p> Full article ">Figure 3
<p>Histopathological alterations observed in lungs of animals treated with nebulized tocilizumab. (<b>A</b>) Group 3 shows slight perivascular inflammatory cells infiltration, OM ×20. (<b>B</b>) Group 1 shows slight perivascular inflammatory cells infiltration, OM ×40. (<b>C</b>) Group 2 shows mild interstitial edema, OM ×40. (<b>D</b>) Group 1 shows mild thickening of alveolar septa, OM ×40. (<b>E</b>) Group 2 shows vacuole in the epithelium, OM ×40. (<b>F</b>) Group 3 shows macrophages in the intra-alveoli, OM ×40.</p> Full article ">Figure 4
<p>Normal liver histological architecture in animals treated with nebulized tocilizumab. (<b>A</b>) Group 1, OM ×10. (<b>B</b>) Group 2, OM ×10. (<b>C</b>) Group 3. (<b>D</b>) Control group, OM ×40.</p> Full article ">
17 pages, 5659 KiB  
Article
Antimicrobial Peptide Screening for Designing Custom Bactericidal Hydrogels
by Matthias Recktenwald, Muskanjot Kaur, Mohammed M. Benmassaoud, Aryanna Copling, Tulika Khanna, Michael Curry, Dennise Cortes, Gilbert Fleischer, Valerie J. Carabetta and Sebastián L. Vega
Pharmaceutics 2024, 16(7), 860; https://doi.org/10.3390/pharmaceutics16070860 - 27 Jun 2024
Viewed by 1352
Abstract
Staphylococcus aureus (S. aureus) is an opportunistic pathogen that lives on surfaces and skin and can cause serious infections inside the body. Antimicrobial peptides (AMPs) are part of the innate immune system and can eliminate pathogens, including bacteria and viruses, and [...] Read more.
Staphylococcus aureus (S. aureus) is an opportunistic pathogen that lives on surfaces and skin and can cause serious infections inside the body. Antimicrobial peptides (AMPs) are part of the innate immune system and can eliminate pathogens, including bacteria and viruses, and are a promising alternative to antibiotics. Although studies have reported that AMP-functionalized hydrogels can prevent bacterial adhesion and biofilm formation, AMP dosing and the combined effects of multiple AMPs are not well understood. Here, three AMPs with different antibacterial properties were synthesized and the soluble minimum inhibitory concentrations (MICs) of each AMP against methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) were determined. Hydrogels with immobilized AMPs at their MIC (DD13-RIP 27.5 µM; indolicidin 43.8 µM; P10 120 µM) were effective in preventing MRSA adhesion and biofilm formation. Checkerboard AMP screens identified synergy between indolicidin (3.1 µM) and P10 (12.5 µM) based on soluble fractional inhibitory concentration indices (FICIs) against MRSA, and hydrogels formed with these AMPs at half of their synergistic concentrations (total peptide concentration, 7.8 µM) were highly efficacious in killing MRSA. Mammalian cells cultured atop these hydrogels were highly viable, demonstrating that these AMP hydrogels are biocompatible and selectively eradicate bacteria, based on soluble checkerboard-screening data. Full article
(This article belongs to the Special Issue Recent Advances in the Prevention and Eradication Strategies for Combating Biofilm-Related Infections)
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Figure 1
<p>Combinatorial effects of soluble AMPs on MSSA strains. Representative checkerboard assay of fractional inhibitory concentrations (FIC) for (<b>A</b>) indolicidin and P10, (<b>B</b>) DD<sub>13</sub>-RIP and P10, and (<b>C</b>) indolicidin and DD<sub>13</sub>-RIP AMP pairs against MSSA. FICs were calculated for each drug (concentration/MIC) and added together for all wells where no growth was observed. The red-colored boxes indicate wells with no bacterial growth (OD<sub>600</sub> &lt; 0.1) and green-colored boxes indicate bacterial growth (OD<sub>600</sub> &gt; 0.1, <a href="#app1-pharmaceutics-16-00860" class="html-app">Figure S1</a>). The box in the bottom right corner contains no drug and serves as a growth control. Yellow-shaded boxes indicate additive interactions (FICI between 0.5–1.0) and blue boxes indicate synergistic interactions (FICI &lt; 0.5).</p> Full article ">Figure 2
<p>Combinatorial effects of soluble AMPs on MRSA strains. Representative checkerboard assay of fractional inhibitory concentrations (FIC) for (<b>A</b>) P10 and indolicidin, (<b>B</b>) P10 and DD<sub>13</sub>-RIP, and (<b>C</b>) indolicidin and DD<sub>13</sub>-RIP AMP pairs against MRSA. FICs were calculated for each drug (concentration/MIC) and added together for all wells where no growth was observed. The red-colored boxes indicate wells with no bacterial growth (OD<sub>600</sub> &lt; 0.1) and green-colored boxes indicate bacterial growth (OD<sub>600</sub> &gt; 0.1, <a href="#app1-pharmaceutics-16-00860" class="html-app">Figure S2</a>). The box in the bottom right corner contains no drug and serves as a growth control. Yellow-shaded boxes indicate additive interactions (FICI between 0.5–1.0) and blue boxes indicate synergistic interactions (FICI &lt; 0.5).</p> Full article ">Figure 3
<p>Examination of MRSA biofilm formation on different AMP-loaded hydrogels. Qualitative images of representative AMP hydrogels at 0.5×, 1×, and 2× MIC for tethered (<b>A</b>) DD<sub>13</sub>-RIP, (<b>B</b>) indolicidin, and (<b>C</b>) P10 AMPs. Scale bars: 1 mm. Quantitative viability analysis of MRSA on (<b>D</b>) DD<sub>13</sub>-RIP, (<b>E</b>) indolicidin, and (<b>F</b>) P10 AMP hydrogels. All the groups were normalized to the results of a control hydrogel group with no AMP. Significance (*** <span class="html-italic">p</span> &lt; 0.001) in viability was established by comparing each test group to a control hydrogel group with no AMP (0×).</p> Full article ">Figure 4
<p>Synergistic effects of P10 and indolicidin on viability of MRSA atop different AMP-loaded hydrogels. (<b>A</b>) Qualitative analysis showing representative hydrogels functionalized with different fractions of the synergistic concentration for indolicidin and P10 (3.125 μM and 12.5 μM, respectively) and stained for live (<b>green</b>) and dead (<b>red</b>) cells. Scale bar: 1 mm. (<b>B</b>) Quantitative analysis of synergistic AMP hydrogel on biofilm viability. Significance (*** <span class="html-italic">p</span> &lt; 0.001) in viability was established by comparing the test group to a control hydrogel group with no AMP (0×).</p> Full article ">Figure 5
<p>Investigation of MRSA cells interacting with AMP-hydrogels. Representative high-resolution (60× magnification) images of hydrogels tethered with (<b>A</b>) no AMP, (<b>B</b>) DD<sub>13</sub>RIP, (<b>C</b>) indolicidin, and (<b>D</b>) P10 at their soluble MIC, and (<b>E</b>) indolicidin and P10 at their synergistic soluble MIC. Scale bars: 100 µm. Quantitative spatial viability analysis of MRSA as a function of hydrogel depth of hydrogels tethered with (<b>F</b>) no AMP, (<b>G</b>) DD<sub>13</sub>RIP, (<b>H</b>) indolicidin, and (<b>I</b>) P10 at their soluble MIC, and (<b>J</b>) indolicidin and P10 at their synergistic soluble MIC.</p> Full article ">Figure 6
<p>Mammalian MSC viability on AMP-loaded hydrogels. (<b>A</b>) Representative live–dead stain of MSCs seeded on AMP hydrogels at 2× MIC for each group. Scale bar: 1 mm. (<b>B</b>) Quantification of the viability of mammalian cells seeded on top of each AMP hydrogel. There were no significant differences in viability observed.</p> Full article ">Figure 7
<p>Schematic of the experimental design. (<b>A</b>) Diagram of 96-well plate array used for high-throughput screening of AMPs at different concentrations to determine additive or synergistic concentrations against MRSA and MSSA. Representative inhibitory (IC<sub>100</sub>), additive, and synergistic combinations are displayed. (<b>B</b>) Chemical components used to synthesize AMP hydrogels. (<b>C</b>) MRSA inoculation, incubation, staining, and imaging process diagram.</p> Full article ">
23 pages, 4798 KiB  
Article
Imiquimod-Loaded Nanosystem for Treatment Human Papillomavirus-Induced Lesions
by Izamara Maocha, Beatriz Rosado, Jéssica Lopes-Nunes, Melanie Lopes, Joana Rolo, Bruno Pires, Eugénia Gallardo, Ana Palmeira-de-Oliveira, José Martinez-de-Oliveira, Rita Palmeira de Oliveira, Rui Medeiros and Carla Cruz
Pharmaceutics 2024, 16(7), 864; https://doi.org/10.3390/pharmaceutics16070864 - 27 Jun 2024
Cited by 2 | Viewed by 1330
Abstract
Human papillomavirus (HPV)-associated cervical cancer is the most common cancer among women worldwide. The treatment options are strongly related to increased infertility in women. Imiquimod (IQ) is an imidazoquinoline, which has proven antiviral effects against persistent HPV infection by activating immune cells via [...] Read more.
Human papillomavirus (HPV)-associated cervical cancer is the most common cancer among women worldwide. The treatment options are strongly related to increased infertility in women. Imiquimod (IQ) is an imidazoquinoline, which has proven antiviral effects against persistent HPV infection by activating immune cells via Toll-like receptors 7/8 when formulated in carriers, like nanogels, for topical use. An effective alternative to conventional therapies is the nanoparticle drug delivery system. We studied lipidic nanoparticles with IQ (Lipo IQ) and functionalized them with a DNA aptamer, AT11 (Lipo IQ AT11), to improve the selectivity for cervical cancer cells combined with the efficacy of essential oils. The formulations showed that the physicochemical properties are adequate for vaginal drug delivery and have antimicrobial activity at higher concentrations (with MIC50 starting from 0.625%). The final formulations exhibited cytotoxicity in cancer cells, enhanced by essential oils without affecting healthy cells, resulting in less than 10% cell viability in HeLa cells and over 60% in NHDF cells. Essential oils potentiate Lipo IQ’s effectiveness, while AT11 increases the selectivity for cervical cancer cells. As suggested by the results of the permeation assay, the formulations were internalized by the cancer cells. Overall, the obtained results suggested that the synergistic effect of the essential oils and the nanosystem potentiate the cytotoxic effect of Lipo IQ and that Lipo IQ AT11 promotes selectivity towards cancer cells. Full article
(This article belongs to the Special Issue The 15th Anniversary of Pharmaceutics—Aptamers as Novel Therapeutics)
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Figure 1
<p>Schematic representation displaying the structure of Lipo IQ AT11.</p> Full article ">Figure 2
<p>Cumulative release of IQ from liposomes in PBS at pH 7.4 (<span class="html-italic">n</span> = 3).</p> Full article ">Figure 3
<p>(<b>A</b>) Relevant and (<b>B</b>) absolute pH buffering capacity of formulations with dispersion in 0.9% NaCl (NaCl columns in gray) and vaginal fluid simulant (VFS columns in black). Titration was performed using HCl (1M) since all formulations had pH &gt; 5 in the dispersion experiments performed in NaCl, and in VFS, NaOH (1M) was used since the pH changed to less than 5 until pH &gt; 9.</p> Full article ">Figure 4
<p>Percentage of relative cell viability of all formulations after 24 h of incubation at 37 °C in (<b>A</b>) HeLa, (<b>B</b>) NHDF, (<b>C</b>) SiHa, and (<b>D</b>) CaSki cell lines. The results correspond to the mean and standard error of three measurements. *, **, and *** represent statistically different values from the control (two-way ANOVA, * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.001, *** <span class="html-italic">p</span> &lt; 0.0001, Dunnett’s multiple comparisons test).</p> Full article ">Figure 4 Cont.
<p>Percentage of relative cell viability of all formulations after 24 h of incubation at 37 °C in (<b>A</b>) HeLa, (<b>B</b>) NHDF, (<b>C</b>) SiHa, and (<b>D</b>) CaSki cell lines. The results correspond to the mean and standard error of three measurements. *, **, and *** represent statistically different values from the control (two-way ANOVA, * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.001, *** <span class="html-italic">p</span> &lt; 0.0001, Dunnett’s multiple comparisons test).</p> Full article ">Figure 5
<p>Comparative dose–response relationship of IQ (BF IQ AT11) in (<b>A</b>) HeLa and (<b>B</b>) NHDF cells after 24 h of incubation at 37 °C.</p> Full article ">Figure 6
<p>Confocal fluorescence images of (<b>A</b>) NHDF, (<b>B</b>) HeLa, (<b>C</b>) SiHa, and (<b>D</b>) CaSki cell lines incubated with IQ liposomes (Lipo IQ AT11), thyme formulation with liposome IQ (TEO IQ AT11), and oregano formulation with liposome IQ (OEO IQ AT11) for 24 h at 37 °C. Cell nuclei are stained with Hoechst 33342 (blue), and Cy5-AT11 is shown in red.</p> Full article ">
20 pages, 2426 KiB  
Article
Evaluation of Polyvinyl Alcohol as Binder during Continuous Twin Screw Wet Granulation
by Phaedra Denduyver, Gudrun Birk, Alessandra Ambruosi, Chris Vervaet and Valérie Vanhoorne
Pharmaceutics 2024, 16(7), 854; https://doi.org/10.3390/pharmaceutics16070854 - 25 Jun 2024
Viewed by 1926
Abstract
Binder selection is a crucial step in continuous twin-screw wet granulation (TSWG), as the material experiences a much shorter residence time (2–40 s) in the granulator barrel compared to batch-wise granulation processes. Polyvinyl alcohol (PVA) 4-88 was identified as an effective binder during [...] Read more.
Binder selection is a crucial step in continuous twin-screw wet granulation (TSWG), as the material experiences a much shorter residence time (2–40 s) in the granulator barrel compared to batch-wise granulation processes. Polyvinyl alcohol (PVA) 4-88 was identified as an effective binder during TSWG, but the potential of other PVA grades—differing in polymerization and hydrolysis degree—has not yet been studied. Therefore, the aim of the current study was to evaluate the potential of different PVA grades as a binder during TSWG. The breakage and drying behavior during the fluidized bed drying of drug-loaded granules containing the PVA grades was also studied. Three PVA grades (4-88, 18-88, and 40-88) were characterized and their attributes were compared to previously investigated binders by Vandevivere et al. through principal component analysis. Three binder clusters could be distinguished according to their attributes, whereby each cluster contained a PVA grade and a previously investigated binder. PVA 4-88 was the most effective binder of the PVA grades for both a good water-soluble and water-insoluble formulation. This could be attributed to its high total surface energy, low viscosity, good wettability of hydrophilic and hydrophobic surfaces, and good wettability by water of the binder. Compared to the previously investigated binders, all PVA grades were more effective in the water-insoluble formulation, as they yielded strong granules (friability below 30%) at lower L/S-ratios. This was linked to the high dispersive surface energy of the high-energy sites on the surface of PVA grades and their low surface tension. During fluidized bed drying, PVA grades proved suitable binders, as the acetaminophen (APAP) granules were dried within a short time due to the low L/S-ratio, at which high-quality granules could be produced. In addition, no attrition occurred, and strong tablets were obtained. Based on this study, PVA could be the preferred binder during twin screw granulation due to its high binder effectiveness at a low L/S-ratio, allowing efficient downstream processing. However, process robustness must be controlled by the included excipients, as PVA grades are operating in a narrow L/S-ratio range. Full article
(This article belongs to the Special Issue Impact of Raw Material Properties on Solid Dosage Form Processes)
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Graphical abstract

Graphical abstract
Full article ">Figure 1
<p>Screw configuration with material flow from left to right. Kneading zones consist of 6 kneading elements (length-to-diameter (L/D) 1/4) in a stagger angle of 60°. Granulation liquid was added just before the first kneading zone.</p> Full article ">Figure 2
<p>The score (<b>a</b>) and loading (<b>b</b>) scatter plot of PC 1 versus PC 2. The different colors refer to the binder clusters, which have comparable binder attributes. Cluster 1 (red) contains PVA 4-88 and SOS CO01; cluster 2 (yellow) has PVA 18-88 and PVP K90; cluster 3 (blue) has PVA 40-88 and HPMC E15; and cluster 4 (green) has maltodextrin 6, HP pea starch, PVP K12, and PVP K30.</p> Full article ">Figure 3
<p>The dispersive (<b>top</b>), specific (<b>middle</b>) and total (<b>bottom</b>) surface energy in the function of the fractional coverage for cluster 1, 2, and 3 binders. The specific and total surface energies of HPMC E15 could not be measured due to weak solvent interactions.</p> Full article ">Figure 4
<p>SME values in function of the L/S-ratio for each binder in mannitol- and DCP-based formulations. L/S-ratios depicted are based on the L/S-ratio range from <a href="#pharmaceutics-16-00854-t001" class="html-table">Table 1</a>.</p> Full article ">Figure 5
<p>Friability in function of the L/S-ratio for each binder in the mannitol- and DCP-based formulations. The L/S-ratios depicted are based on the L/S-ratio range from <a href="#pharmaceutics-16-00854-t001" class="html-table">Table 1</a>.</p> Full article ">Figure 6
<p>Moisture content of the granules in function of drying time. Granules were produced on the L/S-ratio mentioned in <a href="#pharmaceutics-16-00854-t003" class="html-table">Table 3</a>.</p> Full article ">Figure 7
<p>Breakage behavior of APAP granules with PVA grades. Solid line represents the particle size distribution of tray-dried granules (TDG). Dashed and dotted lines represent the particle size distribution after fluid bed drying at first and second drying times, respectively.</p> Full article ">Figure 8
<p>Tensile strength in function of main compaction pressure for studied binders in APAP formulation.</p> Full article ">
21 pages, 3630 KiB  
Article
Investigation of the Electrokinetic Potential of Granules and Optimization of the Pelletization Method Using the Quality by Design Approach
by Azza A. K. Mahmoud, Alharith A. A. Hassan, Dorina Gabriella Dobó, Krisztina Ludasi, László Janovák, Géza Regdon, Jr., Ildikó Csóka and Katalin Kristó
Pharmaceutics 2024, 16(7), 848; https://doi.org/10.3390/pharmaceutics16070848 - 22 Jun 2024
Cited by 1 | Viewed by 3927
Abstract
The preparation of pellets using a high-shear granulator in a rapid single-step is considered a good economic alternative to the extrusion spheronization process. As process parameters and material attributes greatly affect pellet qualities, successful process optimization plays a vital role in producing pellet [...] Read more.
The preparation of pellets using a high-shear granulator in a rapid single-step is considered a good economic alternative to the extrusion spheronization process. As process parameters and material attributes greatly affect pellet qualities, successful process optimization plays a vital role in producing pellet dosage forms with the required critical quality attributes. This study was aimed at the development and optimization of the pelletization technique with the Pro-CepT granulator. According to the quality by design (QbD) and screening design results, chopper speed, the volume of the granulating liquid, binder amount, and impeller speed were selected as the highest risk variables for a two-level full factorial design and central composite design, which were applied to the formula of microcrystalline cellulose, mannitol, and with a binding aqueous polyvinylpyrrolidone solution. The design space was estimated based on physical response results, including the total yield of the required size, hardness, and aspect ratio. The optimized point was tested with two different types of active ingredients. Amlodipine and hydrochlorothiazide were selected as model drugs and were loaded into an optimized formulation. The kinetics of the release of the active agent was examined and found that the results show a correlation with the electrokinetic potential because amlodipine besylate can be adsorbed on the surface of the MCC, while hydrochlorothiazide less so; therefore, in this case, the release of the active agent increases. The research results revealed no significant differences between plain and model drug pellets, except for hydrochlorothiazide yield percent, in addition to acceptable content uniformity and dissolution enhancement. Full article
(This article belongs to the Section Pharmaceutical Technology, Manufacturing and Devices)
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<p>Ishikawa diagram for determining critical factors for direct pelletization by using Pro-CepT granulator (definition and classification of all expected factors).</p> Full article ">Figure 2
<p>Matrix diagram, (<b>A</b>). Effect of quality target profile (QTTP) with critical quality attributes (CQAs), (<b>B</b>). Critical quality attributes (CQAs) with critical material attributes (CMAs)/critical process parameters (CPPs) using three grades: red color—high (H), yellow color—medium (M), and green color—low (L) for direct pelletization techniques.</p> Full article ">Figure 3
<p>Pareto chart effect, (<b>a</b>). Quality target profile (QTTP) with critical quality attributes (CQAs), (<b>b</b>). Critical quality attributes (CQAs) with critical material attributes (CMAs)/critical process parameters (CPPs) using two grades: red color—high (H), and green color—low (L) for direct pelletization techniques.</p> Full article ">Figure 4
<p>Three-dimensional response surface plots showing the effects of impeller speed and chopper speed at constant liquid volume (55 mL) and binder amount (2 g) on (<b>A</b>) aspect ratio, (<b>B</b>) yield percent, and (<b>C</b>) hardness.</p> Full article ">Figure 5
<p>Design space for direct pelletization with Pro-CepT granulator.</p> Full article ">Figure 6
<p><span class="html-italic">T</span>-tests of the physical properties of drug-free pellets: (<b>A1</b>–<b>A3</b>) Amlodipine besylate pellets; (<b>B1</b>–<b>B3</b>) Hydrochlorothiazide pellets.</p> Full article ">Figure 7
<p>Dissolution release profile for amlodipine besylate pellets and hydrochlorothiazide pellets.</p> Full article ">Figure 8
<p>Results of electrokinetic potential measurements (SurPass measurements).</p> Full article ">Figure 9
<p>FT-IR spectroscopy of amlodipine besylate pellets, hydrochlorothiazide hydrochloride pellets, and excipients (MCC, PVP, and mannitol).</p> Full article ">
46 pages, 3156 KiB  
Review
An Overview on the Physiopathology of the Blood–Brain Barrier and the Lipid-Based Nanocarriers for Central Nervous System Delivery
by Francesca Susa, Silvia Arpicco, Candido Fabrizio Pirri and Tania Limongi
Pharmaceutics 2024, 16(7), 849; https://doi.org/10.3390/pharmaceutics16070849 - 22 Jun 2024
Viewed by 2250
Abstract
The state of well-being and health of our body is regulated by the fine osmotic and biochemical balance established between the cells of the different tissues, organs, and systems. Specific districts of the human body are defined, kept in the correct state of [...] Read more.
The state of well-being and health of our body is regulated by the fine osmotic and biochemical balance established between the cells of the different tissues, organs, and systems. Specific districts of the human body are defined, kept in the correct state of functioning, and, therefore, protected from exogenous or endogenous insults of both mechanical, physical, and biological nature by the presence of different barrier systems. In addition to the placental barrier, which even acts as a linker between two different organisms, the mother and the fetus, all human body barriers, including the blood–brain barrier (BBB), blood–retinal barrier, blood–nerve barrier, blood–lymph barrier, and blood–cerebrospinal fluid barrier, operate to maintain the physiological homeostasis within tissues and organs. From a pharmaceutical point of view, the most challenging is undoubtedly the BBB, since its presence notably complicates the treatment of brain disorders. BBB action can impair the delivery of chemical drugs and biopharmaceuticals into the brain, reducing their therapeutic efficacy and/or increasing their unwanted bioaccumulation in the surrounding healthy tissues. Recent nanotechnological innovation provides advanced biomaterials and ad hoc customized engineering and functionalization methods able to assist in brain-targeted drug delivery. In this context, lipid nanocarriers, including both synthetic (liposomes, solid lipid nanoparticles, nanoemulsions, nanostructured lipid carriers, niosomes, proniosomes, and cubosomes) and cell-derived ones (extracellular vesicles and cell membrane-derived nanocarriers), are considered one of the most successful brain delivery systems due to their reasonable biocompatibility and ability to cross the BBB. This review aims to provide a complete and up-to-date point of view on the efficacy of the most varied lipid carriers, whether FDA-approved, involved in clinical trials, or used in in vitro or in vivo studies, for the treatment of inflammatory, cancerous, or infectious brain diseases. Full article
(This article belongs to the Special Issue Nanotechnology-Based Pharmaceutical Treatments)
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<p>Scheme of the BBB neurovascular unit, modified from [<a href="#B10-pharmaceutics-16-00849" class="html-bibr">10</a>,<a href="#B11-pharmaceutics-16-00849" class="html-bibr">11</a>].</p> Full article ">Figure 2
<p>The different molecules’ transport mechanisms in the BBB are divided into paracellular and transcellular.</p> Full article ">Figure 3
<p>Representation of (<b>A</b>) liposomes’ structure made of a lipid bilayer encapsulating an aqueous core and (<b>B</b>) their possible load and functionalization.</p> Full article ">Figure 4
<p>Structure of other synthetic lipid-based delivery systems.</p> Full article ">Figure 5
<p>Representation of extracellular vesicles and their production and release mechanisms.</p> Full article ">
31 pages, 7459 KiB  
Article
Polymeric Microneedles Enhance Transdermal Delivery of Therapeutics
by Hiep X. Nguyen, Thomas Kipping and Ajay K. Banga
Pharmaceutics 2024, 16(7), 845; https://doi.org/10.3390/pharmaceutics16070845 - 22 Jun 2024
Cited by 2 | Viewed by 2184
Abstract
This research presents the efficacy of polymeric microneedles in improving the transdermal permeation of methotrexate across human skin. These microneedles were fabricated from PLGA Expansorb® 50-2A and 50-8A and subjected to comprehensive characterization via scanning electron microscopy, Fourier-transform infrared spectroscopy, and mechanical [...] Read more.
This research presents the efficacy of polymeric microneedles in improving the transdermal permeation of methotrexate across human skin. These microneedles were fabricated from PLGA Expansorb® 50-2A and 50-8A and subjected to comprehensive characterization via scanning electron microscopy, Fourier-transform infrared spectroscopy, and mechanical analysis. We developed and assessed a methotrexate hydrogel for physicochemical and rheological properties. Dye binding, histological examinations, and assessments of skin integrity demonstrated the effective microporation of the skin by PLGA microneedles. We measured the dimensions of microchannels in the skin using scanning electron microscopy, pore uniformity analysis, and confocal microscopy. The skin permeation and disposition of methotrexate were researched in vitro. PLGA 50-8A microneedles appeared significantly longer, sharper, and more mechanically uniform than PLGA 50-2A needles. PLGA 50-8A needles generated substantially more microchannels, as well as deeper, larger, and more uniform channels in the skin than PLGA 50-2A needles. Microneedle insertion substantially reduced skin electrical resistance, accompanied by an elevation in transepidermal water loss values. PLGA 50-8A microneedle treatment provided a significantly higher cumulative delivery, flux, diffusion coefficient, permeability coefficient, and predicted steady-state plasma concentration; however, there was a shorter lag time than for PLGA 50-2A needles, base-treated, and untreated groups (p < 0.05). Conclusively, skin microporation using polymeric microneedles significantly improved the transdermal delivery of methotrexate. Full article
(This article belongs to the Special Issue Microarray Patches for Transdermal Drug Delivery)
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<p>Characterization of methotrexate hydrogel rheology: (<b>a</b>) amplitude sweep, (<b>b</b>) frequency sweep, (<b>c</b>) flow curve, (<b>d</b>) yield stress, (<b>e</b>) temperature-dependent viscosity, (<b>f</b>) thixotropic behavior.</p> Full article ">Figure 2
<p>SEM photographs of microneedles: (<b>a</b>,<b>b</b>) unused and (<b>c</b>,<b>d</b>) used PLGA 50-2A microneedles; (<b>e</b>,<b>f</b>) unused and (<b>g</b>,<b>h</b>) used PLGA 50-8A microneedles. Scale bar: 200 µm.</p> Full article ">Figure 3
<p>SEM photographs of skin tissue: (<b>a</b>) untreated, intact skin, (<b>b</b>) skin porated with PLGA 50-2A microneedles, (<b>c</b>) skin porated with PLGA 50-8A microneedles. Scale bar: 300 µm.</p> Full article ">Figure 4
<p>FTIR spectra: (<b>a</b>) PLGA 50-2A granules and microneedles, (<b>b</b>) PLGA 50-8A granules and microneedles.</p> Full article ">Figure 5
<p>Microscopic photographs of Parafilm M<sup>®</sup> layers porated with PLGA microneedles: (<b>a</b>,<b>b</b>) first layer, (<b>c</b>,<b>d</b>) second layer for PLGA 50-2A microneedles; (<b>e</b>,<b>f</b>) first layer, (<b>g</b>,<b>h</b>) second layer for PLGA 50-8A microneedles.</p> Full article ">Figure 6
<p>Microscopic photographs of microneedles and skin: (<b>a</b>) base substrate, (<b>b</b>) PLGA 50-2A microneedles, (<b>c</b>) PLGA 50-8A microneedles, (<b>d</b>) untreated human skin, skin porated with PLGA 50-2A microneedles employing (<b>e</b>) a spring-loaded applicator, (<b>g</b>) thumb pressure, and skin porated with PLGA 50-8A microneedles employing (<b>f</b>) a spring-loaded applicator, (<b>h</b>) thumb pressure.</p> Full article ">Figure 7
<p>Histological examination of skin tissue: (<b>a</b>,<b>d</b>) untreated, (<b>b</b>,<b>e</b>) treated with PLGA 50-2A microneedles, and (<b>c</b>,<b>f</b>) treated with PLGA 50-8A microneedles. (<b>a</b>–<b>c</b>) Unstained tissues, (<b>d</b>–<b>f</b>) skin tissues stained with hematoxylin and eosin.</p> Full article ">Figure 8
<p>Fluorescent photographs with Pore Permeability Index and distribution of fluorescence intensity of skin: (<b>a</b>–<b>c</b>) skin porated with PLGA 50-2A microneedles, (<b>d</b>–<b>f</b>) skin porated with PLGA 50-8A microneedles.</p> Full article ">Figure 9
<p>Confocal microscopic photographs of skin: (<b>a</b>) treated with PLGA 50-2A microneedles, (<b>b</b>) treated with PLGA 50-8A microneedles.</p> Full article ">Figure 10
<p>Confocal z-stack of microchannel generated by the insertion of PLGA 50-8A microneedles.</p> Full article ">Figure 11
<p>Transepidermal water loss and skin electrical resistance values of untreated (control), base-treated, and PLGA microneedle-porated skin (* denotes statistically significant difference from the untreated and base-treated groups, mean ± SD, n = 4, <span class="html-italic">p</span> &lt; 0.05).</p> Full article ">Figure 12
<p>In vitro transdermal permeation of methotrexate: (<b>a</b>) average cumulative quantity and (<b>b</b>) transdermal flux of methotrexate delivered across untreated, base-treated, and PLGA microneedle-porated skin (* denotes statistically significant difference from the untreated and base-treated groups, mean ± SD, n = 4, <span class="html-italic">p</span> &lt; 0.05).</p> Full article ">Figure 13
<p>Quantities of methotrexate in skin tissue: untreated, base-treated, and PLGA microneedle-porated skin (* denotes statistically significant difference from the untreated and base-treated groups, mean ± SD, n = 4, <span class="html-italic">p</span> &lt; 0.05).</p> Full article ">
13 pages, 3238 KiB  
Article
Derivatization of Hyaluronan to Target Neuroblastoma and Neuroglioma Expressing CD44
by Giau Van Vo, Kummara Madhusudana Rao, Ildoo Chung, Chang-Sik Ha, Seong Soo A. An and Yang H. Yun
Pharmaceutics 2024, 16(6), 836; https://doi.org/10.3390/pharmaceutics16060836 - 20 Jun 2024
Cited by 1 | Viewed by 1380
Abstract
Therapeutics for actively targeting over-expressed receptors are of great interest because the majority of diseased tissues originate from normal cells and do not possess a unique receptor from which they can be differentiated. One such receptor is CD44, which has been shown to [...] Read more.
Therapeutics for actively targeting over-expressed receptors are of great interest because the majority of diseased tissues originate from normal cells and do not possess a unique receptor from which they can be differentiated. One such receptor is CD44, which has been shown to be highly overexpressed in many breast cancers and other types of cancer cells. While CD44 has been documented to express low levels in normal adult neurons, astrocytes, and microglia, this receptor may be overexpressed by neuroblastoma and neuroglioma. If differential expression exists between normal and cancerous cells, hyaluronan (HA) could be a useful carrier that targets carcinomas. Thus, HA was conjugated with resveratrol (HA-R), and its efficacy was tested on cortical–neuroblastoma hybrid, neuroblastoma, and neuroglioma cells. Confocal and flow cytometry showed these cells express CD44 and are able to bind and uptake HA-R. The toxicity of HA-R correlated well with CD44 expression in this study. Therefore, conjugating resveratrol and other chemotherapeutics to HA could minimize the side effects for normal cells within the brain and nervous system and could be a viable strategy for developing targeted therapies. Full article
(This article belongs to the Special Issue Carbohydrate-Based Carriers for Drug Delivery)
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<p>TLC of (1) resveratrol, (2) HA and resveratrol mixture (unconjugated), (3) HA-R, and (4) HA. Migration of resveratrol could be observed for free resveratrol in samples 1 and 2. HA, which did not migrate due to its molecular weight, was observed for groups 2, 3, and 4. (<b>a</b>) TLC was observed using UV light. (<b>b</b>) TLC showed hyaluronan and resveratrol after the application of heat.</p> Full article ">Figure 2
<p>NMR analysis of (<b>a</b>) resveratrol and (<b>b</b>) HA-R. The peaks for the resveratrol group on HA-R were attenuated but present.</p> Full article ">Figure 3
<p>UV spectral analysis of HA-R at (<b>a</b>) various concentrations and (<b>b</b>) the standard curve. The peak wavelength was 305 nm.</p> Full article ">Figure 4
<p>Immunofluorescence and flow cytometry assays showed positive CD44 (green), actin (red), and nuclear (blue) staining for (<b>a</b>,<b>b</b>) cortical–neuroblastoma hybrid (A1G11), (<b>c</b>,<b>d</b>) neuroblastoma (SH-SY5Y), (<b>e</b>,<b>f</b>) brain neuroglioma (H4), (<b>g</b>,<b>h</b>) human embryonic kidney (HEK293), and (<b>i</b>,<b>j</b>) fibroblast (N9). The scale bar is 10 µm. Flow cytometry data (<b>b</b>,<b>d</b>,<b>f</b>,<b>h</b>,<b>j</b>) shows positive CD 44 expression (green) and control (black).</p> Full article ">Figure 4 Cont.
<p>Immunofluorescence and flow cytometry assays showed positive CD44 (green), actin (red), and nuclear (blue) staining for (<b>a</b>,<b>b</b>) cortical–neuroblastoma hybrid (A1G11), (<b>c</b>,<b>d</b>) neuroblastoma (SH-SY5Y), (<b>e</b>,<b>f</b>) brain neuroglioma (H4), (<b>g</b>,<b>h</b>) human embryonic kidney (HEK293), and (<b>i</b>,<b>j</b>) fibroblast (N9). The scale bar is 10 µm. Flow cytometry data (<b>b</b>,<b>d</b>,<b>f</b>,<b>h</b>,<b>j</b>) shows positive CD 44 expression (green) and control (black).</p> Full article ">Figure 5
<p>The uptake of HA-FITC (green), lysosomes (red), and nucleus (blue) (<b>a</b>) cortical–neuroblastoma hybrid (A1G11), (<b>b</b>) neuroblastoma (SH-SY5Y), (<b>c</b>) brain neuroglioma (H4), (<b>d</b>) human embryonic kidney (HEK293), and (<b>e</b>) fibroblast (N9). The scale bar represents 10 nm.</p> Full article ">Figure 6
<p>Cytotoxicity of (<span style="color:red">■</span>) HA-R, (<span style="color:#00B050">▲</span>) resveratrol, and (<span style="color:#2F5496">■</span>) HA for (<b>a</b>) cortical–neuroblastoma hybrid (A1G11), (<b>b</b>) neuroblastoma (SH-SY5Y), (<b>c</b>) brain neuroglioma (H4), (<b>d</b>) human embryonic kidney (HEK293), and (<b>e</b>) fibroblast (N9). IC<sub>50</sub> values (<b>f</b>) for cells exposed to (<span style="color:red">■</span>) HA-R, (<span style="color:#00B050">■</span>) resveratrol, and (<span style="color:#2F5496">■</span>) HA. The groups connected by brackets lack statistical significance, <span class="html-italic">p</span> &gt; 0.05. * Curve fits do not converge onto IC<sub>50</sub> values.</p> Full article ">Figure 6 Cont.
<p>Cytotoxicity of (<span style="color:red">■</span>) HA-R, (<span style="color:#00B050">▲</span>) resveratrol, and (<span style="color:#2F5496">■</span>) HA for (<b>a</b>) cortical–neuroblastoma hybrid (A1G11), (<b>b</b>) neuroblastoma (SH-SY5Y), (<b>c</b>) brain neuroglioma (H4), (<b>d</b>) human embryonic kidney (HEK293), and (<b>e</b>) fibroblast (N9). IC<sub>50</sub> values (<b>f</b>) for cells exposed to (<span style="color:red">■</span>) HA-R, (<span style="color:#00B050">■</span>) resveratrol, and (<span style="color:#2F5496">■</span>) HA. The groups connected by brackets lack statistical significance, <span class="html-italic">p</span> &gt; 0.05. * Curve fits do not converge onto IC<sub>50</sub> values.</p> Full article ">
10 pages, 1763 KiB  
Article
The Distinctive Role of Gluconic Acid in Retarding Percutaneous Drug Permeation: Formulation of Lidocaine-Loaded Chitosan Nanoparticles
by Amnon C. Sintov
Pharmaceutics 2024, 16(6), 831; https://doi.org/10.3390/pharmaceutics16060831 - 19 Jun 2024
Viewed by 860
Abstract
The objective of the present investigation was to evidence the skin retardation phenomenon of lidocaine by gluconic acid as an inactive ingredient involved in citrate-crosslinking chitosan nanoparticles. Lidocaine hydrochloride was loaded in nanoparticles based on chitosan, fabricated by using a water-in-oil microemulsion as [...] Read more.
The objective of the present investigation was to evidence the skin retardation phenomenon of lidocaine by gluconic acid as an inactive ingredient involved in citrate-crosslinking chitosan nanoparticles. Lidocaine hydrochloride was loaded in nanoparticles based on chitosan, fabricated by using a water-in-oil microemulsion as a template and citric acid as an ionic cross-linker. Gluconic acid (pentahydroxy hexanoic acid) was added during the fabrication and compared with caproic acid, a non-hydroxy hexanoic acid. The chitosan nanoparticulate systems were characterized for mean particle size, particle size distribution, and zeta potential. The pentahydroxy hexanoic acid decreased the zeta potential to a significantly lower value than those obtained from both plain citrate and citrate–hexanoic acid formulations. The relatively lower value implies that gluconate ions are partly attached to the nanoparticle’s surface and mask its positively charged groups. It was also noted that the in vitro percutaneous permeation flux of lidocaine significantly decreased when gluconate-containing chitosan nanoparticles were applied, i.e., 6.1 ± 1.5 μg‧cm−2‧h−1 without gluconic acid to 3.4 ± 2.3 μg‧cm−2‧h−1 with gluconic acid. According to this result, it is suggested that gluconate ions played a role in retarding drug permeation through the skin, probably by calcium chelation in the stratum granulosum, which in turn stimulated lamellar body secretion, lipid synthesis, and intracellular release of Ca2+ from the endoplasmic reticulum. Full article
(This article belongs to the Special Issue Drug Delivery of Natural Active Principles: Focus on Topical and Oral Applications, 2nd Edition)
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<p>Penetration and permeation pathways of drugs into and through the skin, and the roles of CPEs (<span class="html-italic">chemical permeation enhancers</span>), CPRs (<span class="html-italic">cutaneous penetration retardants</span>, and PPRs (<span class="html-italic">percutaneous permeation retardants</span>) as modifiers of cutaneous and percutaneous drug delivery.</p> Full article ">Figure 2
<p>Schematic representation of citrate-chitosan nanoparticles. Due to their pentahydroxy groups, gluconate molecules partially bind onto the polysaccharidic particle surface via hydrogen bonds and not through the ionic carboxylate groups only.</p> Full article ">Figure 3
<p>Percutaneous permeation kinetics of lidocaine applied in citrate-crosslinked CHS-NPs and in citrate-crosslinked CHS-NPs/gluconic acid combination.</p> Full article ">Figure 4
<p>Percutaneous permeation kinetics of lidocaine applied in cationic guar NPs and in cationic guar NPs/gluconic acid combination.</p> Full article ">Figure 5
<p>Proposed mechanism of reinforcing the skin barrier without its pre-impairment, by the citrate-crosslinked CHS-NPs/gluconate combination.</p> Full article ">
23 pages, 7648 KiB  
Article
Multistage Nanocarrier Based on an Oil Core–Graphene Oxide Shell
by Immacolata Tufano, Raffaele Vecchione, Valeria Panzetta, Edmondo Battista, Costantino Casale, Giorgia Imparato and Paolo Antonio Netti
Pharmaceutics 2024, 16(6), 827; https://doi.org/10.3390/pharmaceutics16060827 - 18 Jun 2024
Viewed by 1249
Abstract
Potent synthetic drugs, as well as biomolecules extracted from plants, have been investigated for their selectivity toward cancer cells. The main limitation in cancer treatment is the ability to bring such molecules within each single cancer cell, which requires accumulation in the peritumoral [...] Read more.
Potent synthetic drugs, as well as biomolecules extracted from plants, have been investigated for their selectivity toward cancer cells. The main limitation in cancer treatment is the ability to bring such molecules within each single cancer cell, which requires accumulation in the peritumoral region followed by homogeneous spreading within the entire tissue. In the last decades, nanotechnology has emerged as a powerful tool due to its ability to protect the drug during blood circulation and allow enhanced accumulation around the leaky regions of the tumor vasculature. However, the ideal size for accumulation of around 100 nm is too large for effective penetration into the dense collagen matrix. Therefore, we propose a multistage system based on graphene oxide nanosheet-based quantum dots (GOQDs) with dimensions that are 12 nm, functionalized with hyaluronic acid (GOQDs-HA), and deposited using the layer-by-layer technique onto an oil-in-water nanoemulsion (O/W NE) template that is around 100 nm in size, previously stabilized by a biodegradable polymer, chitosan. The choice of a biodegradable core for the nanocarrier is to degrade once inside the tumor, thus promoting the release of smaller compounds, GOQDs-HA, carrying the adsorbed anticancer compound, which in this work is represented by curcumin as a model bioactive anticancer molecule. Additionally, modification with HA aims to promote active targeting of stromal and cancer cells. Cell uptake experiments and preliminary penetration experiments in three-dimensional microtissues were performed to assess the proposed multistage nanocarrier. Full article
(This article belongs to the Special Issue Smart Nanocarriers for Drug Delivery in Cancer Therapy)
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<p>GOQDs-HA morphology and size evaluation, (<b>a</b>) cryo-TEM image, (<b>b</b>) AFM image, (<b>c</b>) height profile, and (<b>d</b>) EDX spectrum.</p> Full article ">Figure 2
<p>XPS survey spectra of (<b>a</b>) graphite; (<b>b</b>) GO<sub>Hummer</sub>; (<b>c</b>) GOQDs-NH<sub>2</sub>; (<b>d</b>) GOQDs-HA.</p> Full article ">Figure 3
<p>Cur release from GOQDs-NH<sub>2</sub> and GOQDs-HA at (<b>a</b>) 37 °C in PBS Buffer pH 7.4, (<b>b</b>) at 37 °C in PBS Buffer pH 5.5, and (<b>c</b>) at 60 °C in PBS buffer pH 7.4.</p> Full article ">Figure 4
<p>DLS size distribution of O/W NE 20% oil and monolayer 1%Oil_0.01%CT.</p> Full article ">Figure 5
<p>Size and ζ potential time stability of GOQDs-HA-CT-NEs.</p> Full article ">Figure 6
<p>(<b>a</b>) DLS size distribution and ζ potential bilayer CT-GOQDs-HA after 180 days; (<b>b</b>) NTA size distribution and frame of nanoparticles detected by NTA camera (insert).</p> Full article ">Figure 7
<p>Multiphoton confocal images of GOQD-HA<sub>Rhod</sub>-Ct<sub>FITC</sub>-NEs (<b>a</b>) FITC fluorescence; (<b>b</b>) Rhodamine fluorescence; (<b>c</b>,<b>d</b>) overlay (the arrows indicate the overlap of the two signals).</p> Full article ">Figure 8
<p>Representative confocal images of HDF cells incubated with GOQD-HA<sub>Rhod</sub> (<b>a</b>,<b>d</b>), CT<sub>FITC</sub>-HA (<b>b</b>,<b>e</b>) and CT-GOQDs-HA<sub>Rhod</sub> (<b>c</b>,<b>f</b>). (<b>g</b>) Experimental data and the linear regression model of fluorescence intensity for GO-QD-HA<sub>Rhod</sub> (black), HA-Ct<sub>FITC</sub>-NEs (dark grey), and GOQDs-HARhod-Ct-NEs (light grey); (<b>h</b>) Internalization of GO-QD-HA<sub>Rhod</sub>, HA-Ct<sub>FITC</sub>-NEs, and GOQDs-HARhod-Ct-NEs in HDF cells. * <span class="html-italic">p</span> &lt; 0.05,** <span class="html-italic">p</span> &lt; 0.01 (n ≥ 3); (<b>i</b>) Cell viability of HDFs cell incubated with 0.01 mg/mL of GO-QD-HA<sub>Rhod</sub> (black), HA-Ct<sub>FITC</sub>-NEs (dark grey) and GOQDs-HARhod-Ct-NEs. No significant differences among cell viabilities were found. Data of internalization and vitality are reported as mean ± SE.</p> Full article ">Figure 9
<p>Representative confocal images of HDF cells incubated with CT<sub>FITC</sub>-HA (<b>a</b>) and CT-GOQDs-HA<sub>Rhod</sub> (<b>e</b>), stained for LAMP2 (<b>b</b>,<b>f</b>) and in transmission (<b>d</b>,<b>h</b>). Merge of CT<sub>FITC</sub>-HA/CT-GOQDs-HA<sub>Rhod</sub> channels and LAMP2 channels (<b>c</b>,<b>g</b>) were used to analyze the co-localization of both systems with lysosomes (<b>c</b>,<b>g</b>). Scale bar, 25 µm. M1 Meander’s overlap coefficient using a threshold for both channels (CT<sub>FITC</sub>-HA/CT-GOQDs-HA<sub>Rhod</sub>) and LAMP2 are reported as mean ± SD (<b>i</b>). <span class="html-italic">p</span> &lt; 0.001 is considered statistically significant (***, Student’s <span class="html-italic">t</span>-test) (n = 9 for CT<sub>FITC</sub>-HA and n = 8 for CT-GOQDs-HA<sub>Rhod</sub>).</p> Full article ">Figure 10
<p>Confocal analysis GOQDs-HA<sub>Rhod</sub> in 3D µTPs incubation at 0.1 mg/mL (<b>top</b>). Fluorescence Intensity Quantification (<b>bottom</b>).</p> Full article ">Scheme 1
<p>Synthesis steps of multistage nanocarrier.</p> Full article ">
20 pages, 2999 KiB  
Article
Acid-Responsive Decomposable Nanomedicine Based on Zeolitic Imidazolate Frameworks for Near-Infrared Fluorescence Imaging/Chemotherapy Combined Tumor Theranostics
by Heze Guo, Vincent Mukwaya, Daikun Wu, Shuhan Xiong and Hongjing Dou
Pharmaceutics 2024, 16(6), 823; https://doi.org/10.3390/pharmaceutics16060823 - 18 Jun 2024
Viewed by 1327
Abstract
Zeolitic imidazolate framework-8 (ZIF-8) nanoparticles (NPs) are gaining traction in tumor theranostics for their effectiveness in encapsulating both imaging agents and therapeutic drugs. While typically, similar hydrophilic molecules are encapsulated in either pure aqueous or organic environments, few studies have explored co-encapsulation of [...] Read more.
Zeolitic imidazolate framework-8 (ZIF-8) nanoparticles (NPs) are gaining traction in tumor theranostics for their effectiveness in encapsulating both imaging agents and therapeutic drugs. While typically, similar hydrophilic molecules are encapsulated in either pure aqueous or organic environments, few studies have explored co-encapsulation of chemotherapeutic drugs and imaging agents with varying hydrophilicity and, consequently, constructed multifunctional ZIF-8 composite NPs for acid-responsive, near-infrared fluorescence imaging/chemotherapy combined tumor theranostics. Here, we present a one-pot method for the synthesis of uniform Cy5.5&DOX@ZIF-8 nanoparticles in mixed solvents, efficiently achieving simultaneous encapsulation of hydrophilic doxorubicin (DOX) and hydrophobic Cyanine-5.5 (Cy5.5). Surface decoration with dextran (Dex) enhanced colloidal stability and biocompatibility. The method significantly facilitated co-loading of Cy5.5 dyes and DOX drugs, endowing the composite NPs with notable fluorescent imaging capabilities and pH-responsive chemotherapy capacities. In vivo near-infrared fluorescence (NIRF) imaging in A549 tumor-bearing mice demonstrated significant accumulation of Cy5.5 at tumor sites due to enhanced permeability and retention (EPR) effects, with fluorescence intensities approximately 48-fold higher than free Cy5.5. Enhanced therapeutic efficiency was observed in composite NPs compared to free DOX, validating tumor-targeted capability. These findings suggest ZIF-8-based nanomedicines as promising platforms for multifunctional tumor theranostics. Full article
(This article belongs to the Topic Application of Nanomaterials and Nanobiotechnology in Cancer)
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<p>TEM images of: (<b>a</b>) Cy5.5&amp;DOX@ZIF-8 composite NPs (<b>c</b>) and dextran-modified Cy5.5&amp;DOX@ZIF-8-Dex composite NPs. Size distributions of: (<b>b</b>) Cy5.5&amp;DOX@ZIF-8 composite NPs and (<b>d</b>) dextran-modified Cy5.5&amp;DOX@ZIF-8-Dex composite NPs. (<b>e</b>) DLS curves of Cy5.5&amp;DOX@ZIF-8 and dextran-modified Cy5.5&amp;DOX@ZIF-8-Dex NPs. (<b>f</b>) Zeta potentials of Cy5.5&amp;DOX@ZIF-8 and dextran-modified Cy5.5&amp;DOX@ZIF-8-Dex NPs.</p> Full article ">Figure 2
<p>(<b>a</b>) DOX release from Cy5.5&amp;DOX@ZIF-8-Dex at different pH values. (<b>b</b>) Cy5.5 release from Cy5.5&amp;DOX@ZIF-8-Dex at various pH values. TEM images of Cy5.5&amp;DOX@ZIF-8-Dex under different pH values for 3 h: (<b>c</b>) pH = 7.4, (<b>d</b>) pH = 5.8, and (<b>e</b>) pH = 4.8. TEM images of Cy5.5&amp;DOX@ZIF-8-Dex under different pH values for 48 h: (<b>f</b>) pH = 7.4, (<b>g</b>) pH = 5.8, and (<b>h</b>) pH = 4.8.</p> Full article ">Figure 3
<p>(<b>a</b>) Nitrogen sorption isotherms of various functional molecule@ZIF-8-Dex composite NPs. (<b>b</b>) Surface areas of various functional molecule@ZIF-8-Dex composite NPs. (<b>c</b>) XRD patterns of various functional molecule@ZIF-8-Dex composite NPs. (<b>d</b>) FTIR spectra of various functional molecule@ZIF-8-Dex composite NPs.</p> Full article ">Figure 4
<p>(<b>a</b>) Cell viabilities of A549 cells incubated with various ZIF-8-Dex composite NPs at various concentrations for 24 h (<span class="html-italic">n</span> = 6, ** <span class="html-italic">p</span> &lt; 0.01, and *** <span class="html-italic">p</span> &lt; 0.001). (<b>b</b>) Cell viability of A549 cells incubated with various ZIF-8-Dex composite NPs at various concentrations for 48 h (<span class="html-italic">n</span> = 6, ** <span class="html-italic">p</span> &lt; 0.01, and *** <span class="html-italic">p</span> &lt; 0.001). (<b>c</b>) Cell sorting analysis of A549 cells that were, respectively, co-incubated with DOX@XIF-8-Dex, Cy5.5@ZIF-8-Dex, and Cy5.5&amp;DOX@ZIF-8-Dex NPs. (<b>d</b>) Confocal microscopy images of A549 cells incubated with Cy5.5&amp;DOX@ZIF-8-Dex NPs for 3 h. The scale bars shown in all images correspond to 10 μm.</p> Full article ">Figure 5
<p>(<b>a</b>) In vivo fluorescent images of free Cy5.5 and Cy5.5&amp;DOX@ZIF-8-Dex NPs in A549 tumor-bearing nude mice. The tumors are shown in red circles at the anterior armpits. (<b>b</b>) In vivo fluorescent images of organs harvested from A549 tumor-bearing nude mice that were, respectively, injected with PBS, free Cy5.5, and Cy5.5&amp;DOX@ZIF-8-Dex NPs. (<b>c</b>) Fluorescence intensities observed in the organs shown in (<b>b</b>) (<span class="html-italic">n</span> = 6, *** <span class="html-italic">p</span> &lt; 0.001).</p> Full article ">Figure 6
<p>(<b>a</b>) Pharmacokinetic behavior of the DOX@ZIF-8 NPs in normal mice. (<b>b</b>) Pharmacokinetic behavior of the Cy5.5&amp;DOX@ZIF-8 NPs in normal mice. (<b>c</b>) Changes in body weights of A549 tumor-bearing mice measured during a three-week period following various treatments. (<b>d</b>) Tumor growth curves of A549 tumor-bearing mice injected with PBS, free DOX, DOX@ZIF-8-Dex, and Cy5.5&amp;DOX@ZIF-8-Dex NPs, respectively. (<b>e</b>) Dissected A549 tumor volumes after treatments (** <span class="html-italic">p</span> &lt; 0.01, and *** <span class="html-italic">p</span> &lt; 0.001). (<b>f</b>) Dissected A549 tumor weights after treatments (** <span class="html-italic">p</span> &lt; 0.01, and *** <span class="html-italic">p</span> &lt; 0.001). (<b>g</b>) H&amp;E staining images of heart, liver, spleen, lung, and kidneys in A549 tumor-bearing mice. The scale bars correspond to 100 μm.</p> Full article ">Scheme 1
<p>(<b>a</b>) Schematic illustration depicting the construction of Cy5.5&amp;DOX@ZIF-8-Dex composite NPs that were simultaneously loaded with the imaging agent Cy5.5 and drug DOX, and the subsequent decoration of dextran on the surfaces of these NPs. (<b>b</b>) Schematic illustration depicting the decomposition of Cy5.5&amp;DOX@ZIF-8 NPs and the releasing of loaded cargoes under an acid environment. (<b>c</b>) Illustration depicting their application as a theranostic delivery system for tumor NIRF imaging and pH-responsive chemotherapy.</p> Full article ">
14 pages, 8407 KiB  
Article
Stability of Multicomponent Antidote Parenteral Formulations for Autoinjectors against Chemical War Agents (Neurotoxics)
by María José Rodríguez Fernández, Daniel Hernández, Brayan Javier Anaya, Dolores R. Serrano and Juan José Torrado
Pharmaceutics 2024, 16(6), 820; https://doi.org/10.3390/pharmaceutics16060820 - 17 Jun 2024
Viewed by 938
Abstract
Combinations of different drugs are formulated in autoinjectors for parenteral administration against neurotoxic war agents. In this work, the effects on the chemical stability of the following three variables were studied: (i) type of drug combination (pralidoxime, atropine, and midazolam versus obidoxime, atropine, [...] Read more.
Combinations of different drugs are formulated in autoinjectors for parenteral administration against neurotoxic war agents. In this work, the effects on the chemical stability of the following three variables were studied: (i) type of drug combination (pralidoxime, atropine, and midazolam versus obidoxime, atropine, and midazolam); (ii) pH (3 versus 4); and (iii) type of elastomeric sealing material (PH 701/50 C BLACK versus 4023/50 GRAY). Syringes were stored at three different temperatures: 4, 25, and 40 °C. Samples were assayed at different time points to study the physical appearance, drug sorption on the sealing elastomeric materials, and drug content in solution. Midazolam was unstable in all tested experimental conditions. Drug adsorption was observed in both types of sealing elastomeric materials and was significantly (p < 0.01) dependent on the lipophilicity of the drug. The most stable formulation was the combination of pralidoxime and atropine at pH 4 with the elastomeric sealing material 4023/50 GRAY. Full article
(This article belongs to the Section Physical Pharmacy and Formulation)
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<p>Chromatograms of API components of formulations 1 (<b>A</b>) and 2 (<b>B</b>). Retention times (minutes) of APIs were 2.1 min for pralidoxime, 2.2 for obidoxime, 3.3 for atropine, and 12.7 for midazolam.</p> Full article ">Figure 2
<p>Example of the two different tested formulations (F1—up in the figure and F2—down), different pHs (3 or 4), and different sealing elastomeric gasket pieces (black color—JEA and gray color—JEN) located in the glass syringes of the autoinjectors after 53 months of storage.</p> Full article ">Figure 3
<p>Visual (up) and micrograph (down) aspect of the two different tested sealing gaskets (JEA left and JEN right) located in the glass syringes of formulation 1 pH 3 of the autoinjectors after 53 months of storage at 25 ± 2 °C.</p> Full article ">Figure 4
<p>Mean results (n = 3) and standard deviation for pralidoxime, obidoxime, atropine, and midazolam within F1 (panel <b>A</b>) and F2 (panel <b>B</b>) formulations stored at 4 ± 2 °C. Key: pH3 JEA (<span class="html-fig-inline" id="pharmaceutics-16-00820-i001"><img alt="Pharmaceutics 16 00820 i001" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i001.png"/></span>), pH3 JEN (<span class="html-fig-inline" id="pharmaceutics-16-00820-i002"><img alt="Pharmaceutics 16 00820 i002" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i002.png"/></span>), pH4 JEA (<span class="html-fig-inline" id="pharmaceutics-16-00820-i003"><img alt="Pharmaceutics 16 00820 i003" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i003.png"/></span>), pH4 JEN (<span class="html-fig-inline" id="pharmaceutics-16-00820-i004"><img alt="Pharmaceutics 16 00820 i004" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i004.png"/></span>).</p> Full article ">Figure 5
<p>Shelf-life prediction for midazolam in F1 formulation at 4 °C. Key: The black dots represent the experimental points while the black solid line represents the predicted drug degradation kinetic from which the shelf-life from each formulation can be extrapolated, being intended as the time in which there is 95% confidence that at least 50% of response is within the specification limits. An upper limit and lower limit of 110 and 90% (dotted line) were taken into consideration for pralidoxime, obidoxime, and midazolam, while 107–93% was considered for atropine.</p> Full article ">Figure 6
<p>Shelf-life prediction for midazolam in F2 formulation at 4 °C. Key: The black dots represent the experimental points while the black solid line represents the predicted drug degradation kinetic from which the shelf-life from each formulation can be extrapolated, being intended as the time in which there is 95% confidence that at least 50% of response is within the specification limits. An upper limit and lower limit of 110 and 90% (dotted line) were taken into consideration for pralidoxime, obidoxime, and midazolam while 107–93% was considered for atropine.</p> Full article ">Figure 7
<p>Mean results (n = 3) and standard deviation for pralidoxime, obidoxime, atropine, and midazolam within F1 (panel <b>A</b>) and F2 (panel <b>B</b>) formulations stored at 25 °C. Key: pH3 JEA (<span class="html-fig-inline" id="pharmaceutics-16-00820-i001"><img alt="Pharmaceutics 16 00820 i001" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i001.png"/></span>), pH3 JEN (<span class="html-fig-inline" id="pharmaceutics-16-00820-i002"><img alt="Pharmaceutics 16 00820 i002" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i002.png"/></span>), pH4 JEA (<span class="html-fig-inline" id="pharmaceutics-16-00820-i003"><img alt="Pharmaceutics 16 00820 i003" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i003.png"/></span>), pH4 JEN (<span class="html-fig-inline" id="pharmaceutics-16-00820-i004"><img alt="Pharmaceutics 16 00820 i004" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i004.png"/></span>).</p> Full article ">Figure 8
<p>Shelf-life prediction for pralidoxime in F1 formulation at 25 °C. Key: The black dots represent the experimental points while the black solid line represents the predicted drug degradation kinetic from which the shelf-life from each formulation can be extrapolated, being intended as the time in which there is 95% confidence that at least 50% of response is within the specification limits. An upper limit and lower limit of 110 and 90% (dotted line) were taken into consideration for pralidoxime, obidoxime, and midazolam while 107–93% was considered for atropine.</p> Full article ">Figure 9
<p>Shelf-life prediction for obidoxime in F2 formulation at 25 °C. Key: The black dots represent the experimental points while the black solid line represents the predicted drug degradation kinetic from which the shelf-life from each formulation can be extrapolated, being intended as the time in which there is 95% confidence that at least 50% of response is within the specification limits. An upper limit and lower limit of 110 and 90% (dotted line) were taken into consideration for pralidoxime, obidoxime, and midazolam while 107–93% was considered for atropine.</p> Full article ">Figure 10
<p>Mean results (n = 3) and standard deviation for pralidoxime, obidoxime, atropine, and midazolam within F1 (panel <b>A</b>) and F2 (panel <b>B</b>) formulations stored at 40 °C. Key: pH3 JEA (<span class="html-fig-inline" id="pharmaceutics-16-00820-i001"><img alt="Pharmaceutics 16 00820 i001" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i001.png"/></span>), pH3 JEN (<span class="html-fig-inline" id="pharmaceutics-16-00820-i002"><img alt="Pharmaceutics 16 00820 i002" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i002.png"/></span>), pH4 JEA (<span class="html-fig-inline" id="pharmaceutics-16-00820-i003"><img alt="Pharmaceutics 16 00820 i003" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i003.png"/></span>), pH4 JEN (<span class="html-fig-inline" id="pharmaceutics-16-00820-i004"><img alt="Pharmaceutics 16 00820 i004" src="/pharmaceutics/pharmaceutics-16-00820/article_deploy/html/images/pharmaceutics-16-00820-i004.png"/></span>).</p> Full article ">
47 pages, 3150 KiB  
Review
Progress in Topical and Transdermal Drug Delivery Research—Focus on Nanoformulations
by Dominique Lunter, Victoria Klang, Adina Eichner, Sanela M. Savic, Snezana Savic, Guoping Lian and Franciska Erdő
Pharmaceutics 2024, 16(6), 817; https://doi.org/10.3390/pharmaceutics16060817 - 16 Jun 2024
Cited by 2 | Viewed by 3480
Abstract
Skin is the largest organ and a multifunctional interface between the body and its environment. It acts as a barrier against cold, heat, injuries, infections, chemicals, radiations or other exogeneous factors, and it is also known as the mirror of the soul. The [...] Read more.
Skin is the largest organ and a multifunctional interface between the body and its environment. It acts as a barrier against cold, heat, injuries, infections, chemicals, radiations or other exogeneous factors, and it is also known as the mirror of the soul. The skin is involved in body temperature regulation by the storage of fat and water. It is an interesting tissue in regard to the local and transdermal application of active ingredients for prevention or treatment of pathological conditions. Topical and transdermal delivery is an emerging route of drug and cosmetic administration. It is beneficial for avoiding side effects and rapid metabolism. Many pharmaceutical, technological and cosmetic innovations have been described and patented recently in the field. In this review, the main features of skin morphology and physiology are presented and are being followed by the description of classical and novel nanoparticulate dermal and transdermal drug formulations. The biophysical aspects of the penetration of drugs and cosmetics into or across the dermal barrier and their investigation in diffusion chambers, skin-on-a-chip devices, high-throughput measuring systems or with advanced analytical techniques are also shown. The current knowledge about mathematical modeling of skin penetration and the future perspectives are briefly discussed in the end, all also involving nanoparticulated systems. Full article
(This article belongs to the Special Issue Nanoparticles for Local Drug Delivery)
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<p>Main cellular elements of the human skin.</p> Full article ">Figure 2
<p>Example of nanoparticle (NP)-based carriers used as topical, dermal and transdermal drug delivery systems.</p> Full article ">Figure 3
<p><b>Top row</b>: Laser light interacts with matter, Stokes scattering provides Raman spectrum, divided into fingerprint and high wavenumber region; <b>Bottom row</b>: laser light focused into different depths of the skin generates one spectrum per skin depth from which the penetration profile is calculated.</p> Full article ">Figure 4
<p>Microscopic PBPK modeling of 4-cyanophenol disposition in SC lipids (<b>a</b>) and corneocytes (<b>b</b>) domains under in vivo exposure to healthy volunteers at 1 min(-------) (•), 5 min (- - -) (<b>×</b>) and 15 min (………) (■). Predicted overall disposition (<b>c</b>) by combining both lipids and corneocytes domains showed good agreement with tape striping data. Figure modified from [<a href="#B183-pharmaceutics-16-00817" class="html-bibr">183</a>] with permission of John Wiley and sons at the License Number 5810360623095.</p> Full article ">Figure 5
<p>Microscopic PBPK modeling of caffeine deposition in SC lipids and corneocytes domains after 20 min application to the chest of health volunteers (above). The predicted effect of hair follicle open (<b>a</b>) and blocked (<b>b</b>) on systemic bioavailability agreed well with experimental data. Figure adopted from [<a href="#B163-pharmaceutics-16-00817" class="html-bibr">163</a>] open access.</p> Full article ">
54 pages, 4234 KiB  
Review
Lipid Biomimetic Models as Simple Yet Complex Tools to Predict Skin Permeation and Drug–Membrane Biophysical Interactions
by Eduarda Fernandes, Carla M. Lopes and Marlene Lúcio
Pharmaceutics 2024, 16(6), 807; https://doi.org/10.3390/pharmaceutics16060807 - 14 Jun 2024
Viewed by 1067
Abstract
The barrier function of the skin is primarily determined by its outermost layer, the Stratum Corneum (SC). The SC consists of corneocytes embedded in a lipid matrix composed mainly of ceramides, cholesterol, and free fatty acids in equimolar proportions and is organised in [...] Read more.
The barrier function of the skin is primarily determined by its outermost layer, the Stratum Corneum (SC). The SC consists of corneocytes embedded in a lipid matrix composed mainly of ceramides, cholesterol, and free fatty acids in equimolar proportions and is organised in a complex lamellar structure with different periodicities and lateral packings. This matrix provides a diffusion pathway across the SC for bioactive compounds that are administered to the skin. In this regard, and as the skin administration route has grown in popularity, there has been an increase in the use of lipid mixtures that closely resemble the SC lipid matrix, either for a deeper biophysical understanding or for pharmaceutical and cosmetic purposes. This review focuses on a systematic analysis of the main outcomes of using lipid mixtures as SC lipid matrix models for pharmaceutical and cosmetic purposes. Thus, a methodical evaluation of the main outcomes based on the SC structure is performed, as well as the main recent developments in finding suitable new in vitro tools for permeation testing based on lipid models. Full article
(This article belongs to the Topic New Challenges in the Cosmetics Industry)
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<p>Schematic illustration of the main differences at the Cers headgroup level. Chemical structures were drawn in MarvinSketch<sup>®</sup> version 5.3.1.</p> Full article ">Figure 2
<p>Lipids constituting the intercellular lipid matrix (ILM) of the <span class="html-italic">Stratum Corneum</span> (SC) are assembled in two crystalline coexistent lamellar phases: a short periodicity phase (SPP) and a long periodicity phase (LLP) and can be laterally packed in orthorhombic, hexagonal, or liquid-like unit cells (created with <a href="http://BioRender.com" target="_blank">BioRender.com</a>).</p> Full article ">Figure 3
<p>(<b>A</b>) Schematic illustration of the sandwich model purposed for a long periodicity phase (LPP) structure. Adapted with permission from [<a href="#B162-pharmaceutics-16-00807" class="html-bibr">162</a>]. Copyright 2007, Springer Nature. (<b>B</b>) Representation of the shape of a Cer[EOS] molecule. Adapted with permission from [<a href="#B161-pharmaceutics-16-00807" class="html-bibr">161</a>]. Copyright 2023, Elsevier. (<b>C</b>) Depiction of the suggested locations of the liquid-like domains in the LPP structure. Adapted with permission from [<a href="#B139-pharmaceutics-16-00807" class="html-bibr">139</a>]. Copyright 2018, Elsevier.</p> Full article ">Figure 4
<p>Schematic illustrations of a short periodicity phase (SPP) structure, highlighting (<b>A</b>) the Cer[EOS] accommodation on SPP. Adapted with permission from [<a href="#B60-pharmaceutics-16-00807" class="html-bibr">60</a>]. Copyright 2009, Elsevier. (<b>B</b>) The reinforcement model. Adapted with permission from [<a href="#B61-pharmaceutics-16-00807" class="html-bibr">61</a>]. Copyright 2005, Springer Nature. (<b>C</b>) The asymmetric lamellae with alternating directions. Adapted with permission from [<a href="#B126-pharmaceutics-16-00807" class="html-bibr">126</a>]. Copyright 2014, American Chemical Society.</p> Full article ">Figure 5
<p>Schematic diagram of the cholesterol (Chol) influence on intercellular lipid matrix organisation. Adapted with permission from [<a href="#B125-pharmaceutics-16-00807" class="html-bibr">125</a>]. Copyright 2022, Elsevier.</p> Full article ">Figure 6
<p>Schematic representation of each approach described to obtain <span class="html-italic">Stratum Corneum</span> lipid mixture-based surrogates for in vitro permeation evaluation. (<b>A</b>) Spraying by airbrush. Adapted with permission from [<a href="#B58-pharmaceutics-16-00807" class="html-bibr">58</a>]. Copyright 2006, Elsevier. (<b>B</b>) Skin-Parallel Artificial Membrane Permeability Assay (Skin-PAMPA<sup>TM</sup>). Adapted with permission from [<a href="#B197-pharmaceutics-16-00807" class="html-bibr">197</a>]. Copyright 2013, Elsevier. (<b>C</b>) Phospholipid Vesicle-based Permeation Assay (PVPA). Adapted with permission from [<a href="#B189-pharmaceutics-16-00807" class="html-bibr">189</a>]. Copyright 2013, Wiley.</p> Full article ">
17 pages, 2761 KiB  
Review
Development, Optimization, and Clinical Relevance of Lactoferrin Delivery Systems: A Focus on Ocular Delivery
by Erika Ponzini, Gloria Astolfi, Rita Grandori, Silvia Tavazzi and Piera Versura
Pharmaceutics 2024, 16(6), 804; https://doi.org/10.3390/pharmaceutics16060804 - 14 Jun 2024
Cited by 1 | Viewed by 1508
Abstract
Lactoferrin (Lf), a multifunctional protein found abundantly in secretions, including tears, plays a crucial role in ocular health through its antimicrobial, immunoregulatory, anti-inflammatory, and antioxidant activities. Advanced delivery systems are desirable to fully leverage its therapeutic potential in treating ocular diseases. The process [...] Read more.
Lactoferrin (Lf), a multifunctional protein found abundantly in secretions, including tears, plays a crucial role in ocular health through its antimicrobial, immunoregulatory, anti-inflammatory, and antioxidant activities. Advanced delivery systems are desirable to fully leverage its therapeutic potential in treating ocular diseases. The process of Lf quantification for diagnostic purposes underscores the importance of developing reliable, cost-effective detection methods, ranging from conventional techniques to advanced nano-based sensors. Despite the ease and non-invasiveness of topical administration for ocular surface diseases, challenges such as rapid drug elimination necessitate innovations, such as Lf-loaded contact lenses and biodegradable polymeric nanocapsules, to enhance drug stability and bioavailability. Furthermore, overcoming ocular barriers for the treatment of posterior segment disease calls for nano-formulations. The scope of this review is to underline the advancements in nanotechnology-based Lf delivery methods, emphasizing the pivotal role of multidisciplinary approaches and cross-field strategies in improving ocular drug delivery and achieving better therapeutic outcomes for a wide spectrum of eye conditions. Full article
(This article belongs to the Special Issue Application Progress of Lactoferrin in Biomedicine)
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<p>Structure of diferric human lactoferrin [<a href="#B5-pharmaceutics-16-00804" class="html-bibr">5</a>]. Fe<sup>3+</sup> ions are highlighted as 3D spheres.</p> Full article ">Figure 2
<p>Schematic representation of the main protective functions of Lf. Abbreviations: Lf: lactoferrin; IFN: interferon.</p> Full article ">Figure 3
<p>Schematic representation of the eye.</p> Full article ">Figure 4
<p>Lf delivery systems for topical ophthalmic administration. PLGA: poly(lactic-co-glycolic) acid.</p> Full article ">
21 pages, 4436 KiB  
Article
Benznidazole-Loaded Polymeric Nanoparticles for Oral Chemotherapeutic Treatment of Chagas Disease
by Lucas Resende Dutra Sousa, Thays Helena Chaves Duarte, Viviane Flores Xavier, Aline Coelho das Mercês, Gabriel Maia Vieira, Maximiliano Delany Martins, Cláudia Martins Carneiro, Viviane Martins Rebello dos Santos, Orlando David Henrique dos Santos and Paula Melo de Abreu Vieira
Pharmaceutics 2024, 16(6), 800; https://doi.org/10.3390/pharmaceutics16060800 - 13 Jun 2024
Viewed by 1171
Abstract
Chagas disease (CD) is a worldwide public health problem. Benznidazole (BZ) is the drug used to treat it. However, in its commercial formulation, it has significant side effects and is less effective in the chronic phase of the infection. The development of particulate [...] Read more.
Chagas disease (CD) is a worldwide public health problem. Benznidazole (BZ) is the drug used to treat it. However, in its commercial formulation, it has significant side effects and is less effective in the chronic phase of the infection. The development of particulate systems containing BZ is therefore being promoted. The objective of this investigation was to develop polymeric nanoparticles loaded with BZ and examine their trypanocidal impact in vitro. Two formulas (BNP1 and BNP2) were produced through double emulsification and freeze drying. Subsequent to physicochemical and morphological assessment, both formulations exhibited adequate yield, average particle diameter, and zeta potential for oral administration. Cell viability was assessed in H9C2 and RAW 264.7 cells in vitro, revealing no cytotoxicity in cardiomyocytes or detrimental effects in macrophages at specific concentrations. BNP1 and BNP2 enhanced the effect of BZ within 48 h using a treatment of 3.90 μg/mL. The formulations notably improved NO reduction, particularly BNP2. The findings imply that the compositions are suitable for preclinical research, underscoring their potential as substitutes for treating CD. This study aids the quest for new BZ formulations, which are essential in light of the disregard for the treatment of CD and the unfavorable effects associated with its commercial product. Full article
(This article belongs to the Special Issue Nanoparticles: Technology and Therapeutic Applications in Human Infectious and Parasitic Diseases)
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<p>Partial chemical reaction to obtain acetylated PEG 4000.</p> Full article ">Figure 2
<p>Average particle diameter and PDI of particulate systems before and after freeze drying. Results expressed as the mean of triplicates ± standard deviation. a: significant difference in relation to the size of BNP2 before freeze drying and in relation to the size of BNP1 after freeze drying; b: significant difference in relation to the size of BNP2 after freeze drying; c: significant difference in relation to the PDI of BNP2 before freeze drying; d: significant difference in relation to the PDI of BNP2 after freeze drying.</p> Full article ">Figure 3
<p>Morphology of the BZ and of the nanospheres (P1, P2, BNP1 and BNP2) obtained by scanning electron microscopy. The white scale bars represent 200 nm or 300 nm.</p> Full article ">Figure 4
<p>Morphology of the BZ and of the nanospheres (P1, P2, BNP1 and BNP2) obtained by atomic force microscopy. The white scale bars represent 200 nm or 2 μm.</p> Full article ">Figure 5
<p>Standard analytical curve prepared with BZ.</p> Full article ">Figure 6
<p>Cell viability of H9C2 cardiomyocytes treated for 24 h (<b>A</b>) and 48 h (<b>B</b>) and of RAW 264.7 macrophages treated for 24 h (<b>C</b>) and 48 h (<b>D</b>) with BZ, P1, P2, BNP1 and BNP2. DMSO 5%: negative control; *: concentration corresponding to the concentration of polymers used in BNP1 and BNP2; a: significant difference in relation to BZ; b: significant difference in relation to P1; c: significant difference in relation to P2. Results expressed as the mean of triplicates ± standard deviation.</p> Full article ">Figure 7
<p>Number of infected cells (<b>A</b>) and number of amastigotes in 100 cells treated for 24 h (<b>B</b>). Number of infected cells (<b>C</b>) and number of amastigotes in 100 cells treated for 48 h (<b>D</b>). DMSO 5%: negative control; *: concentration corresponding to the concentration of polymers used in BNP1 and BNP2; a: significant difference in relation to untreated; b: significant difference in relation to BZ. Results expressed as the mean of triplicates ± standard deviation.</p> Full article ">Figure 8
<p>Representative photomicrographs of the treatments with the samples observed 24 h and 48 h post-treatment. Untreated, P1 and P2: intense or moderate parasitic load; BZ: low parasitic load; BNP1 and BNP2: low parasitic load. The black scale bars represent 10 μm.</p> Full article ">Figure 9
<p>NO measurement in the supernatant of RAW 264.7 macrophages treated for 24 h (<b>A</b>) and 48 h (<b>B</b>) with BZ, P1, P2, BNP1 and BNP2. DMSO 5%: negative control; *: concentration corresponding to the concentration of polymers used in BNP1 and BNP2; a: significant difference in relation to the untreated stimulated with LPS + IFN- γ control. Results expressed as the mean of triplicates ± standard deviation.</p> Full article ">
19 pages, 4927 KiB  
Article
Synthesis of Gd-DTPA Carborane-Containing Compound and Its Immobilization on Iron Oxide Nanoparticles for Potential Application in Neutron Capture Therapy
by Ilya V. Korolkov, Alexander Zaboronok, Kairat A. Izbasar, Zhangali A. Bekbol, Lana I. Lissovskaya, Alexandr V. Zibert, Rafael I. Shakirzyanov, Luiza N. Korganbayeva, Haolan Yang, Eiichi Ishikawa and Maxim V. Zdorovets
Pharmaceutics 2024, 16(6), 797; https://doi.org/10.3390/pharmaceutics16060797 - 12 Jun 2024
Viewed by 1274
Abstract
Cancer is one of the leading causes of global mortality, and its incidence is increasing annually. Neutron capture therapy (NCT) is a unique anticancer modality capable of selectively eliminating tumor cells within normal tissues. The development of accelerator-based, clinically mountable neutron sources has [...] Read more.
Cancer is one of the leading causes of global mortality, and its incidence is increasing annually. Neutron capture therapy (NCT) is a unique anticancer modality capable of selectively eliminating tumor cells within normal tissues. The development of accelerator-based, clinically mountable neutron sources has stimulated a worldwide search for new, more effective compounds for NCT. We synthesized magnetic iron oxide nanoparticles (NPs) that concurrently incorporate boron and gadolinium, potentially enhancing the effectiveness of NCT. These magnetic nanoparticles underwent sequential modifications through silane polycondensation and allylamine graft polymerization, enabling the creation of functional amino groups on their surface. Characterization was performed using Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), energy dispersive X-ray (EDX), dynamic light scattering (DLS), thermal gravimetric analysis (TGA), and transmission electron microscopy (TEM). ICP-AES measurements indicated that boron (B) content in the NPs reached 3.56 ppm/mg, while gadolinium (Gd) averaged 0.26 ppm/mg. Gadolinium desorption was observed within 4 h, with a peak rate of 61.74%. The biocompatibility of the NPs was confirmed through their relatively low cytotoxicity and sufficient cellular tolerability. Using NPs at non-toxic concentrations, we obtained B accumulation of up to 5.724 × 1010 atoms per cell, sufficient for successful NCT. Although limited by its content in the NP composition, the Gd amount may also contribute to NCT along with its diagnostic properties. Further development of the NPs is ongoing, focusing on increasing the boron and gadolinium content and creating active tumor targeting. Full article
(This article belongs to the Special Issue Development of Novel Tumor-Targeting Nanoparticles, 2nd Edition)
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<p>Scheme of DTPA modification and reaction with GDD.</p> Full article ">Figure 2
<p>Scheme of synthesis and reduction of 2,3-(3-nitrophenyl)-4-(isopropyl-o-carboranyl)hydrindone (GDD-NO<sub>2</sub>).</p> Full article ">Figure 3
<p>Calibration curve of DTPA-GDD.</p> Full article ">Figure 4
<p>Calibration curve of acid orange.</p> Full article ">Figure 5
<p>Scheme of Fe<sub>3</sub>O<sub>4</sub> modification and Gd-DTPA-GDD immobilization.</p> Full article ">Figure 6
<p>FTIR of Fe<sub>3</sub>O<sub>4</sub> at different stages of modification and adsorption of DTPA-GDD.</p> Full article ">Figure 7
<p>TEM of Fe<sub>3</sub>O<sub>4</sub>-TMSPM-PAlAm/Gd-DTPA-GDD.</p> Full article ">Figure 8
<p>The zeta potential of the obtained nanoparticles.</p> Full article ">Figure 9
<p>X-ray diffraction patterns of the obtained samples. 1—Fe<sub>3</sub>O<sub>4</sub>, 2—Fe<sub>3</sub>O<sub>4</sub>-TMSPM, 3—Fe<sub>3</sub>O<sub>4</sub>-TMSPM-PAlAm, 4—Fe<sub>3</sub>O<sub>4</sub>-TMSPM-PAlAm/Gd-DTPA-GDD.</p> Full article ">Figure 10
<p>TGA analysis of Fe<sub>3</sub>O<sub>4</sub> NPs at different stages of modification for Fe<sub>3</sub>O<sub>4</sub> (<b>a</b>), Fe<sub>3</sub>O<sub>4</sub>-TMSPM (<b>b</b>), Fe<sub>3</sub>O<sub>4</sub>-TMSPM-PAlAm (<b>c</b>), and Fe<sub>3</sub>O<sub>4</sub>-TMSPM-PAlAm/Gd-DTPA-GDD (<b>d</b>) samples.</p> Full article ">Figure 11
<p>The release of Gd-DTPA-GDD in a phosphate-buffered saline (PBS) solution.</p> Full article ">Figure 12
<p>T98G cell proliferation after incubation with NPs.</p> Full article ">Figure 13
<p>Fe, B, and Gd accumulation per 10<sup>6</sup> T98G cells after incubation with NPs.</p> Full article ">
12 pages, 3160 KiB  
Review
Nano-Delivery of Immunogenic Cell Death Inducers and Immune Checkpoint Blockade Agents: Single-Nanostructure Strategies for Enhancing Immunotherapy
by Yujeong Moon, Hanhee Cho and Kwangmeyung Kim
Pharmaceutics 2024, 16(6), 795; https://doi.org/10.3390/pharmaceutics16060795 - 12 Jun 2024
Cited by 3 | Viewed by 1374
Abstract
Cancer immunotherapy has revolutionized oncology by harnessing the patient’s immune system to target and eliminate cancer cells. However, immune checkpoint blockades (ICBs) face limitations such as low response rates, particularly in immunologically ‘cold’ tumors. Enhancing tumor immunogenicity through immunogenic cell death (ICD) inducers [...] Read more.
Cancer immunotherapy has revolutionized oncology by harnessing the patient’s immune system to target and eliminate cancer cells. However, immune checkpoint blockades (ICBs) face limitations such as low response rates, particularly in immunologically ‘cold’ tumors. Enhancing tumor immunogenicity through immunogenic cell death (ICD) inducers and advanced drug delivery systems represents a promising solution. This review discusses the development and application of various nanocarriers, including polymeric nanoparticles, liposomes, peptide-based nanoparticles, and inorganic nanoparticles, designed to deliver ICD inducers and ICBs effectively. These nanocarriers improve therapeutic outcomes by converting cold tumors into hot tumors, thus enhancing immune responses and reducing systemic toxicity. By focusing on single-nanoparticle systems that co-deliver both ICD inducers and ICBs, this review highlights their potential in achieving higher drug concentrations at tumor sites, improving pharmacokinetics and pharmacodynamics, and facilitating clinical translation. Future research should aim to optimize these nanocarrier systems for better in vivo performance and clinical applications, ultimately advancing cancer immunotherapy. Full article
(This article belongs to the Special Issue Nanomedicines in Cancer Therapy)
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<p>Schematic illustration of the action mechanism of nanoparticles encapsulating ICD and ICB agents in cancer immunotherapy.</p> Full article ">Figure 2
<p>(<b>A</b>) Preparation of DOX/P-12-encapsulated cell-penetration peptide octaarginine (R8) conjugated lipid/PLGA nanoparticles and their mechanism of action. (<b>B</b>) TEM images of LPN (<b>top</b>) and LPN-30-R8<sup>2k</sup> (<b>bottom</b>) with hydrodynamic volumes of 100 nm and 125 nm, respectively. (<b>C</b>) Enhanced cellular uptake of LPN-30-R8<sup>2k</sup>@DP in CT26 cells compared to LPN@DP. (<b>D</b>) Expression of CRT as DAMPs due to DOX release from LPN-30-R8<sup>2k</sup>@DP. (<b>E</b>) Improved tumor targeting efficiency of LPN-30-R8<sup>2k</sup>@DP via passive targeting in CT26-tumor bearing mice. The white dashed line indicates the location of the tumor. (<b>F</b>) Enhanced tumor growth suppression using LPN-30-R8<sup>2k</sup>@DP compared to other groups with a dosage of 3 mg dox/kg and 5 mg P-12/kg. (<b>G</b>) CD8<sup>+</sup> T cell staining in excised tumor tissue from the LPN-30-R8<sup>2k</sup>@DP treated group. The student’s <span class="html-italic">t</span>-test was applied for statistical significance. (* <span class="html-italic">p</span> &lt; 0.05). Reproduced with permission [<a href="#B55-pharmaceutics-16-00795" class="html-bibr">55</a>]. Copyright: ACS publications, 2022.</p> Full article ">Figure 3
<p>(<b>A</b>) Preparation of DOX/IDO1-encapsulated liposomes surface-modified with CD44 and PD-L1 aptamers, and their mode of action. (<b>B</b>) The size of the nanostructures confirmed by DLS and TEM images with an average size of 183 nm. (<b>C</b>) Released DOX from the liposomes induced ATP release as DAMPs in MDA-MB-231 cells. (<b>D</b>) Aptm[DOX/IDO1] demonstrated enhanced tumor targeting in 4T1-tumor-bearing mice compared to Lipm[DOX/IDO1], due to a dual-targeting mechanism via passive and active targeting. (<b>E</b>) Effective tumor suppression was observed in the Aptm[DOX/IDO1]-treated group in the 4T1 tumor xenograft model. (<b>F</b>) Immune activation was evident in the excised tumor tissue from the Aptm[DOX/IDO1]-treated group. One-way analysis of variance (ANOVA) with Dunnett’s test was applied for statistical significance (* <span class="html-italic">p</span> &lt; 0.05, *** <span class="html-italic">p</span> &lt; 0.005). Reproduced with permission [<a href="#B58-pharmaceutics-16-00795" class="html-bibr">58</a>]. Copyright: Elsevier, 2022.</p> Full article ">Figure 4
<p>(<b>A</b>) Preparation of self-assembled peptide-derived prodrug nanoparticles capable of releasing DOX in the presence of cathepsin B. (<b>B</b>) The size of the self-assembled nanoparticles was confirmed by DLS and TEM with an average size of 157.4 ± 12.1 nm, while no particle formation was observed in the presence of cathepsin B. (<b>C</b>) CRT expression was observed as DAMPs due to the specific release of DOX in 4T1 cancer cells. (<b>D</b>) Enhanced tumor-targeting efficiency of nanoparticles via the EPR effect in the 4T1 tumor xenograft model at 150–200 mm<sup>3</sup> volume. (<b>E</b>) Improved tumor suppression due to the release of DOX and PD-L1 blockade peptide with a dosage of 3 mg DOX/kg. (<b>F</b>) Immune analysis (CD8<sup>+</sup> cytotoxic T cell, regulatory T cell) of excised tumor tissue following tumor suppression evaluation. Reproduced with permission [<a href="#B61-pharmaceutics-16-00795" class="html-bibr">61</a>]. One-way analysis of variance (ANOVA) with Tukey-Kramer <span class="html-italic">posthoc</span> test was applied for statistical significance (** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.005). Copyright: Ivyspring international Publisher, 2022.</p> Full article ">
22 pages, 11192 KiB  
Article
Identification and Characterization of Critical Processing Parameters in the Fabrication of Double-Emulsion Poly(lactic-co-glycolic) Acid Microparticles
by Elizabeth R. Bentley, Stacia Subick, Michael Pezzillo, Stephen C. Balmert, Aidan Herbert and Steven R. Little
Pharmaceutics 2024, 16(6), 796; https://doi.org/10.3390/pharmaceutics16060796 - 12 Jun 2024
Viewed by 1995
Abstract
In the past several decades, polymeric microparticles (MPs) have emerged as viable solutions to address the limitations of standard pharmaceuticals and their corresponding delivery methods. While there are many preclinical studies that utilize polymeric MPs as a delivery vehicle, there are limited FDA-approved [...] Read more.
In the past several decades, polymeric microparticles (MPs) have emerged as viable solutions to address the limitations of standard pharmaceuticals and their corresponding delivery methods. While there are many preclinical studies that utilize polymeric MPs as a delivery vehicle, there are limited FDA-approved products. One potential barrier to the clinical translation of these technologies is a lack of understanding with regard to the manufacturing process, hindering batch scale-up. To address this knowledge gap, we sought to first identify critical processing parameters in the manufacturing process of blank (no therapeutic drug) and protein-loaded double-emulsion poly(lactic-co-glycolic) acid MPs through a quality by design approach. We then utilized the design of experiments as a tool to systematically investigate the impact of these parameters on critical quality attributes (e.g., size, surface morphology, release kinetics, inner occlusion size, etc.) of blank and protein-loaded MPs. Our results elucidate that some of the most significant CPPs impacting many CQAs of double-emulsion MPs are those within the primary or single-emulsion process (e.g., inner aqueous phase volume, solvent volume, etc.) and their interactions. Furthermore, our results indicate that microparticle internal structure (e.g., inner occlusion size, interconnectivity, etc.) can heavily influence protein release kinetics from double-emulsion MPs, suggesting it is a crucial CQA to understand. Altogether, this study identifies several important considerations in the manufacturing and characterization of double-emulsion MPs, potentially enhancing their translation. Full article
(This article belongs to the Section Nanomedicine and Nanotechnology)
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<p>Process diagram illustrating emulsion process and potential critical processing parameters in double-emulsion solvent evaporation microparticle fabrication. Critical processing parameters can exist in all steps of fabrication, including the primary emulsion (W/O), secondary emulsion (W/O/W), and solvent evaporation steps.</p> Full article ">Figure 2
<p>Single-factor plots demonstrating the significant relationship between microparticle size and significant critical processing parameters (<span class="html-italic">p</span> &lt; 0.05), including (Parameter A) inner aqueous phase volume (<span class="html-italic">n</span> = 200), (Parameter B) solvent volume (<span class="html-italic">n</span> = 200), (Parameter C) PLGA amount (<span class="html-italic">n</span> = 200), (Parameter E) homogenization time (<span class="html-italic">n</span> = 200), and (Parameter F) concentration of surfactant in the outer aqueous phase (<span class="html-italic">n</span> = 200). The black solid line represents the expected trend, and the blue dotted line illustrates the confidence bands for that trend, as predicted by the RSM.</p> Full article ">Figure 3
<p>Aqueous phase volume (μL) and solvent volume (mL) interact to impact microparticle size. (<b>A</b>) A 3D surface (left) and contour plot (right) illustrating the overall trends of the impact of aqueous phase volume and solvent volume on microparticle size. (<b>B</b>) Model and experimental data examining size trends at specific levels of aqueous phase volume and solvent volume. Other preparation conditions for these batches are as follows: 200 mg of polymer, 3000 rpm homogenization speed, 1 min homogenization time, 600 rpm stirring speed, 55% sonication amplitude, 2% outer aqueous phase concentration, and 3 h solvent evaporation. * Indicates significant difference in microparticle size (<span class="html-italic">p</span> &lt; 0.05).</p> Full article ">Figure 4
<p>Solvent volume (mL) and polymer amount (mg) interact to impact microparticle size. (<b>A</b>) A 3D surface (<b>left</b>) and contour plot (<b>right</b>) illustrating overall trends of the impact of polymer and solvent volume on microparticle size. (<b>B</b>) Model and experimental data examining size trends at specific levels of polymer amount and solvent volume. Other preparation conditions for these batches are as follows: 200 μL inner aqueous phase volume, 3000 rpm homogenization speed, 1 min homogenization time, 600 rpm stirring speed, 55% sonication amplitude, 2% outer aqueous phase concentration, and 3 h solvent evaporation. * Indicates significant difference in microparticle size (<span class="html-italic">p</span> &lt; 0.05). ns indicates non-significant difference in microparticle size.</p> Full article ">Figure 5
<p>Interaction between inner aqueous phase volume and solvent volume impacts release and internal structure of rhCCL22-MPs. (<b>A</b>) Cumulative release (ng rhCCL22/mg MP) profiles of rhCCL22-MPs formulated with each inner aqueous phase volume (200, 500, and 800 μL) at each solvent volume (2, 4, and 8 mL) (<span class="html-italic">n</span> = 3). (<b>B</b>) Scanning electron microscopy (SEM) images of microparticle cross-sections. SEM images taken at 1.5 kx. Scale bar = 10 μm. (<b>C</b>) Diagram depicting features of microparticle internal structure and associated measurements. (<b>D</b>) Microparticle inner occlusion diameter (μm) and polymer matrix composition (% polymer) measurements (<span class="html-italic">n</span> = 3–4). Other preparation conditions for these batches are as follows: 200 mg of PLGA, 3000 rpm homogenization speed, 1 min homogenization time, 55% sonication amplitude, 600 rpm stirring speed, 2% outer aqueous phase concentration, and 3 h solvent evaporation. * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001, **** <span class="html-italic">p</span> &lt; 0.0001, ns indicates non-significant difference.</p> Full article ">Figure 6
<p>Interaction between polymer amount and solvent volume impacts release kinetics and internal structure of rhCCL22-MPs. (<b>A</b>) Cumulative release (%) profiles of rhCCL22-MPs formulated with each polymer amount (200, 400, and 600 mg) at each solvent volume (2, 4, and 8 mL) (<span class="html-italic">n</span> = 3). (<b>B</b>) Scanning electron microscopy (SEM) images of microparticle cross-sections. SEM images taken at 1.5 kx. Scale bar = 10 μm. (<b>C</b>) Microparticle inner occlusion diameter (μm) and polymer matrix composition (% polymer) measurements (<span class="html-italic">n</span> = 3–4). Other preparation conditions for these batches are as follows: 200 μL IA phase volume, 3000 rpm homogenization speed, 1 min homogenization time, 55% sonication amplitude, 600 rpm stirring speed, 2% outer aqueous phase concentration, and 3 h solvent evaporation. * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001, ns indicates non-significant difference.</p> Full article ">Figure 7
<p>Interaction between solvent volume and polymer amount impacts rhCCL22 release kinetics. Cumulative rhCCL22 release (ng rhCCL22/mg MP) from microparticles formulated with (<b>A</b>) 2 mL and (<b>B</b>) 4 mL of solvent. Other preparation conditions for these batches are as follows: 3000 rpm homogenization speed, 1 min homogenization time, 55% sonication amplitude, 600 rpm stirring speed, 2% outer aqueous phase concentration, and 3 h solvent evaporation.</p> Full article ">Figure 8
<p>Formulations A, B, and C exhibit some similarities and differences in CQAs. (<b>A</b>) Cumulative release curves (ng rhCCL22/mg MP) for formulations A, B, and C. (<b>B</b>) Scanning electron microscopy images demonstrating spherical surface morphology. Images were taken at 1.2 kx. Scale bars = 10 μm. (<b>C</b>) Volume-weighted size distributions of formulations A, B, and C. <span class="html-italic">n</span> = 10,000 particles. Data from formulation A (control) adapted from Fisher et al., <span class="html-italic">Sci. Adv</span>. (2019). [<a href="#B22-pharmaceutics-16-00796" class="html-bibr">22</a>].</p> Full article ">
14 pages, 2993 KiB  
Article
Evaluation of Novel Nasal Mucoadhesive Nanoformulations Containing Lipid-Soluble EGCG for Long COVID Treatment
by Nicolette Frank, Douglas Dickinson, Garrison Lovett, Yutao Liu, Hongfang Yu, Jingwen Cai, Bo Yao, Xiaocui Jiang and Stephen Hsu
Pharmaceutics 2024, 16(6), 791; https://doi.org/10.3390/pharmaceutics16060791 - 11 Jun 2024
Cited by 1 | Viewed by 1643
Abstract
Following recovery from the acute infection stage of the SARS-CoV-2 virus (COVID-19), survivors can experience a wide range of persistent Post-Acute Sequelae of COVID-19 (PASC), also referred to as long COVID. According to the US National Research Action Plan on Long COVID 2022, [...] Read more.
Following recovery from the acute infection stage of the SARS-CoV-2 virus (COVID-19), survivors can experience a wide range of persistent Post-Acute Sequelae of COVID-19 (PASC), also referred to as long COVID. According to the US National Research Action Plan on Long COVID 2022, up to 23.7 million Americans suffer from long COVID, and approximately one million workers may be out of the workforce each day due to these symptoms, leading to a USD 50 billion annual loss of salary. Neurological symptoms associated with long COVID result from persistent infection with SARS-CoV-2 in the nasal neuroepithelial cells, leading to inflammation in the central nervous system (CNS). As of today, there is no evidence that vaccines or medications can clear the persistent viral infection in olfactory mucosa. Recently published clinical data demonstrate that only 5% of long COVID anosmia patients have fully recovered during the past 2 years, and 10.4% of COVID patients are still symptomatic 18 months post-infection. Our group demonstrated that epigallocatechin-3-gallate-monopalmitate (EC16m) nanoformulations possess strong antiviral activity against human coronavirus, suggesting that this green-tea-derived compound in nanoparticle formulations could be developed as an intranasally delivered new drug targeting the persistent SARS-CoV-2 infection, as well as inflammation and oxidative stress in the CNS, leading to restoration of neurologic functions. The objective of the current study was to evaluate the mucociliary safety of the EC16m nasal nanoformulations and their efficacy against human coronavirus. Methods: Nanoparticle size and Zeta potential were measured using the ZetaView Nanoparticle Tracking Analysis system; mucociliary safety was determined using the MucilAir human nasal model; contact antiviral activity and post-infection inhibition against the OC43 viral strain were assessed by the TCID50 assay for cytopathic effect on MRC-5 cells. Results: The saline-based EC16 mucoadhesive nanoformulations containing 0.005 to 0.02% w/v EC16m have no significant difference compared to saline (0.9% NaCl) with respect to tissue integrity, cytotoxicity, and cilia beat frequency. A 5 min contact resulted in 99.9% inactivation of β-coronavirus OC43. OC43 viral replication was inhibited by >90% after infected MRC-5 cells were treated with the formulations. Conclusion: The saline-based novel EC16m mucoadhesive nasal nanoformulations rapidly inactivated human coronavirus with mucociliary safety properties comparable to saline, a solution widely used for nasal applications. Full article
(This article belongs to the Special Issue Nanoparticle-Mediated Targeted Drug Delivery Systems)
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<p>Schematic illustration of sample application and end-point measurements of MucilAire 3D human nasal model.</p> Full article ">Figure 2
<p>(<b>A</b>) Size distribution of particles in FC. (<b>B</b>) Size distribution of particles in FD The size distribution profile for one representative sample/formulation determined by NTA is shown. (<b>C</b>) Zeta potential and distribution of FC. (<b>D</b>) Zeta potential and distribution of FD.</p> Full article ">Figure 3
<p>Cell viability (MTT) assay results for the four nanoformulations in comparison to saline as vehicle control and untreated cell control. The assay was conducted in 48-well tissue culture plates with confluent HCT-8 cells in each well (<span class="html-italic">n</span> = 3). Select <span class="html-italic">p</span> values are shown; ns: not significant (<span class="html-italic">p</span> &gt; 0.05). Arrows point to columns with differences.</p> Full article ">Figure 4
<p>Impact on tissue integrity of the nanoformulations in comparison to saline (vehicle). Select <span class="html-italic">p</span> values are shown; ns: not significant (<span class="html-italic">p</span> &gt; 0.05). Arrows indicate significant difference between pairs of columns. Brackets show groups of columns with no significant difference to indicated column to adjacent columns.</p> Full article ">Figure 5
<p>Cytotoxicity induced by the nanoformulations in comparison to untreated control, saline (vehicle), and positive control (10% Triton X-100). Select <span class="html-italic">p</span> values are shown; ns: not significant (<span class="html-italic">p</span> &gt; 0.05). Arrows indicate significant difference between pairs of columns. Brackets show groups of columns with no significant difference to indicated column to adjacent columns.</p> Full article ">Figure 6
<p>Cilia beating frequency measurements for the four nanoformulations in comparison to saline (vehicle) after two days of applications based on twice daily 30 min application/each schedule. FA was the only nanoformulation associated with significantly reduced CBF among the formulations. Select <span class="html-italic">p</span> values are shown; ns: not significant (<span class="html-italic">p</span> &gt; 0.05). Arrows indicate significant difference between pairs of columns. Brackets show groups of columns with no significant difference to indicated column to adjacent columns.</p> Full article ">Figure 7
<p>Mean log<sub>10</sub> reduction in OC43 infectivity after incubation with FC and FD for 5 and 10 min (V: vehicle control). The results are from three independent TCID50 assays for the nanoformulations. Select <span class="html-italic">p</span> values are shown; ns: not significant (<span class="html-italic">p</span> &gt; 0.05).</p> Full article ">Figure 8
<p>Mean log<sub>10</sub> reduction in OC43 infectivity after post-infection incubation with FC and FD for 5 min (VC, VD: vehicle controls for FC and FD, respectively). The results are from three independent TCID50 assays for the nanoformulations.</p> Full article ">
26 pages, 4460 KiB  
Article
A Physiologically Based Pharmacokinetic (PBPK) Study to Assess the Adjuvanticity of Three Peptides in an Oral Vaccine
by Leonor Saldanha, Ülo Langel and Nuno Vale
Pharmaceutics 2024, 16(6), 780; https://doi.org/10.3390/pharmaceutics16060780 - 8 Jun 2024
Viewed by 912
Abstract
Following up on the first PBPK model for an oral vaccine built for alpha-tocopherol, three peptides are explored in this article to verify if they could support an oral vaccine formulation as adjuvants using the same PBPK modeling approach. A literature review was [...] Read more.
Following up on the first PBPK model for an oral vaccine built for alpha-tocopherol, three peptides are explored in this article to verify if they could support an oral vaccine formulation as adjuvants using the same PBPK modeling approach. A literature review was conducted to verify what peptides have been used as adjuvants in the last decades, and it was noticed that MDP derivatives have been used, with one of them even being commercially approved and used as an adjuvant when administered intravenously in oncology. The aim of this study was to build optimized models for three MDP peptides (MDP itself, MTP-PE, and murabutide) and to verify if they could act as adjuvants for an oral vaccine. Challenges faced by peptides in an oral delivery system are taken into consideration, and improvements to the formulations to achieve better results are described in a step-wise approach to reach the most-optimized model. Once simulations are performed, results are compared to determine what would be the best peptide to support as an oral adjuvant. According to our results, MTP-PE, the currently approved and commercialized peptide, could have potential to be incorporated into an oral formulation. It would be interesting to proceed with further in vivo experiments to determine the behavior of this peptide when administered orally with a proper formulation to overcome the challenges of oral delivery systems. Full article
(This article belongs to the Special Issue Approaches to Individualized Drug Therapy Based on Population Pharmacometrics)
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<p>Steps to build optimized MDP model.</p> Full article ">Figure 2
<p>Steps to build optimized MTP-PE model.</p> Full article ">Figure 3
<p>Steps to build optimized murabutide model.</p> Full article ">Figure 4
<p>MDP simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0; (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0.</p> Full article ">Figure 4 Cont.
<p>MDP simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0; (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0.</p> Full article ">Figure 4 Cont.
<p>MDP simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0; (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0.</p> Full article ">Figure 5
<p>MTP-PE simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0; (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0, (<b>g</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>h</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm. And volume of 512 mL, (<b>i</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>j</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm and volume of 512 mL.</p> Full article ">Figure 5 Cont.
<p>MTP-PE simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0; (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0, (<b>g</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>h</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm. And volume of 512 mL, (<b>i</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>j</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm and volume of 512 mL.</p> Full article ">Figure 5 Cont.
<p>MTP-PE simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0; (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0, (<b>g</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>h</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm. And volume of 512 mL, (<b>i</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>j</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm and volume of 512 mL.</p> Full article ">Figure 5 Cont.
<p>MTP-PE simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0; (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0, (<b>g</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>h</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm. And volume of 512 mL, (<b>i</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>j</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm and volume of 512 mL.</p> Full article ">Figure 5 Cont.
<p>MTP-PE simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0; (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0, (<b>g</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>h</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm. And volume of 512 mL, (<b>i</b>) Plasma concentration with capsule on dosage form, dose 4 mg and volume 200 mL, Peff 1.0; and particle size of 2 μm (<b>j</b>) Absorption and dissolution profile with capsule on dosage form, dose 4 mg and volume 200 mL and Peff 1.0 and particle size of 2 μm and volume of 512 mL.</p> Full article ">Figure 6
<p>Murabutide simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 7 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 7 g and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 7 mg and volume 200 mL and Peff 0.6 (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 7 mg and volume 200 mL and Peff 0.6.</p> Full article ">Figure 6 Cont.
<p>Murabutide simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 7 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 7 g and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 7 mg and volume 200 mL and Peff 0.6 (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 7 mg and volume 200 mL and Peff 0.6.</p> Full article ">Figure 6 Cont.
<p>Murabutide simulation results for the model optimization. (<b>a</b>) Plasma concentration without experimental data (ADMET predictions only); (<b>b</b>) Absorption and dissolution profile without experimental data (ADMET predictions only); (<b>c</b>) Plasma concentration with capsule on dosage form, dose 7 mg and volume 200 mL; (<b>d</b>) Absorption and dissolution profile with capsule on dosage form, dose 7 g and volume 200 mL, (<b>e</b>) Plasma concentration with capsule on dosage form, dose 7 mg and volume 200 mL and Peff 0.6 (<b>f</b>) Absorption and dissolution profile with capsule on dosage form, dose 7 mg and volume 200 mL and Peff 0.6.</p> Full article ">Figure 7
<p>(<b>a</b>) Compartmental absorption without experimental data (ADMET predictor only); (<b>b</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL (<b>c</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL and Peff of 1.0.</p> Full article ">Figure 8
<p>MTP-PE (<b>a</b>) Compartmental absorption without experimental data (ADMET predictor only); (<b>b</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL (<b>c</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL and Peff of 1.0 (<b>d</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL and Peff of 1.0 and particle size of 2 μm (<b>e</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL.</p> Full article ">Figure 8 Cont.
<p>MTP-PE (<b>a</b>) Compartmental absorption without experimental data (ADMET predictor only); (<b>b</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL (<b>c</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL and Peff of 1.0 (<b>d</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL and Peff of 1.0 and particle size of 2 μm (<b>e</b>) Compartmental absorption with capsule on dosage form, dose 4 mg and volume 200 mL.</p> Full article ">Figure 9
<p>Murabutide (<b>a</b>) Compartmental absorption without experimental data (ADMET predictor only); (<b>b</b>) Compartmental absorption with capsule on dosage form, dose 7 mg and volume 200 mL (<b>c</b>) Compartmental absorption with capsule on dosage form, dose 7 mg and volume 200 mL and Peff 0.6.</p> Full article ">Scheme 1
<p>Impact of adjuvants on innate and adaptative immune system.</p> Full article ">
21 pages, 6251 KiB  
Article
Preparation and Evaluation of Inhalable Microparticles with Improved Aerodynamic Performance and Dispersibility Using L-Leucine and Hot-Melt Extrusion
by Jin-Hyuk Jeong, Ji-Su Kim, Yu-Rim Choi, Dae Hwan Shin, Ji-Hyun Kang, Dong-Wook Kim, Yun-Sang Park and Chun-Woong Park
Pharmaceutics 2024, 16(6), 784; https://doi.org/10.3390/pharmaceutics16060784 - 8 Jun 2024
Viewed by 1280
Abstract
Dry-powder inhalers (DPIs) are valued for their stability but formulating them is challenging due to powder aggregation and limited flowability, which affects drug delivery and uniformity. In this study, the incorporation of L-leucine (LEU) into hot-melt extrusion (HME) was proposed to enhance dispersibility [...] Read more.
Dry-powder inhalers (DPIs) are valued for their stability but formulating them is challenging due to powder aggregation and limited flowability, which affects drug delivery and uniformity. In this study, the incorporation of L-leucine (LEU) into hot-melt extrusion (HME) was proposed to enhance dispersibility while simultaneously maintaining the high aerodynamic performance of inhalable microparticles. This study explored using LEU in HME to improve dispersibility and maintain the high aerodynamic performance of inhalable microparticles. Formulations with crystalline itraconazole (ITZ) and LEU were made via co-jet milling and HME followed by jet milling. The LEU ratio varied, comparing solubility, homogenization, and aerodynamic performance enhancements. In HME, ITZ solubility increased, and crystallinity decreased. Higher LEU ratios in HME formulations reduced the contact angle, enhancing mass median aerodynamic diameter (MMAD) size and aerodynamic performance synergistically. Achieving a maximum extra fine particle fraction of 33.68 ± 1.31% enabled stable deep lung delivery. This study shows that HME combined with LEU effectively produces inhalable particles, which is promising for improved drug dispersion and delivery. Full article
(This article belongs to the Special Issue Dry Powders for Pulmonary Delivery: Device and Formulation Technologies for an Enhanced Lung Deposition, 2nd Edition)
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<p>SEM micrographs of (<b>A</b>) raw materials, (<b>B</b>) JMs, and (<b>C</b>) HMEs. Magnification for samples was 250× and 1000×, respectively.</p> Full article ">Figure 2
<p>DSC thermogram of (<b>A</b>) raw materials, (<b>B</b>) PMs, (<b>C</b>) JMs, (<b>D</b>) HMEs.</p> Full article ">Figure 3
<p>XRD pattern of (<b>A</b>) raw materials, (<b>B</b>) PMs, (<b>C</b>) JMs, and (<b>D</b>) HMEs. Colors represent the following: ITZ is red, MAN is blue, and LEU is green.</p> Full article ">Figure 4
<p>Percentage deposition in each stage of NGI: (<b>A</b>) JMs and (<b>B</b>) HMEs. Aerodynamic performance according to LEU ratio in NGI: (<b>C</b>) ED, (<b>D</b>) FPF, and (<b>E</b>) eFPF (mean ± SD, n = 3). <sup>###</sup> ANOVA, <span class="html-italic">p</span>-value &lt; 0.001 compared with JM-L0; <sup>#</sup> ANOVA, <span class="html-italic">p</span>-value &lt; 0.05, compared with JM-L0.1; * ANOVA, <span class="html-italic">p</span>-value &lt; 0.05 compared with JM-L1; **** ANOVA, <span class="html-italic">p</span>-value &lt; 0.0001 compared with JM-L10.</p> Full article ">Figure 5
<p>Solubility of raw ITZ, JMs, HMEs (mean ± SD, n = 3). **** ANOVA: HME-L10 vs. JM-L10, <span class="html-italic">p</span>-value &lt; 0.0001; ** ANOVA: JM-L10 vs. raw ITZ, <span class="html-italic">p</span>-value 0. 0035.</p> Full article ">Figure 6
<p>The in vitro drug release profiles of JMs and HMEs according to LEU ratios in Franz diffusion cells: (<b>A</b>) L0, (<b>B</b>) L0.1, (<b>C</b>) L1, and (<b>D</b>) L10 (mean ± SD, n = 3).</p> Full article ">Figure 7
<p>Water sorption/desorption isotherms for ITZ and formulations at 25 °C: (<b>A</b>) JM-L0, JM-L10, and (<b>B</b>) HME-L0, HME-L10.</p> Full article ">Figure 8
<p>Comparison of wettability through contact angle measurement (mean ± SD, n = 3).</p> Full article ">Figure 9
<p>Binary images of Raman microscopy: (<b>A</b>) JMs and (<b>B</b>) HMEs. Dispersiveness analysis of Raman microscopy: (<b>C</b>) ITZ dispersibility of JMs and HMEs, and (<b>D</b>) LEU dispersibility of JMs and HMEs (mean ± SD, n = 3). <sup>††††</sup> ANOVA, <span class="html-italic">p</span>-value &lt; 0.0001 compared with JM-L0.1; <sup>#</sup> ANOVA, <span class="html-italic">p</span>-value &lt; 0.05 compared with JM-L10; *** ANOVA, <span class="html-italic">p</span>-value &lt; 0.05 compared with JM-L10.</p> Full article ">
17 pages, 2526 KiB  
Article
Geometry-Driven Fabrication of Mini-Tablets via 3D Printing: Correlating Release Kinetics with Polyhedral Shapes
by Young-Jin Kim, Yu-Rim Choi, Ji-Hyun Kang, Yun-Sang Park, Dong-Wook Kim and Chun-Woong Park
Pharmaceutics 2024, 16(6), 783; https://doi.org/10.3390/pharmaceutics16060783 - 8 Jun 2024
Cited by 1 | Viewed by 1232
Abstract
The aim of this study was to fabricate mini-tablets of polyhedrons containing theophylline using a fused deposition modeling (FDM) 3D printer, and to evaluate the correlation between release kinetics models and their geometric shapes. The filaments containing theophylline, hydroxypropyl cellulose (HPC), and EUDRAGIT [...] Read more.
The aim of this study was to fabricate mini-tablets of polyhedrons containing theophylline using a fused deposition modeling (FDM) 3D printer, and to evaluate the correlation between release kinetics models and their geometric shapes. The filaments containing theophylline, hydroxypropyl cellulose (HPC), and EUDRAGIT RS PO (EU) could be obtained with a consistent thickness through pre-drying before hot melt extrusion (HME). Mini-tablets of polyhedrons ranging from tetrahedron to icosahedron were 3D-printed using the same formulation of the filament, ensuring equal volumes. The release kinetics models derived from dissolution tests of the polyhedrons, along with calculations for various physical parameters (edge, SA: surface area, SA/W: surface area/weight, SA/V: surface area/volume), revealed that the correlation between the Higuchi model and the SA/V was the highest (R2 = 0.995). It was confirmed that using 3D- printing for the development of personalized or pediatric drug products allows for the adjustment of drug dosage by modifying the size or shape of the drug while maintaining or controlling the same release profile. Full article
(This article belongs to the Special Issue Pharmaceutical Applications of 3D Printing)
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<p>Photographs of filament being extruded from the HME (<b>A</b>,<b>B</b>), SEM images of the cross-section of extruded filament (<b>C</b>,<b>D</b>), and sequential filament thickness of the long and short axes, measured at intervals of 20 mm. ((<b>A</b>,<b>C</b>,<b>E</b>) non-pre-drying, (<b>B</b>,<b>D</b>,<b>F</b>) pre-drying).</p> Full article ">Figure 2
<p>Drug release profiles of the TP filaments in phosphate buffer (pH 6.0). Each of the values represents the mean ± standard deviation (<span class="html-italic">n</span> = 3).</p> Full article ">Figure 3
<p>(<b>A</b>) DSC thermal curves, (<b>B</b>) PXRD curves, and (<b>C</b>) FT-IR spectra of HPC, EU, stearic acid, theophylline, PM, filaments, and mini-tablets.</p> Full article ">Figure 4
<p>Morphology images of the 3D-printed polyhedron mini-tablets (from left: tetrahedron, hexahedron, octahedron, dodecahedron, icosahedron). (<b>A</b>) SEM images and (<b>B</b>) Micro-CT images.</p> Full article ">Figure 5
<p>Photographs of 3D-printed mini-tablets by dissolution times. (From top: tetrahedron, hexahedron, octahedron, dodecahedron, and icosahedron).</p> Full article ">Figure 6
<p>(<b>A</b>) Drug release profiles and (<b>B</b>) Higuchi (square root) kinetics profiles of the polyhedrons in phosphate buffer (pH 6.0).</p> Full article ">Figure 7
<p>Various physical parameters (edge, SA, SA/V, SA/W) of polyhedrons fitted to the release constant (<span class="html-italic">K<sub>H</sub></span>) of the Higuchi model.</p> Full article ">
27 pages, 6940 KiB  
Article
Evidence That a Peptide-Drug/p53 Gene Complex Promotes Cognate Gene Expression and Inhibits the Viability of Glioblastoma Cells
by Ana Neves, Tânia Albuquerque, Rúben Faria, Cecília R. A. Santos, Eric Vivès, Prisca Boisguérin, Diana Carneiro, Daniel F. Bruno, Maria D. Pavlaki, Susana Loureiro, Ângela Sousa and Diana Costa
Pharmaceutics 2024, 16(6), 781; https://doi.org/10.3390/pharmaceutics16060781 - 8 Jun 2024
Viewed by 1289
Abstract
Glioblastoma multiform (GBM) is considered the deadliest brain cancer. Conventional therapies are followed by poor patient survival outcomes, so novel and more efficacious therapeutic strategies are imperative to tackle this scourge. Gene therapy has emerged as an exciting and innovative tool in cancer [...] Read more.
Glioblastoma multiform (GBM) is considered the deadliest brain cancer. Conventional therapies are followed by poor patient survival outcomes, so novel and more efficacious therapeutic strategies are imperative to tackle this scourge. Gene therapy has emerged as an exciting and innovative tool in cancer therapy. Its combination with chemotherapy has significantly improved therapeutic outcomes. In line with this, our team has developed temozolomide–transferrin (Tf) peptide (WRAP5)/p53 gene nanometric complexes that were revealed to be biocompatible with non-cancerous cells and in a zebrafish model and were able to efficiently target and internalize into SNB19 and U373 glioma cell lines. The transfection of these cells, mediated by the formulated peptide-drug/gene complexes, resulted in p53 expression. The combined action of the anticancer drug with p53 supplementation in cancer cells enhances cytotoxicity, which was correlated to apoptosis activation through quantification of caspase-3 activity. In addition, increased caspase-9 levels revealed that the intrinsic or mitochondrial pathway of apoptosis was implicated. This assumption was further evidenced by the presence, in glioma cells, of Bax protein overexpression—a core regulator of this apoptotic pathway. Our findings demonstrated the great potential of peptide TMZ/p53 co-delivery complexes for cellular transfection, p53 expression, and apoptosis induction, holding promising therapeutic value toward glioblastoma. Full article
(This article belongs to the Special Issue Recent Advances in Nanotechnology Therapeutics)
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<p>Cellular viability of HA1800 cells after 24 h, 48 h, and 72 h of transfection mediated by the different peptide/pDNA complexes developed at N/P ratio of 1 (using 1 µg pDNA). Cells not transfected were used as a positive control and cells treated with naked pDNA and TMZ drug were used as controls. Data were obtained from six independent measurements (mean ± SD, n = 6).</p> Full article ">Figure 2
<p>Cellular viability of CTX-TNA2 cells after 24 h, 48 h, and 72 h of transfection mediated by the different peptide/pDNA complexes developed at N/P ratio of 1 (using 1 µg pDNA). Cells not transfected were used as a positive control and cells treated with naked pDNA and TMZ drug were used as controls. Data were obtained from six independent measurements (mean ± SD, n = 6).</p> Full article ">Figure 3
<p>(<b>A</b>) Effects of TMZ drug in <span class="html-italic">Danio rerio</span> zebrafish embryo–larval hatching success, malformations, and survival over time after exposure to a range of concentrations of 0–5.15 µM of TMZ drug. (<b>B</b>) Larval stereomicroscope images of control (<b>a</b>), DMSO solvent (<b>b</b>), and concentration of 1 mg TMZ drug/L (<b>c</b>,<b>d</b>) after 96 h of exposure. Scale bar = 1000 μm. ns—non-significant (<span class="html-italic">p</span> &gt; 0.5).</p> Full article ">Figure 4
<p>(<b>A</b>) Effects of plasmid DNA (pDNA) in <span class="html-italic">Danio rerio</span> zebrafish embryo–larval hatching success, malformations, and survival over time after exposure to a range of concentrations of 0–1 mg of pDNA/L. (<b>B</b>) Larval stereomicroscope images of control (<b>a</b>), and concentration of 1 mg pDNA/L (<b>b</b>,<b>c</b>) after 96 h of exposure. Scale bar = 1000 μm. ns—non-significant (<span class="html-italic">p</span> &gt; 0.5).</p> Full article ">Figure 5
<p>(<b>A</b>) Effects of TMZ/WRAP5 peptide in <span class="html-italic">Danio rerio</span> zebrafish embryo–larval hatching success, malformations, and survival over time after exposure to a range of concentrations of 0–6.5 mg of TMZ/WRAP5 peptide/L. (<b>B</b>) Larval stereomicroscope images of control (<b>a</b>), and concentration of 6.5 mg TMZ/WRAP5 peptide/L (<b>b</b>,<b>c</b>) after 96 h of exposure. Scale bar = 1000 μm. ns—non-significant (<span class="html-italic">p</span> &gt; 0.5).</p> Full article ">Figure 6
<p>(<b>A</b>) Effects of TMZ/Tf-WRAP5 peptide in <span class="html-italic">Danio rerio</span> zebrafish embryo–larval hatching success, malformations, and survival in time after exposure to a range of concentrations of 0–2 mg of TMZ/Tf-WRAP5 peptide/L. (<b>B</b>) Larval stereomicroscope images of control (<b>a</b>), and concentration of 2 mg TMZ/Tf-WRAP5 peptide/L (<b>b</b>,<b>c</b>) after 96 h of exposure. Scale bar = 1000 μm. ns—non-significant (<span class="html-italic">p</span> &gt; 0.5).</p> Full article ">Figure 7
<p>(<b>A</b>) Effects of Tf-WRAP5/pDNA at N/P ratio of 1 in <span class="html-italic">Danio rerio</span> zebrafish embryo–larval hatching success, malformations, and survival over time after exposure to a range of concentrations of 0–1 mg of pDNA/L. (<b>B</b>) Larval stereomicroscope images of control (<b>a</b>), and concentration of 2 mg pDNA/L (<b>b</b>,<b>c</b>) after 96 h of exposure. Scale bar = 1000 μm. ns—non-significant (<span class="html-italic">p</span> &gt; 0.5).</p> Full article ">Figure 8
<p>(<b>A</b>) Effects of TMZ/Tf-WRAP5/pDNA at N/P ratio of 1 in <span class="html-italic">Danio rerio</span> zebrafish embryo–larval hatching success, malformations, and survival over time after exposure to a range of concentrations of 0–1 mg of pDNA/L. (<b>B</b>) Larval stereomicroscope images of control (<b>a</b>), and concentration of 1 mg pDNA/L (<b>b</b>,<b>c</b>) after 96 h of exposure. Scale bar = 1000 μm. ns—non-significant (<span class="html-italic">p</span> &gt; 0.5).</p> Full article ">Figure 9
<p>Fluorescence confocal microscopy study. Representative live cell images of SNB19 cells after 4 h of transfection mediated by the different peptide/pDNA complexes developed at N/P ratio of 1 (using 1 µg pDNA). DAPI dye stained the nuclei while FITC labeled pDNA green. Panels (<b>A1</b>–<b>E1</b>) indicate labeled pDNA-FITC, panels (<b>A2</b>–<b>E2</b>) show nuclei stained by DAPI, panels (<b>A3</b>–<b>E3</b>) correspond to merged images, and panels (<b>A4</b>–<b>E4</b>) present a 4× zoom of the highlighted regions. Non-transfected cells (<b>A</b>) and cells transfected with the developed complexes: WRAP5/pDNA-FITC (<b>B</b>), Tf-WRAP5/pDNA-FITC (<b>C</b>), TMZ/WRAP5/pDNA-FITC (<b>D</b>), and TMZ/Tf-WRAP5/pDNA-FITC (<b>E</b>). Scale bar = 50 μm.</p> Full article ">Figure 10
<p>Fluorescence confocal microscopy study. Representative live cell images of U373 cells after 4 h of transfection mediated by the different peptide/pDNA complexes developed at N/P ratio of 1 (using 1 µg pDNA). DAPI dye stained the nuclei while FITC labeled pDNA green. Panels (<b>A1</b>–<b>E1</b>) indicate labeled pDNA-FITC, panels (<b>A2</b>–<b>E2</b>) show nuclei stained by DAPI, panels (<b>A3</b>–<b>E3</b>) correspond to merged images, and panels (<b>A4</b>–<b>E4</b>) present a 4× zoom of the highlighted regions. Non-transfected cells (<b>A</b>) and cells transfected with the developed complexes: WRAP5/pDNA-FITC (<b>B</b>), Tf-WRAP5/pDNA-FITC (<b>C</b>), TMZ/WRAP5/pDNA-FITC (<b>D</b>), and TMZ/Tf-WRAP5/pDNA-FITC (<b>E</b>). Scale bar = 50 μm.</p> Full article ">Figure 11
<p>p53 protein levels in SNB19 and U373 cells after 48 h of transfection mediated by the different peptide/pDNA complexes developed at N/P ratio of 1 (using 1 µg pDNA). Data were obtained from six independent measurements (mean ± SD, n = 6) and analyzed by one-way ANOVA, followed by the Bonferroni test. **** <span class="html-italic">p</span> &lt; 0.001; ** <span class="html-italic">p</span> &lt; 0.021; ns—non-significant (<span class="html-italic">p</span> &gt; 0.5).</p> Full article ">Figure 12
<p>Cellular viability of SNB19 and U373 cells after 24 and 48 h of transfection mediated by the different peptide/pDNA complexes developed at N/P ratio of 1 (using 1 µg pDNA). Cells not transfected were used as a positive control and cells treated with naked pDNA and TMZ drug were used as controls. Data were obtained from six independent measurements (mean ± SD, n = 6) and analyzed by one-way ANOVA, followed by the Bonferroni test. The dotted line represents the limit accepted for a substance to be considered as non-cytotoxic. **** <span class="html-italic">p</span> &lt; 0.001; ns—non-significant (<span class="html-italic">p</span> &gt; 0.5).</p> Full article ">Figure 13
<p>Caspase-3 (above) and caspase-9 (below) activity in U87, SNB19, and U373 cells after transfection (48 h) mediated by the different peptide/pDNA complexes developed at N/P ratio of 1 (using 1 µg pDNA). Positive control refers to the study with staurosporine (1 μM), while negative control considers untreated cells. Data were obtained from three independent measurements (mean ± SD, n = 3) and analyzed by one-way ANOVA, followed by the Bonferroni test. **** <span class="html-italic">p</span> &lt; 0.001; ** <span class="html-italic">p</span> &lt; 0.021.</p> Full article ">
15 pages, 4235 KiB  
Article
Intranasal Delivery of Cell-Penetrating Therapeutic Peptide Enhances Brain Delivery, Reduces Inflammation, and Improves Neurologic Function in Moderate Traumatic Brain Injury
by Yaswanthi Yanamadala, Ritika Roy, Afrika Alake Williams, Navya Uppu, Audrey Yoonsun Kim, Mark A. DeCoster, Paul Kim and Teresa Ann Murray
Pharmaceutics 2024, 16(6), 774; https://doi.org/10.3390/pharmaceutics16060774 - 7 Jun 2024
Cited by 1 | Viewed by 1433
Abstract
Following traumatic brain injury (TBI), secondary brain damage due to chronic inflammation is the most predominant cause of the delayed onset of mood and memory disorders. Currently no therapeutic approach is available to effectively mitigate secondary brain injury after TBI. One reason is [...] Read more.
Following traumatic brain injury (TBI), secondary brain damage due to chronic inflammation is the most predominant cause of the delayed onset of mood and memory disorders. Currently no therapeutic approach is available to effectively mitigate secondary brain injury after TBI. One reason is the blood–brain barrier (BBB), which prevents the passage of most therapeutic agents into the brain. Peptides have been among the leading candidates for CNS therapy due to their low immunogenicity and toxicity, bioavailability, and ease of modification. In this study, we demonstrated that non-invasive intranasal (IN) administration of KAFAK, a cell penetrating anti-inflammatory peptide, traversed the BBB in a murine model of diffuse, moderate TBI. Notably, KAFAK treatment reduced the production of proinflammatory cytokines that contribute to secondary injury. Furthermore, behavioral tests showed improved or restored neurological, memory, and locomotor performance after TBI in KAFAK-treated mice. This study demonstrates KAFAK’s ability to cross the blood–brain barrier, to lower proinflammatory cytokines in vivo, and to restore function after a moderate TBI. Full article
(This article belongs to the Special Issue Advances in Delivering Protein and Peptide Therapeutics, 2nd Edition)
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<p>Concentration-dependent uptake of fluorescently labeled peptides. Various concentrations of each peptide were incubated with BMVECs (10,000 per well) for 4 h with 20, 30, 50, and 75 µM of FITC-KAFAK (K20–K75), FITC-AIP-1 (A20–A75), and FITC-L57-AIP-1 (L20–L75). All three peptides exhibited concentration-dependent cellular uptake. However, intensity of internalized FITC-AIP-1 was significantly lower for all concentrations (<span class="html-italic">p</span> &lt; 0.01). Even the highest concentration of FITC-AIP-1 had a significantly lower fluorescence intensity than the lowest concentration of KAFAK (A75 vs. K20, <span class="html-italic">p</span> &lt; 0.01). While the intensity for FITC-L57-AIP-1 was higher than FITC-AIP-1 for each concentration level (<span class="html-italic">p</span> &lt; 0.01), it was also significantly lower than concentration of FITC-KAFAK at each concentration level (<span class="html-italic">p</span> &lt; 0.01). However, 75 µM FITC-L57-AIP-1 produced about the same fluorescence as 20 µM of KAFAK (downward arrows in figure). A total of 120 cells (<span class="html-italic">n</span> = 6 wells, 20 cells per well) were analyzed per concentration of each peptide using Image J software. Mean fluorescence intensity was normalized to that of control cells with no peptide (autofluorescence). Normalized fluorescence intensity was compared using ANOVA with a Bonferroni correction for multiple comparisons. (Data are represented as the mean ± 2 standard deviations).</p> Full article ">Figure 2
<p>The cell viability of BMVECs was measured using an ATP assay after 24 h of incubation with different peptide concentrations. FITC-AIP-1 (AIP) and FITC-L57-AIP-1 (L57) did not elicit cytotoxicity between 10–500 µM. KAFAK treatment from 10 to 100 µM did not significantly affect cell viability, and reduced cell viability was observed at 250 and 500 µM. Data are expressed as % of untreated control (mean ± SD, n = 3 wells/concentration of each peptide * <span class="html-italic">p</span> &lt; 0.05).</p> Full article ">Figure 3
<p>Intranasal delivery of KAFAK to the brain. Mice were treated with 500 µM RITC-KAFAK. They were perfused four hours after treatment and brain sections were stained with DAPI to visualize cell nuclei. Images (<b>A</b>–<b>C</b>) are of a representative section showing that RITC-KAFAK permeated the olfactory bulb. Images (<b>D</b>–<b>F</b>) are of a representative section from the cerebral cortex. Images in the first column show fluorescently labeled KAFAK in the olfactory bulb (<b>A</b>) and cortex (<b>D</b>), with DAPI counterstaining shown in the second column (<b>B</b>,<b>E</b>). The KAFAK and DAPI images are merged in the third column (<b>C</b>,<b>F</b>). Scale bars are 100 µm.</p> Full article ">Figure 4
<p>Mean normalized rotarod performance on day 2, 5, and 7 for TBI-vehicle (TBI), Sham-vehicle (Sham), and TBI-KAFAK (KAFAK) groups. (<b>A</b>) shows the performance for all mice, while (<b>B</b>,<b>C</b>) represent the performance of female and male mice, respectively. (Mean ± SD, n = 8 for each group in (<b>A</b>) and n = 4 for each group in (<b>B</b>,<b>C</b>); * <span class="html-italic">p</span> &lt; 0.05 Sham versus KAFAK and TBI, ** <span class="html-italic">p</span> &lt; 0.01 Sham versus KAFAK and TBI, # <span class="html-italic">p</span> &lt; 0.05 TBI versus KAFAK, ## <span class="html-italic">p</span> &lt; 0.01 TBI versus KAFAK, †† <span class="html-italic">p</span> &lt; 0.01 Sham versus TBI).</p> Full article ">Figure 5
<p>Mean mNSS for Sham-vehicle (Sham), TBI-KAFAK treated (KAFAK), and TBI-vehicle treated (TBI) groups on day 2, 5, and 7 (days after injury). Plots show mNSS results for (<b>A</b>) all mice, (<b>B</b>) female mice, and (<b>C</b>) male mice. (Data are expressed as the mean ± SD, n = 8 for each group in (<b>A</b>) and n = 4 for each group in (<b>B</b>,<b>C</b>); ** <span class="html-italic">p</span> &lt; 0.01 Sham versus KAFAK and TBI, # <span class="html-italic">p</span> &lt; 0.05 TBI versus KAFAK, ## <span class="html-italic">p</span> &lt; 0.01 TBI versus KAFAK, † <span class="html-italic">p</span> &lt; 0.05 Sham versus TBI, †† <span class="html-italic">p</span> &lt; 0.01 Sham versus TBI).</p> Full article ">Figure 6
<p>KAFAK treatment rescued memory deficits associated with a moderate TBI. The mean discrimination index of the novel object recognition test for the TBI-vehicle group (TBI) was significantly lower than both Sham-vehicle (Sham) and TBI-KAFAK treated (KAFAK) groups seven days after injury (n = 8/group, mean of the discrimination index ± 2SD, ** <span class="html-italic">p</span> &lt; 0.01 for TBI versus KAFAK and for TBI versus Sham).</p> Full article ">Figure 7
<p>IN administration of KAFAK (KAFAK IN) after TBI reduced levels of key proinflammatory cytokines, TNF, IL-1β, and IL-6, to sham levels. ELISA results show a significant increase in all three cytokines in injured, vehicle-treated mice (TBI-vehicle) versus sham-injured, vehicle treated mice (Sham). A significant reduction in TNF and IL-1β was observed in the KAFAK IN group versus the TBI-vehicle group. In contrast, no difference was observed between the TBI-vehicle group and the group that received an intraperitoneal delivery of KAFAK (KAFAK IP). (Data shown represent mean pg/mg tissue ± 2SD, n = 5/group, * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01).</p> Full article ">
21 pages, 1087 KiB  
Article
Predicting ADMET Properties from Molecule SMILE: A Bottom-Up Approach Using Attention-Based Graph Neural Networks
by Alessandro De Carlo, Davide Ronchi, Marco Piastra, Elena Maria Tosca and Paolo Magni
Pharmaceutics 2024, 16(6), 776; https://doi.org/10.3390/pharmaceutics16060776 - 7 Jun 2024
Cited by 2 | Viewed by 1332
Abstract
Understanding the pharmacokinetics, safety and efficacy of candidate drugs is crucial for their success. One key aspect is the characterization of absorption, distribution, metabolism, excretion and toxicity (ADMET) properties, which require early assessment in the drug discovery and development process. This study aims [...] Read more.
Understanding the pharmacokinetics, safety and efficacy of candidate drugs is crucial for their success. One key aspect is the characterization of absorption, distribution, metabolism, excretion and toxicity (ADMET) properties, which require early assessment in the drug discovery and development process. This study aims to present an innovative approach for predicting ADMET properties using attention-based graph neural networks (GNNs). The model utilizes a graph-based representation of molecules directly derived from Simplified Molecular Input Line Entry System (SMILE) notation. Information is processed sequentially, from substructures to the whole molecule, employing a bottom-up approach. The developed GNN is tested and compared with existing approaches using six benchmark datasets and by encompassing regression (lipophilicity and aqueous solubility) and classification (CYP2C9, CYP2C19, CYP2D6 and CYP3A4 inhibition) tasks. Results show the effectiveness of our model, which bypasses the computationally expensive retrieval and selection of molecular descriptors. This approach provides a valuable tool for high-throughput screening, facilitating early assessment of ADMET properties and enhancing the likelihood of drug success in the development pipeline. Full article
(This article belongs to the Section Pharmacokinetics and Pharmacodynamics)
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<p>Example of how adjacency matrices are extracted from molecular SMILE. For each type of bond (i.e., single, double, triple or aromatic), a specific adjacency matrix is derived in order to focus on molecular substructures.</p> Full article ">Figure 2
<p>Schematic representation of the GNN adopted. The architecture is organized as a stack of three main modules, each with a specific function.</p> Full article ">Figure 3
<p>Schematic representation of the implemented five-fold cross validation. At each step, one fold (orange) is used as an external test set; the remaining four are used for training and validation. And <math display="inline"><semantics> <mrow> <mn>20</mn> <mo>%</mo> </mrow> </semantics></math> of the four folds are used as validation data.</p> Full article ">Figure 4
<p>Distributions of regression variables in two benchmark datasets. Histograms of Lipophilicity AZ panel (<b>A</b>) and AqSolDB panel (<b>B</b>) data.</p> Full article ">Figure 5
<p>Example of the weighting strategy adopted for both regression tasks. Panel (<b>A</b>) shows the weights introduced for training the GNN on LogD prediction. Panel (<b>B</b>) focuses on LogS. For both tasks, <math display="inline"><semantics> <mi>α</mi> </semantics></math> was set to 0.55.</p> Full article ">Figure 6
<p>Models used in the ablation study to benchmark the implemented GNN architecture. Panel (<b>A</b>) illustrates the ‘Whole Molecule’ GNN, which does not consider molecular substructures. Panel (<b>B</b>) represents the ‘Convolutional’ GNN, in which the attention mechanism for the entire molecule is replaced by a graph convolutional (GC) layer.</p> Full article ">Figure 7
<p>Results of the ablation study on the regression tasks.</p> Full article ">Figure 8
<p>Results of the ablation study on the classification tasks.</p> Full article ">
26 pages, 1438 KiB  
Review
Design of Chitin Cell Culture Matrices for 3D Tissue Engineering: The Importance of Chitin Types, Solvents, Cross-Linkers, and Fabrication Techniques
by Turna Basak and Julia L. Shamshina
Pharmaceutics 2024, 16(6), 777; https://doi.org/10.3390/pharmaceutics16060777 - 7 Jun 2024
Cited by 3 | Viewed by 1432
Abstract
This review focuses on factors and the fabrication techniques affecting the microarchitecture of tissue engineering scaffolds from the second most abundant biopolymer, chitin. It emphasizes the unique potentiality of this polymer in tissue engineering (TE) applications and highlights the variables important to achieve [...] Read more.
This review focuses on factors and the fabrication techniques affecting the microarchitecture of tissue engineering scaffolds from the second most abundant biopolymer, chitin. It emphasizes the unique potentiality of this polymer in tissue engineering (TE) applications and highlights the variables important to achieve tailored scaffold properties. First, we describe aspects of scaffolds’ design, and the complex interplay between chitin types, solvent systems, additives, and fabrication techniques to incorporate porosity, with regard to best practices. In the following section, we provide examples of scaffolds’ use, with a focus on in vitro cell studies. Finally, an analysis of their biodegradability is presented. Our review emphasizes the potentiality of chitin and the pressing need for further research to overcome existing challenges and fully harness its capabilities in tissue engineering. Full article
(This article belongs to the Section Pharmaceutical Technology, Manufacturing and Devices)
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<p>Structures of chitin (<b>a</b>), chitosan (<b>b</b>), and glucosamine glucan (<b>c</b>).</p> Full article ">Figure 2
<p>(<b>a</b>) Molecular models of the crystal structure of <span class="html-italic">α</span>-chitin (<b>left</b>) and <span class="html-italic">β</span>-chitin (<b>right</b>) showing the antiparallel and parallel chain directionality. Reprinted with permission from ref [<a href="#B38-pharmaceutics-16-00777" class="html-bibr">38</a>]. Copyright © 2021 The Royal Society Publishing; (<b>b</b>) Structure of chitin vs. chitosan; (<b>c</b>) Pattern of acetylation (PA): the sequence of <span class="html-italic">β</span>-1,4-linked glucosamine (deacetylated) and <span class="html-italic">N</span>-acetylglucosamine (acetylated) units. Reprinted with permission from ref. [<a href="#B41-pharmaceutics-16-00777" class="html-bibr">41</a>]. Copyright © 2020 American Chemical Society; (<b>d</b>) Schematic representation of high- and low-molecular-weight (Mw) chitin [<a href="#B42-pharmaceutics-16-00777" class="html-bibr">42</a>,<a href="#B43-pharmaceutics-16-00777" class="html-bibr">43</a>,<a href="#B44-pharmaceutics-16-00777" class="html-bibr">44</a>].</p> Full article ">Figure 3
<p>Hydrogen bonding network in α-chitin.</p> Full article ">
14 pages, 2264 KiB  
Article
Heparin Dosing Regimen Optimization in Veno-Arterial Extracorporeal Membrane Oxygenation: A Pharmacokinetic Analysis
by Julien Lanoiselée, Jérémy Mourer, Marie Jungling, Serge Molliex, Lise Thellier, Julien Tabareau, Emmanuelle Jeanpierre, Emmanuel Robin, Sophie Susen, Benoit Tavernier, André Vincentelli, Edouard Ollier and Mouhamed Djahoum Moussa
Pharmaceutics 2024, 16(6), 770; https://doi.org/10.3390/pharmaceutics16060770 - 6 Jun 2024
Viewed by 971
Abstract
Background. Unfractionated heparin is administered in patients undergoing veno-arterial extracorporeal membrane oxygenation (VA-ECMO). Anticoagulation monitoring is recommended, with an anti-activated factor X (anti-Xa) targeting 0.3 to 0.7 IU/mL. Owing to heparin’s heterogeneous pharmacokinetic properties, anti-Xa is unpredictable, generating a challenge in anticoagulation practices. [...] Read more.
Background. Unfractionated heparin is administered in patients undergoing veno-arterial extracorporeal membrane oxygenation (VA-ECMO). Anticoagulation monitoring is recommended, with an anti-activated factor X (anti-Xa) targeting 0.3 to 0.7 IU/mL. Owing to heparin’s heterogeneous pharmacokinetic properties, anti-Xa is unpredictable, generating a challenge in anticoagulation practices. The aim of this study was to build a pharmacokinetic model of heparin accounting for potential confounders, and derive an optimized dosing regimen for a given anti-Xa target. Methods. Adult patients undergoing VA-ECMO were included between January 2020 and June 2021. Anticoagulation was managed with an initial 100 IU/kg heparin loading dose followed by a continuous infusion targeting 0.2 to 0.7 IU/mL anti-Xa. The data were split into model development and model validation cohorts. Statistical analysis was performed using a nonlinear mixed effects modeling population approach. Model-based simulations were performed to develop an optimized dosing regimen targeting the desired anti-Xa. Results. A total of 74 patients were included, with 1703 anti-Xa observations. A single-compartment model best fitted the data. Interpatient variability for distribution volume was best explained by body weight, C-reactive protein and ECMO indication (post-cardiotomy shock or medical cardiogenic shock), and interpatient variability for elimination clearance was best explained by serum creatinine and C-reactive protein. Simulations using the optimized regimen according to these covariates showed accurate anti-Xa target attainment. Conclusion. In adult patients on VA-ECMO, heparin’s effect increased with serum creatinine and medical indication, whereas it decreased with body weight and systemic inflammation. We propose an optimized dosing regimen accounting for key covariates, capable of accurately predicting a given anti-Xa target. Full article
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<p>Goodness of fit plots for the development cohort. (<b>Top panels</b>): observations versus predictions. The black line represents the identity line. Blue circles represent the observed anti-Xa versus the corresponding predicted anti-Xa. Red circles represent the censored data. The yellow line represents the trend line. (<b>Left panel</b>): plot of the observed anti-Xa (IU/mL) versus population predicted (no random component). (<b>Right panel</b>): plot of the observed anti-Xa versus individual predicted anti-Xa (with random component). (<b>Bottom panels</b>): NPDE versus time and population predictions. The black line represents the identity line. NPDE, normalized prediction distribution errors.</p> Full article ">Figure 2
<p>Optimized dosing regimen estimated to reach a 0.5 IU/mL target anti-Xa. Loading dose depended on body weight, CRP and ECMO indication ((<b>top row</b>): medical indication, (<b>middle row</b>): post-cardiotomy indication). Continuous IV (intravenous) maintenance dose (<b>bottom row</b>) depended on serum creatinine and CRP. (<b>Left column</b>): CRP 5 mg/L. (<b>Middle column</b>): CRP 100 mg/L. (<b>Right column</b>): CRP 400 mg/L. Red line represents the median dose to reach the target. Dark, average and light blue shaded areas correspond to the interpatient variability intervals estimated in our model (50%, 70% and 90%, respectively). The loading dose increased with increasing body weight and CRP, and with post-cardiotomy indication. The maintenance dose decreased with renal impairment and decreasing CRP.</p> Full article ">Figure 3
<p>Simulations of the anti-Xa time courses using the final PK model with focusing on CRP. Simulations were performed for a medical patient weighing 80 kg with a serum creatinine of 115 µmol/L. Red line: CRP 5 mg/L. Green line: CRP 100 mg/L. Blue line: CRP 400 mg/L. The black lines correspond to the recommended 0.3–0.7 IU/mL target anti-Xa interval. Dark, average and light blue shaded areas correspond to the interpatient variability intervals estimated in our model (50%, 70% and 90%, respectively). (<b>Top panels</b>): simulations according to an 8000 IU (100 IU/kg) loading dose immediately followed by a 1200 IU/h (15 IU/kg/h) without adaptation to body weight or renal function. (<b>Bottom panels</b>): simulations according to our optimized dosing regimen. (<b>Bottom left panel</b>): 2500 IU loading dose followed by a 750 IU/h maintenance dose. (<b>Bottom middle panel</b>): 4000 IU loading dose followed by a 1500 IU/h maintenance dose. (<b>Bottom right panel</b>): 5500 IU loading dose followed by a 2200 IU/h maintenance dose.</p> Full article ">
23 pages, 19779 KiB  
Article
Optimizing mRNA-Loaded Lipid Nanoparticles as a Potential Tool for Protein-Replacement Therapy
by Rocío Gambaro, Ignacio Rivero Berti, María José Limeres, Cristián Huck-Iriart, Malin Svensson, Silvia Fraude, Leah Pretsch, Shutian Si, Ingo Lieberwirth, Stephan Gehring, Maximiliano Cacicedo and Germán Abel Islan
Pharmaceutics 2024, 16(6), 771; https://doi.org/10.3390/pharmaceutics16060771 - 6 Jun 2024
Cited by 1 | Viewed by 3606
Abstract
Lipid nanoparticles (LNPs) tailored for mRNA delivery were optimized to serve as a platform for treating metabolic diseases. Four distinct lipid mixes (LMs) were formulated by modifying various components: LM1 (ALC-0315/DSPC/Cholesterol/ALC-0159), LM2 (ALC-0315/DOPE/Cholesterol/ALC-0159), LM3 (ALC-0315/DSPC/Cholesterol/DMG-PEG2k), and LM4 (DLin-MC3-DMA/DSPC/Cholesterol/ALC-0159). LNPs exhibited stability and homogeneity [...] Read more.
Lipid nanoparticles (LNPs) tailored for mRNA delivery were optimized to serve as a platform for treating metabolic diseases. Four distinct lipid mixes (LMs) were formulated by modifying various components: LM1 (ALC-0315/DSPC/Cholesterol/ALC-0159), LM2 (ALC-0315/DOPE/Cholesterol/ALC-0159), LM3 (ALC-0315/DSPC/Cholesterol/DMG-PEG2k), and LM4 (DLin-MC3-DMA/DSPC/Cholesterol/ALC-0159). LNPs exhibited stability and homogeneity with a mean size of 75 to 90 nm, confirmed by cryo-TEM and SAXS studies. High mRNA encapsulation (95–100%) was achieved. LNPs effectively delivered EGFP-encoding mRNA to HepG2 and DC2.4 cell lines. LNPs induced cytokine secretion from human peripheral blood mononuclear cells (PBMCs), revealing that LM1, LM2, and LM4 induced 1.5- to 4-fold increases in IL-8, TNF-α, and MCP-1 levels, while LM3 showed minimal changes. Reporter mRNA expression was observed in LNP-treated PBMCs. Hemotoxicity studies confirmed formulation biocompatibility with values below 2%. In vivo biodistribution in mice post intramuscular injection showed significant mRNA expression, mainly in the liver. The modification of LNP components influenced reactogenicity, inflammatory response, and mRNA expression, offering a promising platform for selecting less reactogenic carriers suitable for repetitive dosing in metabolic disease treatment. Full article
(This article belongs to the Special Issue Advances in Nanotechnology-Based Drug Delivery Systems)
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<p>Scheme of the different LNPs from different lipid mixes (LMs) and the modification of each component (created with Biorender). The replacement of each component of LM1 was highlighted with different colors: LM2 (DSPC for DOPE in violet), LM3 (ALC 0159 for DMG-PE-2000 in orange) and LM4 (ALC 0315 for Dlin-MC3-DMA in red).</p> Full article ">Figure 2
<p>Distribution of the main LNP components by molar fraction: GenVoy, commercial lipid mix provided by Precision NanoSystems, Onpattro (patisiran), BNT162b2 (Comirnaty) Pfizer–BioNTech COVID vaccine, mRNA-1273 Moderna COVID vaccine, CVnCoV Curevac COVID vaccine candidate, and lipid mixes presented in this manuscript (LM 1 to 4).</p> Full article ">Figure 3
<p>Mean size (<b>a</b>), polydispersity index (PDI) (<b>b</b>), zeta potential (<b>c</b>), and mRNA encapsulation efficiency (EE) (<b>d</b>) of the different LNP formulations. Comparison with a commercial formulation (GenVoy) is shown. The graphs represent the mean (n = 3) ± SD.</p> Full article ">Figure 4
<p>CryoTEM images of the different LNP formulations. Arrows indicate the presence of particular LNP characteristics and irregularities in their structure: “blebs” (green arrows) and internal defects (blue arrows).</p> Full article ">Figure 5
<p>SAXS profiles of the different LNP formulations. The <b>left</b> column shows the Log plot of the experimental SAXS pattern in (symbol) and the fitted curve in continuous line. The <b>right</b> column shows the electron density profile obtained from the bilayer model.</p> Full article ">Figure 6
<p>Stability of the formulations after storage at 4 °C for 1 month. The mean size (<b>a</b>), polydispersity (PDI) (<b>b</b>), zeta potential (<b>c</b>), and mRNA encapsulation efficiency (EE) (<b>d</b>) were measured every week. The graphs represent the mean (n = 3) ± SD.</p> Full article ">Figure 7
<p>HepG2 cells transfection by the different LM formulations. The percentage of EGFP+ cells (<b>a</b>), the mean fluorescence intensity (MFI), (<b>b</b>) and fluorescence microscopy images (<b>c</b>) were analyzed. The transfection was studied and quantified by the presence of EGFP+ cells (by fluorescence microscopy and flow cytometry). The graphs represent the mean (n = 3) ± SD. Abbreviations: C−: untreated cells; C+: cells treated with <span class="html-italic">EGFP</span> mRNA loaded into lipofectamine; GV: commercial formulation; LM: own lipid mixes.</p> Full article ">Figure 8
<p>DC 2.4 mouse dendritic cell line transfection by the different LM formulations. The EGFP+ cells (<b>a</b>), the mean EGFP fluorescence (<b>b</b>), and the cell viability by 7-AAD (<b>c</b>) were determined by flow cytometry. The release of TNF-α by LNP-stimulated DC was measured by CBA (<b>d</b>). One-way ANOVA following Fisher’s LSD test was used to compare among groups. ns: not significant, <span class="html-italic">p</span> &lt; 0.05 (*) and <span class="html-italic">p</span> &lt; 0.0001 (****). MFI: mean fluorescence intensity. Abbreviations: C−: untreated cells; GV: commercial formulation; LM: own lipid mixes.</p> Full article ">Figure 9
<p>PBMC transfection by the different LM formulations. The mRNA transfection was studied by two techniques: the expression of <span class="html-italic">EGFP</span> mRNA detected by flow cytometry (<b>a</b>,<b>b</b>) and the expression of <span class="html-italic">Luc</span> mRNA quantified using a Luciferase kit (<b>c</b>). PBMCs were transfected with 8 µg/mL of mRNA and in the presence of ApoE (1 µg/mL). The graphs represent the mean (n = 5) ± SD. One-way ANOVA following Fisher’s LSD test was used to compare among groups. <span class="html-italic">p</span> &lt; 0.05 (*), <span class="html-italic">p</span> &lt; 0.01 (**), <span class="html-italic">p</span> &lt; 0.001 (***), and <span class="html-italic">p</span> &lt; 0.0001 (****). MFI: mean fluorescence intensity. Abbreviations: C−: untreated cells; LM: own lipid mixes.</p> Full article ">Figure 10
<p>Cytokine secretion of TNF-α, MCP-1, IL-8, IL-1β, IFN-γ, IL-6, IP-10, and IL-4 from LNP-stimulated PBMCs determined by cytometric bead assay. * The results are expressed as folds against the negative control. PBMCs were exposed to LNPs with an equivalent dose of 8 µg/mL of mRNA for 24 h. The graphs represent the mean (n = 5) ± SD. One-way ANOVA following Fisher’s LSD test was used to compare among groups. ns: not significant, <span class="html-italic">p</span> &lt; 0.05 (*), <span class="html-italic">p</span> &lt; 0.01 (**), <span class="html-italic">p</span> &lt; 0.001 (***), and <span class="html-italic">p</span> &lt; 0.0001 (****). Abbreviations: C−: untreated cells; LM: own lipid mixes.</p> Full article ">Figure 11
<p>Activation markers of the CD11c+ population from PBMCs after LNP stimulation with the different formulations, determined by flow cytometry analysis. The graphs represent the mean (n = 3) ± SD. One-way ANOVA following Fisher’s LSD test was used to compare among groups. ns: not significant, <span class="html-italic">p</span> &lt; 0.05 (*) and <span class="html-italic">p</span> &lt; 0.01 (**). Abbreviations: C−: untreated cells; LM: own lipid mixes; MFI: mean fluorescent intensity, folds vs. control (no treated).</p> Full article ">Figure 12
<p>The in vivo biodistribution of the LNPs determined in a C57BL/6-naïve mice model after i.m. injection with the LNP formulations delivering <span class="html-italic">Luc</span> mRNA. The graphs represent the region of interest (ROI) mean n = 5 ± SD, except for LM1-LNP, in which n = 4. One-way ANOVA following Fisher’s LSD test was used to compare among groups. ns: not significant, <span class="html-italic">p</span> &lt; 0.01 (**), <span class="html-italic">p</span> &lt; 0.0001 (****). Abbreviations: PBS: phosphate-buffered-saline-treated mice; LM: mice treated with our own LNP–lipid mixes.</p> Full article ">Figure 13
<p>The in vivo biodistribution of the LNPs at organ level in a C57BL/6-naïve mice model after i.m. injection with the LNP formulations delivering <span class="html-italic">Luc</span> mRNA. The graphs represent the mean with n = 5 ± SD, except for LM1, in which n = 4. One-way ANOVA following Fisher’s LSD test was used to compare among groups. ns: not significant, <span class="html-italic">p</span> &lt; 0.01 (**), <span class="html-italic">p</span> &lt; 0.001 (***), and <span class="html-italic">p</span> &lt; 0.0001 (****).</p> Full article ">
18 pages, 809 KiB  
Review
The Integration of Advanced Drug Delivery Systems into Conventional Adjuvant Therapies for Peri-Implantitis Treatment
by Iria Seoane-Viaño, Mariola Seoane-Gigirey, Carlos Bendicho-Lavilla, Luz M. Gigirey, Francisco J. Otero-Espinar and Santiago Seoane-Trigo
Pharmaceutics 2024, 16(6), 769; https://doi.org/10.3390/pharmaceutics16060769 - 5 Jun 2024
Viewed by 1763
Abstract
Despite the high success rates of dental implants, peri-implantitis is currently the most common complication in dental implantology. Peri-implantitis has an inflammatory nature, it is associated with the accumulation of plaque in the peri-implant tissues, and its evolution can be progressive depending on [...] Read more.
Despite the high success rates of dental implants, peri-implantitis is currently the most common complication in dental implantology. Peri-implantitis has an inflammatory nature, it is associated with the accumulation of plaque in the peri-implant tissues, and its evolution can be progressive depending on various factors, comorbidities, and poor oral health. Prophylaxis and different treatment methods have been widely discussed in recent decades, and surgical and non-surgical techniques present both advantages and disadvantages. In this work, a literature review of different studies on the application of adjuvant treatments, such as local and systemic antibiotics and antiseptic treatments, was conducted. Positive outcomes have been found in the short (up to one year after treatment) and long term (up to ten years after treatment) with combined therapies. However, there is still a need to explore new therapies based on the use of advanced drug delivery systems for the effective treatment of peri-implantitis in the long term and without relapses. Hence, micro- and nanoparticles, implants, and injectable hydrogels, among others, should be considered in future peri-implantitis treatment with the aim of enhancing overall therapy outcomes. Full article
(This article belongs to the Special Issue Advanced Materials Science and Technology in Drug Delivery)
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<p>(<b>A</b>) Peri-implant health, (<b>B</b>) peri-implant mucositis, and (<b>C</b>) peri-implantitis.</p> Full article ">Figure 2
<p>Drug delivery systems for peri-implantitis management: microparticles, nanoparticles, nanofibers, implantable systems, hydrogels, and coatings.</p> Full article ">
13 pages, 2982 KiB  
Article
Nanoliposomes Permeability in a Microfluidic Drug Delivery Platform across a 3D Hydrogel
by Corentin Peyret, Aleka Manousaki, Sabine Bouguet-Bonnet, Emmanuel Stratakis, Laura Sanchez-Gonzalez, Cyril J.F. Kahn and Elmira Arab-Tehrany
Pharmaceutics 2024, 16(6), 765; https://doi.org/10.3390/pharmaceutics16060765 - 4 Jun 2024
Viewed by 1192
Abstract
Nanoliposomes are nano-sized vesicles that can be used as drug delivery carriers with the ability to encapsulate both hydrophobic and hydrophilic compounds. Moreover, their lipid compositions facilitate their internalization by cells. However, the interaction between nanoliposomes and the membrane barrier of the human [...] Read more.
Nanoliposomes are nano-sized vesicles that can be used as drug delivery carriers with the ability to encapsulate both hydrophobic and hydrophilic compounds. Moreover, their lipid compositions facilitate their internalization by cells. However, the interaction between nanoliposomes and the membrane barrier of the human body is not well-known. If cellular tests and animal testing offer a solution, their lack of physiological relevance and ethical concerns make them unsuitable to properly mimic human body complexity. Microfluidics, which allows the environment of the human body to be imitated in a controlled way, can fulfil this role. However, existing models are missing the presence of something that would mimic a basal membrane, often consisting of a simple cell layer on a polymer membrane. In this study, we investigated the diffusion of nanoliposomes in a microfluidic system and found the optimal parameters to maximize their diffusion. Then, we incorporated a custom made GelMA with a controlled degree of substitution and studied the passage of fluorescently labeled nanoliposomes through this barrier. Our results show that highly substituted GelMA was more porous than lower substitution GelMA. Overall, our work lays the foundation for the incorporation of a hydrogel mimicking a basal membrane on a drug delivery microfluidic platform. Full article
(This article belongs to the Special Issue Novel Applications of Modern Excipients in Advanced Pharmaceutical Products)
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<p>Global experimental pattern. (<b>A</b>) Synthesis protocol for GelMA. (<b>B</b>) Protocol for GelMA integration in the chip, chip assembly, and permeability assay. The black arrows indicate flow direction, beige rectangle symbolizes the GelMA membrane. Two syringe pumps were used at two different flow rates; fluids were injected in the microfluidic chip with a flow rate ratio of 2: (a) 30 µL/min for Dil nanoliposomes in PBS and (b) 15 µL/min for PBS. Chip outputs (c) and (d) were collected from both the bottom and top channel every hour for absorbance measurement by UV–Vis spectroscopy.</p> Full article ">Figure 2
<p><sup>1</sup>H NMR spectra of the unmodified gelatin, G45, and G70, respectively. (<b>a</b>) 7.23–7.5 ppm: aromatic amino acid peak used as reference. (<b>b</b>) 3–3.2 ppm: lysine methylene protons’ peak. The area under these peaks were used to calculate the degree of substitution.</p> Full article ">Figure 3
<p>Porosity of GelMA. (<b>A</b>,<b>B</b>) Representative SEM images of the G45 and G70 surfaces. Scale bar: 2 µm, respectively. (<b>C</b>) Pore size for G45 and G70, average of n = 460 *** = relevance &gt; 99.99% (for <span class="html-italic">p</span> value &lt; 0.001).</p> Full article ">Figure 4
<p>Chemical structure of Dil.</p> Full article ">Figure 5
<p>Physico-chemical characterization via DLS of nanoliposomes (<b>A</b>) and Dil nanoliposomes (<b>B</b>). (<b>C</b>) absorption spectra of Dil-loaded nanoliposomes.</p> Full article ">
19 pages, 3674 KiB  
Article
Antitumoral-Embedded Biopolymeric Spheres for Implantable Devices
by Valentina Grumezescu, Oana Gherasim, Bianca Gălățeanu and Ariana Hudiță
Pharmaceutics 2024, 16(6), 754; https://doi.org/10.3390/pharmaceutics16060754 - 3 Jun 2024
Viewed by 852
Abstract
The bioactive surface modification of implantable devices paves the way towards the personalized healthcare practice by providing a versatile and tunable approach that increase the patient outcome, facilitate the medical procedure, and reduce the indirect or secondary effects. The purpose of our study [...] Read more.
The bioactive surface modification of implantable devices paves the way towards the personalized healthcare practice by providing a versatile and tunable approach that increase the patient outcome, facilitate the medical procedure, and reduce the indirect or secondary effects. The purpose of our study was to assess the performance of composite coatings based on biopolymeric spheres of poly(lactide-co-glycolide) embedded with hydroxyapatite (HA) and methotrexate (MTX). Bio-simulated tests performed for up to one week evidenced the gradual release of the antitumor drug and the biomineralization potential of PLGA/HA-MTX sphere coatings. The composite materials proved superior biocompatibility and promoted enhanced cell adhesion and proliferation with respect to human preosteoblast and osteosarcoma cell lines when compared to pristine titanium. Full article
(This article belongs to the Special Issue Polymers Enhancing Bioavailability in Drug Delivery, 3rd Edition)
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<p>Graphic representation of the MTX’s calibration curve at 220 nm (<b>a</b>) and the release profile of MTX from PLGA/HA-MTX sphere coatings under bio-simulated conditions (<b>b</b>).</p> Full article ">Figure 2
<p>ATR-FTIR of PLGA/HA-MTX sphere coatings, before and after testing under bio-simulated conditions.</p> Full article ">Figure 3
<p>SEM micrographs of initial PLGA/HA-MTX sphere coatings (<b>a</b>,<b>b</b>), SEM image (<b>c</b>) and corresponding EDS spectrum (<b>e</b>), overlapped EDS map (<b>d</b>) and individual EDS maps (<b>f</b>–<b>j</b>) of initial PLGA/HA-MTX sphere coatings.</p> Full article ">Figure 4
<p>SEM micrographs of PLGA/HA-MTX sphere coatings after testing under bio-simulated conditions.</p> Full article ">Figure 5
<p>EDS maps (<b>a<sub>1</sub></b>–<b>d<sub>1</sub></b>,<b>e</b>,<b>f</b>) and EDS spectra (<b>a<sub>2</sub></b>–<b>d<sub>2</sub></b>) of PLGA/HA-MTX sphere coatings after testing under bio-simulated conditions at 24 h (<b>a<sub>1</sub></b>,<b>a<sub>2</sub></b>), 48 h (<b>b<sub>1</sub></b>,<b>b<sub>2</sub></b>), 72 h (<b>c<sub>1</sub></b>,<b>c<sub>2</sub></b>), 96 h (<b>d<sub>1</sub></b>,<b>d<sub>2</sub></b>), and 168 h (<b>e</b>,<b>f</b>), and graphic representation of the mass variation in PLGA/HA-MTX sphere coatings after testing under bio-simulated conditions (<b>g</b>).</p> Full article ">Figure 6
<p>Graphical representation of cell viability and proliferation of (<b>a</b>) human preosteoblasts hFOB 1.19 and (<b>b</b>) human osteosarcoma Saos-2 cells after 24 h and 72 h of contact with PLGA/HA and PLGA/HA-MTX coatings as revealed by the MTT assay (* <span class="html-italic">p</span> value ≤ 0.05, ** <span class="html-italic">p</span> value ≤ 0.01, *** <span class="html-italic">p</span> value ≤ 0.001, **** <span class="html-italic">p</span> value ≤ 0.0001).</p> Full article ">Figure 7
<p>Fluorescence micrographs revealing live (green) and dead (red) (<b>a</b>) human preosteoblasts hFOB 1.19 and (<b>b</b>) human osteosarcoma Saos-2 cells after 24 h and 72 h of contact with the non-coated samples, PLGA/HA coatings, and PLGA/HA-MTX coatings (magnification 10×).</p> Full article ">Figure 8
<p>Fluorescence micrographs of the cell cytoskeleton of (<b>a</b>) human preosteoblasts hFOB 1.19 and (<b>b</b>) human osteosarcoma Saos-2 cells after 24 h and 72 h of contact with the non-coated samples, PLGA/HA coatings and PLGA/HA-MTX coatings, after staining of the actin filaments with phalloidin-FITC (green) and cell nuclei with DAPI (blue), (magnification 10×).</p> Full article ">
28 pages, 6006 KiB  
Article
Design and Biocompatibility of Biodegradable Poly(octamethylene suberate) Nanoparticles to Treat Skin Diseases
by Dragana P. C. de Barros, Luís P. Fonseca, Luís G. Gonçalves, Diogo S. Serrano and Abel Oliva
Pharmaceutics 2024, 16(6), 753; https://doi.org/10.3390/pharmaceutics16060753 - 3 Jun 2024
Viewed by 977
Abstract
Biodegradable aliphatic polyester formulations as carriers for topical drug delivery show the potential to encapsulate structurally different therapeutic compounds. Poly(octamethylene suberate) (POS) nanoparticles (POS-NPs) were used as a matrix to encapsulate four therapeutic molecules used to treat skin disorders: caffeine (CF), quercetin (QR), [...] Read more.
Biodegradable aliphatic polyester formulations as carriers for topical drug delivery show the potential to encapsulate structurally different therapeutic compounds. Poly(octamethylene suberate) (POS) nanoparticles (POS-NPs) were used as a matrix to encapsulate four therapeutic molecules used to treat skin disorders: caffeine (CF), quercetin (QR), hydrocortisone (HC), and adapalene (AD). Hydrophobicity and chemical structure of bioactive compounds (BCs) influenced the physicochemical stability of drug-loaded nanoparticles. The particle size of drug-loaded nanoparticles was between 254.9 nm for the CF-POS-NP and 1291.3 for QR-POS-NP. Particles had a negative charge from −27.6 mV (QR) to −49.2 mV (HC). Drug loading content for all BC-POS-NPs varies between 36.11 ± 1.48% (CF-POS-NP) and 66.66 ± 4.87% (AD-POS-NP), and their entrapment efficiency is relatively high (28.30 ± 1.81% and 99.95 ± 0.04%, respectively). Calorimetric analysis showed the appearance of polymorphism for AD- and HC-loaded systems and the drug’s complete solubilisation into all nanoparticle formulations. FTIR and NMR spectra showed apparent drug incorporation into the polymer matrix of NPs. The encapsulation of BCs enhanced the antioxidative effect. The prepared POS nanoparticles’ cytotoxicity was studied using two dermal cell lines, keratinocyte (HaCaT) cells and fibroblasts (HDFn). The nanoparticle cytotoxic effect was more substantial on HaCaT cell lines. A reconstructed human epidermis (RHE) was successfully used to investigate the penetration of polymeric NPs. Based on permeation and histology studies, HC-POS-NPs and CF-POS-NPs were shown not to be suitable for dermal applications with the explored drug concentrations. AD presents a high permeation rate and no toxic impact on RHE. Full article
(This article belongs to the Special Issue Topical Drug Carriers: Recent Advances and Future Challenges)
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<p>Chemical structures of caffeine (CF, CAS 58-08-2), hydrocortisone (HC, CAS 50-23-7), quercetin (QR, CAS 117-39-5), and adapalene (AD, CAS 106685-40-9) and POS (Mw = 13,806 g mol<sup>−1</sup>).</p> Full article ">Figure 2
<p>SEC spectra were obtained from the synthesis of poly(octylmethylene suberate) (POS) for the sample at time zero (green line) with an average Mw of 180 g mol<sup>−1</sup> for residence time (RT) = 11.2 min, corresponding to a mixture of octanedioic acid and 1,8-octanediol for equimolar monomer concentrations (0.5 M) and sample after 48 h of polycondensation (blue line) at 55 °C with synthesised polyesters corresponding to residence time of 8.33 min (average Mw = 13,806 g mol<sup>−1</sup>). The peak RT of 12.6 min corresponds to water in the partially air-dried samples.</p> Full article ">Figure 3
<p>Schematic overview of RHE model generation [<a href="#B50-pharmaceutics-16-00753" class="html-bibr">50</a>].</p> Full article ">Figure 4
<p>Cytotoxic effect of POSsolvent, POSwater, and POSminiemulsions on (<b>a</b>) keratinocytes (HaCaT) and (<b>b</b>) fibroblasts (HDFn); medium (DMEM) without FBS supplement was used as control.</p> Full article ">Figure 5
<p>Physicochemical characteristics for POS-NP and BC-POS-NPs; BC—bioactive compound; QR—quercetin, HC—hydrocortisone; CF—caffeine; AD—adapalene; NP—nanoparticle; 0.5 wt% of BC; Span 80.</p> Full article ">Figure 6
<p>DSC thermogram of POS-NP and POS pure polymer; temperature ranges from 0 °C to 100 °C and from 100 °C to 0 °C.</p> Full article ">Figure 7
<p>DSC thermogram for POS-NP and BC-POS-NPs: (<b>a</b>) from 0 °C to 100 °C and from 100 °C to 0 °C; (<b>b</b>) from 0 °C to 400 °C. * The temperature regime for AD-POS-NP (<b>a</b>) from −10 °C to 100 °C.</p> Full article ">Figure 8
<p>DSC thermogram for pure compounds.</p> Full article ">Figure 9
<p>DSC thermogram for blends of POS pure polymer and blends of pure BCs (BC: CF, QR, HC, and AD) with POS pure polymer: temperature range from 0 °C to 100 °C and from 100 °C to 0 °C; BC concentration 0.5 wt%.</p> Full article ">Figure 10
<p>FTIR spectra of pure bioactive compounds (BC: CF, QR, AD, HC) and BC-POS-NPs.</p> Full article ">Figure 11
<p><sup>1</sup>H NMR spectra of BC-POS-NPs (BC: AD, HC, CF; CDCl3, 400 MHz, 25 °C) (<b>A</b>), and QR-POS-NP (DMSO-d6, 400 MHz, 25 °C) (<b>B</b>).</p> Full article ">Figure 12
<p>Antioxidant activity of pure BC used in the preparation of the polymeric nanoparticles BC-POS-NP (BC, 0.5 wt%), and of a pure BC solution (5 mg mL<sup>−1</sup>).</p> Full article ">Figure 13
<p>Cytocompatibility (viability %) of keratinocytes (HaCaT) for POS-NP, CF-POS-NP, QR-POS-NP, AD-POS-NP, and HC-POS-NP; D-dilution factor; D-NPs dilution; D/1, D/10, D/20, D/30, D/40, D/50 correspond to NP concentrations 25, 2.5, 1.25, 0.83, 0.63, 0.50, and 0.25 mg/mL<sup>−1</sup> expressed by amount of total polymer. The dashed line presents the 70% cell viability limit for cell cytotoxicity (ISO 10993-5:2009 [<a href="#B60-pharmaceutics-16-00753" class="html-bibr">60</a>]).</p> Full article ">Figure 14
<p>Cytocompatibility (viability, %) of fibroblasts (HDFn) for POS-NP, CF-POS-NP, QR-POS-NP, AD-POS-NP, and HC-POS-NP; D-dilution factor; D-NPs dilution; D/1, D/10, D/20, D/30, D/40, D/50 correspond NPs concentration 25, 2.5, 1.25, 0.83, 0.63, 0.50, and 0.25 mg/mL<sup>−1</sup> expressed by amount of total polymer. The dashed line presents the 70% cell viability limit for cell cytotoxicity (ISO 10993-5:2009 [<a href="#B60-pharmaceutics-16-00753" class="html-bibr">60</a>]).</p> Full article ">Figure 15
<p>In vitro permeation profiles of BC-POS-NP. Cumulative amounts released from NPs in an RHE model. Plots were obtained from five experiments with each tested compound running under the same experimental conditions with three replicates. Error bars represent the mean ± SD value.</p> Full article ">Figure 16
<p>Comparative histological aspect of BC-POS-NP permeation on RHE. RHE—reconstructed human epidermis; RHE-POS—reconstructed human epidermis with POS-NP (<b>A</b>), QR-RHE-POS and AD-RHE-POS—reconstructed human epidermis with AD and QR loaded POS-NPs (<b>B</b>), HC-RHE-POS and CF-RHE-POS—reconstructed human epidermis with HC and CF loaded POS-NPs (<b>C</b>); SC—stratum corneum; hematoxylin–eosin staining; Nikon Eclipse TE2000-S bright field microscope (Nikon instruments, Melville, NY, USA) magnification 20×.</p> Full article ">
31 pages, 2372 KiB  
Review
Translational Challenges in Drug Therapy and Delivery Systems for Treating Chronic Lower Extremity Wounds
by Danny Aljamal, Priya S. Iyengar and Tammy T. Nguyen
Pharmaceutics 2024, 16(6), 750; https://doi.org/10.3390/pharmaceutics16060750 - 2 Jun 2024
Cited by 1 | Viewed by 1300
Abstract
Despite several promising preclinical studies performed over the past two decades, there remains a paucity of market-approved drugs to treat chronic lower extremity wounds in humans. This translational gap challenges our understanding of human chronic lower extremity wounds and the design of wound [...] Read more.
Despite several promising preclinical studies performed over the past two decades, there remains a paucity of market-approved drugs to treat chronic lower extremity wounds in humans. This translational gap challenges our understanding of human chronic lower extremity wounds and the design of wound treatments. Current targeted drug treatments and delivery systems for lower extremity wounds rely heavily on preclinical animal models meant to mimic human chronic wounds. However, there are several key differences between animal preclinical wound models and the human chronic wound microenvironment, which can impact the design of targeted drug treatments and delivery systems. To explore these differences, this review delves into recent new drug technologies and delivery systems designed to address the chronic wound microenvironment. It also highlights preclinical models used to test drug treatments specific for the wound microenvironments of lower extremity diabetic, venous, ischemic, and burn wounds. We further discuss key differences between preclinical wound models and human chronic wounds that may impact successful translational drug treatment design. Full article
(This article belongs to the Special Issue Advances in the Development of Cutting-Edge Drug Delivery Systems for the Effective Treatment of Chronic Skin Wounds, 2nd Edition)
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<p>Types of chronic non-healing wounds by underlying etiology. The underlying etiology for chronic wounds can be identified based on location on the lower extremity and distinguishing wound characteristics [<a href="#B2-pharmaceutics-16-00750" class="html-bibr">2</a>].</p> Full article ">Figure 2
<p>Stages of normal wound healing. The temporal four stages of normal wound healing progresses from hemostasis to inflammation, proliferation, and remodeling. The expected length of time in each wound healing stages is indicated. Image created in BioRender, agreement number HL26S82IJQ.</p> Full article ">Figure 3
<p>Common features of the chronic wound microenvironment. Wounds that fail to heal after 6–8 weeks are considered chronic. Chronic wounds may stem from different etiologies; however, they often have shared features that prevent progression towards healing. Chronic wounds often have a layer of biofilm that prevents epithelialization. Furthermore, the chronic wound microenvironment is hypoxic. The up arrow indicates an increase in ROS, inflammatory macrophages, and metalloproteases (MMP). The microenvironment can lead to a reduction in angiogenesis, cytokine, and growth factor signaling, and reduced anti-inflammatory macrophage recruitment as indicated by the down arrow. Image created in BioRender, agreement number ZU26R8×4PV.</p> Full article ">Figure 4
<p>Chronic Lower Extremity Wound Drug Delivery Systems. Common wound drug delivery systems used for chronic lower extremity wounds. Image created in BioRender, agreement number BQ26U95IML.</p> Full article ">
34 pages, 2873 KiB  
Review
Chitosan Nanoparticles for Intranasal Drug Delivery
by Hossein Omidian, Erma J. Gill, Sumana Dey Chowdhury and Luigi X. Cubeddu
Pharmaceutics 2024, 16(6), 746; https://doi.org/10.3390/pharmaceutics16060746 - 31 May 2024
Cited by 2 | Viewed by 2126
Abstract
This manuscript explores the use of nanostructured chitosan for intranasal drug delivery, targeting improved therapeutic outcomes in neurodegenerative diseases, psychiatric care, pain management, vaccination, and diabetes treatment. Chitosan nanoparticles are shown to enhance brain delivery, improve bioavailability, and minimize systemic side effects by [...] Read more.
This manuscript explores the use of nanostructured chitosan for intranasal drug delivery, targeting improved therapeutic outcomes in neurodegenerative diseases, psychiatric care, pain management, vaccination, and diabetes treatment. Chitosan nanoparticles are shown to enhance brain delivery, improve bioavailability, and minimize systemic side effects by facilitating drug transport across the blood–brain barrier. Despite substantial advancements in targeted delivery and vaccine efficacy, challenges remain in scalability, regulatory approval, and transitioning from preclinical studies to clinical applications. The future of chitosan-based nanomedicines hinges on advancing clinical trials, fostering interdisciplinary collaboration, and innovating in nanoparticle design to overcome these hurdles and realize their therapeutic potential. Full article
(This article belongs to the Special Issue Recent Advances in Nanoparticles for Mucosal Drug Delivery)
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<p>Ex vivo permeability studies of RH loaded from PLGA and PLGA/chit NPs across sheep nasal mucosa. (<b>A</b>) Permeability studies across nasal mucosa at several time points (PLGA NPs in red color, PLGA/chit NPs in blue color). Results are presented as mean value ± SEM, n ≥ 3. Histopathological studies of (<b>B</b>) untreated and (<b>C</b>) treated nasal mucosa with RH loaded PLGA NPs and (<b>D</b>) RH loaded PLGA/chit NPs after 4 h incubation. Scale bar: 100 μm, Adopted with permission [<a href="#B11-pharmaceutics-16-00746" class="html-bibr">11</a>].</p> Full article ">Figure 2
<p>Evaluation of mast cell activation by nanoparticles. (<b>A</b>) Degranulation of the mast cell line HMC-1 was evaluated by a β-hex release assay. Cells were stimulated with C48/80 at 20, 40 or 80 μg/mL in solution or incorporated in nanoparticles. Blank Chi NP and Chi/Alg NP were used as controls at the same concentration of nanoparticles tested with C48/80 loaded particles. Not only Chi-C48/80 NP but also Chi NP induced a higher % of β-hex release than C48/80 in solution and Chi/Alg-C48/80 NP, indicating the intrinsic ability of Chi NP to activate mast cells. Data are representative of three independent experiments performed in triplicate or quadruplicate, mean ± SD, n = 3. Symbols above bars indicate the differences relative to C48/80 in solution, * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001 2-way ANOVA (<b>B</b>) Confocal images of HMC-1 before (<b>B1</b>) and 2 min after treatment with FITC labeled Chi-C48/80 NP (green) at a dose corresponding to 40 μg/mL of C48/80 in Tyrode’s solution (<b>B2</b>). Cells were labeled with Hoechst 33,342 (blue) for nuclei and with Alexa Fluor<sup>®</sup> 594 WGA to identify cell membranes. Images showed that Chi-C48/80 immediately adsorbed on the cell surface, Adopted with permission [<a href="#B69-pharmaceutics-16-00746" class="html-bibr">69</a>].</p> Full article ">Figure 3
<p>Immunization and sampling timeline (<b>A</b>); changes in body weight of mice (<b>B</b>); BSA-specific serum IgG (<b>C</b>–<b>E</b>) and nasal lavage fluid (NLF) IgA (<b>F</b>) antibody responses; all results are reported as mean (SD) (n = 6, * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001) [<a href="#B16-pharmaceutics-16-00746" class="html-bibr">16</a>].</p> Full article ">Figure 4
<p>Plasma glucose levels in rabbits following nasal administration (at pH 7.4) of: A, insulin-loaded PE3gC24 nanoparticles suspended in PBS; B, insulin-PE3gC24 PBS suspension; C, insulin in PBS. Mean ± SD, n = 6. * Statistically significant differences from control insulin solution (<span class="html-italic">p</span> &lt; 0.05), Adopted with permission [<a href="#B42-pharmaceutics-16-00746" class="html-bibr">42</a>].</p> Full article ">Figure 5
<p>Impact of HPD and HPD/NPs on LPS-induced inflammation in mice. At 3 h post-LPS, PBS (vehicle), HPD, or HPD/NPs were nasally administered to mice. Lung tissues were collected at 24 h post-LPS challenge. (<b>A</b>) Expression levels of IL-1β and (<b>B</b>) IL-6 in mouse plasma at 24 h post-LPS challenge (n = 4/group; ** <span class="html-italic">p</span> &lt; 0.001 vs. PBS vehicle and vs. HPD). (<b>C</b>) Representative micrographs of H&amp;E-stained lung tissue cross-sections at 24 h post-LPS challenge. Scale bar, 1 mm (<b>upper row</b>) or 100 μm (<b>lower row</b>) [<a href="#B117-pharmaceutics-16-00746" class="html-bibr">117</a>].</p> Full article ">
11 pages, 5736 KiB  
Article
Co-Amorphization of Acemetacin with Basic Amino Acids as Co-Formers for Solubility Improvement and Gastric Ulcer Mitigation
by Jiayue Hou, Peixu Zhao, Yanfei Wang, Xiwei Jiang and Qiang Fu
Pharmaceutics 2024, 16(6), 745; https://doi.org/10.3390/pharmaceutics16060745 - 31 May 2024
Viewed by 1062
Abstract
Acemetacin (ACM) is a new non-steroidal anti-inflammatory drug with anti-inflammatory, analgesic, and antipyretic effects. However, the poor water solubility and gastrointestinal side effects limit its use. Recently, the co-amorphous (CAM) strategy has attracted great interest to improve solubility for poorly water-soluble drugs, and [...] Read more.
Acemetacin (ACM) is a new non-steroidal anti-inflammatory drug with anti-inflammatory, analgesic, and antipyretic effects. However, the poor water solubility and gastrointestinal side effects limit its use. Recently, the co-amorphous (CAM) strategy has attracted great interest to improve solubility for poorly water-soluble drugs, and basic amino acids have the potential to protect the gastrointestinal tract. In order to develop a highly efficient and low-toxic ACM formulation, we prepared ACM CAM systems, with basic amino acids (lysine, arginine, and histidine) as co-formers, using a cryo-milling method. The solid-state behaviors of the ACM CAM systems were characterized by polarizing light microscopy, differential scanning calorimetry, and powder X-ray diffraction. Fourier transform infrared spectroscopy and molecular docking were carried out to understand the formation mechanism. Moreover, the gastro-protective effects of ACM CAM systems were evaluated in a rat gastric ulcer model. The results demonstrated that the CAM systems improved the dissolution rates of ACM compared with the neat amorphous counterpart. Furthermore, ACM CAM systems are significantly effective in mitigating the ACM-induced gastric ulcer in rats, and the ulcer inhibition rates were almost 90%. More importantly, this study provided a useful method for mitigating drug-induced gastrointestinal damage and broadened the applications of drug–amino acid CAM systems. Full article
(This article belongs to the Special Issue Formulation of Poorly Water-Soluble Drugs)
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Graphical abstract

Graphical abstract
Full article ">Figure 1
<p>Molecular structures of (<b>A</b>) ACM and (<b>B</b>) Lys, Arg, and His.</p> Full article ">Figure 2
<p>The PLM images of (<b>A</b>) ACM, (<b>B</b>) PM-Lys, (<b>C</b>) PM-Arg, (<b>D</b>) PM-His, (<b>E</b>) ACM-AM, (<b>F</b>) ACM-Lys, (<b>G</b>) ACM-Arg, and (<b>H</b>) ACM-His. The scale bar represents 100 μm.</p> Full article ">Figure 3
<p>PXRD patterns of (a) ACM-AM, (b) ACM, (c) ACM-Lys, (d) PM-Lys, (e) ACM-Arg, (f) PM-Arg, (g) ACM-His, and (h) PM-His.</p> Full article ">Figure 4
<p>DSC thermograms (<b>A</b>) and the second heating DSC curves (<b>B</b>) of ACM-His, ACM-Lys, ACM-Arg, PM-His, PM-Lys, PM-Arg, and ACM.</p> Full article ">Figure 5
<p>FTIR spectra of (a) ACM-AM, (b) ACM, (c) ACM-Lys, (d) PM-Lys, (e) ACM-Arg, (f) PM-Arg, (g) ACM-His, and (h) PM-His.</p> Full article ">Figure 6
<p>The molecular docking of the interaction between ACM and (<b>A</b>) Lys, (<b>B</b>) Arg, and (<b>C</b>) His. Red represents oxygen atoms, blue for nitrogen atoms, gray for hydrogen atoms, and the green dotted lines indicate hydrogen bonds formed between atoms.</p> Full article ">Figure 7
<p>Dissolution profiles of ACM, ACM-AM, PMs, and ACM CAM systems.</p> Full article ">Figure 8
<p>IDR curves (<b>A</b>) and the normalized dissolution rates (<b>B</b>) of ACM, ACM-AM, PMs, and ACM CAM systems (<span class="html-italic">n</span> = 3, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001, **** <span class="html-italic">p</span> &lt; 0.0001 vs. ACM and #### <span class="html-italic">p</span> &lt; 0.0001 vs. PM groups).</p> Full article ">Figure 9
<p>Stomach anatomy images (<b>A</b>) and the inhibition rate (<b>B</b>) of (<b>a</b>) ACM, (<b>b</b>) PM-Lys, (<b>c</b>) PM-Arg, (<b>d</b>) PM-His, (<b>e</b>) PBS, (<b>f</b>) CAM-Lys, (<b>g</b>) CAM-Arg, and (<b>h</b>) CAM-His. The scale bar represents 1 cm (<span class="html-italic">n</span> = 4, **** <span class="html-italic">p</span> &lt; 0.0001 vs. ACM and ## <span class="html-italic">p</span> &lt; 0.001 vs. PM groups).</p> Full article ">Figure 10
<p>Histopathological examinations of (<b>A</b>) ACM, (<b>B</b>) PM-Lys, (<b>C</b>) PM-Arg, (<b>D</b>) PM-His, (<b>E</b>) PBS, (<b>F</b>) CAM-Lys, (<b>G</b>) CAM-Arg, and (<b>H</b>) CAM-His. The scale bar represents 50 μm.</p> Full article ">
28 pages, 12497 KiB  
Review
Atomic Force Microscopy for the Study of Cell Mechanics in Pharmaceutics
by Henrik Siboni, Ivana Ruseska and Andreas Zimmer
Pharmaceutics 2024, 16(6), 733; https://doi.org/10.3390/pharmaceutics16060733 - 29 May 2024
Viewed by 1184
Abstract
Cell mechanics is gaining attraction in drug screening, but the applicable methods have not yet become part of the standardized norm. This review presents the current state of the art for atomic force microscopy, which is the most widely available method. The field [...] Read more.
Cell mechanics is gaining attraction in drug screening, but the applicable methods have not yet become part of the standardized norm. This review presents the current state of the art for atomic force microscopy, which is the most widely available method. The field is first motivated as a new way of tracking pharmaceutical effects, followed by a basic introduction targeted at pharmacists on how to measure cellular stiffness. The review then moves on to the current state of the knowledge in terms of experimental results and supplementary methods such as fluorescence microscopy that can give relevant additional information. Finally, rheological approaches as well as the theoretical interpretations are presented before ending on additional methods and outlooks. Full article
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<p>Research trend analysis of cell mechanics up to and including the year 2023. Note the logarithmic y-axis. The data are from Clarivate Web of Science. © Copyright Clarivate 2024. All rights reserved.</p> Full article ">Figure 2
<p>Illustration of nanoindentation by AFM. (<b>a</b>) The piezo moves the tip toward the cell at a speed <math display="inline"><semantics> <mrow> <mover accent="true"> <mrow> <mi>v</mi> </mrow> <mo>→</mo> </mover> </mrow> </semantics></math> while it is still out of contact. (<b>b</b>) The tip indents a distance <math display="inline"><semantics> <mrow> <mi>δ</mi> </mrow> </semantics></math> into the cell. The cantilever is correspondingly bent at a distance <math display="inline"><semantics> <mrow> <mi>x</mi> </mrow> </semantics></math>. The piezo has moved a distance <math display="inline"><semantics> <mrow> <msub> <mrow> <mi>Z</mi> </mrow> <mrow> <mi mathvariant="normal">p</mi> </mrow> </msub> </mrow> </semantics></math>. (<b>c</b>) Generic sketch of a force–indentation curve with the approach (blue) and the subsequent retraction (red). This illustration was created using <a href="http://BioRender.com" target="_blank">BioRender.com</a>.</p> Full article ">Figure 3
<p>Cells are ideally elastic (green) when stress and strain are proportional, which is the case at small deformations of the cell cortex. However, past a certain load, the cell’s inhomogeneity causes deviations from this ideality (yellow) [<a href="#B30-pharmaceutics-16-00733" class="html-bibr">30</a>]. At a sufficiently high strain, the cell ruptures completely (blue). The red curve is a generic stress–strain curve based on [<a href="#B28-pharmaceutics-16-00733" class="html-bibr">28</a>]. The cell illustrations were created with <a href="http://BioRender.com" target="_blank">BioRender.com</a>.</p> Full article ">Figure 4
<p>Illustration of the different views of indentation at a depth <math display="inline"><semantics> <mrow> <mi>δ</mi> </mrow> </semantics></math>. (<b>a</b>) Simple compression of a spring. (<b>b</b>) Even compression of a cylinder with a positive Poisson ratio. (<b>c</b>) Realistic uneven compression of a cell. This illustration was created using <a href="http://BioRender.com" target="_blank">BioRender.com</a>.</p> Full article ">Figure 5
<p>Meta-analysis on the reviewed experimental papers. (<b>a</b>) Fractions of studies performed by similar pharmaceutical institutes. (<b>b</b>) Studies published in pharmaceutical vs. non-pharmaceutical journals. (<b>c</b>) Fraction of cells seeded on a hard substrate (glass) vs. a soft substrate. (<b>d</b>) Fraction of studies including rheological measurements.</p> Full article ">Figure 6
<p>Stiffness changes in cells treated with one of eight different chemicals. While an effect could be measured, the relative variability was also large. Large dots represent group means while small dots represent each individual measured cell. Control (C) and treated (A) groups measured on the same day have the same color. Reproduced with permission from [<a href="#B69-pharmaceutics-16-00733" class="html-bibr">69</a>].</p> Full article ">Figure 7
<p>Using stiffness to track the uptake of gold nanoparticles. The stiffness change could be correlated with receptor expression. Adapted with permission from [<a href="#B15-pharmaceutics-16-00733" class="html-bibr">15</a>]. Copyright 2024 American Chemical Society.</p> Full article ">Figure 8
<p>Combined force volume mapping and fluorescence microscopy show a correlation between nanofibers and increased stiffness. (<b>a</b>) Differential Interference Contrast image. (<b>b</b>) Fluorescence image showing the position of the nanofibers. (<b>c</b>) Topographical map. (<b>d</b>) Stiffness map with stiffness peaks in the same location as the nanofibers. Reproduced from [<a href="#B59-pharmaceutics-16-00733" class="html-bibr">59</a>] under the Creative Commons license CC BY 4.0 DEED.</p> Full article ">Figure 9
<p>Recommended procedure for achieving significant cell mechanics results with AFM. This graphic was created using <a href="http://BioRender.com" target="_blank">BioRender.com</a>.</p> Full article ">Figure 10
<p>Time-dependent measurements for cell rheology approach (blue), pause (green), and retract (red). In the top row are the force–time curves, while the bottom row shows the piezo positions over time. (<b>a</b>) The force-time representation of a regular force-distance measurement (as in <a href="#pharmaceutics-16-00733-f002" class="html-fig">Figure 2</a>c). (<b>b</b>) Creep compliance where the force is kept constant. (<b>c</b>) Stress relaxation where the piezo position is constant. (<b>d</b>) An oscillatory measurement where the piezo oscillates and the force therefore also oscillates. The illustration is based on [<a href="#B2-pharmaceutics-16-00733" class="html-bibr">2</a>,<a href="#B43-pharmaceutics-16-00733" class="html-bibr">43</a>] and was created using <a href="http://BioRender.com" target="_blank">BioRender.com</a>.</p> Full article ">Figure 11
<p>Qualitative illustrations of the select mechanical models for cells. (<b>a</b>) Ideal elastic. (<b>b</b>) Spring-dashpot. (<b>c</b>) Polymer network. (<b>d</b>) Poroelastic. (<b>e</b>) Semi-permeable water balloon. (<b>f</b>) Soft-glassy. (<b>g</b>) Finite element analysis where the colors represent strain values. (<b>a</b>–<b>f</b>) were created using BioRender.com, while (<b>f</b>) was adapted from [<a href="#B92-pharmaceutics-16-00733" class="html-bibr">92</a>] under the Creative Commons license CC BY 3.0 Deed.</p> Full article ">Figure 12
<p>(<b>a</b>) A spring represents elasticity in the model as described by Hooke’s law (Equation (6)). (<b>b</b>) A dashpot represents viscosity as described by Equation (29). The two can be combined (<b>c</b>) in series as the Maxwell model or (<b>d</b>) in parallel as the Kelvin-Voight model. Larger combinations include (<b>e</b>) Standard Linear Solid (SLS) and (<b>f</b>) the five-element Maxwell model. Illustration based on [<a href="#B29-pharmaceutics-16-00733" class="html-bibr">29</a>].</p> Full article ">Figure 13
<p>Master curves reported by Fabry et al. for (<b>a</b>) normalized stiffness <math display="inline"><semantics> <mrow> <mi>G</mi> </mrow> </semantics></math> and (<b>b</b>) the loss tangent <math display="inline"><semantics> <mrow> <mi>η</mi> </mrow> </semantics></math> as a function of <math display="inline"><semantics> <mrow> <mi>x</mi> </mrow> </semantics></math>. Filled-in squares (■) represent control cells while the other data point shapes represent treatmeant with different drugs and yet all lie on the same line. Each color represents a distinct cell line. Adapted with permission from [<a href="#B97-pharmaceutics-16-00733" class="html-bibr">97</a>].</p> Full article ">Figure 14
<p>Knowledge of the timescales for the various processes can be used to interpret results. Adapted with modifications with permission from [<a href="#B2-pharmaceutics-16-00733" class="html-bibr">2</a>].</p> Full article ">Figure 15
<p>Plot of the viscoelastic parameters over time for cells treated with paclitaxel (PTX) and doxorubicin (DOX). Here, <math display="inline"><semantics> <mrow> <msub> <mrow> <mi>b</mi> </mrow> <mrow> <mn>0</mn> </mrow> </msub> </mrow> </semantics></math> and <math display="inline"><semantics> <mrow> <msub> <mrow> <mi>b</mi> </mrow> <mrow> <mn>2</mn> </mrow> </msub> </mrow> </semantics></math> refer to the viscosities of the two Maxwell elements. A clear difference from the control past the 4 h mark can be observed in all parameters, but less so for Young’s modulus. Adapted with permission from [<a href="#B63-pharmaceutics-16-00733" class="html-bibr">63</a>].</p> Full article ">Figure 16
<p>Automatic measurement of thousands of cells before and after treatment with caspofungin. (<b>a</b>) The multi-cell experimental setup. (<b>b</b>) Cell stiffness before treatment. (<b>c</b>) Cell stiffness after treatment. Note that the authors in this experiment describe the cell stiffness with a spring constant (Equation (5)). Adapted with permission from [<a href="#B106-pharmaceutics-16-00733" class="html-bibr">106</a>].</p> Full article ">Figure 17
<p>Mapping Young’s modulus, the cell nucleus becomes more pronounced for larger indentation depths: (<b>left</b>) 50–200 nm, (<b>middle</b>) 200–400 nm, and (<b>right</b>) 400–850 nm. The graphic was provided by Ricardo Garcia with data from [<a href="#B108-pharmaceutics-16-00733" class="html-bibr">108</a>].</p> Full article ">Figure 18
<p>Selection of different experimental techniques for cell mechanics and imaging along with typical forces and length scales. (<b>A</b>) Optical tweezers, (<b>B</b>) atomic force microscopy, (<b>C</b>) bio-MEMS, (<b>D</b>) elastography, (<b>E</b>) fluorescence resonance energy transfer (FRET), (<b>F</b>) super resolution microscopy, (<b>G</b>) particle tracking micro-rheology (PTM), (<b>H</b>) microgel cellular force measurement, (<b>I</b>) PAINT, (<b>J</b>) DNA hairpin-based sensors, (<b>K</b>) Brillouin microscopy, and (<b>L</b>) traction force microscopy. Reproduced from [<a href="#B8-pharmaceutics-16-00733" class="html-bibr">8</a>].</p> Full article ">Figure 19
<p>In the model by Shneider [<a href="#B122-pharmaceutics-16-00733" class="html-bibr">122</a>], AFM for cell mechanics in pharmaceutics is now in stage two of its development as a scientific field. Created using <a href="http://BioRender.com" target="_blank">BioRender.com</a>.</p> Full article ">
18 pages, 10969 KiB  
Article
Porous Chitosan/Hydroxyapatite Composite Microspheres for Vancomycin Loading and Releasing
by Meng-Ying Wu, Yi-Ting Kuo, I-Fang Kao and Shiow-Kang Yen
Pharmaceutics 2024, 16(6), 730; https://doi.org/10.3390/pharmaceutics16060730 - 29 May 2024
Cited by 1 | Viewed by 975
Abstract
Porous chitosan/hydroxyapatite (Chi-HAp) composite microspheres were prepared in an aqueous solution containing chitosan, calcium nitrate, and ammonium dihydrogen phosphate by using a hydrothermal method at various temperatures. The investigation indicated that temperature significantly impacted the final product’s appearance. Hydroxyapatite (HAp) coupled with dicalcium [...] Read more.
Porous chitosan/hydroxyapatite (Chi-HAp) composite microspheres were prepared in an aqueous solution containing chitosan, calcium nitrate, and ammonium dihydrogen phosphate by using a hydrothermal method at various temperatures. The investigation indicated that temperature significantly impacted the final product’s appearance. Hydroxyapatite (HAp) coupled with dicalcium phosphate dihydrate (DCPD) flakes were obviously found at 65 and 70 °C, while the latter gradually disappeared at higher temperatures. Conversely, synthesis at 90 °C led to smaller particle sizes due to the broken chitosan chains. The microspheres synthesized at 75 °C were selected for further analysis, revealing porous structures with specific surface areas of 36.66 m2/g, pores ranging from 3 to 100 nm, and pore volumes of 0.58 cm3/g. Vancomycin (VCM), an antibiotic, was then absorbed on and released from the microspheres derived at 75 °C, with a drug entrapment efficiency of 20% and a release duration exceeding 20 days. The bacteriostatic activity of the VCM/composite microspheres against Staphylococcus aureus increased with the VCM concentration and immersion time, revealing a stable inhibition zone diameter of approximately 4.3 mm from 24 to 96 h, and this indicated the retained stability and efficacy of the VCM during the encapsulating process. Full article
(This article belongs to the Special Issue Design of Mesoporous Materials for Biomedical Application)
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<p>(<b>a</b>) Chitosan structure and (<b>b</b>) vancomycin structure.</p> Full article ">Figure 2
<p>An illustration of the loading process.</p> Full article ">Figure 3
<p>Supposed formation mechanism of the porous chitosan-hydroxyapatite (Chi-HAp) microspheres.</p> Full article ">Figure 4
<p>SEM micrographs of the Chi-HAp microspheres synthesized at (<b>a</b>) 65 °C, (<b>b</b>) 70 °C, (<b>c</b>) 75 °C, (<b>d</b>) 80 °C, (<b>e</b>) 85 °C, and (<b>f</b>) 90 °C.</p> Full article ">Figure 5
<p>Particle sizes of the Chi-HAp synthesized at 65 °C, 70 °C, 75 °C, 80 °C, 85 °C, and 90 °C.</p> Full article ">Figure 6
<p>FE-SEM micrographs of the Chi-HAp with a synthesis temperature of 75 °C.</p> Full article ">Figure 7
<p>EDS spectrum of the Chi-HAp microspheres.</p> Full article ">Figure 8
<p>XRD pattern of the Chi-HAp compared with ICDD file no. 86-1199 and no. 72-0713.</p> Full article ">Figure 9
<p>Nitrogen adsorption/desorption isotherm derived from the BET method.</p> Full article ">Figure 10
<p>Distribution of the BJH desorption pore sizes.</p> Full article ">Figure 11
<p>TGA/DSC graphics of (<b>a</b>) chitosan, (<b>b</b>) commercial HAp, and (<b>c</b>) Chi-HAp in air.</p> Full article ">Figure 12
<p>FTIR spectra acquired from the commercial HAp, chitosan, and Chi-HAp, respectively.</p> Full article ">Figure 13
<p>(<b>a</b>) The calibration curve of the VCM concentrations. (<b>b</b>) The VCM release curves from VCM/Chi-HAp and Chi-VCM/Chi-Hap, respectively, and the right inserted profile is the enlargement of the Chi-VCM/Chi-HAp for the initial 24 h.</p> Full article ">Figure 14
<p>The morphologies of the Chi-HAp after drug release for (<b>a</b>) 1 h, (<b>b</b>) 8 h, (<b>c</b>) 12 h, (<b>d</b>) 7 days, and (<b>e</b>) 28 days.</p> Full article ">Figure 14 Cont.
<p>The morphologies of the Chi-HAp after drug release for (<b>a</b>) 1 h, (<b>b</b>) 8 h, (<b>c</b>) 12 h, (<b>d</b>) 7 days, and (<b>e</b>) 28 days.</p> Full article ">Figure 15
<p>A schematic of the drug-release process involving (<b>a</b>) the dissolution of the dried VCM on the surface of a microsphere in 4 h, (<b>b</b>) the swelling and degradation of the chitosan on the surface of a microsphere before day 7, and (<b>c</b>) the swelling and degradation of the chitosan within the mesopores and/or macropores of a microsphere after day 7. The big arrows indicate the drug release process and the small ones show the releasing directions.</p> Full article ">Figure 16
<p>Bacterial inhibition zones around the holes that were initially filled with 70 μL of elution fluids containing the VCM released from the Chi-HAp microspheres.</p> Full article ">Figure 17
<p>The bacterial inhibition zones of the VCM released from the Chi-VCM/Chi-HAp microspheres into the PBS solution corresponded to the cumulative concentrations.</p> Full article ">
17 pages, 3570 KiB  
Review
Review of Prodrug and Nanodelivery Strategies to Improve the Treatment of Colorectal Cancer with Fluoropyrimidine Drugs
by Santu Sarkar, Sezgin Kiren and William H. Gmeiner
Pharmaceutics 2024, 16(6), 734; https://doi.org/10.3390/pharmaceutics16060734 - 29 May 2024
Cited by 1 | Viewed by 1495
Abstract
Fluoropyrimidine (FP) drugs are central components of combination chemotherapy regimens for the treatment of colorectal cancer (CRC). FP-based chemotherapy has improved survival outcomes over the last several decades with much of the therapeutic benefit derived from the optimization of dose and delivery. To [...] Read more.
Fluoropyrimidine (FP) drugs are central components of combination chemotherapy regimens for the treatment of colorectal cancer (CRC). FP-based chemotherapy has improved survival outcomes over the last several decades with much of the therapeutic benefit derived from the optimization of dose and delivery. To provide further advances in therapeutic efficacy, next-generation prodrugs and nanodelivery systems for FPs are being developed. This review focuses on recent innovative nanodelivery approaches for FP drugs that display therapeutic promise. We summarize established, clinically useful FP prodrug strategies, including capecitabine, which exploit tumor-specific enzyme expression for optimal anticancer activity. We then describe the use of FP DNA-based polymers (e.g., CF10) for the delivery of activated FP nucleotides as a nanodelivery approach with proven activity in pre-clinical models and with clinical potential. Multiple nanodelivery systems for FP delivery show promise in CRC pre-clinical models and we review advances in albumin-mediated FP delivery, the development of mesoporous silica nanoparticles, emulsion-based nanoparticles, metal nanoparticles, hydrogel-based delivery, and liposomes and lipid nanoparticles that display particular promise for therapeutic development. Nanodelivery of FPs is anticipated to impact CRC treatment in the coming years and to improve survival for cancer patients. Full article
(This article belongs to the Special Issue Recent Advances in Drug Delivery for the Treatment of Colorectal Cancer)
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<p>(<b>A</b>) Structure of 5-FU and leucovorin and components of FOLFOX and FOLFIRI. (<b>B</b>) Schematic demonstration of TS inhibition of 5FU resulting in DNA damage. (<b>C</b>) Limitations of 5FU treatment.</p> Full article ">Figure 2
<p>(<b>A</b>) Schematic depiction of Fuplatin design and mechanism of activity. (<b>B</b>) Determination of Pt accumulation in HCT116 cells after treatment with different concentrations of Pt-containing drugs. (<b>C</b>) Percentage apoptosis measurement after treatment with equimolar amounts of 5-FU, oxaliplatin, and their mixture. (<b>D</b>) Structure and release mechanism of prodrug FDU-DB-NO2 under hypoxia condition. Cell viability of MCF-7 cells after treatment with FDU-DB-NO2 and other controls under normoxic and hypoxic conditions for (<b>E</b>) 24 h and (<b>F</b>) 48 h [<a href="#B40-pharmaceutics-16-00734" class="html-bibr">40</a>,<a href="#B42-pharmaceutics-16-00734" class="html-bibr">42</a>]. [Reproduced with permission].</p> Full article ">Figure 3
<p>DNA-based fluoropyrimidine polymers dual target TS and DNA topoisomerase 1 (Top1). (<b>A</b>) Treatment of HCT-116 cells with equal concentrations of F10 and 5-FU followed by CldU and IdU incorporation. (<b>B</b>) Analysis of corresponding DNA fiber images. Significant reductions in (<b>C</b>) fork velocity and (<b>D</b>) terminal forks by F10 (10 nM) than 5-FU (10 nM). (<b>E</b>) Evaluation of replication stress-associated proteins of HCT-116 cells after 2 h of exposure to F10 (10 μm). (<b>F</b>) Western blot showing the detection of TS classic complex (TS CC) formation after treatment with F10, CF10, and 5-FU at specified concentrations for 24 h. (<b>G</b>) Representative immunofluorescence images with an antibody specific for Top1 cleavage complexes (Top1cc—green with DAPI (blue) nuclear staining) at indicated concentrations of F10, CF10, and 5-FU for 24 h and Topotecan for 1 h, with quantification graphs to the right [<a href="#B54-pharmaceutics-16-00734" class="html-bibr">54</a>,<a href="#B56-pharmaceutics-16-00734" class="html-bibr">56</a>]. (***, <span class="html-italic">p</span> &lt; 0.001, ****, <span class="html-italic">p</span> &lt; 0.0001) [Reproduced with permission].</p> Full article ">Figure 4
<p>Pictorial demonstration of passive tumor targeting of nanoparticles through the EPR effect [<a href="#B67-pharmaceutics-16-00734" class="html-bibr">67</a>] and examples of 5-FU nanodelivery systems discussed herein. [Reproduced with permission].</p> Full article ">Figure 5
<p>(<b>A</b>) Schematic demonstrating the synthesis of MS@5-FU^Azo and the release of 5-FU in the presence of a reducing agent (sodium dithionate). (<b>B</b>) %Release of 5-FU from MS@5-Fu^Azo at pH 1.2, 7.2, and in the presence of 100 μg/mL sodium dithionate, pH 7.2. (<b>C</b>) Cell viability analysis after incubation with MS@5-Fu^Azo for 48 h on HT-29 cells. (<b>D</b>) SEM image of 5FU-LV NPs. (<b>E</b>) Cumulative release of 5FU and LV from NPs in the presence of gastrointestinal fluids. (<b>F</b>) Cell viability of different doses of 5FU, LV, and 5FU-LV NPs after incubation with HT-29 cells. *** <span class="html-italic">p</span> &lt; 0.001 in comparison to control and blank NPs. ΔΔ <span class="html-italic">p</span> &lt; 0.01, ΔΔΔ <span class="html-italic">p</span> &lt; 0.001 in comparison to same dose of free 5-FU and LV. [<a href="#B74-pharmaceutics-16-00734" class="html-bibr">74</a>,<a href="#B78-pharmaceutics-16-00734" class="html-bibr">78</a>]. [Reproduced with permission].</p> Full article ">Figure 6
<p>(<b>A</b>) Schematic demonstration of PLGA-1, 3-diaminopropane-folic acid nanodrug delivery system [<a href="#B81-pharmaceutics-16-00734" class="html-bibr">81</a>]. (<b>B</b>) Comparison of the cytotoxicity effect of 5-FU, 5-FU-PLGA, and 5-FU-PLGA-1, 3-diaminopropane-folic acid nanoparticles to HT-29 cells [<a href="#B81-pharmaceutics-16-00734" class="html-bibr">81</a>]. (<b>C</b>) Cellular uptake of bare MNCs and Arg@MNCs in HCT 116 cells [<a href="#B83-pharmaceutics-16-00734" class="html-bibr">83</a>]. (<b>D</b>) Cytotoxicity effect of 5-FU, 5-FU@MNCs, and 5-FU-Arg@MNCs on HCT 116 [<a href="#B83-pharmaceutics-16-00734" class="html-bibr">83</a>]. (<b>E</b>) T1 and (<b>F</b>) T2 vs. Mn concentration for bare MNCs and Arg@MNCs [<a href="#B83-pharmaceutics-16-00734" class="html-bibr">83</a>]. (<b>G</b>) T1-weighted MR images (left) and T2-weighted MR images (right) of the MNC and Arg@MNC aqueous solutions at various Mn concentrations [<a href="#B83-pharmaceutics-16-00734" class="html-bibr">83</a>]. (<b>H</b>) Schematic representation of action mechanism of 5FU and folate-liposomal 5FU in HeLa cell. [<a href="#B84-pharmaceutics-16-00734" class="html-bibr">84</a>]. (<b>I</b>) Changes in tumor volume and (<b>J</b>) photographs of tumor after treatment with control, 5-FU, and folate liposomal 5-FU [<a href="#B84-pharmaceutics-16-00734" class="html-bibr">84</a>]. [Reproduced with permission].</p> Full article ">
12 pages, 1988 KiB  
Article
Structural Optimization of Carboxy-Terminal Phenylalanine-Modified Dendrimers for T-Cell Association and Model Drug Loading
by Hiroya Shiba, Tomoka Hirose, Akinobu Sakai, Ikuhiko Nakase, Akikazu Matsumoto and Chie Kojima
Pharmaceutics 2024, 16(6), 715; https://doi.org/10.3390/pharmaceutics16060715 - 27 May 2024
Viewed by 1181
Abstract
Dendrimers are potent nanocarriers in drug delivery systems because their structure can be precisely controlled. We previously reported that polyamidoamine (PAMAM) dendrimers that were modified with 1,2-cyclohexanedicarboxylic acid (CHex) and phenylalanine (Phe), PAMAM-CHex-Phe, exhibited an effective association with various immune cells, including T-cells. [...] Read more.
Dendrimers are potent nanocarriers in drug delivery systems because their structure can be precisely controlled. We previously reported that polyamidoamine (PAMAM) dendrimers that were modified with 1,2-cyclohexanedicarboxylic acid (CHex) and phenylalanine (Phe), PAMAM-CHex-Phe, exhibited an effective association with various immune cells, including T-cells. In this study, we synthesized various carboxy-terminal Phe-modified dendrimers with different linkers using phthalic acid and linear dicarboxylic acids to determine the association of these dendrimers with Jurkat cells, a T-cell model. PAMAM-n-hexyl-Phe demonstrated the highest association with Jurkat T-cells. In addition, dendri-graft polylysine (DGL) with CHex and Phe, DGL-CHex-Phe, was synthesized, and its association with Jurkat cells was investigated. The association of DGL-CHex-Phe with T-cells was higher than that of PAMAM-CHex-Phe. However, it was insoluble in water and thus it is unsuitable as a drug carrier. Model drugs, such as protoporphyrin IX and paclitaxel, were loaded onto these dendrimers, and the most model drug molecules could be loaded into PAMAM-CHex-Phe. PTX-loaded PAMAM-CHex-Phe exhibited cytotoxicity against Jurkat cells at a similar level to free PTX. These results suggest that PAMAM-CHex-Phe exhibited both efficient T-cell association and drug loading properties. Full article
(This article belongs to the Special Issue Dendrimers: A Themed Issue in Honor of Professor Donald A. Tomalia on the Occasion of His 85th Birthday for His Outstanding Achievements in Advancing the Field of Dendrimers)
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Graphical abstract
Full article ">Figure 1
<p>Structures of (<b>a</b>) carboxy-terminal phenylalanine (Phe)-modified PAMAM dendrimers used in this study and (<b>b</b>) DGL of G2.</p> Full article ">Figure 2
<p>Synthetic scheme of carboxy-terminal phenylalanine (Phe)-modified PAMAM dendrimers and DGL with CHex (<b>top</b>), Ph (<b>middle</b>), and C<sub>n</sub> (<b>bottom</b>) linkers. MeO-C<sub>n</sub>-COOH are monomethyl adipate (<span class="html-italic">n</span> = 4), monomethyl suberate (<span class="html-italic">n</span> = 6), and monomethyl sebacate (<span class="html-italic">n</span> = 8), respectively.</p> Full article ">Figure 3
<p>Association of dendrimers with Jurkat cells. (<b>a</b>) Fluorescence intensity of PAMAM-R-Phe normalized to PAMAM-CHex. R means CHex, Ph and C<sub>n</sub> linkers. (<b>b</b>) Fluorescence intensity of DGL-CHex and DGL-CHex-Phe normalized to PAMAM-CHex. * <span class="html-italic">p</span> &lt; 0.05 vs. PAMAM-CHex.</p> Full article ">Figure 4
<p>Structure of carboxy-terminal phenylalanine (Phe)-modified PAMAM dendrimers via different linkers with four termini. (<b>a</b>) PAMAM-CHex-Phe, (<b>b</b>) PAMAM-Ph-Phe, and (<b>c</b>) PAMAM-C<sub>6</sub>-Phe. White, black, blue, and red balls correspond to hydrogen, carbon, nitrogen, and oxygen atoms, respectively.</p> Full article ">Figure 5
<p>Adsorption of carboxy-terminal phenylalanine (Phe)-modified dendrimers to liposomes via different linkers after the 3 h-incubation.</p> Full article ">Figure 6
<p>Chemical structure of (<b>a</b>) PpIX and (<b>b</b>) PTX.</p> Full article ">Figure 7
<p>Loading of (<b>a</b>) PpIX and (<b>b</b>) PTX into PAMAM-CHex-Phe, PAMAM-Ph-Phe, and PAMAM-C<sub>6</sub>-Phe in water.</p> Full article ">
13 pages, 635 KiB  
Review
Current Progress in Magnetic Resonance-Guided Focused Ultrasound to Facilitate Drug Delivery across the Blood-Brain Barrier
by Ying Meng, Lorraine V. Kalia, Suneil K. Kalia, Clement Hamani, Yuexi Huang, Kullervo Hynynen, Nir Lipsman and Benjamin Davidson
Pharmaceutics 2024, 16(6), 719; https://doi.org/10.3390/pharmaceutics16060719 - 27 May 2024
Cited by 2 | Viewed by 1799
Abstract
This review discusses the current progress in the clinical use of magnetic resonance-guided focused ultrasound (MRgFUS) and other ultrasound platforms to transiently permeabilize the blood-brain barrier (BBB) for drug delivery in neurological disorders and neuro-oncology. Safety trials in humans have followed on from [...] Read more.
This review discusses the current progress in the clinical use of magnetic resonance-guided focused ultrasound (MRgFUS) and other ultrasound platforms to transiently permeabilize the blood-brain barrier (BBB) for drug delivery in neurological disorders and neuro-oncology. Safety trials in humans have followed on from extensive pre-clinical studies, demonstrating a reassuring safety profile and paving the way for numerous translational clinical trials in Alzheimer’s disease, Parkinson’s disease, and primary and metastatic brain tumors. Future directions include improving ultrasound delivery devices, exploring alternative delivery approaches such as nanodroplets, and expanding the application to other neurological conditions. Full article
(This article belongs to the Special Issue Ultrasound-Mediated Blood–Brain Barrier Disruption and Drug Delivery)
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Figure 1

Figure 1
<p>Ultrasound-mediated drug delivery systems and mechanisms. (<b>A</b>) Three systems in use are illustrated. The ExAblate system utilizes a helmet lined with a phased array of ultrasound transducers separated from the scalp by cooled degassed water and operated within an MRI. The SonoCloud is an implanted device placed through a burr hole in the skull either at the time of a tumour resection surgery, or with an independently planned procedure. It is powered through a transdermal needle connected to an external power supply for each treatment. NaviFUS is a multi-channel array, like the ExAblate system, but utilizes a smaller surface area and is not performed within an MRI. (<b>B</b>) When microbubbles pass through sonicated tissue, they undergo cavitation, causing mechanical forces on the capillary wall, astrocytic endfeet, and pericytes, causing the temporary opening of the blood-brain barrier, allowing larger molecules to pass through into the parenchyma. This figure was adapted with permission from Figures 2 and 3 in Meng et al. (2021) [<a href="#B11-pharmaceutics-16-00719" class="html-bibr">11</a>].</p> Full article ">
17 pages, 3465 KiB  
Article
In Vitro and In Vivo Synergetic Radiotherapy with Gold Nanoparticles and Docetaxel for Pancreatic Cancer
by Abdulaziz Alhussan, Nolan Jackson, Norman Chow, Ermias Gete, Nicole Wretham, Nancy Dos Santos, Wayne Beckham, Cheryl Duzenli and Devika B. Chithrani
Pharmaceutics 2024, 16(6), 713; https://doi.org/10.3390/pharmaceutics16060713 - 26 May 2024
Cited by 2 | Viewed by 1578
Abstract
This research underscores the potential of combining nanotechnology with conventional therapies in cancer treatment, particularly for challenging cases like pancreatic cancer. We aimed to enhance pancreatic cancer treatment by investigating the synergistic effects of gold nanoparticles (GNPs) and docetaxel (DTX) as potential radiosensitizers [...] Read more.
This research underscores the potential of combining nanotechnology with conventional therapies in cancer treatment, particularly for challenging cases like pancreatic cancer. We aimed to enhance pancreatic cancer treatment by investigating the synergistic effects of gold nanoparticles (GNPs) and docetaxel (DTX) as potential radiosensitizers in radiotherapy (RT) both in vitro and in vivo, utilizing a MIA PaCa-2 monoculture spheroid model and NRG mice subcutaneously implanted with MIA PaCa-2 cells, respectively. Spheroids were treated with GNPs (7.5 μg/mL), DTX (100 nM), and 2 Gy of RT using a 6 MV linear accelerator. In parallel, mice received treatments of GNPs (2 mg/kg), DTX (6 mg/kg), and 5 Gy of RT (6 MV linear accelerator). In vitro results showed that though RT and DTX reduced spheroid size and increased DNA DSBs, the triple combination of DTX/RT/GNPs led to a significant 48% (p = 0.05) decrease in spheroid size and a 45% (p = 0.05) increase in DNA DSBs. In vivo results showed a 20% (p = 0.05) reduction in tumor growth 20 days post-treatment with (GNPs/RT/DTX) and an increase in mice median survival. The triple combination exhibited a synergistic effect, enhancing anticancer efficacy beyond individual treatments, and thus could be employed to improve radiotherapy and potentially reduce adverse effects. Full article
(This article belongs to the Special Issue Nanoparticle-Mediated Diagnostics and Drug Delivery Therapy)
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<p>Illustration depicting the integration of nanotechnology, chemotherapy, and radiotherapy (RT) as a combined approach for treating pancreatic cancer, applied both in vitro and in vivo.</p> Full article ">Figure 2
<p>Uptake and retention of gold nanoparticles in pancreatic cancer spheroids. (<b>A</b>) The measured uptake of 7.5 µg/mL GNPs in MIA PaCa-2 monoculture spheroids treated with GNPs alone and GNPs combined with DTX as determined by ICP-MS. *** indicates a significance level of <span class="html-italic">p</span> &lt; 0.001. (<b>B</b>) Cell cycle data for monoculture spheroid cells treated with DTX over a span of 3 days. (<b>C</b>) Confocal images showing the uptake and retention of GNPs, depicted in red, over a 3-day period in MIA PaCa-2 monoculture spheroids. These are categorized into three columns: GNPs only (first row) and GNPs plus DTX (second row).</p> Full article ">Figure 3
<p>Post-treatment size of monoculture spheroids. (<b>A</b>,<b>B</b>) Tracking the normalized size of monoculture spheroids across 14 days after treatment, with (<b>A</b>) indicating no radiation (0 Gy) and (<b>B</b>) showing the effect of 2 Gy radiation. (<b>C</b>) Brightfield images of monoculture spheroids captured 14 days following treatment, with a scale bar set to 200 µm.</p> Full article ">Figure 4
<p>Impact of DTX on in vivo tumor tissues. (<b>A</b>) The concentration of GNPs per gram of tumor tissue over time, comparing untreated tissues and those treated with DTX. Statistical significance is denoted as ** for <span class="html-italic">p</span> &lt; 0.01, and *** for <span class="html-italic">p</span> &lt; 0.001. (<b>B</b>) A cell cycle assay showing the changes over time in untreated tumor tissue and tissue treated with DTX. (<b>C</b>) Darkfield images of 4 µm sections of tumor tissues. These included untreated tissues and tissues treated with DTX. Scale bar set at 40 µm.</p> Full article ">Figure 5
<p>In vivo assessment of GNP/DTX/RT treatment. (<b>A</b>,<b>B</b>) The normalized tumor volume in mice post-treatment, with (<b>A</b>) illustrating results without radiation and (<b>B</b>) showing results with radiation. The data, representing an average normalized tumor volume, were obtained from at least five mice and are expressed as mean ± standard deviation, demonstrating the impact of different treatment strategies on tumor growth reduction. (<b>C</b>,<b>D</b>) The Kaplan–Meier survival curves for female NRG mice inoculated with MiaPaCa2 are shown without radiation (<b>C</b>) and with radiation (<b>D</b>), comparing the survival rates post-treatment with various strategies. The inset provides the median survival times of the mice in days following tumor incubation.</p> Full article ">Figure 6
<p>Irradiation effects on monoculture spheroids. (<b>A</b>) The size of normalized irradiated monoculture spheroids for 14 days following treatment, providing a longitudinal view of spheroid response to irradiation. (<b>B</b>) Brightfield images of these irradiated monoculture spheroids captured 14 days after treatment, with a scale bar of 200 µm. (<b>C</b>) Quantifying the average number of 53BP1 foci per cell in irradiated 2D monocultures. Statistical significance is denoted as ** for <span class="html-italic">p</span> &lt; 0.01, and *** for <span class="html-italic">p</span> &lt; 0.001. (<b>D</b>) Confocal microscopy images showcasing the repair protein 53BP1 within the nuclei of MIA PaCa-2 monoculture cells. A scale bar of 20 µm is included for size comparison. In these images, cell nuclei are colored blue, and 53BP1 foci are represented as green dots.</p> Full article ">
24 pages, 7549 KiB  
Article
Focused Ultrasound-Mediated Disruption of the Blood–Brain Barrier for AAV9 Delivery in a Mouse Model of Huntington’s Disease
by Bernie S. Owusu-Yaw, Yongzhi Zhang, Lilyan Garrett, Alvin Yao, Kai Shing, Ana Rita Batista, Miguel Sena-Esteves, Jaymin Upadhyay, Kimberly Kegel-Gleason and Nick Todd
Pharmaceutics 2024, 16(6), 710; https://doi.org/10.3390/pharmaceutics16060710 - 24 May 2024
Viewed by 1401
Abstract
Huntington’s disease (HD) is a monogenic neurodegenerative disorder caused by a cytosine–adenine–guanine (CAG) trinucleotide repeat expansion in the HTT gene. There are no cures for HD, but the genetic basis of this disorder makes gene therapy a viable approach. Adeno-associated virus (AAV)-miRNA-based therapies [...] Read more.
Huntington’s disease (HD) is a monogenic neurodegenerative disorder caused by a cytosine–adenine–guanine (CAG) trinucleotide repeat expansion in the HTT gene. There are no cures for HD, but the genetic basis of this disorder makes gene therapy a viable approach. Adeno-associated virus (AAV)-miRNA-based therapies have been demonstrated to be effective in lowering HTT mRNA; however, the blood–brain barrier (BBB) poses a significant challenge for gene delivery to the brain. Delivery strategies include direct injections into the central nervous system, which are invasive and can result in poor diffusion of viral particles through the brain parenchyma. Focused ultrasound (FUS) is an alternative approach that can be used to non-invasively deliver AAVs by temporarily disrupting the BBB. Here, we investigate FUS-mediated delivery of a single-stranded AAV9 bearing a cDNA for GFP in 2-month-old wild-type mice and the zQ175 HD mouse model at 2-, 6-, and 12-months. FUS treatment improved AAV9 delivery for all mouse groups. The delivery efficacy was similar for all WT and HD groups, with the exception of the zQ175 12-month cohort, where we observed decreased GFP expression. Astrocytosis did not increase after FUS treatment, even within the zQ175 12-month group exhibiting higher baseline levels of GFAP expression. These findings demonstrate that FUS can be used to non-invasively deliver an AAV9-based gene therapy to targeted brain regions in a mouse model of Huntington’s disease. Full article
(This article belongs to the Section Drug Delivery and Controlled Release)
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Graphical abstract

Graphical abstract
Full article ">Figure 1
<p>Schematic diagram of FUS-AAV delivery procedure. Mice were treated unilaterally with FUS targeting four regions in the right striatum. Following FUS treatment, mice were injected intravenously with 5.5 × 10<sup>11</sup> vg/mouse (2.2 × 10<sup>10</sup> vg/g) of ss-AAV9-U6-miR10150-CBA-GFP and MRI images were acquired before and after gadolinium injections to confirm BBB opening in the targeted brain region. Three weeks after treatment, mice were euthanized, and the brains were cut into coronal sections. The sections were stained for Hoechst (nuclei), GFP, NeuN (neurons), Iba1 (microglia), GFAP (reactive astrocytes), and S100β (astrocytes) to assess GFP area coverage and cell-type specificity of GFP expression. MB, microbubbles; IV, intravenous; MRI, magnetic resonance imaging; Gad, gadolinium; IF, immunofluorescence.</p> Full article ">Figure 2
<p>Diagram of AAV9-U6-miRNA10150-CBA-GFP construct.</p> Full article ">Figure 3
<p>BBB opening confirmation with T1-weighted contrast-enhanced MRI. (<b>A</b>) Gadolinium-enhanced T1-weighted coronal images (presented as percent change before and after gadolinium injections) showing the location and extent of BBB opening for each animal in the WT 2-month, zQ175 2-month, and zQ175 6-month-old groups, acquired with a 7T MRI scanner. The images presented for the zQ175 12-month cohort were acquired with a 3T scanner. The MRI images demonstrate an increase in BBB permeability in the FUS-treated hemisphere (indicated with black arrows) compared to the untreated hemisphere. (<b>B</b>) The Gad-enhanced T1-weighted images were co-registered to a mouse brain template to determine percent signal change in the striatum (outlined in red). (<b>C</b>) Percent signal change in the striatum of WT 2-month (<span class="html-italic">n</span> = 6), zQ175 2-month (<span class="html-italic">n</span> = 6), zQ175 6-month (<span class="html-italic">n</span> = 6), and zQ175 12-month-old mice (<span class="html-italic">n</span> = 6). Two-way ANOVA and Tukey’s post hoc test for multiple comparisons were used for statistical analysis. * <span class="html-italic">p</span> &lt; 0.05, *** <span class="html-italic">p</span> &lt; 0.001.</p> Full article ">Figure 4
<p>FUS treatment enhances gene delivery in sonicated hemisphere. Mice were treated with FUS targeting four areas in the right striatum and then injected with ss-AAV9-U6-miR10150-CBA-GFP intravenously. Coronal sections stained for NeuN (purple), GFP (green), and Hoechst (not shown) represent one section for each FUS-treated mouse. GFP expression was detected in the cortex and striatum of the FUS-treated hemisphere by immunofluorescence. The red boxes indicate missing sections due to the death of mice zQ175_11 and zQ175_5. Magnification, 20×; scale bar represents 500 µm. WT 2-month (<span class="html-italic">n</span> = 6), zQ175 2-month (<span class="html-italic">n</span> = 5), zQ175 6-month (<span class="html-italic">n</span> = 5), and zQ175 12-month (<span class="html-italic">n</span> = 6).</p> Full article ">Figure 5
<p>GFP expression in the striatum and cortex following FUS treatment. Representative coronal sections of heterogenous GFP expression in WT 2-month, zQ175 2-month, zQ175 6-month, and zQ175 12-month-old mouse brains following FUS treatment and intravenous injection of ss-AAV9-U6-miR10150-CBA-GFP. The dashed white boxes indicate regions of interest (ROIs) in the cortex and striatum that were zoomed into for figures (<b>E</b>–<b>L</b>). (<b>A</b>–<b>D</b>) Example whole-brain images showing GFP expression in the cortex and striatum inside the sonicated hemisphere of WT and zQ175 HD mice; Magnification, 20×. (<b>E</b>–<b>H</b>) Zoomed ROI of GFP expression in the FUS-treated cortex of WT 2-month, zQ175 2-month, zQ175 6-month, and zQ175 12-month mice; (<b>I</b>–<b>L</b>) zoomed ROI of GFP expression in the FUS-treated striatum of WT 2-month, zQ175 2-month, zQ175 6-month, and zQ175 12-month-old mice. Scale bar represents 500 µm (<b>A</b>–<b>D</b>) and 100 µm (<b>E</b>–<b>L</b>).</p> Full article ">Figure 6
<p>FUS-mediated delivery of ss-AAV9-U6-miR10150-CBA-GFP. (<b>A</b>) Representative images of whole-brain coronal sections stained for GFP (green), NeuN (purple), Iba1 (brown), GFAP (cyan), and S100β (red) from WT 2-month, zQ175 2-month, zQ175 6-month, and zQ175 12-month-old mice following FUS treatment and intravenous administration of ss-AAV9-U6-miR10150-CBA-GFP at 5.5 <span class="html-italic">×</span> 10<sup>11</sup> vg/mouse. Magnification, 20×; scale bar represents 500 µm. (<b>B</b>) Quantification of GFP coverage by the percent area of each hemisphere with GFP signal above threshold. WT 2-month (<span class="html-italic">n</span> = 6), zQ175 2-month (<span class="html-italic">n</span> = 5), zQ175 6-month (<span class="html-italic">n</span> = 5) and zQ175 12-month (<span class="html-italic">n</span> = 6). Data were analyzed using a two-way ANOVA followed by Tukey’s multiple comparisons test. * <span class="html-italic">p</span> &lt; 0.05, *** <span class="html-italic">p</span> &lt; 0.001, ****<span class="html-italic">p</span> &lt; 0.0001. (<b>C</b>) Analysis of GFP area coverage using an image processing script in MATLAB. An ROI was drawn around each hemisphere, then the two hemispheres were overlayed with a mask, and then % GFP area was quantified in each hemisphere based on GFP signal above threshold.</p> Full article ">Figure 7
<p>GFP expression in neurons in the FUS-treated hemisphere following AAV administration. Confocal microscopic analysis of Hoechst (blue), GFP (green), and NeuN (magenta) expression in (<b>A</b>) the cortex and (<b>B</b>) striatum of the FUS-treated hemisphere of WT 2-month, zQ175 2-month, zQ175 6-month and zQ175 12-month-old mice injected intravenously with ss-AAV9-U6-miR10150-CBA-GFP at 5.5 <span class="html-italic">×</span> 10<sup>11</sup> vg/mouse. Yellow arrows highlight examples of colocalization of GFP (green) with the neuronal marker NeuN (magenta). Scale bar represents 50 µm. (<b>C</b>) Percentage of GFP<sup>+</sup> cells that are neurons in the striatum and cortex. (<b>D</b>) Percentage of Hoechst cells that are GFP<sup>+</sup> and NeuN<sup>+</sup> in the striatum and cortex. WT 2-month (<span class="html-italic">n</span> = 6), zQ175 2-month (<span class="html-italic">n</span> = 5), zQ175 6-month (<span class="html-italic">n</span> = 5), and zQ175 12-month (<span class="html-italic">n</span> = 6). Data were analyzed using a two-way ANOVA followed by Tukey’s multiple comparisons test. * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001. Data shown are mean ± SEM.</p> Full article ">Figure 8
<p>GFP expression in astrocytes in the FUS-treated hemisphere following AAV administration. Confocal microscopic analysis of Hoechst (blue), GFP (green), and S100β (red) staining in the (<b>A</b>) cortex and (<b>B</b>) striatum of the FUS-treated hemisphere of WT 2-month, zQ175 2-month, zQ175 6-month, and zQ175 12-month-old mice injected intravenously with ss-AAV9-U6-miR10150-CBA-GFP at 5.5 <span class="html-italic">×</span> 10<sup>11</sup> vg/mouse. Yellow arrows highlight colocalization of GFP (green) with the astrocytic marker S100β (red). Scale bar represents 50 µm. (<b>C</b>) Percentage of GFP<sup>+</sup> cells that are astrocytes in the striatum and cortex. (<b>D</b>) Percentage of Hoechst<sup>+</sup> cells that are GFP<sup>+</sup> and S100b<sup><span class="html-small-caps">+</span></sup> cells in the striatum and cortex. WT 2-month (<span class="html-italic">n</span> = 6), zQ175 2-month (<span class="html-italic">n</span> = 5), zQ175 6-month (<span class="html-italic">n</span> = 5), and zQ175 12-month (<span class="html-italic">n</span> = 6)-old mice. Data were analyzed using a two-way ANOVA followed by Tukey’s multiple comparisons test. * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01. Data shown are mean ± SEM.</p> Full article ">Figure 9
<p>GFAP fluorescence intensity. (<b>A</b>) Quantification of GFAP fluorescence in the cortex and (<b>B</b>) striatum of FUS-treated WT 2-month (<span class="html-italic">n</span> = 6), zQ175 2-month (<span class="html-italic">n</span> = 5), zQ175 6-month (<span class="html-italic">n</span> = 5), and zQ175 12-month (<span class="html-italic">n</span> = 6) mice following AAV delivery. * <span class="html-italic">p</span> &lt; 0.05, **** <span class="html-italic">p</span> &lt; 0.0001. Data shown are mean ± SEM.</p> Full article ">Figure 10
<p>Iba1 expression in the cortex and striatum of WT and HD mice after 3 weeks following treatment. (<b>A</b>–<b>D</b>) Representative whole-brain images of Iba1 (magenta) and GFP staining for each experimental group. Magnification 20. Confocal microscopic analysis of Iba1 (magenta) staining in the cortex and striatum of the FUS-treated hemisphere and contralateral side in (<b>A</b>,<b>E</b>–<b>H</b>) WT 2-month, (<b>B</b>,<b>I</b>–<b>L</b>) zQ175 2-month, (<b>C</b>,<b>M</b>–<b>P</b>) zQ175 6-month, and (<b>D</b>,<b>Q</b>–<b>T</b>) zQ175 12-month mice. Scale bar represents 500 µm for (<b>A</b>–<b>D</b>) and 50 µm for the rest.</p> Full article ">
27 pages, 4655 KiB  
Review
Complex Emulsions as an Innovative Pharmaceutical Dosage form in Addressing the Issues of Multi-Drug Therapy and Polypharmacy Challenges
by Naresh Yandrapalli
Pharmaceutics 2024, 16(6), 707; https://doi.org/10.3390/pharmaceutics16060707 - 24 May 2024
Viewed by 1502
Abstract
This review explores the intersection of microfluidic technology and complex emulsion development as a promising solution to the challenges of formulations in multi-drug therapy (MDT) and polypharmacy. The convergence of microfluidic technology and complex emulsion fabrication could herald a transformative era in multi-drug [...] Read more.
This review explores the intersection of microfluidic technology and complex emulsion development as a promising solution to the challenges of formulations in multi-drug therapy (MDT) and polypharmacy. The convergence of microfluidic technology and complex emulsion fabrication could herald a transformative era in multi-drug delivery systems, directly confronting the prevalent challenges of polypharmacy. Microfluidics, with its unparalleled precision in droplet formation, empowers the encapsulation of multiple drugs within singular emulsion particles. The ability to engineer emulsions with tailored properties—such as size, composition, and release kinetics—enables the creation of highly efficient drug delivery vehicles. Thus, this innovative approach not only simplifies medication regimens by significantly reducing the number of necessary doses but also minimizes the pill burden and associated treatment termination—issues associated with polypharmacy. It is important to bring forth the opportunities and challenges of this synergy between microfluidic-driven complex emulsions and multi-drug therapy poses. Together, they not only offer a sophisticated method for addressing the intricacies of delivering multiple drugs but also align with broader healthcare objectives of enhancing treatment outcomes, patient safety, and quality of life, underscoring the importance of dosage form innovations in tackling the multifaceted challenges of modern pharmacotherapy. Full article
(This article belongs to the Special Issue Micro- and Nano-Emulsions as Drug Delivery Systems)
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Figure 1

Figure 1
<p>Transformation of traditional, multiple-dosage-form prescription into a single dosage form using microfluidics. In the schematic above, each active ingredient/dosage form is depicted using different colours. These active ingredient solutions can be transitioned into a complex emulsion with each ingredient occupying different compartments that form the complex emulsion. In this case, four different active ingredients can be encapsulated in a single complex emulsion.</p> Full article ">Figure 2
<p>Microfluidic flow geometries and emulsion types. (<b>a</b>) Schematic representation of different microfluidic geometries typically used to produce droplets—T-junction/co-flow/flow-focusing methodologies. (<b>b</b>) Different types of complex emulsions and schematic representations of their production methods using PDMS-based microfluidics. The blue colour and yellow colours represent different immiscible fluids, i.e., aqueous or oil or gas.</p> Full article ">Figure 3
<p>Double-emulsion-type complex emulsion production using microfluidics. (<b>a</b>,<b>b</b>) Double-emulsion-templated production of liposomes with high precision in size uniformity and encapsulation efficiency (Reprinted/adapted with permission from Ref. [<a href="#B22-pharmaceutics-16-00707" class="html-bibr">22</a>]. 2021, Springer Nature)”. (<b>c</b>) Highly uniform liposomes containing different encapsulated; from left to right: plasmid DNA, small unilamellar vesicles, cells, (Reprinted/adapted with permission from Ref. [<a href="#B22-pharmaceutics-16-00707" class="html-bibr">22</a>]. 2021, Springer Nature) and double emulsions with graphitic carbon nitride nanoparticles within the hydrophobic phase produced using PDMS-based microfluidics (Reprinted/adapted with permission from Ref. [<a href="#B20-pharmaceutics-16-00707" class="html-bibr">20</a>]. 2020, John Wiley and Sons). (<b>d</b>) Formation of microfluidic W/O/W using prefabricated W/O emulsion (mechanical agitation) with dual drug encapsulation for oral drug delivery (Reprinted/adapted with permission from Ref. [<a href="#B24-pharmaceutics-16-00707" class="html-bibr">24</a>]. 2015, American Chemical Society). (<b>e</b>) Microfluidic production of W/O/W emulsions with dual drug encapsulation along with drug-loaded nanoparticles for site-specific release properties after polymerization to treat inflammatory bowel disease delivery (Reprinted/adapted with permission from Ref. [<a href="#B32-pharmaceutics-16-00707" class="html-bibr">32</a>]. 2021, Royal Society of Chemistry).</p> Full article ">Figure 4
<p>Nested emulsions. (<b>a</b>) PDMS-based production methodology for nested emulsions with increasing number of inner compartments, from one to five (all compartments are produced using same inner fluid). Inset showing the micrographs of the nested emulsions (Reprinted/adapted with permission from Ref. [<a href="#B78-pharmaceutics-16-00707" class="html-bibr">78</a>]. 2023, Naresh Yandrapalli). (<b>b</b>) Nested emulsions produced from two different inner solutions: red, Rhodamine B; green, Wright stain (Reprinted/adapted with permission from Ref. [<a href="#B65-pharmaceutics-16-00707" class="html-bibr">65</a>]. 2018, Elsevier). (<b>c</b>) Nested emulsions with prefabricated polymersomes with three different inner compartments (Reprinted/adapted with permission from Ref. [<a href="#B63-pharmaceutics-16-00707" class="html-bibr">63</a>]. 2011, American Chemical Society). (<b>d</b>) Layered emulsions with multiple hydrophobic and hydrophilic compartments produced using sequential layering of W/O to W/OW to W/O/W/O to W/O/W/O/W and their eventual conversion into double layered microcapsules with interesting cargo release properties (Reprinted/adapted with permission from Ref. [<a href="#B68-pharmaceutics-16-00707" class="html-bibr">68</a>]. 2018, Wiley and Sons). (<b>e</b>) Layered emulsions produced through encapsulation of preformed emulsions or polymersomes encapsulating different cargoes (shown in red and green colour) (Reprinted/adapted with permission from Ref. [<a href="#B63-pharmaceutics-16-00707" class="html-bibr">63</a>]. 2011, American Chemical Society). (<b>f</b>) Multi-layered emulsion showing triple membrane structure after polymerization, inset showing sequential disintegration of membranes leading to cargo release (Reprinted/adapted with permission from Ref. [<a href="#B63-pharmaceutics-16-00707" class="html-bibr">63</a>]. 2011, American Chemical Society).</p> Full article ">Figure 5
<p>Drug release kinetics in double and complex emulsions. (<b>a</b>) Osmotic-pressure-induced drug release from W/O/W double-emulsion-based PLGA drug delivery vehicles (Reprinted/adapted with permission from Ref. [<a href="#B81-pharmaceutics-16-00707" class="html-bibr">81</a>]. 2017, Wiley and Sons). (<b>b</b>) Photo-switchable drug delivery carriers for programmable drug delivery in the presence of UV illumination (Reprinted/adapted with permission from Ref. [<a href="#B86-pharmaceutics-16-00707" class="html-bibr">86</a>]. 2023, American Chemical Society). (<b>c</b>) Layered-emulsion-based drug delivery vehicles showing both sequential and simultaneous dual drug delivery (Reprinted/adapted with permission from Ref. [<a href="#B67-pharmaceutics-16-00707" class="html-bibr">67</a>]. 2018, Wiley and Sons). (<b>d</b>) Osmotic-pressure-induced burst release of gold nanoparticles from polymerized nested emulsions (Reprinted/adapted with permission from Ref. [<a href="#B25-pharmaceutics-16-00707" class="html-bibr">25</a>]. 2022, Wiley and Sons).</p> Full article ">Figure 6
<p>Challenges in addressing complex emulsions as a pharmaceutical formulation. Stability of the emulsions dictated by the concentrations of electrolytes and surfactants inducing emulsion destabilization via coalescence and osmotic pressure differences. Leakage of drugs from the emulsions renders the applications of the complex emulsions, especially passive leakage via diffusion and permeation to surfactant membranes. Emulsion formulation including the rheology for enhanced stability and choice of drug components to prevent adverse reactions is essential. Scalability: industrial-scale production is necessary for wider utilization and applications of the complex emulsions, which could be achieved through parallelization ((Reprinted/adapted with permission from Ref. [<a href="#B113-pharmaceutics-16-00707" class="html-bibr">113</a>]. 2012, Royal Society of Chemistry) of the devices or 3D printing methodologies ((Reprinted/adapted with permission from Ref. [<a href="#B65-pharmaceutics-16-00707" class="html-bibr">65</a>]. 2018, Elsevier).</p> Full article ">
32 pages, 5508 KiB  
Review
Drug Development for Alzheimer’s and Parkinson’s Disease: Where Do We Go Now?
by Lisa Sequeira, Sofia Benfeito, Carlos Fernandes, Inês Lima, Joana Peixoto, Catarina Alves, Cláudia Sofia Machado, Alexandra Gaspar, Fernanda Borges and Daniel Chavarria
Pharmaceutics 2024, 16(6), 708; https://doi.org/10.3390/pharmaceutics16060708 - 24 May 2024
Cited by 4 | Viewed by 1855
Abstract
Neurodegenerative diseases (NDs) are a set of progressive, chronic, and incurable diseases characterized by the gradual loss of neurons, culminating in the decline of cognitive and/or motor functions. Alzheimer’s disease (AD) and Parkinson’s disease (PD) are the most common NDs and represent an [...] Read more.
Neurodegenerative diseases (NDs) are a set of progressive, chronic, and incurable diseases characterized by the gradual loss of neurons, culminating in the decline of cognitive and/or motor functions. Alzheimer’s disease (AD) and Parkinson’s disease (PD) are the most common NDs and represent an enormous burden both in terms of human suffering and economic cost. The available therapies for AD and PD only provide symptomatic and palliative relief for a limited period and are unable to modify the diseases’ progression. Over the last decades, research efforts have been focused on developing new pharmacological treatments for these NDs. However, to date, no breakthrough treatment has been discovered. Hence, the development of disease-modifying drugs able to halt or reverse the progression of NDs remains an unmet clinical need. This review summarizes the major hallmarks of AD and PD and the drugs available for pharmacological treatment. It also sheds light on potential directions that can be pursued to develop new, disease-modifying drugs to treat AD and PD, describing as representative examples some advances in the development of drug candidates targeting oxidative stress and adenosine A2A receptors. Full article
(This article belongs to the Special Issue The Therapy of Alzheimer’s Disease: Towards a New Generation of Drugs)
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Graphical abstract

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<p>Primary brain regions affected in the most common neurodegenerative diseases. Adapted from [<a href="#B6-pharmaceutics-16-00708" class="html-bibr">6</a>].</p> Full article ">Figure 2
<p>Major hallmarks of AD. (<b>A</b>) Degeneration of cholinergic neurons in the basal forebrain. (<b>B</b>) Formation of senile plaques and neurofibrillary tangles. Adapted from [<a href="#B52-pharmaceutics-16-00708" class="html-bibr">52</a>].</p> Full article ">Figure 3
<p>Major hallmarks of Parkinson’s disease. (<b>A</b>) Degeneration of nigrostriatal dopaminergic neurons. (<b>B</b>) Formation of Lewy inclusions. Adapted from [<a href="#B80-pharmaceutics-16-00708" class="html-bibr">80</a>,<a href="#B81-pharmaceutics-16-00708" class="html-bibr">81</a>].</p> Full article ">Figure 4
<p>Enzymes and transporters involved in the synthesis, storage, and metabolism of acetylcholine. Abbreviations: Ach, acetylcholine; AchE, acetylcholinesterase; BchE, butyrylcholinesterase; ChT1, high-affinity choline transporter; VAT, vesicular acetylcholine transporter. Adapted from [<a href="#B110-pharmaceutics-16-00708" class="html-bibr">110</a>,<a href="#B113-pharmaceutics-16-00708" class="html-bibr">113</a>].</p> Full article ">Figure 5
<p>Drugs in clinical use for AD treatment.</p> Full article ">Figure 6
<p>Enzymes and transporters involved in the synthesis, storage, and metabolism of dopamine. Abbreviations: 3-MT, 3-methoxytyramine; AADC, aromatic amino acid decarboxylase; COMT, catechol-O-methyltransferase; DA, dopamine; DAT, dopamine transporter; DOPAC, 3,4-Dihydroxyphenylacetic acid; HVA, homovanillic acid; L-DOPA, L-3,4-dihydroxyphenylalanine; MAO-A, monoamine oxidase A; MAO-B, monoamine oxidase B; TH, tyrosine hydroxylase; VMAT2, vesicular monoamine transporter. Adapted from [<a href="#B130-pharmaceutics-16-00708" class="html-bibr">130</a>,<a href="#B131-pharmaceutics-16-00708" class="html-bibr">131</a>].</p> Full article ">Figure 7
<p>Drugs in clinical use for PD treatment.</p> Full article ">Figure 8
<p>Pathological events of neurodegenerative diseases associated with increased oxidative stress.</p> Full article ">Figure 9
<p>Formation of ROS in mitochondria. Abbreviations: ADP, adenosine diphosphate; ATP, adenosine triphosphate; CAT, catalase; Cyt C, Cytochrome C; GPx, Glutathione peroxidase; I, complex I; II, complex II; III, complex III; IV, complex IV; NADH, Nicotinamide adenine dinucleotide; Q, coenzyme Q10; SOD1, superoxide dismutase 1; SOD2, superoxide dismutase 2; V, complex V (ATP synthase). Adapted from [<a href="#B119-pharmaceutics-16-00708" class="html-bibr">119</a>,<a href="#B220-pharmaceutics-16-00708" class="html-bibr">220</a>].</p> Full article ">Figure 10
<p>Generation of ROS during enzymatic (dashed line) and non-enzymatic (plain line) DA decomposition. Abbreviations: ALDH, aldehyde dehydrogenase; COMT, catechol O-methyltransferase; DA, dopamine; DA-quinone, dopamine quinone; MAO, monoamine oxidase, NAD(P)H, nicotinamide adenine dinucleotide (phosphate). Adapted from [<a href="#B229-pharmaceutics-16-00708" class="html-bibr">229</a>].</p> Full article ">Figure 11
<p>(<b>A</b>) Mitochondrial uptake of lipophilic cations. (<b>B</b>) Representative examples of lipophilic cations to target bioactive molecules to mitochondria. Adapted from [<a href="#B255-pharmaceutics-16-00708" class="html-bibr">255</a>,<a href="#B257-pharmaceutics-16-00708" class="html-bibr">257</a>].</p> Full article ">Figure 12
<p>Schematic representation of G protein-coupled adenosine receptors.</p> Full article ">Figure 13
<p>Chemical structures of selective A<sub>2A</sub> AR antagonists evaluated in clinical trials.</p> Full article ">
34 pages, 1507 KiB  
Review
Process Simulation of Twin-Screw Granulation: A Review
by Tony Bediako Arthur and Nejat Rahmanian
Pharmaceutics 2024, 16(6), 706; https://doi.org/10.3390/pharmaceutics16060706 - 24 May 2024
Cited by 1 | Viewed by 3851
Abstract
Twin-screw granulation has emerged as a key process in powder processing industries and in the pharmaceutical sector to produce granules with controlled properties. This comprehensive review provides an overview of the simulation techniques and approaches that have been employed in the study of [...] Read more.
Twin-screw granulation has emerged as a key process in powder processing industries and in the pharmaceutical sector to produce granules with controlled properties. This comprehensive review provides an overview of the simulation techniques and approaches that have been employed in the study of twin-screw granulation processes. This review discusses the major aspects of the twin-screw granulation process which include the fundamental principles of twin-screw granulation, equipment design, process parameters, and simulation methodologies. It highlights the importance of operating conditions and formulation designs in powder flow dynamics, mixing behaviour, and particle interactions within the twin-screw granulator for enhancing product quality and process efficiency. Simulation techniques such as the population balance model (PBM), computational fluid dynamics (CFD), the discrete element method (DEM), process modelling software (PMS), and other coupled techniques are critically discussed with a focus on simulating twin-screw granulation processes. This paper examines the challenges and limitations associated with each simulation approach and provides insights into future research directions. Overall, this article serves as a valuable resource for researchers who intend to develop their understanding of twin-screw granulation and provides insights into the various techniques and approaches available for simulating the twin-screw granulation process. Full article
(This article belongs to the Special Issue Pharmaceutical Solids: Advanced Manufacturing and Characterization)
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<p>A schematic of a continuous twin-screw granulation [<a href="#B2-pharmaceutics-16-00706" class="html-bibr">2</a>].</p> Full article ">Figure 2
<p>Twin-screw element, from Aftab (2018) [<a href="#B31-pharmaceutics-16-00706" class="html-bibr">31</a>].</p> Full article ">Figure 3
<p>Schematic of simulation workflow.</p> Full article ">
17 pages, 4839 KiB  
Article
Risperidone-Loaded Nasal Thermosensitive Polymeric Micelles: Quality by Design-Based Formulation Study
by Bence Sipos, Gábor Katona and Ildikó Csóka
Pharmaceutics 2024, 16(6), 703; https://doi.org/10.3390/pharmaceutics16060703 - 24 May 2024
Cited by 1 | Viewed by 1132
Abstract
The current research aims to develop thermosensitive polymeric micelles loaded with risperidone for nasal administration, emphasizing the added benefits of their thermosensitive behavior under nasal conditions. An initial risk assessment facilitated the advanced development process, confirming that the key indicators of thermosensitivity were [...] Read more.
The current research aims to develop thermosensitive polymeric micelles loaded with risperidone for nasal administration, emphasizing the added benefits of their thermosensitive behavior under nasal conditions. An initial risk assessment facilitated the advanced development process, confirming that the key indicators of thermosensitivity were suitable for nasal application. The polymeric micelles exhibited an average size of 118.4 ± 3.1 nm at ambient temperature and a size of 20.47 ± 1.2 nm at 36.5 °C, in both cases in monodisperse distribution. Factors such as pH and viscosity did not significantly impact these parameters, demonstrating appropriate nasal applicability. The model formulations showed a rapid, burst-like drug release profile in vitro, accompanied by a quick and high permeation rate at nasal conditions. Overall, the Quality by Design-based risk assessment process led to the development of an advanced drug delivery system capable of administering risperidone through the nasal cavity. Full article
(This article belongs to the Special Issue Nasal Drug Delivery: Challenges and Future Opportunities)
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Graphical abstract
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<p>Interdependence rating amongst CQA–QTPP (<b>A</b>) and CQA–CMA/CPP elements (<b>B</b>).</p> Full article ">Figure 2
<p>Result of the risk assessment process. The severity scores of each Critical Quality Attribute (<b>A</b>) and Critical Material Attribute/Critical Process Parameter (<b>B</b>).</p> Full article ">Figure 3
<p>Determination of LCST via dynamic light scattering measurements. The decrease in micelle size was registered upon the increase in temperature from 25 to 40 °C. The red lines indicate the proper range for thermosensitive systems regarding the LCST value. Data are presented as means ± SD (<span class="html-italic">n</span> = 3).</p> Full article ">Figure 4
<p>Micelle size (expressed as average hydrodynamic diameter) (<b>A</b>) and micelle size distribution (expressed as polydispersity index) (<b>B</b>) at 36.5 °C. Data are presented as means ± SD (<span class="html-italic">n</span> = 3).</p> Full article ">Figure 5
<p>Change in micelle size (expressed as average hydrodynamic diameter) in the temperature interval of 25–40 °C. Data are presented as means ± SD (<span class="html-italic">n</span> = 3).</p> Full article ">Figure 6
<p>The 3D surface plots of the factorial design. The effect of the independent factors on micelle size (<b>A</b>) and micelle size distribution (polydispersity index) (<b>B</b>).</p> Full article ">Figure 7
<p>The effect of pH on the micelle size (<b>A</b>) and the micelle size distribution (<b>B</b>). The effect of viscosity on the micelle size (<b>C</b>) and the micelle size distribution (<b>D</b>). Data are presented as means ± SD (<span class="html-italic">n</span> = 3).</p> Full article ">Figure 8
<p>The effect of pH on thermodynamic solubility (<b>A</b>), on encapsulation efficiency (<b>B</b>), and the effect of viscosity on thermodynamic solubility (<b>C</b>) and encapsulation efficiency (<b>D</b>). Data are presented as means ± SD (<span class="html-italic">n</span> = 3).</p> Full article ">Figure 9
<p>In vitro drug release study (<b>A</b>) and in vitro drug permeation study (<b>B</b>) with the model formulations compared to the initial RIS. Both measurements took place at simulated nasal conditions. Data are presented as means ± SD (<span class="html-italic">n</span> = 3).</p> Full article ">
41 pages, 2380 KiB  
Review
Insight into the Functional Dynamics and Challenges of Exosomes in Pharmaceutical Innovation and Precision Medicine
by Anu Sharma, Anita Yadav, Aparajita Nandy and Subhadip Ghatak
Pharmaceutics 2024, 16(6), 709; https://doi.org/10.3390/pharmaceutics16060709 - 24 May 2024
Cited by 5 | Viewed by 2038
Abstract
Of all the numerous nanosized extracellular vesicles released by a cell, the endosomal-originated exosomes are increasingly recognized as potential therapeutics, owing to their inherent stability, low immunogenicity, and targeted delivery capabilities. This review critically evaluates the transformative potential of exosome-based modalities across pharmaceutical [...] Read more.
Of all the numerous nanosized extracellular vesicles released by a cell, the endosomal-originated exosomes are increasingly recognized as potential therapeutics, owing to their inherent stability, low immunogenicity, and targeted delivery capabilities. This review critically evaluates the transformative potential of exosome-based modalities across pharmaceutical and precision medicine landscapes. Because of their precise targeted biomolecular cargo delivery, exosomes are posited as ideal candidates in drug delivery, enhancing regenerative medicine strategies, and advancing diagnostic technologies. Despite the significant market growth projections of exosome therapy, its utilization is encumbered by substantial scientific and regulatory challenges. These include the lack of universally accepted protocols for exosome isolation and the complexities associated with navigating the regulatory environment, particularly the guidelines set forth by the U.S. Food and Drug Administration (FDA). This review presents a comprehensive overview of current research trajectories aimed at addressing these impediments and discusses prospective advancements that could substantiate the clinical translation of exosomal therapies. By providing a comprehensive analysis of both the capabilities and hurdles inherent to exosome therapeutic applications, this article aims to inform and direct future research paradigms, thereby fostering the integration of exosomal systems into mainstream clinical practice. Full article
(This article belongs to the Special Issue Extracellular Vesicle-Based Drug Delivery Systems)
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<p>Exosome-based therapy in clinical trials (as listed on <a href="http://clinicaltrials.gov" target="_blank">clinicaltrials.gov</a> as accessed on 20 April 2024).</p> Full article ">Figure 2
<p>Illustration of the protein corona formation around exosomes.</p> Full article ">Figure 3
<p>Timeline outlining the advancement in exosome-based therapy.</p> Full article ">Figure 4
<p>A schematic representation of the different strategies used for modifying the exosomal surface for targeted therapy and imaging applications. These strategies include labelling with antibodies, fluorescent dyes, radiolabeling, and other engineering techniques to enhance delivery specificity.</p> Full article ">
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