TW201206457A - Reishi polysaccharide-based compositions and methods for treatment of cancer - Google Patents
Reishi polysaccharide-based compositions and methods for treatment of cancer Download PDFInfo
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Description
201206457 /、 發明說明： 【發明所屬之技術領域】 本發療癌症之方法及组成物。具體而言 更 ，ΕΜΤ來治療跡絲及移轉之方法 萃 ^癌及=i:有’用於調節ΕΜΤ及抑制癌症病程，包括 取二。〒；D，盆之（Ganoderma lucidum，Reishi 或 Ling_Zhi) 【先前技術】 _1皮=’舉例而言，攝護腺癌、乳癌、大腸癌、肺癌、 4的二印巢癌、脾喊癌、睪丸癌、胸腺癌科，係特徵為上皮 ί t加速生長之疾病。起初此加速生長造成腫瘤形成。最 會發生移轉至不同器官位置。雖然已進行各種癌症的診斷 和>口療，这些疾病仍然造成顯著的死亡率。 肺癌是癌症死亡率的首要原因。在美國，2006年106,374男 性，9〇,〇8〇女性被診斷出肺癌，且89,243男性及69,356女性死於 肺癌。（U.S.癌症統計工作組。美國癌症統計：1999至2〇〇6年發 病率和死亡率基於網路的報告。亞特蘭大（GA):衛生署和人類 服務部，疾病控制和預防中心，以及國家癌症研究所2〇1〇年。） 肺癌被粗分為小細胞肺癌（SCLC，包含2〇%之肺癌），及非 小細胞肺癌（NSCLC ’包含80%之肺癌）。非小細胞肺癌的腺癌亞 型，表最常見的肺癌病理類型。中晚期NSCLC患者具轉移性疾病 且#斷後平均存活4-5個月，存活1年者不到1〇% Q(Shamia sv，201206457 /, DESCRIPTION OF THE INVENTION: TECHNICAL FIELD OF THE INVENTION The present invention relates to a method and composition for treating cancer. Specifically, the method of treating silk and transferring it is to extract cancer and =i: there is a 'used to regulate sputum and inhibit the course of cancer, including taking two. 〒; D, Pot (Ganoderma lucidum, Reishi or Ling_Zhi) [Prior Art] _1 skin = 'for example, prostate cancer, breast cancer, colorectal cancer, lung cancer, 4 of the second nest cancer, spleen call cancer, test pills Cancer, thymic cancer, is characterized by epithelial ί t accelerated growth. At first this accelerated growth caused tumor formation. It is most likely to move to a different organ location. Although various cancer diagnoses and >oral treatments have been performed, these diseases still cause significant mortality. Lung cancer is the leading cause of cancer mortality. In the United States, 106,374 men, 9 weeks, and 〇8 women were diagnosed with lung cancer in 2006, and 89,243 men and 69,356 women died of lung cancer. (US Cancer Statistics Working Group. US Cancer Statistics: Web-based reports of morbidity and mortality from 1999 to 2002. Atlanta (GA): Department of Health and Human Services, Centers for Disease Control and Prevention, and National Cancer The Institute is 2〇1〇.) Lung cancer is roughly classified into small cell lung cancer (SCLC, containing 2% of lung cancer), and non-small cell lung cancer (NSCLC 'containing 80% of lung cancer). Adenocarcinoma subtypes of non-small cell lung cancer, the most common pathological type of lung cancer. Patients with advanced NSCLC have metastatic disease and have an average survival of 4-5 months after the break, and less than 1% of the patients who survived for 1 year (Shamia sv,
BellDW ’ SettlemanJ ’ & HaberDA (2007)肺癌之表皮生長因子 受體突變。Nat Rev Cancer 7 (3) : 169-181。） 在全世界，乳癌佔女性癌症發病之丨〇 4%，為第二最常見之非 皮膚癌症之類型（第一為肺癌）及第五大癌症死亡的常見原因。 有許多不同類型的乳癌，不同階段（擴散）、侵略性 '和遺傳結構； 生存率與這些因素有很大關係。2009年，在美國有192,370 (女性） 201206457 和1，910 (男性）新發病例，和4〇，i7〇 (女性）和440 (男性）人 死於乳癌。其中一個潛在的治療癌症治療策略，是抑制移轉。 靈芝（Ganoderma lucidum，Reishi 或 Ling-Zhi)，一種腐生真 菌’早已被傳統中醫建議用做膳食補充劑（Wang Y_Y，et al. (2002 ) 靈芝多醣之免疫調節和抗腫瘤活性之研究：含海藻醣之醣蛋白部 分之活性的功能與蛋白質分析。Bioorganic & Medicinal Chemistry 10 (4): 1057-1062)。靈芝LZ-8之蛋白質靈芝-8對全身過敏性反 應有正面效果’且已被用做治療肝癌及預防糖尿病。（Kin〇 K. et al. ’ J. Biol. Chem. 1989 ; 264 ( 1 ) : 472-8)。 先前研究已證實’靈芝之葡聚餹萃取物衍生之多聽（E〇rp)， 因為醣蛋白部分含有具β·1，3_聯結之多醣骨架主鏈或具^^丨，#•聯 結之聚甘露醣主鏈，發揮了多種免疫功能和抗腫瘤活性（Chen H-S ’ et al. (2004)靈芝多醣之免疫調節和抗腫瘤活性之研究。 Bioorganic & Medicinal Chemistry 12 (21 ) ： 5595-5601 ； Hsu H-Y » etal. 2004)靈芝多醣萃取物誘發細胞激素表現，經由TLR4_調節 之蛋白質激酶訊息傳遞途徑。J Immunol 173 ( 10) : 5989-5999)。 舉例而言，來自靈芝之多醣抑制腫瘤生長和誘導各種炎性細胞激 素之表現。因此，多醣化合物可能是一種抗腫瘤劑的基礎上，能 夠增強宿主的防禦系統（Cao Q-Z & Lin Z-B ( 2006 )靈芝多聽肽 能抑制人類肺癌細胞内血管内皮細胞之生長和VEGF之誘發QLifeBellDW 'Settleman J ' & HaberDA (2007) Epidermal Growth Factor Receptor Mutation in Lung Cancer. Nat Rev Cancer 7 (3): 169-181. Breast cancer accounts for 4% of all cancers worldwide, and is the second most common type of non-skin cancer (the first is lung cancer) and the fifth leading cause of cancer death. There are many different types of breast cancer, different stages (diffusion), aggressiveness, and genetic structure; survival rates are strongly related to these factors. In 2009, there were 192,370 (female) 201206457 and 1,910 (male) new cases in the United States, and 4〇, i7〇 (female) and 440 (male) people died of breast cancer. One of the potential treatment strategies for cancer treatment is to inhibit migration. Ganoderma lucidum (Reishi or Ling-Zhi), a saprophytic fungus, has long been recommended as a dietary supplement by traditional Chinese medicine (Wang Y_Y, et al. (2002) Study on immunomodulation and antitumor activity of Ganoderma lucidum polysaccharides: containing algae Functional and protein analysis of the activity of the glycoprotein portion of the sugar. Bioorganic & Medicinal Chemistry 10 (4): 1057-1062). Ganoderma lucidum LZ-8 protein Ganoderma lucidum-8 has a positive effect on systemic allergic reactions and has been used to treat liver cancer and prevent diabetes. (Kin〇 K. et al.' J. Biol. Chem. 1989; 264 (1): 472-8). Previous studies have confirmed that 'German's Glucosinolate extract derived from Espor (E〇rp), because the glycoprotein part contains a polysaccharide backbone with β·1,3_ linkage or has a ^^丨,#• linkage The polymannose backbone exerts a variety of immune functions and antitumor activities (Chen HS ' et al. (2004) Immunomodulation and antitumor activity of Ganoderma lucidum polysaccharides. Bioorganic & Medicinal Chemistry 12 (21 ) : 5595-5601 Hsu HY » et al. 2004) Ganoderma lucidum polysaccharide extract induces cytokine expression via a TLR4_ regulated protein kinase signaling pathway. J Immunol 173 (10): 5989-5999). For example, polysaccharides derived from Ganoderma lucidum inhibit tumor growth and induce the expression of various inflammatory cytokines. Therefore, the polysaccharide compound may be an anti-tumor agent that enhances the host's defense system (Cao Q-Z & Lin Z-B (2006) Ganoderma lucidum polypeptide can inhibit the growth of vascular endothelial cells and VEGF induced QLife in human lung cancer cells.
Sciences 78 (13)· 1457-1463)。雖然已發現靈芝可抑制轉錄因子 NF-κΒ 及 AP-1’ 並減少 VEGF 及 TGF-βΙ 之分泌（Stanley G，Harvey K，Slivova V，Jiang J ’ & Sliva D ( 2005 )靈芝透過抑制 VEGF 及 TGF-βΙ從前列腺癌細胞分泌來抑制血管生成。BiochemicalSciences 78 (13)· 1457-1463). Although Ganoderma lucidum has been found to inhibit the transcription factors NF-κΒ and AP-1' and reduce the secretion of VEGF and TGF-βΙ (Stanley G, Harvey K, Slivava V, Jiang J ' & Sliva D (2005) Ganoderma lucidum inhibits VEGF and TGF-βΙ is secreted from prostate cancer cells to inhibit angiogenesis.
Biophysical Research Communications 330( 1 ): 46-52)。然而，EORP 之抗轉移功能的分子機制很不明朗。 TGF-β在癌症生物學扮演雙重角色，亦即，腫瘤抑制及促癌 作用。TGF-β抑制上皮、内皮細胞和造血細胞譜系增生，作為腫 瘤抑制因子。在腫瘤發展過程中，腫瘤可抵抗TGF-β中介的生長 抑制並過度表達TGF-β。在這個階段，TGF-β促進腫瘤生長及移 201206457 轉（Dumont N & Arteaga CL ( 2003 )瞄準TGF-β在人類腫瘤形成 之信息傳導網路。Cancer Cell 3( 6): 531-536; Siegel PM & Massague J(2003 )TGF-β在體内平衡和癌症之抑制細胞生長和凋亡的行為。 Nat Rev Cancer 3(11)： 807-820 ； Wakefield LM & Roberts AB( 2002 ) TGF-β信息傳遞：對腫瘤形成積極和消極影響。Current Opinion inBiophysical Research Communications 330(1): 46-52). However, the molecular mechanism of EORP's anti-metastatic function is unclear. TGF-β plays a dual role in cancer biology, namely, tumor suppression and cancer promotion. TGF-β inhibits epithelial, endothelial and hematopoietic lineage proliferation as a tumor suppressor. During tumor development, tumors are resistant to TGF-β-mediated growth inhibition and overexpress TGF-β. At this stage, TGF-β promotes tumor growth and shifts 201206457 rpm (Dumont N & Arteaga CL (2003) targets TGF-β signaling networks in human tumor formation. Cancer Cell 3(6): 531-536; Siegel PM & Massague J (2003) TGF-beta in vivo balances and inhibits cell growth and apoptosis in cancer. Nat Rev Cancer 3(11): 807-820; Wakefield LM & Roberts AB (2002) TGF- Beta information transmission: positive and negative effects on tumor formation. Current Opinion in
Genetics & Development 12 (1 ) : 22-29)。當癌前病變進展，TGF-β 之角色從腫瘤抑制因子變成腫瘤促進因子。由TGF-β誘導的上皮· 間質轉化（epithelial-mesenchymal transition, EMT )，早期上皮腫瘤 轉化為侵入性、轉移性腫瘤（Thiery JP (2002)腫瘤進程之上皮-間質轉化。Nat Rev Cancer 2 (6) : 442-454)。細胞可進血管内入 淋巴或血管’允許其被動傳輸至遠處器官（Id.)。 上皮-間質轉化（EMT)是一個重要的癌細胞表型改變，其觸 發入侵和移轉。Snail，一個鋅指結構轉錄因子，被認為有參與上 皮-間質轉化。它已被證明可減弱細胞週期，並在外渗、傳播和血 管内週期（extravasation ’ dissemination and intravasation cycle)期 間提供腫瘤細胞生存優勢（Vega S，et al. (2004) Snail阻斷細胞 週期並抗細胞死亡。Genes & Development 18.( 10) : li3i_1143)。 上皮-間質轉化的相關細胞轉化及細胞交互作用發生於非小細 胞肺癌（NSCLC)、大腸癌（CRC) ’及胰臟癌。在上皮_間質轉化 期間，上皮細胞下調其細胞間附著，喪失頂端_基底極性，並進行 細胞形態的變化，從立方型單層至分散、梭形纖維母細胞樣細胞。 分化標記之表現轉換了如E-鈣黏素之細胞-細胞接合蛋白成句 上素ίίίΪ接蛋白之間質標記。再者，靜止細胞轉換為能 =過碰絲胃人侵之雜細胞的。上皮_間質轉化在致癌 轉方面很重要（Janda Ε，et al. (2002) Ras和TGF-β協同詷伙卜 塑^移轉：Ras訊息傳遞途徑之剖析。了 cel丨二即156 【發明内容】 需要以能調節上皮間質轉化（EMT)之藥劑來增進治療如肺 201206457 癌之固態腫瘤之方法。 果。靈芝F3部分，於靈芝F3部分對肺腺癌病程及移轉之效 誘發之EMT。此外了^體外A549人類NSCLC細胞系由TGF-(31 模型肺臟與肝臟之實部分抑制活體内帶有LLC之小鼠 F3部分可抑制轉之表現。本揭露内容揭露了 ’靈芝 本發明係有_ 腫瘤有關之EMT及移轉。 或多個子部分種ff組成物，其包含1芝萃取物之一 以（〇調節上皮、·間質=夕二''之=靈芝F3部分）係足 上可接受之賦形劑。 及兩者a疋，及可選的，一醫樂 組織在=:、容子部分可由以下獲得:均質化靈芝 ί==5Γ.2±α2並過遽以獲得透明靈芝粗萃取物；以 並獲^化取^物向流過濾法，以移除低分子量成分 廊中’該靈芝F3部分顯示如圖6之肌C輪 端海藻，靈芝F3部分包含㈣或醣肽，其包含終 亡發，，揭露驗預防、治療或降低細胞巾與癌症有關事項 包含：將—包含靈芝F3部分之組成物與該細胞接觸， 部分具有如圖6之roLC輪廓，又其巾馳成物包 3 1靈之F3部分以能（a)調節上皮·間質轉化（EMT)，戋（匕） 降低肺癌細胞之移轉及入侵’或兩者皆是。某些具體實施例中， 該細胞係哺乳動物細胞。某些具體實施例中，該哺乳動物細 在人體中。 在某些方面，該癌細胞包含神經母細胞瘤、黑色素瘤、非何 杰金氏淋巴瘤、Epstein-Barr相關淋巴瘤、何杰金氏淋巴瘤、視網 膜胚細胞瘤、小細胞肺癌、腦瘤、血癌、表皮樣癌、攝護腺癌、 腎細胞癌、移形細胞癌、乳癌、卵巢癌、肺癌、大腸癌、肝癌、 201206457 胃癌，及其他腸胃道癌症。 在某些方面，該癌細胞包含一固態腫瘤，其選自非小細胞肺 癌（NSCLC)、胰臟癌、大腸癌、乳癌及肝癌。 在某些方面，上皮-間質轉化（EMT)之調節包含降低一或多 個選自下列之症狀··喪失上皮-細胞標記；由上皮腫瘤細胞造成之 喪失細胞極性及細胞-接合蛋白；獲得蛋白質間質-細胞標記。 在某些方面，上皮-間質轉化（EMT)之調節包含消耗EMT， 其具一或多個選自下列之效果：轉化纖維母細胞至上皮形態；上 調主要上皮細胞標記表現；及下調間質細胞標記表現。 在某些方面，該EMT係由EMT-相關訊息傳遞途徑所誘發， 該訊息傳遞途徑係經由受體所驅動，如血小板衍生生長因子受體 (PDGFR);纖維母細胞生長因子受體（FGFR) ; cMET ; TGFBR ; IGF-1R ;及選自PI3K ' AKT及mTOR之政酶。 某些具體實施例中，該EMT係由TGF-βΙ所誘發。某些具體 實施例中，EMT降低包含TGF-βΙ-中介之訊息傳遞之抑制，&包 含一或多個選自下列之效果：（i)降低TGF-βΙ產生，（ϋ)下調 TGF-β受體Π表現，（出）減少Smad2/3填酸化，及（iv )減少如沾 蛋白質表現。 某些具體實施例中，該上皮-細胞標記係選自£_鈣黏素及 連環蛋白。某些具體實施例中，該蛋白質間質_細胞標記係'選自中 間絲蛋白、纖維連接蛋白、及Ν-舞黏素。 在某些方面’肺癌細胞之移轉及入侵之降低包含抗轉移性效 某些具體貝施例中，§亥肺癌細胞係非小細胞肺癌（Nsclc ) ^胞。某些具體實施例巾’該抗轉移性效果包含降低上 (MET)並藉此降低上皮腫瘤細胞在離原發腫瘤遙遠處之增生 某些紐實施财’該蚊F3部分雜練±有效量之 Ϊ =合併制該劑選自_、阿《、紫杉醇、道諾黴素: '。上黴素、放線菌，D、博來徵素、νρΐ6、腫瘤壞死因子.’、長春 ，狄、長春驗、雙乳乙基亞硝脲（cammstine)、祕笨丙胺酸、 201206457 ，填酿胺、氮芥苯丁酸、馬利蘭、環己亞硝、盤尼西林、紅黴素、 女莫西林、頭抱坐林、亞胺培南、氮浠内酿胺、舒巴坦、利奈嗤 胺、慶大黴素、磺胺甲異噁唑、萬古黴素、塞普沙辛、梭鏈孢酸、 曲美普林、硝基甲嘧唑乙醇、氣林絲菌素、莫螢菌素、兩性黴素B、 立汎黴素、氟康唑，或其等之組合物。 斤某些具體實施例中，該靈芝F3部分係與治療上有效量之順_ 雙氨雙氣鉑（II) (CDDP或順鉑）合併施用。某些具體實施例中， 該順巧係以腹腔内施用。某些具體實施例中，施用靈芝F3部分及 順鉑藉由一或多個（a)降低肺腫瘤多發性，及（b)降低腫瘤尺 寸，來降低遠處轉移，其中該腫瘤多發性或尺寸被降低了較單獨 施用順鉑達成更多的量。 一 、今某些方面，該靈芝F3部分係與抗癌療法合併施用，該抗癌 療法係選自於藥物、荷爾蒙、基因治療組成物、放射性核種、營 養。〇、外科程序、輻射程序或其等之組合。某些 該靈芝分之_録抗絲狄·之彳f f j + 、某些具體實施例中，該靈芝F3部分係同時或依序以口服、靜 脈’主射n續、肌肉注射、腹腔内、滴鼻或經皮方法施用。 由較佳具體實施例之說明及以下的圖式，將清楚呈現本發明 ，上述及其他態樣，雖然在不脫離本揭露内容新穎概念之精神盥 範圍下仍可做成變化例及修飾例。 〃 【實施方式】 本發縣日种之_通常具有在本技術領域巾、在本發明内 t中、及各用語所在之特定内容中的原始意^某些用來描 發明之詞彙將在以下或在發明說明書中討論，以對專門人員 有關於t發明之描述的附加指南。為了方便，某些詞彙可能合 ，目標示，例如使料體及/或引號。醒目標示之使用並二一 詞彙之範圍及意義；-詞彙之範圍及意義在_容中係相^， 無論是否以醒目標示。可被理解的是，同樣的事情可用—種以 的方式說明。因此’可使用其他語言及同義字驗任何_或多個 201206457 此處討論的詞彙，且一詞彙是否在此處被詳盡闡述或討論並不賦 予任何特殊的意義。提供某些詞彙的同義字。一或多個同義字之 敘述並不排除其他同義字之使用。在本發明說明書任一處，實例 的使用包括此處所討論的任何詞彙的例子，係僅用於解說，並不 限制本發明或舉例用辭彙之範圍及意義。同樣地，本發明並不受 本發明說明書中所給的各種具體實施例。 除非另有定義，此處所有使用之技術及科學名詞具有與本發 明所屬技術領域中熟習此藝者所一般性了解者相同的含意。如有 矛盾的情形，以本文件（包括定義）為準。 士，瘤」乙詞，以本說明書及中請範圍目的而 =胸、前列腺、結腸，、騰腺、肝、胃、膀胱、= 卵巢、子宮内膜料）、大腦和骨髓之腫瘤；黑色素 瘤書= 抑制腫瘤入侵，造成一 圍目使2防止腫瘤發展」乙詞，以本說明書及申請範 亡，防止腫瘤形成腫塊。 狀疋而吕、成腫瘤細胞死 「抗癌藥劑」可對個體内癌症 _ 癌細胞、誘導癌細胞;周亡、減少例“’殺死 腫瘤或癌細胞、促腫瘤线，減少血液供應到 ϊίi，ίΐ r個體壽命。更—般地說，這則且成ΐ 的組成物或製劑同時接觸而實π中=:r::不！ 201206457 個表現構築，而另一組成物包括第二劑，如放射性核種或抗癌藥。 F3靈芝多醣萃取物展現抗腫瘤活性。本揭露内容揭露了靈芝 F3組成物在非小細胞肺癌（NSCLC)中調節上皮-間質轉化（EMT) 並減少移轉及入侵之功能。 本揭露内容之靈芝F3部分組成物逆轉了人細胞系 A549之EMT。以TGF-βΙ預培養A549細胞’接著以靈芝F3組成 物處理，造成TGF-βΙ誘發之EMT之消耗，包括（i)轉化纖維母 細巧至上皮形態’（ϋ )上調主要上皮細胞標記表現：E-鈣黏素及γ-連裱蛋白，（iii)下調間質細胞標記表現：中間絲蛋白及N—鈣黏素。 不受理論的約束，所建議可改變EMT之機制可基於靈芝打 部分抑制了 TGF-βΙ中介之訊息傳遞之觀察，其經由〇)降低 TJJF-βΙ產生，（u)下調TGF-β受體Π表現，（出）減少Smad2/3 破酸化’或（iv)減少Snail蛋白質表現。 又，鼠類Lewis肺癌（LLC)活體外模型，靈芝打部分降低 細胞移轉、細胞人侵及g落形成。蚊F3部分在NSCLC方面的 進一步治療用途，係基於口服及注射靈芝F3部分對 :小鼠中移轉之效果。特定而言，LLC七細胞經二靜== 才土入小鼠。七天後’以腹腔注射CDDP (〇5mg/g)同時每兩天以 腹腔注射EORP (〇.〇75mg/g)或每天以口服E〇Rp (〇〇75 處理小鼠4週。以蚊F3部分對LLC異種鍾小鼠尾靜脈腹腔 左射已在統δ十上及明顯地降地了肺癌移轉，相較於控制 。 又，靈芝F3部分及順鉑之結合處理顯著地降低了在肺部的 小結之標記及尺寸。在賴外添加妓F3部分增強了轉至 肝之抑制。我們目前的結果證實，在動物模型中，靈芝Β 定義與EMT相關表型及其逆轉過程以及肺癌 ^ 演關鍵角色。 •上皮間質轉化（EMT)及癌症 已觀察到患者腫瘤樣本中，上皮_細胞標記之喪失 標記之獲得，特別是在固態麵之前緣或人侵前端，如非小細^ 201206457 肺癌（NSCLC)、賴癌、直腸癌，及肝癌。此等上皮樣及間質樣 細胞標記表型之改變無瘤絲之減有關。上皮·細胞標記（例 如E-_占素、γ·連環蛋白，及其他）之喪失與腫瘤之疾病病程轉 移潛力有關。顯鋪細胞可透過與ΕΜΤ錢之特定生物路徑之活 化而去分化，藉此獲得㈣及人侵之能力。因此，已在實驗上觀 察到有關ΕΜΤ及正常||官發展者，亦被認為可在随腫瘤之 病程-轉錄重絲雛’藉此上皮麵細胞喪失細胞極性及細胞 接合蛋白，且同時獲得蛋白質間質·細胞標記（例如中間絲蛋白、 纖維連接$白’及其他）及與間質細胞有關之訊息傳遞活性，在 細胞不著地（anchorage-independent)環境促進移轉及存活，並最 終移轉至遠處。 間質樣腫瘤細胞以增生潛力為代價獲得移轉能力。如同在正 常EMT中，腫瘤細胞中病理EMT起因於細胞轉錄重編程導致其 轉化成間質樣細胞表現型，由EMT有關訊息傳遞途徑促進，其部 为透過異#存活彳§號經經由受體及調節激酶所驅動，受體如企小 板衍生生長因子受體（PDGFR);纖維母細胞生長因子受體 (FGFR) ; cMET ; TGFfiR ; IGF-1R ;調節激酶如 pI3K，akt 及 mTOR。細胞改變’其造成由癌症病理emt驅動之更間質樣狀態， 被認為在與不良預後有關之疾病病程扮演重要角色。 相反的，間質上皮轉化（MET)，藉此間質腫瘤細胞改變成更 上皮樣表現型，被§忍為是被需要來再生一增生狀態並形成類似原 發腫瘤之巨轉移至遠處。據推測，MET幫助遠離原發腫瘤處之上 皮腫瘤細胞之增生及生長。Genetics & Development 12 (1 ) : 22-29). As precancerous lesions progress, the role of TGF-β changes from a tumor suppressor to a tumor promoting factor. Epithelial-mesenchymal transition (EMT) induced by TGF-β, early epithelial tumors are transformed into invasive, metastatic tumors (Thiery JP (2002) Tumor progression epithelial-mesenchymal transition. Nat Rev Cancer 2 (6) : 442-454). Cells can enter the blood vessels into the lymph nodes or blood vessels 'allowing them to be passively transmitted to distant organs (Id.). Epithelial-mesenchymal transition (EMT) is an important cancer cell phenotypic change that triggers invasion and migration. Snail, a zinc finger structural transcription factor, is thought to be involved in epithelial-mesenchymal transition. It has been shown to attenuate the cell cycle and provide tumor cell survival advantages during extravasation, dissemination and intravasation cycles (Vega S, et al. (2004) Snail blocks cell cycle and fights cells Death. Genes & Development 18.(10) : li3i_1143). Cellular transformation and cell interaction of epithelial-mesenchymal transition occur in non-small cell lung cancer (NSCLC), colorectal cancer (CRC)', and pancreatic cancer. During epithelial-mesenchymal transition, epithelial cells down-regulate their intercellular attachment, lose apical basal polarity, and undergo cell morphology changes from cubic monolayers to dispersed, spindle-shaped fibroblast-like cells. The expression of the differentiation marker converts the cell-cell junction protein such as E-cadherin into a serotonin. In addition, the resting cells are converted into cells that can pass through the stomach and invade the stomach. Epithelial-mesenchymal transition is important in carcinogenesis (Janda Ε, et al. (2002) Ras and TGF-β synergy 詷 卜 ^ 移 : : : : : : : : : : : Ra Ra Ra Ra Ra Ra Ra Ra Ra Ra Ra 156 156 156 Contents] It is necessary to use a drug that can regulate epithelial-mesenchymal transition (EMT) to improve the treatment of solid tumors such as lung 201206457. Fruit. F3 part of Ganoderma lucidum is induced in the course of lung adenocarcinoma and its effect on the transfer of lung adenocarcinoma. EMT. In addition, the in vitro A549 human NSCLC cell line is inhibited by TGF- (31 model lung and liver parts inhibiting the F3 part of the mouse with FL in vivo. The disclosure reveals that 'Ganoderma lucidum There are _ tumor-related EMT and transfer. Or multiple sub-species ff composition, which contains one of the 1 zhi extract (〇 上 上 上 · · 间 间 夕 夕 夕 夕 夕 = = = = = = = = = = = = Acceptable excipients, and both a, and optionally, a medical organization in the =:, the part of the volume can be obtained by homogenizing Ganoderma lucidum ί==5Γ.2±α2 and passing through to obtain transparent Ganoderma lucidum Crude extract; to remove and reduce the flow to remove the low In the sub-components gallery, the F3 part of the Ganoderma lucidum shows the muscle C-wheel seaweed as shown in Figure 6. The Ganoderma F3 part contains (4) or glycopeptide, which contains the final hair loss, and exposes the prevention, treatment or reduction of cell towels and cancer. The method comprises: contacting the composition comprising the F3 part of Ganoderma lucidum with the cell, partially having the outline of the roLC as shown in Fig. 6, and further comprising the F3 part of the stalk of the stalk of the stalk to enable (a) to regulate the epithelial/mesenchymal transition ( EMT), 戋(匕) reduces the transfer and invasion of lung cancer cells' or both. In some embodiments, the cell line is a mammalian cell. In some embodiments, the mammal is fine in the human body. In some aspects, the cancer cell comprises neuroblastoma, melanoma, non-Hodgkin's lymphoma, Epstein-Barr-associated lymphoma, Hodgkin's lymphoma, retinoblastoma, small cell lung cancer, brain Tumor, blood cancer, epidermoid carcinoma, prostate cancer, renal cell carcinoma, metastatic cell carcinoma, breast cancer, ovarian cancer, lung cancer, colon cancer, liver cancer, 201206457 gastric cancer, and other gastrointestinal cancers. In some aspects, the cancer Cell contains one A solid tumor selected from the group consisting of non-small cell lung cancer (NSCLC), pancreatic cancer, colorectal cancer, breast cancer, and liver cancer. In certain aspects, the regulation of epithelial-mesenchymal transition (EMT) comprises reducing one or more selected from the group consisting of Symptoms · Loss of epithelial-cell markers; loss of cell polarity and cell-conjugating proteins by epithelial tumor cells; acquisition of protein interstitial-cell markers. In some aspects, regulation of epithelial-mesenchymal transition (EMT) involves consumption of EMT , having one or more effects selected from the group consisting of transforming fibroblasts to epithelial morphology; up-regulating the expression of major epithelial cells; and down-regulating the expression of mesenchymal cells. In certain aspects, the EMT is induced by an EMT-related signaling pathway driven by a receptor, such as a platelet-derived growth factor receptor (PDGFR); a fibroblast growth factor receptor (FGFR) cMET; TGFBR; IGF-1R; and a political enzyme selected from the group consisting of PI3K 'AKT and mTOR. In certain embodiments, the EMT is induced by TGF-[beta]. In certain embodiments, the EMT reduces the inhibition of TGF-βΙ-mediated signaling, & contains one or more effects selected from the group consisting of: (i) reducing TGF-βΙ production, and (ϋ) downregulating TGF-β Receptor Π shows, (out) reduces Smad2/3 acidification, and (iv) reduces protein expression. In certain embodiments, the epithelial-cell marker is selected from the group consisting of £-cadherin and catenin. In certain embodiments, the protein interstitial-cell marker line is selected from the group consisting of a middle filament protein, a fibronectin, and a scorpion-mucosa. In some respects, the reduction of lung cancer cell migration and invasion includes anti-metastatic efficacy. In some specific examples, the CYH lung cancer cell line is non-small cell lung cancer (Nsclc) cells. In some embodiments, the anti-metastatic effect comprises lowering the upper (MET) and thereby reducing the proliferation of epithelial tumor cells distant from the primary tumor. Ϊ = combined preparation of the agent selected from _, A, paclitaxel, daunorubicin: '. Upper mycin, actinomycetes, D, bleeder, νρΐ6, tumor necrosis factor. ', Changchun, Di, Changchun, double emulsion ethyl nitrosourea (cammstine), secret alanine, 201206457, filling amine , nitrogen mustard butyl butyric acid, malilan, cyclohexyl nitrite, penicillin, erythromycin, female moxilin, cephalopod, imipenem, arsenic, sulbactam, linabamide, qingda Streptomycin, sulfamethoxazole, vancomycin, cepsporin, fusidic acid, trimetine, nitromethoxazole, lincomycin, morocin, amphotericin B , ubiquitin, fluconazole, or a combination thereof. In certain embodiments, the Ganoderma lucidum F3 moiety is administered in combination with a therapeutically effective amount of cis-diammine-pivoxil (II) (CDDP or cisplatin). In some embodiments, the sequence is administered intraperitoneally. In certain embodiments, administration of the Ganoderma lucidum F3 moiety and cisplatin reduces distant metastasis by one or more (a) reducing lung tumor polymorphism, and (b) reducing tumor size, wherein the tumor is multiple or sized A greater amount is achieved than when cisplatin is administered alone. In some aspects, the Ganoderma lucidum F3 moiety is administered in combination with an anti-cancer therapy selected from the group consisting of a drug, a hormone, a gene therapy composition, a radioactive nucleus, and a nutrient. A combination of sputum, surgical procedures, radiation procedures, or the like. In some specific embodiments, the Ganoderma lucidum F3 part is simultaneously or sequentially administered orally, intravenously, intravenously, intramuscularly, intraperitoneally, nasally. Or administered by transdermal methods. The invention will be apparent from the following description of the preferred embodiments and the appended claims. 〃 [Embodiment] The genre of this county has the original meaning in the technical field, the t in the present invention, and the specific content in which the terms are used. Or discussed in the description of the invention, with additional guidance for the description of the invention by the specialist. For convenience, certain terms may be combined, and the target is shown, for example, to make the material and/or quotation marks. The purpose of waking up the target and the scope and meaning of the vocabulary; - the scope and meaning of the vocabulary in the _ 容 中 相 ^, whether or not to show the goal. It can be understood that the same thing can be explained in a way. Therefore, any vocabulary and synonymous words can be used to test any _ or more of the vocabulary discussed herein, and whether a vocabulary is elaborated or discussed here does not give any special meaning. Provide synonyms for certain words. The recitation of one or more synonyms does not exclude the use of other synonyms. The use of examples, including any examples of the terms discussed herein, is intended to be illustrative, and not to limit the scope and meaning of the invention. As such, the invention is not limited to the specific embodiments disclosed herein. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those of ordinary skill in the art. In case of conflicts, this document (including definitions) will prevail. "Shi, tumor" in the form of this manual and the scope of the request = chest, prostate, colon, gonad, liver, stomach, bladder, = ovary, endometrial), brain and bone marrow tumor; melanoma Book = inhibition of tumor invasion, resulting in a periscope to prevent the development of tumors." This article and application for death, to prevent tumor formation of lumps.疋 疋 吕, into tumor cells dead "anti-cancer agents" can be cancer in individuals _ cancer cells, induced cancer cells; weeks of death, reduce cases "kill cancer or cancer cells, promote tumor lines, reduce blood supply to ϊίi , ΐ 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体 个体Such as radionuclides or anticancer drugs. F3 Ganoderma lucidum polysaccharide extract exhibits anti-tumor activity. The disclosure discloses that Ganoderma lucidum F3 composition regulates epithelial-mesenchymal transition (EMT) and reduces migration in non-small cell lung cancer (NSCLC). Invasion function. The partial composition of Ganoderma lucidum F3 in this disclosure reverses the EMT of human cell line A549. Pre-culture of A549 cells with TGF-βΙ, followed by treatment with Ganoderma lucidum F3 composition, resulting in TGF-βΙ-induced EMT consumption, including (i) Integrity of the fibrillar to the epithelial morphology '(ϋ) up-regulated the main epithelial cell markers: E-cadherin and γ-catenin, (iii) down-regulation of mesenchymal cells: intermediate filament protein and N-calcium Muscle. Constraints, the proposed mechanism for altering EMT can be based on the observation that Ganoderma lucidum partially inhibits TGF-βΙ-mediated signaling, which reduces TJJF-βΙ production via 〇) and (u) down-regulates TGF-β receptor Π expression, ( () Reduce Smad2/3 deacidification' or (iv) reduce Snail protein performance. Also, in a mouse Lewis lung cancer (LLC) in vitro model, Ganoderma lucidum partially reduced cell turnover, cell invasion and g fall formation. Further therapeutic use in NSCLC is based on the effect of oral and injection of F3 on the F3 part: the effect of transfer in mice. In particular, LLC seven cells are injected into the mice via two static == soil. Seven days later, 'intraperitoneal injection CDDP (〇5mg/g) was injected intraperitoneally with EORP (〇.〇75mg/g) every two days or daily with E〇Rp (〇〇75 treated mice for 4 weeks. Mosquito F3 part to LLC heterogeneous mice The left ventricle of the tail vein has been transferred to the δ10 and significantly reduced lung cancer, compared with the control. In addition, the combination of Ganoderma lucidum F3 and cisplatin significantly reduced the marking and size of the nodules in the lungs. Adding 妓F3 to the outside of the sputum enhances the inhibition of liver transfer. The previous results confirmed that in animal models, Ganoderma lucidum defines the EMT-related phenotype and its reversal process as well as the key role of lung cancer. • Epithelial-mesenchymal transition (EMT) and cancer have been observed in patients with tumor samples, epithelial-cell Obtaining the loss of the marker, especially at the leading edge of the solid surface or the front of the human invasion, such as non-small ^ 201206457 lung cancer (NSCLC), diarrhea, rectal cancer, and liver cancer. These epithelial and interstitial-like cell markers The type of change is related to the reduction of tumor-free silk. The loss of epithelial-cell markers (such as E-_ocyanin, γ-catenin, and others) is associated with the disease metastasis potential of the tumor. The display cells can be dedifferentiated through the activation of specific biological pathways for saving money, thereby obtaining (4) and the ability of human invasion. Therefore, it has been experimentally observed that sputum and normal || official development, is also considered to be able to transcribe the heavy filaments in the pathogenesis of the tumor, thereby losing the cell polarity and cell junction proteins, and simultaneously obtaining the protein. Interstitial-cell markers (eg, intermediate filament proteins, fiber-linked whites and others) and signaling activities associated with mesenchymal cells promote transfer and survival in an anchorage-independent environment and ultimately Go to the distance. Interstitial tumor cells acquire transfer capacity at the expense of proliferative potential. As in normal EMT, pathological EMT in tumor cells results from the transcriptional reprogramming of cells leading to their transformation into a mesenchymal-like phenotype, which is facilitated by EMT-related signaling pathways, which are transmitted through the receptor And regulated by kinases, receptors such as small plate-derived growth factor receptor (PDGFR); fibroblast growth factor receptor (FGFR); cMET; TGFfiR; IGF-1R; regulatory kinases such as pI3K, akt and mTOR. Cellular changes, which cause a more mesenchymal state driven by the cancer pathology, are thought to play an important role in the course of the disease associated with poor prognosis. Conversely, mesenchymal epithelial transformation (MET), whereby interstitial tumor cells change to a more epithelial-like phenotype, is ought to be required to regenerate a proliferative state and form a giant metastasis similar to the primary tumor to distant. It is speculated that MET helps to proliferate and grow away from epithelial tumor cells at the primary tumor.
•靈芝F3部分逆轉人類A549NSCLC中由TGF_pi誘發之EMT 據報導，A549細胞發生了因應TGF-βΙ從立方形變化到呈長 梭形，並顯示減少上皮標記表現及增強間質標記表現（Kasai H，• Ganoderma lucidum F3 partially reverses TGF_pi-induced EMT in human A549 NSCLC. A549 cells have undergone a change from a cuboid to a long fusiform shape in response to TGF-βΙ, and have been shown to reduce epithelial marker expression and enhance interstitial marker expression (Kasai H,
Allen J ’ Mason R，Kamimura T，& Zhang Z ( 2005 ) TGF-betal 誘 么人顯肺泡上皮成間質細胞轉化（EMT)。Respiratory Research 6 (1 ) ·· 56)。細胞經TGF-βΙ處理72小時顯示細胞間接觸減少且由 201206457 圓形變成梭樣間質表現型（圖1A，比較圖a及圖b_d)。生長於正 常培養基之平坦、上皮狀A549細胞並無因靈芝F3部分處理而改 變（圖1A ’圖a ’及〇 °因此’我們估計，TGF_pi所需要來起始 A549中之EMT的最佳濃度與時間為以高達5ng/mi之TGF-pi培 養24小時。間質標s己，包括中間絲蛋白及N_每黏素，被TGF—y 處理上調，而上皮標記，包括E-鈣黏素及連環蛋白，則被下調 (圖1Β，比較欄3及欄1)。研究經及靈芝打部分共處 理細胞72小時之表現型。以靈芝部分處理A549廢止了由 TGF-βΙ誘發之EMT，如細胞形態改變所示（圖1A，比較圖 及圖f_h)，及EMT標記，包括E_鈣黏素、γ_連環蛋白、N_妈黏素 及中間絲蛋白（圖1B ’比較襴3與攔4)。生長於正常培養基之 A549細胞之EMT標記並無因靈芝F3部分處理而改變（圖m， 比較攔1與攔2)。由於細胞運動增強是一個眾所周知的與EMT 相關表型變化’以移轉分析法分析細胞運動的變化。測試靈芝F3 部分對TGF-βΙ誘發之A549細胞移轉之效果。結果顯示，靈芝F3 刀抑制了 TGF-βΙ誘發之A549細胞移轉至聚碳酸醋多孔膜下方 (圖1C)。靈芝F3部分逆轉了 A549細胞中TGF-βΙ誘發之EMT 特性。 •靈芝F3部分抑制A549中TGF-pl誘發有關EMT之訊息傳遞 由於癌細胞往往會增加其活性TGF-βΙ之產生，其不僅觸發 EMT並允許細胞變成具侵入性（Derynck R，Akhurst RJ，& Balmain A( 2001 )腫瘤中TGF-β訊息傳遞之抑制及癌症病程。Nat Genet 29 (2): 117-129)，我們研究了在A549細胞做靈芝F3部分處理後 TGF-βΙ之產生及其#EMT之關聯。結果表明，靈芝F3部分可在 6小時或24小時抑制A549細胞中TGF-βΙ產生（圖2A)。為了探 索其中靈芝F3部分抑制TGF-βΙ誘導的EMT ’我們測得的效果靈 芝F3部分早期TGF-βΙ誘導的信號事件在A549細胞的機制。主 要的TGF-βΙ誘發之訊息傳遞途徑為Smad訊息傳遞。因為smad2/3 碟酸化是TGF-βΙ誘發之上皮_間質轉化所需。因此，我們研究了 12 201206457 J部分對TGF_pi誘發之Smad2/3磷酸化之效果。我們發現 ，3部分之處理於3小時降低了 TGF_pi誘發之Smad2/3磷酸 (圖2B，比較攔3與攔4)。已知Snail及Slug可調節E_妈黏素 =錄作用。兩個轉錄因子皆因應進行见邮誘發之EMT之細胞 —之TGF-βΙ而被誘發（Can〇 A，过al (2〇〇〇)。轉錄因子仏⑹ 3由抑制E-!弓黏素表現來控制上皮間質轉化（EMTs)。碰⑽ 1〇12(2)·76-83)。為定義靈芝F3部分逆轉TGF-βΙ誘發之EMT =機制’以西方墨點料究Snail及Slug之蛋白質濃度。當以靈 f F3,部分處理，其阻斷了 TGF_pi路徑下游受體，A549細胞中 nail被減少了。然而’ Slug並不隨靈芝F3部分變化（圖2C，比 較襴3與襴4)。 •靈芝F3部分抑制LLC之入侵及菌落形成 因為，發現活體外靈芝F3部分可逆轉TGF—y誘發之 EMT」接著在帶有LLC之小鼠模型研究活體内靈芝F3部分之抗 移，效果=先，我們測試靈芝F3部分對LLC細胞之細胞移轉、 ^侵，及菌落形成之抗移轉效果。在靈芝F3部分濃度為12〇μδΛη1 時，LLC細胞之入侵被顯著地減少至33 8%，相較於未經處理之 控，組（:，p<0·01)(圖 3A)。此外，靈芝 F3 部分（120pg/ml) 之效果顯著減少40%菌落形成，相較於控制組孔（*，p<〇 〇5)(圖 3B)。因此，靈芝F3部分可減少LLC細胞入侵及菌落形成。 •靈芝F3部分結合順鉑對帶有LLC之小鼠之抑制效果 /為研究靈芝F3部分在小鼠模型活體内之抗轉移性能力，經由 ^靜脈注射LLC將LLC細胞導入(：57BL/6小鼠。注射7天後， 選擇小鼠以2日之間隔腹腔注射PBS或靈芝F3部分，或口服 靈芝F3部分。我們發現，靈芝F3部分（腹腔注射）顯著地抑制 肺腫瘤多發性（26.8±4.9 v.s. 7.0±1.4，ρ<〇·〇〇ι )及肺病變之腫瘤體 積（146.5±65.9 v.s. 78·2±8.0，ρ<〇.〇5)，相較於僅處理 pBS (圖 4Α)。在靈芝F3部分口服組觀察到相似的變化，且此組顯著的抑 制肺腫瘤多發性（9.7±4_0v.s.26.8±4.9，ρ<〇.〇5)及肺病變之腫瘤 13 201206457 芝F3部分之έ士入r胺狄、±以、予1表第7天。而CDDP及靈 對昭ϋ 射）進一步降低了肺腫瘤多發性（姐8Allen J ’ Mason R, Kamimura T, & Zhang Z (2005) TGF-betal induces alveolar epithelial to become interstitial cell transformation (EMT). Respiratory Research 6 (1 ) ·· 56). Treatment of cells with TGF-βΙ for 72 hours showed a decrease in intercellular contact and a circular change from 201206457 to a shuttle-like stroma phenotype (Fig. 1A, comparing graph a and panel b_d). The flat, epithelial A549 cells grown in normal medium did not change due to partial treatment with Ganoderma lucidum F3 (Fig. 1A 'Fig. a' and 〇°' therefore we estimate that the optimal concentration of EMT required to initiate A549 in TGF_pi is The time was 24 hours of TGF-pi up to 5 ng/mi. The interstitial label, including the intermediate filament protein and N_ per mucin, was up-regulated by TGF-y treatment, while the epithelial markers, including E-cadherin and The catenin was down-regulated (Fig. 1Β, comparison column 3 and column 1). The phenotype of the co-treated cells was treated with Ganoderma lucidum for 72 hours. Partial treatment with Ganoderma lucidum A549 abolished EMT induced by TGF-βΙ, such as cells. Morphological changes (Fig. 1A, comparison and f_h), and EMT markers, including E_cadherin, γ_catenin, N_mamumu and intermediate filament proteins (Fig. 1B 'Comparative 襕 3 and 4 4 The EMT marker of A549 cells grown in normal medium was not changed by the partial treatment of Ganoderma lucidum F3 (Fig. m, comparing 1 and 2). Since cell movement enhancement is a well-known phenotypic change associated with EMT Analysis of changes in cell movement by transformation analysis. Testing Ganoderma F3 The effect of TGF-βΙ-induced A549 cell transfer. The results showed that Ganoderma lucidum F3 knife inhibited the transfer of TGF-βΙ-induced A549 cells to the polycarbonate porous membrane (Fig. 1C). Ganoderma lucidum F3 partially reversed A549 cells. TGF-βΙ-induced EMT characteristics • Ganoderma lucidum F3 partially inhibits TGF-pl-induced EMT signaling in A549 Since cancer cells tend to increase the production of their active TGF-βΙ, which not only triggers EMT but also allows cells to become invasive ( Derynck R, Akhurst RJ, & Balmain A (2001) Inhibition of TGF-β signaling in tumors and the course of cancer. Nat Genet 29 (2): 117-129), we studied the partial treatment of Ganoderma lucidum F3 in A549 cells. The relationship between the production of TGF-βΙ and its #EMT. The results showed that the F3 part of Ganoderma lucidum could inhibit the production of TGF-βΙ in A549 cells at 6 hours or 24 hours (Fig. 2A). In order to explore the inhibition of TGF-βΙ induced by Ganoderma lucidum F3 EMT 'We measured the effect of G. lucidum F3 on early TGF-βΙ-induced signaling events in A549 cells. The main TGF-βΙ-induced signaling pathway is Smad signaling. Because smad2/3 acidification is induced by TGF-βΙ Epithelial-mesenchymal transition required. Therefore, we studied the effect of 12 201206457 Part J on TGF_pi-induced Smad2/3 phosphorylation. We found that 3-part treatment reduced TGF_pi-induced Smad2/3 phosphate at 3 hours (Fig. 2B, compare 3 and stop 4). It is known that Snail and Slug can regulate E_Mamumu = recording effect. Both transcription factors were induced in response to TGF-βΙ, a cell that was evoked by EMT (Can〇A, over al (2〇〇〇). Transcription factor 仏(6) 3 was inhibited by E-! To control epithelial-mesenchymal transition (EMTs). Touch (10) 1〇12(2)·76-83). To define the GEM F3 partial reversal of TGF-βΙ-induced EMT = mechanism' Western protein spots to study the protein concentrations of Snail and Slug. When treated with a f f3, partial blocking, which blocks the receptor downstream of the TGF_pi pathway, the nail is reduced in A549 cells. However, 'Sulug does not change with Ganoderma F3 (Figure 2C, compare 襕3 and 襕4). • Ganoderma lucidum F3 partially inhibits LLC invasion and colony formation because it was found that F3 part of Ganoderma lucidum can reverse TGF-y-induced EMT. Then, in the mouse model with LLC, the anti-migration of Ganoderma lucidum F3 was studied. We tested the anti-transfer effect of G3 of Ganoderma lucidum on cell migration, invasion, and colony formation of LLC cells. When the concentration of Ganoderma lucidum F3 was 12 〇μδΛη1, the invasion of LLC cells was significantly reduced to 33 8%, compared to untreated control, group (:, p < 0·01) (Fig. 3A). In addition, the effect of the Ganoderma lucidum F3 fraction (120 pg/ml) significantly reduced colony formation by 40% compared to the control group wells (*, p < 〇 〇 5) (Fig. 3B). Therefore, Ganoderma lucidum F3 can reduce LLC cell invasion and colony formation. • Ganoderma lucidum F3 partially binds cisplatin to mice with LLC. In order to study the anti-metastatic ability of Ganoderma lucidum F3 in vivo in mouse models, LLC cells were introduced via intravenous injection of LLC (: 57BL/6 small After 7 days of injection, the mice were selected to intraperitoneally inject PBS or Ganoderma F3 fraction at 2 days, or oral F3 of Ganoderma lucidum. We found that Ganoderma lucidum F3 (intraperitoneal injection) significantly inhibited multiple tumors of lung tumors (26.8 ± 4.9) Vs 7.0±1.4, ρ<〇·〇〇ι) and tumor volume of lung lesions (146.5±65.9 vs 78·2±8.0, ρ<〇.〇5) compared to pBS only (Fig. 4Α). A similar change was observed in the oral administration group of Ganoderma lucidum F3, and this group significantly inhibited multiple tumors of the lung tumor (9.7±4_0v.s.26.8±4.9, ρ<〇.〇5) and tumors of lung lesions 13 201206457 The gentleman enters r-amine, ±, and gives the first day of the seventh day. CDDP and the spirit of Zhao Zhao) further reduce the multiple tumors of the lung (Sister 8
(7.〇±,8 f,<BS :5.2 ’ P<0.05)，相較於僅處理 CDDP (圖 4A)。 ，、、 驗。ί 步，分析i則試小鼠中之LLC移轉，進行組織染色實 織之料#顯不了腫瘤在第28天移轉至肺泡肺袓 $ = ’其在以靈芝F3部分處理制失（圖4b )。我們發=、， 靈之F3 刀（腹腔注射）或靈芝打部分（口服 入 ^ 了，移轉至肺泡肺組織，相較於僅以瞻處、f 靈芝F3部分（腹腔注射）或靈;^%(7. 〇 ±, 8 f, <BS : 5.2 ' P < 0.05) compared to only CDDP (Fig. 4A). ,,, and inspection. ί step, analysis i test the LLC transfer in the mouse, the tissue staining of the actual weaving material # can not show the tumor on the 28th day transferred to the alveolar lung 袓 $ = ' it is treated with Ganoderma lucidum F3 part of the loss (Figure 4b). We send =,, F3 knife (intraperitoneal injection) or Ganoderma lucidum (partially into the ^, transferred to the alveolar lung tissue, compared to only the prospect, f Ganoderma F3 part (intraperitoneal injection) or spirit; ^ %
處理(”圖40^二抑制了軸移轉至肝組織’她於僅以CDDP i些數據表_部分的蚊及有效CDDP 之組合也可能降低遠處轉移。 盆他ίΐϊΐΐΐ明方法之有效性’可能需要結合抗癌組成物及 •^明二^效〉。療超增生疾病之藥劑。此等組合治療可發生於本 咖在相同的原位注射方案内結合基 偏ΐί減擇的’可在本發明範_使驗合治療， 施用〗施用治療藥劑以外，藉由以本發明方法原位 她用一或多個治療藥劑。 番+匕學治療及放射線治療藥劑之腫瘤細胞代表了臨床腫瘤學 :題。目刖癌症研究的一個目標是要設法藉由結合基因治療 化療及放射線治療的療效。舉例而言，當以反轉錄病毒載 將單純泡療-胸腺激酶⑽_tK)基因傳送至腦瘤，可成功 的3對抗病毒藥更昔洛韋（ganddovir)之易感性（Culver，etal， 。本發明设想，除了其他促細胞凋亡或細胞週期調節劑外， =F3治療可相似地合併用於化學治療、放射線治療、或免疫治 療干預。 14 201206457 歸 藥劑絲ΐ療可以數分鐘至數週之間隔在其他 ΐίΐϊ"7別送入細胞，—般會確保—段顯著時間不超出欠Ϊ ，相接觸更小時内將細胞與兩麵式皆互 長時間來顯著治療，數天（2、3、4、5、6 3、4、5、6、7或8)等各施用間之時效。或7)至數週（卜2、 大約^峨其⑽叙紐練如输合併施用。 玲斷手術某些情型會進行手術’包括預防性手術， 二:卞或刀』手術，根治性手術和姑息性手術。根治 二以5併其他療法的癌症治療，例如本 興二疋 =丰荷爾蒙治療、基因治療、免疫二 及包括將全部或部分癌變組織被物理性移除切除 =微觀控制的手術（莫氏手術）。進—步設想本發明=併表面= 癌、初癌，或偶然量之正常組織之切除來使用。 在癌變_、_、或腫狀全部或部分 以附加的抗癌療法灌注、直接注射或區=3 70成此、/σ療。此治療可重複，舉例而言，每1、2、3'4、5、 f 7 天’或每卜 2、3、4、及 5 週或每 1、2'3、4、5、6、7、、8'、 、10、11、或12月。這些治療亦可為不同劑量。 使用細稍本㈣之靈芝F3料纽成物合併 ^細士;體及二=:===: i县增生細胞對細胞週亡誘導因子 =其他細胞激素；F42K *其他細胞激素二物或 ΜΠΜ、ΜΡ·1β、M〇m、驗似，及其__。本發明^ 15 201206457 步没想細胞表面受體或其配體如Fas/Fas配體、DR4咬 DR5/TRAIL之上調將增強本發明之誘發細胞凋亡能力，其透過通 過對超增生細胞建立自分泌或旁分泌作用。藉由提升GAp連結數 來增加細胞間訊息傳遞將增加對鄰近超增生細胞族群之抗超增生 效果。其他具體實施例中，細胞生長抑制劑或分化劑可用於&本 發明合併使用以改善治療的抗超增生效果。細胞吸附抑制劑被認 為可改善本發明之效用。細胞吸附抑制劑之實例為局部黏合激酶 (FAKs )抑制劑及洛維丁（ Lovastatin )。本發明亦設想其他可增 加超增生細胞對細胞凋亡之易感性之藥劑，如抗體c225，可用來 與本發明合併使用以改善治療效果。 靈芝F 3部分可與其他用於治療癌症之化學治療劑合併製劑或 合併施用，包括烷化劑、抗代謝物抗拮劑、抗癌抗生素、及植物 ，生之抗癌藥劑。”烧化劑，，之實例包括含氮芥子油、含氮芥子油_N_ 氧化物鹽酸鹽、苯丁酸氮芥（chlorambutyl)、環礙醯胺、依弗醯胺、 沙奥特帕、卡波醌（carboquone)、甲苯磺酸胺丙磺酯（improsulfan tosylate )、二曱磺酸丁酯、鹽酸尼莫司汀（nimustine hydrochloride)、二溴甘露醇（mitobronitol)、氮芬苯丙胺酸、氮稀 β坐胺（dacarbazine )、雷諾氮芥（ranimustine )、雌二醇氮芬碌酸鈉、 二亞乙基蜜胺（triethylenemelamine )、雙氣乙基亞；g肖脲 (cammstine)、環己亞硝、鏈尿佐菌素、哌血生（pip〇br〇man)、 乙環氧咬（etoglucid)、卡舶定、順紐、米舶（mib〇piatin)、奈達 鉑（nedaplatin )、奥沙利鉑（oxaiipiatin )、六曱基蜜胺altretamine )、 氨莫司汀（ambamustine )、二溴螺銨鹽酸鹽（dibrospidium hydrochloride )、佛替姆丁（ f〇temustine )、潑尼氮芥 (prednimustine)、〇票〇密替派（pUmitepa)、rib〇mustin、替莫0坐胺 (temozolomide)、treosulphan、氯乙環填酰胺（trophosphamide)、 净司他丁斯酉旨（zinostatin stimalamer)、卡波酿（carboquone)、阿 多來新（adozelesin)、半胱胺亞石肖脲（CyStemustine)、及比折來新 (bizelesin)。π抗代謝物”之實例包括硫醇嘌呤、6_硫醇嘌呤核糖 音、硫肌苷（thioinosine)、胺甲葉酸、依諾他賓（enocitabine)、 201206457 阿拉伯糖基胞嘧啶、阿拉伯糖基胞嘧啶十八烷基磷酸钠、環胞芽 鹽酸鹽（ancitabine hydrochloride)、5-FU藥物（例如，氟尿嘴咬、 喃氟咬（tegafUr)、UFT、5'-去氧-5-氟尿苦、卡莫氟（carmofUr)、 加洛他濱（gallocitabine)、乙嘧替氟（emmitefUr))、氨基蝶呤 (aminopterine )、留可佛林鈣、片錠、甘氨硫嘌呤（butocine )、亞 葉酸鈣（folinate calcium )、左亞葉酸鈣（levofolinate calcium )、克 拉屈濱（cladribine)、乙嘧替氣（emitefUr)、氟達拉濱（fludarabine)、 吉西他濱（gemcitabine)、羥基尿素（hydroxycarbamide)、喷司他 丁（ pentostatin )、°比曲克辛（piritrexim )、蛾去氧尿甘、丙脉腙 (mitoguazone )、σ塞唾吱林（thiazophrine )，及氨莫司汀 (ambamustine)等等。"抗癌抗生素”之實例包括放線菌素。、放 線菌素-C、絲裂黴素-C、色黴素-A3、鹽酸博來黴素、硫酸博來黴 素、硫酸培來黴素（peplomycin sulfate)、鹽酸道諾黴素、鹽酸阿 黴素、鹽酸阿柔比星（aclarubicin hydrochloride )、鹽酸畢拉魯比辛、 鹽酸依畢魯比辛、新抑癌素、光神黴素（mithramycin)、肉瘤黴素 (sarcomycin )、嗜癌素（carzinophilin )、来托坦（mitotane ) ’、鹽 酸佐柔比星（zorubicin hydrochloride)、鹽酸邁杜慧酮、及鹽酸艾 達魯比辛等等。"植物衍生之抗癌藥劑”包括磷酸依托泊甙 (etoposide phosphate)、硫酸長春鹼、硫酸長春新鹼、硫酸長春地 辛（vindesine sulfate)、坦尼坡赛、紫杉醇、多西泰索（加⑽咖丨）、 及長春瑞濱（vinorelbine)等等。 可與靈芝F3組成物合併施用之例示藥物為順鉑。順鉑已被廣 泛用來治療癌症如轉移性紅癌或㈣癌，晚期膀胱癌，頭部或 頸部癌，子宮頸癌，肺癌或其他腫瘤。順鉑（CDDp)可被單 贼與靈芝F3藥劑合併，以臨床應用之有效劑量5_2〇mg/m2，在 全部二，病程中每三週五天。例示劑量可$ 〇5以2、 l.Omg/m、1.5〇mg/m、U5mg/m2,mg/m2、3 〇mg/m2、4 〇 5Omg/m_、lOmg/m、20mg/m2、50mg/m2。當然’所有這此漸^量 皆為例示用’任何在這些點之間的劑量亦可用於本發明 可為口服吸收’因此必須經靜脈注射、皮下注射、腫瘤内或腹腔 17 201206457 内輸送。 治療5本發明合併使用或與任何前述之其他癌症 列療某些癌症時翻，如乳癌、前 激素或雌激素之效果。此治療通常蒙如q 使用，作為-種治療選項或降低轉移的風險個-他癌症治療結合 輸送靈物以能達到所欲目的之方式被 外用、直腸給^等。^一有效路徑施用，如口服、 靈長類動物、寵物，如描、狗、家纽人類、女性人類、 到所欲結宿主。有效量是指能達 規地決定，例如，進行劑量反應實驗、=量可常 素來選擇量，包括靈芝萃取物施，於各種因 動物之年齡胞二?：之病人或 見，2克’ 3克，5〇〇毫克25券笪 宅見2.5克’ 1 $粉末，靈芝F3糊，或靈芝F3輪成 =靈芝 ，取決於受者的需要Ϊ製備方Ϊ有效成 以夂括t月可與夕樣賦形劑一起使用。本發明之靈芝F3細出榀 以各種有_式，包括但秘 =F3組成物可 ，i:明:==物何形式被施 二ί 用任何標準_‘二二.1:腔乃： 道、内動脈、爾_内等使用直:何成陰 和其他傳統的組成物。 、从末、磨石τ、孔液、萃取物， 201206457 « 成分太ίϊ或ΐ活性，包括_形式，或作為食物錄料添加劑。 时ί ϊί 频實施射，喊物崎何合適的形式被經 f ϋϊ ’例如’丸劑’膠囊劑，顆_，或懸浮劑。 ，、且成物柯結合任何醫藥上可射_。「醫藥上可接受之載 ’ 任何醫藥載劑’如標準載劑，如，Remin_:藥劑 ，Lippincott Williams & w.jk.ns _ Tw^ ^ 括;二二5):述者。合適的載劑實例在本領域為習知，可包 之顧，衝液、水、乳化液如油/水2匕和tLi ，，、他載劑可能還包括無菌溶液、片劑，醫荜包衣片和#囊二、 此等載劑典型地含有賦形劑如殿粉、牛奶、糖醫樂某：二=。、Treatment ("Fig. 40^2 inhibited the shift of the axis to the liver tissue". Her combination of mosquitoes and effective CDDP with only CDDP i data sheets may also reduce distant metastases. Pottery's effectiveness of the method' It may be necessary to combine an anti-cancer composition with a drug that cures hyperproliferative diseases. These combination therapies can occur in the same in situ injection protocol that the coffee can be combined with in the same in situ injection protocol. In the present invention, in addition to the administration of the therapeutic agent, one or more therapeutic agents are used in situ by the method of the present invention. The tumor cells of the therapeutic and radiotherapy agents represent clinical oncology. : The goal of witnessing cancer research is to try to combine the efficacy of chemotherapy with chemotherapy and radiation therapy. For example, when retrovirus is used to deliver the simple bubble-thymidine (10)_tK gene to brain tumors, Succeeded in the susceptibility of the antiviral drug ganciclovir (Culver, etal). The present invention contemplates that =F3 treatment can be similarly combined, among other pro-apoptotic or cell cycle regulators. For chemotherapy, radiation therapy, or immunotherapeutic intervention. 14 201206457 Pharmacological treatment can be used for several minutes to several weeks at other intervals, and will not be sent to the cells, as will ensure that the period of time does not exceed the deficiencies In the hour after the contact, the cells and the two sides are treated for a long time to be treated significantly, and the aging time between the administrations is several days (2, 3, 4, 5, 63, 4, 5, 6, 7, or 8). Or 7) to several weeks (Bu 2, about ^ 峨 ( ( ( ( ( 纽 纽 纽 纽 纽 纽 纽 纽 纽 纽 纽 纽 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲 玲Surgery and palliative surgery. Radical treatment with 5 and other therapies for cancer, such as Ben Xing Er 疋 = Feng hormone treatment, gene therapy, immunity II and including physical removal of all or part of cancerous tissue removed = microscopic control Surgery (Mohs surgery). Further envisaged the invention = and surface = cancer, primary cancer, or accidental amount of normal tissue resection for use. In cancerous _, _, or lumps in whole or in part with additional anti-cancer Therapy perfusion, direct injection or zone = 3 70% / / σ treatment. This treatment can be repeated, for example, every 1, 2, 3 '4, 5, f 7 days ' or every 2, 3, 4, and 5 weeks or every 1, 2 '3, 4 , 5, 6, 7, 8', 10, 11, or December. These treatments can also be different doses. Use the fine (4) Ganoderma lucidum F3 material to merge with the fines; body and two =: ===: i county proliferating cells on cell death inducer = other cytokines; F42K * other cytokines two or ΜΠΜ, ΜΡ · 1β, M 〇 m, test, and __. The present invention ^ 15 201206457 It is not thought that cell surface receptors or their ligands such as Fas/Fas ligands, DR4 bite DR5/TRAIL upregulation will enhance the ability of the present invention to induce apoptosis by establishing autocrine or paracrine effects on hyperproliferative cells. . Increasing the number of GAp linkages to increase intercellular communication will increase the anti-hyperproliferative effect on adjacent hyperproliferative cell populations. In other embodiments, cytostatic or differentiation agents can be used in conjunction with the present invention to improve the anti-hyperproliferative effect of the treatment. Cell adsorption inhibitors are believed to improve the utility of the present invention. Examples of cell adsorption inhibitors are topical adhesion kinase (FAKs) inhibitors and lovastatin. The present invention also contemplates other agents that increase the susceptibility of hyperproliferative cells to apoptosis, such as antibody c225, which can be used in conjunction with the present invention to improve therapeutic effects. The Ganoderma lucidum F 3 moiety can be combined with other chemotherapeutic agents for treating cancer or combined, including alkylating agents, antimetabolite anti-antagonists, anti-cancer antibiotics, and plant, anti-cancer agents. Examples of the "burning agent" include nitrogen-containing mustard oil, nitrogen-containing mustard oil _N_oxide hydrochloride, chlorambutyl, guanamine, ephingamine, Shaotpa, card Carboquone, improsulfan tosylate, butyl sulfonate, nimustine hydrochloride, mitobronitol, azathioprine, nitrogen Betacarbamate, ranimustine, estradiol sodium sulphonate, triethylenemelamine, bis-ethylidene; g-chammine, cyclohexylnitrite , streptozotocin, pip〇br〇man, etoglucid, caproden, shunnu, mib〇piatin, nedaplatin, oxa Oxaiipiatin, hexamethyl melamine altretamine, ambastamustine, dibrospidium hydrochloride, f〇temustine, prednimustine ), 〇 〇 〇 〇 (pUmitepa), rib〇mustin, temo 0 amine (temozo Lomide), treosulphan, trophosphamide, zinostatin stimalamer, carboquone, adozelesin, cysteine sulphate (CyStemustine), And bizelesin. Examples of π antimetabolites include thiol sulfonium, 6-thiol ribose, thioinosine, amine leucovorin, enocitabine, 201206457 Arab Glycosyl cytosine, arabinosylcytosine octadecyl phosphate, ancitabine hydrochloride, 5-FU drugs (eg, fluoropurine bite, tegafUr, UFT, 5 '-Deoxy-5-fluorouric acid, carmofUr, gallocitabine, emmitefUr), aminopterine, leucovorin calcium, tablets , butocine, folinate calcium, levofolinate calcium, cladribine, emitefUr, fludarabine, gemcitabine (gemcitabine), hydroxy urea ( Hydroxycarbamide), pentostatin, piritrexim, moth deoxyuridine, mitoguazone, thiazophrine, and ambamustine and many more. Examples of "anticancer antibiotics" include actinomycin, actinomycin-C, mitomycin-C, chromomycin-A3, bleomycin hydrochloride, bleomycin sulfate, and petomycin sulfate. (peplomycin sulfate), daunorubicin hydrochloride, doxorubicin hydrochloride, aclarubicin hydrochloride, biraubicin hydrochloride, ibrubicin hydrochloride, neopotoma, mithramycin ( Mithramycin), sarcomycin, carzinophilin, mitotane, zorubicin hydrochloride, dextromethorphan hydrochloride, and idarubicin hydrochloride "Plant-derived anticancer agents" include etoposide phosphate, vinblastine sulfate, vincristine sulfate, vindesine sulfate, tannippoise, paclitaxel, and docetaxel ( Plus (10) curry), and vinorelbine (vinorelbine) and so on. An exemplary drug that can be administered in combination with a Ganoderma Lucidum F3 composition is cisplatin. Cisplatin has been widely used to treat cancer such as metastatic red or (4) cancer, advanced bladder cancer, head or neck cancer, cervical cancer, lung cancer or other tumors. Cisplatin (CDDp) can be combined with a single thief and a Ganoderma lucidum F3 agent for an effective dose of 5 2 〇 mg/m 2 for clinical use, in all two, every three days of the disease. The exemplary dose can be 〇5 to 2, l.Omg/m, 1.5〇mg/m, U5mg/m2, mg/m2, 3 〇mg/m2, 4 〇5Omg/m_, lOmg/m, 20mg/m2, 50mg /m2. Of course, all of this is an exemplary use. Any dose between these points can also be used in the present invention for oral absorption. Therefore, it must be delivered intravenously, subcutaneously, intratumorally or intraperitoneally. Treatment 5 The invention combines or uses any of the foregoing other cancers to treat certain cancers, such as breast cancer, pro-hormone or estrogen. This treatment is usually used as a treatment option or to reduce the risk of metastasis - his cancer treatment combined with transporting the spirit to be used externally, rectal, etc. in a manner that achieves the desired purpose. ^ An effective route of administration, such as oral, primate, pets, such as tracing, dogs, New Zealand humans, female humans, to the desired host. The effective amount refers to the ability to reach the regulatory decision, for example, the dose response experiment, the amount of the amount can be used to select the amount, including the Ganoderma lucidum extract, in the age of various animal-like cells: or see, 2 grams of '3 grams , 5 〇〇 mg 25 coupons, see 2.5 grams of '1 $ powder, Ganoderma lucidum F3 paste, or Ganoderma lucidum F3 round = Ganoderma lucidum, depending on the needs of the recipient, the preparation of the square is effective to include the t month and the evening The agent is used together. The Ganoderma lucidum F3 of the present invention has a variety of _, including but the secret = F3 composition, i: Ming: == What is the form of the object is applied by any standard _'二二.1: cavity is: Road , internal artery, _ _, etc. use straight: He Chengyin and other traditional compositions. , from the end, grindstone τ, pore liquid, extract, 201206457 « Ingredients too ϊ or ΐ activity, including _ form, or as a food recording additive. When the ί ϊί frequency is fired, the appropriate form of shouting saki is taken by f ϋϊ ', for example, 'pill' capsule, _, or suspension. , and the combination of the substance and the ke can be combined with any medicine. "Pharmaceutically acceptable 'any pharmaceutical carrier' such as a standard carrier, eg, Remin_: Pharmacy, Lippincott Williams & w.jk.ns _ Tw^ ^ Included; 22:5): The appropriate load Examples of agents are well known in the art and may be included in the compositions, including flushing, water, emulsions such as oil/water 2 and tLi, and other carriers may also include sterile solutions, tablets, medical dressing tablets and # Capsule 2, these carriers typically contain excipients such as powder, milk, sugar, and medicine: two =.
St由硬:Ξ或其鹽類、硬脂酸鎂或硬脂酸鈣、滑石粉、植物脂 °此等載劑亦可包括香味及顏色添加劑或 二含此等載劑被以廣為人知的傳統方法製劑。 ^而s與組成物一起被製劑的賦形劑係適用於口服且不與該组 成物或其他活性成分起有害反應。 /、"/、、、 合適的醫藥上可接受細包括但不限於水、鹽溶 膠硬'、ί醇 '明膠、碳水化合物如乳糖、直鍵殿粉 二1更月曰觳鎂/月石、石夕酸、粘性石蠟、香料油、脂肪酸星 ΐίίίΤ t、㈣=維素、_基甲基纖維素、甲基纖維素、 =本曰知（octoxynol) 9、油醇、聚維酮、丙二醇單硬 j基硫酸納、山梨_、硬騎、黃#膠、黃轉及其衍1物、 ’和雜項成分如微晶_素、檸檬酸、 糖 鳴、乳酸、乳糖、氣化鎂、偏雜，粉及=夜體 ，組成物亦可與其他活性成份—妙劑，如抗氧化^維生 素（A、C、抗壞血酸、B’s，如m、硫胺素、β6，維生素恥、b 19 201206457 群、生物素、膽驗、於驗酸、泛酸、B12、氰錄胺和/或B2、D、 D2、D3、骨化醇、E，如生育紛、核黃素、κ、K1、。 實例 。。無意j艮制本發明之範圍，有關於本發明之具體實施例之例示 具，設備，方法及其等之相關結果提供如下。值得注意的是， 才不題，子;f示題可為便於閱讀方便而用於實例中，但不應限制本發 =之範圍。再者’此處提出及揭露某些理論；然而，只要本發明 ，在根據本發明下實施而無考慮到特定理論或行為流程圖，不論 這些理論正確與否，都不應限制本發明之範圍。 實例1 :細胞系 A549，一種人類肺腺癌細胞系，得自於生物資源保存及研究 中心（BCRC，新竹’台灣）。將細胞培養在培養液中， 其補充有10%胎牛血清及2mM L-麩胺酸（Life Technologies， Inc.’MD)。以胰蛋白酶_EDTA將吸附的細胞剝離。鼠類Lewis肺 癌（LLC，H-2b，ATCC : CRL-1642)得自於 BCRC 並培養在 RPMI 1640 及 Dulbecco 改進之 Eagle 培養基（GIBCO-LifeSt is hard: strontium or its salts, magnesium stearate or calcium stearate, talc, vegetable fats. These carriers may also include flavors and color additives or two traditional methods known as these carriers. preparation. The excipients in which s is formulated with the composition are suitable for oral administration and do not adversely react with the composition or other active ingredients. /, "/,,, Suitable pharmaceutically acceptable fines include, but are not limited to, water, salt sol hard ', yol' gelatin, carbohydrates such as lactose, direct bond temple powder, 2, 1 month, magnesium, moonstone , ashes, viscous paraffin, perfume oil, fatty acid star ΐίίΤ t, (4) = vegan, _ methyl cellulose, methyl cellulose, = octoxynol 9, oleyl alcohol, povidone, propylene glycol Single hard j-based sodium sulphate, Yamanashi _, hard riding, yellow #胶, yellow turn and its derivatives, 'and miscellaneous ingredients such as microcrystalline 素, citric acid, sugar, lactic acid, lactose, magnesium gas, partial Miscellaneous, powder and = night body, the composition can also be combined with other active ingredients - such as antioxidants ^ vitamins (A, C, ascorbic acid, B's, such as m, thiamine, β6, vitamin shame, b 19 201206457 group , biotin, biliary test, acid test, pantothenic acid, B12, cyanamide and / or B2, D, D2, D3, bone alcohol, E, such as fertility, riboflavin, κ, K1. Examples. The scope of the present invention is not intended to be exhaustive, and examples of the specific embodiments of the present invention, apparatus, methods, and the like are provided as It is worth noting that the subject can be used in the examples for convenience of reading, but should not limit the scope of this issue. Furthermore, some theories are proposed and disclosed here; however, As long as the invention is practiced in accordance with the present invention without taking into account a particular theory or behavioral flow diagram, the scope of the invention should not be limited, whether or not the theory is correct. Example 1: Cell line A549, a human lung adenocarcinoma cell Department, obtained from the Center for Conservation and Research of Biological Resources (BCRC, Hsinchu 'Taiwan). The cells were cultured in culture medium supplemented with 10% fetal bovine serum and 2 mM L-glutamic acid (Life Technologies, Inc.'MD The adsorbed cells were stripped by trypsin_EDTA. Mouse Lewis lung carcinoma (LLC, H-2b, ATCC: CRL-1642) was obtained from BCRC and cultured in RPMI 1640 and Dulbecco's modified Eagle medium (GIBCO-Life).
Technologies)，其補充有15机之NaHC〇3及ι〇%胎牛血清 (FBS)。所有細胞被維持在37τ在一 5%c〇2_95%空氣之潮濕環 境。 實例2:試劑和抗醴 重組人類 TGF-βΙ (R&D)，抗-E-弼黏素（BD bioscience)， 抗-N-鈣黏素（BD bioscience )，抗-中間絲蛋白（Sigma)，抗个 連環蛋白（Santa Cruz)，抗-磷酸-Smad2/3 (Santa Cruz)，抗-snail (Abeam)，抗-slug (Cell Signaling) ’ 抗-TGF-PRII (Santa Cruz)， 抗-β-肌動蛋白（Santa Cruz)。 實例3:小鼠 雄性C57BL/6 (H-2b ; 6-8週大）小鼠購自於國家實驗動物 中心（台北）。小鼠在陽明大學實驗動物照護及使用委員會核准下 20 201206457 養育。 實例4 :靈芝部分粉末之製餚 靈芝粗萃取物之製備：將靈芝子實體研磨 將這些子實體顆粒加入〇.麵預熱至8〇。。 〇 ;立’ 公聽液），於崎㈣混合物溫和縣 液至至溫並以32%HC1調整pH至7.2±0,2。將辦石r= 升溶液）添加至該溶液，勝，接著i^慮過慮 工= 靈芝粗萃取物。此靈芝粗萃取物之顯示許多不 HPLC (高壓液相層析）圖形如圖5所示。 里 芝7部分之製備：使用1GGKDt"向流過濾、法系統 jlhpore C〇rp〇ratlon，USA)進一步純化該靈芝粗萃取物以 量部分、成分及就丨。純化之f芝F3部分之肌C 圖形如圖6所不。 、、乾燥靈芝F3部分粉末：以標準凍乾程序從靈芝打部分純化 溶液移除水。 此處所揭露之靈芝F3部分相同於美國專利^^〇. 7，135，183圖j 中之部分-3，155至205 mL，該案之全文以引用的方式併入本文 中。用來比較這些部分之方法為HPLC指紋圖形及細胞激素 圖形。 靈芝F3部分組成物：將靈芝F3部分粉末溶解於二次蒸餾 H20或PBS中。於25°C攪拌溶液30分鐘，並於4°C離心（1800 X，）10分鐘。將上清液（溶解率為50%)使用〇 22μπ1無菌過濾 器（Millipore，Inc. ’ Temecula，CA)過滤’如文獻所述（Hsu Η-Υ， etal. (2004)靈芝多醣萃取物透過TLR4_調節蛋白質激酶訊息傳 遞途徑誘發細胞激素表現。jImmun〇l丨73 ( 1〇) : 5989-5999)。 實例5 :全細胞萃取物之製備 為了分析蛋白質表現，培養細胞（4 X 105於6 cm組織培養 盤）並以靈芝F3部分（60，120及180 pg/ml)或空載體處理。以 含l%Na3V04之冷PBS潤洗細胞一次，以40 μΐ裂解緩衝液將細 21 201206457 胞原位裂解’裂解之細胞被置於冰上10分鐘，接著於pc 12〇〇〇χ rpm離心10分鐘。使用Bradford蛋白質分析法（Bi〇_Racj， Hercules ’ CA)以牛血清白蛋白做為基準來決定上清溶液之蛋白 質濃度。 實例6:西方墨點法分析 在培養期後收集細胞並添加裂解緩衝液。等量之蛋白質被裝 入10% SDS-PAGE並透過濕轉印墨點系統移轉至Imm〇bil〇n⑧ PVDF 膜上（Blot Electrophoretic Transfer Cell，Bio-Red ;移轉緩衝 液：25 mM Tris-base ’ 192 mM 甘胺酸 ’ 〇.i〇/0 SDS 及 20%甲醇， 250mA，100分鐘）’用於西方墨點分析。 實例7 :活髖外細胞移轉、入侵及軟瘦脂菌落形成分析 使用 6.5-mm Costar Transwell® chambers (8 μιη 孔徑； Coming ’ NY)進行細胞移轉分析。在llc細胞以靈芝F3部分 (120pg/ml)處理後 ’ 4 X 1〇4 細胞/2〇〇μΐ 被種在 Transwell® 室中。 使用Tmnswell®過濾器之細胞入侵分析被覆以適當的Matrigd(g) (Becton Dickinson ’ Franklin Lakes，NJ)。在 24 小時培養後，將 該過濾器溫和地從室中移除並以棉籤掃除上表面之無侵入性細 胞、。將細胞以曱醇固定10分鐘。將移轉至膜下側之細胞以劉氏染 色法染色（Handsel Technologies，Inc.，台北，台灣）。於 ιοοχ 放 大倍率顯微鏡下檢測細胞。為做軟瓊脂菌落形成分析研究，LL(： 細胞被置入具上層為0.33%瓊脂、底層為〇.5%瓊脂（N〇Weagar， Becton Dickinson )、含 1 〇% FBS 之 DMEM6 孔盤中（每孔！ 500 細 胞）。培養細胞16天並計算菌落數。 v 實例8:活«内移轉分析 LLC細胞（2x105細胞）經由側尾靜脈注射植入C57bl/6 小鼠。隨機將小鼠分為六個處理組：、靈芝F3部分（腹腔注 射）、靈芝F3部分（口服）、順-雙氨雙氣鉑（n)(CDDp〇r順 /靈芝F3部分（腹腔注射），或CDDP/靈芝F3部分（腹腔注 射）。第7天對小鼠腹腔注射CDDP (〇.5mg/g)。同時，對小鼠每 22 201206457 兩天腹腔注射靈芝F3部分（〇.〇75mg/g)，或每天口服靈芝部 分（0.15 mg/g)。4週後犧牲小鼠，檢驗所有器官移轉形成的情形。 將肺移除並固定在10〇/〇三聚曱醛中。計算肺腫瘤菌落之數量及腫 瘤體積。腫瘤體積以長X寬X高xO.5236 (mm3)計算，參照文獻 (Santini D > Vincenzi B > Tonini G > Scarpa S » & Baldi A ( 2003 ) Correspondence Re : E。Corey et al.，唑來膦酸展現對攝護腺癌 轉移成骨和溶骨之抑制效果。Clin. Cancer Res。，9: 295_306, 2003。Clinical Cancer Research 9 ( 8 ) : 3215-3215 )。代表性之肺 腫瘤被移除、固定並包埋在石蠟中。將包埋之組織以H&E染色用 來組織分析。 y' 實例9 :统計分析 數據以平均數±標準差表示’且統計上顯㈣學^t·測試來檢 本說明書中所有引用之文獻及專利申請案以引用的方式併入 本文中’猶如每獨立文獻或專利φ請案被特定且獨立地 用的方式併入。 =前述發明已以明確理解之目的藉由圖說及實例詳盡地敛 二j t心此一行業技藝人士依本發明之教示，可明顯知道某種 ^。、修飾是可在轉_之巾請糊範圍之簡絲嘴下被做成 【圖式簡單說明】 内容成1明書之部分並被包括以進—步證明本揭露 由==== 23 201206457 顯示上皮形態。B-D，A549細胞與TGF-βΙ ( 1，2及5 ng/ml) — 起培養72小時。E-H ’以含各種濃度TGF-βΙ (〇，1，2及5 ng/ml) 之120 pg/nil靈之F3部分處理細胞72小時（放大倍率X2〇〇)。 (IB) A549 細胞（1x105 細胞/ml)與 5 ng/ml 之 TGF-βΙ —起培 養24小時。24小時後’以靈芝F3部分（120 刺激24小 時。蒐集總細胞裂解液，以西方墨點法偵測EMT標記。（ic)在 與或不與TGF-βΙ (5 ng/ml) —起培養24小時後，以靈芝F3部 分（120 pg/ml)處理A549細胞（2x105細胞/總）24小時。在處 理後’細胞被殖於Transwell系統。由Transwell系統以以聚碳酸 酯過濾器（孔徑，8 μιη)測量細胞移轉。藉由在顯微鏡下計量移 動至聚碳酸I旨多孔膜下方之細胞數來量化Α549細胞之移轉能 力。 圖2顯示’靈芝F3部分（顯示為EORP)配方降低了 A549細 胞中TGFpl訊息傳遞途徑。（2A)將A549細胞與或不與靈芝F3 部分一起培養在培養基中6及24小時。蒐集培養液以測量 TGF-βΙ，其由ELISA測定。數據以平均數土標準差表示。* :控 制組及經靈芝F3部分處理之A549細胞之間的顯著不同，p< 0.05。將A549細胞與或不與TGF-βΙ —起培養24小時後，以或 不以靈芝F3部分處理3小時。以或不以TGF-βΙ ( 5 ng/ml)處理 A549細胞24小時，接著置換新鮮培養液。接著，以靈芝F3部 分（120 pg/ml)處理A549細胞3小時。（2B)鱗酸化Smad2/3 之表現、（2C) Snail及Slug之表現以西方墨點法偵測之。 圖3顯示靈芝F3部分（顯示為EORP)配方抑制了 A549及 LLC-1細胞之移轉、入侵及菌落形成。（3A)以靈芝F3部分（12〇 pg/ml)處理LLC-1細胞（4 X 104細胞/ 200μ1) 18小時。由 Transwell系統以以聚碳酸酯過濾器（孔徑，8 μιη)測量細胞移轉 及細胞入侵。藉由在顯微鏡下計量入侵至聚碳酸酯多孔膜下方之 細胞數來量化Α549細胞之移轉及入侵能力。（3Β)將LLC-1辦 胞鋪在具上層為0.33%瓊脂、底層為0.5%瓊脂、含10% FBS之 DMEM 6孔盤中。培養細胞16天並計量菌落數。顯示之數據為爹 24 201206457 複實驗的三重樣本之平均數+標準差，在三個獨立 似結果。* p<0.05及**ρ<〇.〇ι對照控制組。 又于相 圖4顯示，結合CDDP之靈芝F3部分（顯示為E〇Rp 方抑制了腫瘤移轉。將llc-1細胞經由尾靜脈注射植入小f二 "組n=8)。7天後，以CDDP(0.5mg/g)腹腔注射處理小氣乳2 以靈芝F3部分（0.075 mg/g)每2天腹腔注射處理，或以靈芝F3 部分（〇.〇75mg/g)每日口服投藥進行4週。（4A)左圖為肺腫瘤 夕重性’右圖為肺病變之腫瘤體積（mm3)。* 〇1 及***ρ<0·001對照控制組或CDDP組。（4Β)上圖為肺；中圖為 以HE染色之肺切片：Χ20放大倍率；下圖為以册染色之肺切片： x 400放大倍率。（4C)上圖為he染色之肝切片：χ1〇〇放大倍 率；下圖為HE染色之肝切片：x4〇〇放大倍率。 ° 圖5顯示靈芝粗萃取物之HPLC圖樣，使用尺寸篩除管（Tosoh TSK-GEL G5000PW) ° 圖6顯示經純化之靈芝F3部分之HPLC圖樣，使用尺寸篩 除管（TosohTSK-GELG5000PW)。 【主要元件符號說明】 盔Technologies), which is supplemented with 15 machines of NaHC〇3 and ι〇% fetal bovine serum (FBS). All cells were maintained at 37 τ in a humid environment of 5% c〇2_95% air. Example 2: Reagents and anti-tuberculosis recombinant human TGF-βΙ (R&D), anti-E-mucin (BD bioscience), anti-N-cadherin (BD bioscience), anti-intermediate silk protein (Sigma) , anti-catenin (Santa Cruz), anti-phospho-Smad2/3 (Santa Cruz), anti-snail (Abeam), anti-slug (Cell Signaling) 'anti-TGF-PRII (Santa Cruz), anti-beta - Actin (Santa Cruz). Example 3: Mice Male C57BL/6 (H-2b; 6-8 weeks old) mice were purchased from the National Laboratory Animal Center (Taipei). The mice were raised under the approval of the Experimental Animal Care and Use Committee of Yangming University 20 201206457. Example 4: Ganoderma lucidum partial powder preparation Ganoderma lucidum crude extract preparation: Ganoderma lucidum fruiting body grinding These fruiting body granules were added to the enamel surface to preheat to 8 Torr. . 〇;立's public hearing liquid, Yuqi (four) mixture mild county liquid to the temperature and adjust the pH to 7.2 ± 0, 2 with 32% HC1. Add the stone r = liter solution) to the solution, win, and then consider the work = Ganoderma lucidum crude extract. The display of this crude extract of Ganoderma lucidum is not shown in Figure 5. Preparation of 7 parts of Lizhi: The crude extract of Ganoderma lucidum was further purified by using 1GGKDt"flow filtration, system jlhpore C〇rp〇ratlon, USA). The muscle C pattern of the purified F3 F3 part is shown in Fig. 6. , Drying Ganoderma F3 Partial Powder: Partially purified solution from Ganoderma Lucidum was removed by standard lyophilization procedure. The Ganoderma lucidum F3 portion disclosed herein is identical to the portion of the U.S. Patent No. 7,135,183, the entire disclosure of which is incorporated herein by reference. The methods used to compare these parts are HPLC fingerprints and cytokine graphics. Ganoderma lucidum F3 partial composition: The Ganoderma lucidum F3 partial powder is dissolved in a secondary distillation H20 or PBS. The solution was stirred at 25 ° C for 30 minutes and centrifuged (1800 X,) for 10 minutes at 4 °C. The supernatant (dissolution rate 50%) was filtered using a 〇22μπ1 sterile filter (Millipore, Inc. 'Temecula, CA) as described in the literature (Hsu Η-Υ, et al. (2004) Ganoderma lucidum polysaccharide extract through TLR4 _ regulate protein kinase signaling pathway induces cytokine expression. jImmun〇l丨73 (1〇): 5989-5999). Example 5: Preparation of whole cell extracts To analyze protein expression, cells (4 X 105 in 6 cm tissue culture plates) were cultured and treated with Ganoderma lucidum F3 fraction (60, 120 and 180 pg/ml) or empty vector. The cells were washed once with cold PBS containing 1% Na3V04, and the cells of the fine 21 201206457 cells were lysed in 40 μM lysis buffer. The lysed cells were placed on ice for 10 minutes, followed by centrifugation at pc 12 rpm. minute. The protein concentration of the supernatant solution was determined using Bradford protein assay (Bi〇_Racj, Hercules ' CA) based on bovine serum albumin. Example 6: Western blot analysis The cells were harvested after the incubation period and lysis buffer was added. Equal amounts of protein were loaded onto a 10% SDS-PAGE and transferred to a Imm〇bil〇n8 PVDF membrane via a wet transfer dot system (Blot Electrophoretic Transfer Cell, Bio-Red; Transfer Buffer: 25 mM Tris- Base '192 mM glycine' 〇.i〇/0 SDS and 20% methanol, 250 mA, 100 minutes) 'for western blot analysis. Example 7: Analysis of live hip extracellular migration, invasion, and soft leptin colony formation Cell transfer assays were performed using 6.5-mm Costar Transwell® chambers (8 μιη pore size; Coming ' NY). After treatment with the Ganoderma lucidum F3 fraction (120 pg/ml), the '4 X 1〇4 cells/2〇〇μΐ were planted in the Transwell® chamber. Cell invasion assays using a Tmnswell® filter were coated with the appropriate Matrigd(g) (Becton Dickinson ' Franklin Lakes, NJ). After 24 hours of incubation, the filter was gently removed from the chamber and the non-invasive cells on the upper surface were swabbed with a cotton swab. The cells were fixed with sterol for 10 minutes. Cells that were transferred to the underside of the membrane were stained by Liu staining (Handsel Technologies, Inc., Taipei, Taiwan). The cells were examined under a large magnification microscope under ιοοχ. For the analysis of soft agar colony formation, LL (: cells were placed in DMEM6 well plates with 0.33% agar on the top layer, 〇.5% agar (N〇Weagar, Becton Dickinson), 1% F% FBS ( Each well! 500 cells). The cells were cultured for 16 days and the number of colonies was counted. v Example 8: Live «internal transfer analysis LLC cells (2x105 cells) were injected into C57bl/6 mice via lateral tail vein injection. For the six treatment groups: Ganoderma F3 (intraperitoneal injection), Ganoderma F3 (oral), cis-diammine-p-platinum (n) (CDDp〇r/Ganoderma F3 (intraperitoneal injection), or CDDP/Ganoderma lucidum Part F3 (intraperitoneal injection). CDDP (〇.5mg/g) was intraperitoneally injected into mice on day 7. At the same time, mice were intraperitoneally injected with Ganoderma lucidum F3 (〇.〇75mg/g) every day for 22 201206457, or daily Oral Ganoderma lucidum fraction (0.15 mg/g). After 4 weeks, the mice were sacrificed to test the formation of all organs. The lungs were removed and fixed in 10 〇 / 〇 trimeric furfural. Calculate the number of lung tumor colonies and Tumor volume. Tumor volume is calculated as length X width X height xO.5236 (mm3), reference (Santini D > Vincenzi B > Tonini G > Scarpa S » & Baldi A ( 2003 ) Correspondence Re : E. Corey et al., Zoledronic acid exhibits inhibitory effects on metastatic osteogenesis and osteolysis of prostate cancer. Clin. Cancer Res., 9: 295_306, 2003. Clinical Cancer Research 9 (8): 3215-3215). Representative lung tumors were removed, fixed and embedded in paraffin. The embedded tissue was stained with H&E. Used to organize the analysis. y' Example 9: Statistical analysis data is expressed as mean ± standard deviation 'and statistically significant (4) learning ^ t · test to check all references and patent applications in this specification are incorporated by reference. In this article, 'as if each independent document or patent φ request is specifically and independently used. = The foregoing invention has been explicitly understood by the illustrations and examples. According to the teachings of the invention, it is obvious that a certain kind of decoration can be made under the simple mouth of the towel, and the content is made into a part of the book and is included. Step proof of this disclosure by ==== 23 201206457 Epithelial morphology .B-D, A549 cells with TGF-βΙ (1,2 and 5 ng / ml) - 72 hours onwards. E-H' treatment of cells with 120 pg/nil of F3 fraction containing various concentrations of TGF-βΙ (〇, 1, 2 and 5 ng/ml) for 72 hours (magnification X2〇〇). (IB) A549 cells (1 x 105 cells/ml) were incubated with 5 ng/ml of TGF-β(R) for 24 hours. After 24 hours, the F3 fraction of Ganoderma lucidum was stimulated for 24 hours. The total cell lysate was collected and the EMT marker was detected by Western blotting. (ic) was cultured with or without TGF-βΙ (5 ng/ml). After 24 hours, A549 cells (2 x 105 cells/total) were treated with Ganoderma F3 fraction (120 pg/ml) for 24 hours. After treatment, the cells were colonized in the Transwell system. The Transwell system was used with a polycarbonate filter (aperture, 8 μιη) was measured for cell transfer. The transfer ability of Α549 cells was quantified by metering the number of cells moving under the microporous membrane under the microscope. Figure 2 shows that the Ganoderma lucidum F3 fraction (shown as EORP) formulation was reduced. TGFpl message transduction pathway in A549 cells. (2A) A549 cells were cultured in medium with or without Ganoderma lucidum F3 fraction for 6 and 24 hours. Culture medium was collected to measure TGF-βΙ, which was determined by ELISA. The standard deviation is expressed.*: Significant difference between the control group and A549 cells treated with Ganoderma lucidum F3, p< 0.05. A549 cells were cultured with or without TGF-β for 24 hours, with or without Ganoderma lucidum F3 Partially processed for 3 hours. A549 cells were not treated with TGF-βΙ (5 ng/ml) for 24 hours, followed by replacement of fresh medium. Then, A549 cells were treated with Ganoderma F3 fraction (120 pg/ml) for 3 hours. (2B) Scalified Smad2/3 Performance, (2C) Snail and Slug were detected by Western blotting. Figure 3 shows that Ganoderma lucidum F3 (shown as EORP) inhibits the migration, invasion and colony formation of A549 and LLC-1 cells. Treatment of LLC-1 cells (4 X 104 cells / 200 μl) for 18 hours with Ganoderma lucidum F3 fraction (12 〇pg/ml). Cell transfer and cells were measured by Transwell system with polycarbonate filter (pore size, 8 μιη) Invasion. The migration and invasion ability of Α549 cells were quantified by measuring the number of cells invaded under the porous membrane of polycarbonate under the microscope. (3Β) The LLC-1 cells were spread on the upper layer with 0.33% agar and the bottom layer was 0.5% agar, DMEM 6-well plate containing 10% FBS. The cells were cultured for 16 days and the number of colonies was counted. The data shown is 平均24 201206457 The average of the triple samples of the repeated experiments + standard deviation, in three independent results. * p < 0.05 and ** ρ < 〇. 〇 ι control group. Also in phase diagram 4 , the CD3-derived Ganoderma lucidum F3 moiety (shown as E〇Rp side inhibited tumor metastasis. The llc-1 cells were injected via the tail vein into the small f two " group n=8). After 7 days, CDDP (0.5) Mg/g) intraperitoneal injection of small air lotion 2 was administered intraperitoneally every 2 days with Ganoderma lucidum F3 fraction (0.075 mg/g) or daily for 4 weeks with Ganoderma lucidum F3 fraction (〇.〇75 mg/g). (4A) The left picture shows the lung tumor. The right picture shows the tumor volume (mm3) of the lung disease. * 〇1 and ***ρ<0·001 control control group or CDDP group. (4Β) The upper picture shows the lung; the middle picture shows the lung section stained with HE: Χ20 magnification; the lower picture shows the lung section sliced by the book: x 400 magnification. (4C) The upper panel shows hepatic sections stained with he: χ1〇〇 magnification; the lower panel is HE-stained liver section: x4〇〇 magnification. ° Figure 5 shows the HPLC pattern of the crude extract of Ganoderma lucidum, using a size screening tube (Tosoh TSK-GEL G5000PW) ° Figure 6 shows the HPLC pattern of the purified Ganoderma lucidum F3 part, using a size screening tube (TosohTSK-GELG5000PW). [Main component symbol description] Helmet
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| CN106124683A (en) * | 2016-08-26 | 2016-11-16 | 中国兽医药品监察所 | A kind of method of multiple antimicrobial component in veterinary drug of detection simultaneously |
| TWI608846B (en) * | 2014-03-13 | 2017-12-21 | 益生生技開發股份有限公司 | Combination therapy for ameliorating adverse side effects caused by chemotherapy |
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| TWI599370B (en) * | 2013-07-26 | 2017-09-21 | 中央研究院 | Antibody-mediated anti-tumor activity induced by reishi mushroom polysaccharides |
| US20220125805A1 (en) * | 2019-02-07 | 2022-04-28 | Universidad Central Del Caribe | Biologically active ganoderma lucidum compounds and synthesis of anticancer derivatives; ergosterol peroxide probes for cellular localization |
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| US7135183B1 (en) * | 2001-08-06 | 2006-11-14 | Academia Sinica | Immuno-modulating antitumor activities of Ganoderma lucidum (Reishi) polysaccharides |
| US20050032882A1 (en) * | 2002-03-06 | 2005-02-10 | Sophie Chen | Botanical extract compositions and methods of use |
| ES2373663T3 (en) * | 2003-09-08 | 2012-02-07 | Genyous Biomed International Inc. | COMPOSITIONS OF BOTANICAL EXTRACTS FOR CANCER THERAPY. |
| US20070071766A1 (en) * | 2005-09-23 | 2007-03-29 | Ko Jiunn L | Compositions comprising fungal immunomodulatory protein and use thereof |
| WO2007136857A2 (en) * | 2006-05-19 | 2007-11-29 | The Johns Hopkins University | Hox compositions and methods |
| WO2009111067A2 (en) * | 2008-03-07 | 2009-09-11 | Osi Pharmaceuticals, Inc. | Methods for the identification of agents that inhibit mesenchymal -like tumor cells or their formation |
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| US10493126B2 (en) | 2014-03-13 | 2019-12-03 | Yeastern Biotech Co., Ltd. | Combination therapy for ameliorating adverse side effects caused by chemotherapy |
| CN106124683A (en) * | 2016-08-26 | 2016-11-16 | 中国兽医药品监察所 | A kind of method of multiple antimicrobial component in veterinary drug of detection simultaneously |
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