WO2008119771A2

WO2008119771A2 - Quinoline-s-carboxylic acid derivatives as tyrosine kinase inhibitors

Info

Publication number: WO2008119771A2
Application number: PCT/EP2008/053763
Authority: WO
Inventors: Jacob Westman; Johan Wannberg; Natalia Nekhotiaeva; Ulrika BÄCKMAN
Original assignee: Clanotech Ab
Priority date: 2007-03-30
Filing date: 2008-03-28
Publication date: 2008-10-09
Also published as: WO2008119771A3

Abstract

A quinoline-3-carboxyderivative of formula (I), a method for preparing it and use ofthe derivativein therapy, in particular for the treatment and prevention of cell proliferative disorders or cell differentiation disorders.

Description

Compounds, use and methods

Field of the invention

The present invention relates to quinoline derivatives and to the use thereof in therapy. More particularly, the present invention relates to quinoline-3-carboxylic acid derivatives acting as tyrosine kinase inhibitors for treatment and prevention of cell proliferative disorders or cell differentiation disorders, disorders that are associated with abnormal tyrosine kinase activities.

Background of the invention Angiogenesis, the outgrowth of new capillaries from pre-existing vessels, is essential for embryonic development, organ formation, tissue regeneration, and remodeling (1). It also contributes to the development and progression of a variety of pathological conditions, including tumor growth and metastasis, cardiovascular diseases, diabetic retinopathy, rheumatoid arthritis, macular degeneration-related disorders and psoriasis (2). Angiogenesis and vasculogenesis are com- plex multistep processes that include proliferation, migration and differentiation of endothelial cells, degradation of the extracellular matrix, tube formation, and sprouting of new capillary branches (3, 11). The complexity of the angiogenic processes suggests the existence of multiple controls of the system, which can be transiently switched on and off. A switch of the angiogenic phenotype in tissues is thought to depend on a local change of the balance between angiogenic stimulators and inhibitors (4).

Among many described angiogenic factors, vascular endothelial growth factor (VEGF)/ vascular permeability factor is one of the best-characterized positive regulators with its distinct specificity for vascular endothelial cells (5-7). The biological actions of VEGF include stimulation of endo- thelial cell proliferation, migration, differentiation, tube formation, increase of vascular permeability, and maintenance of vascular integrity (8-11, 28). The angiogenic responses induced by VEGF are mediated by two structurally related tyrosine kinase receptors, VEGFR-I (FIt-I) and VEGFR-2 (KDR or FIk-I), both of which are expressed primarily on vascular cells of the endothelial lineage (8, 12, 13).

VEGFR-I and VEGFR-2 belong to a class of enzymes that catalyze the transfer of the terminal phosphate of adenosine triphosphate to tyrosine residues in protein substrates and therefore are called tyrosine kinases. Tyrosine kinases are believed, by the way of substrate phosphorylation, to play critical roles in signal transduction for a number of cell functions. Though the exact mechanism of signal transduction is still unclear, tyrosine kinases have been shown to be important contributing factors in cell proliferation, carcinogenesis and cell differentiation. Tyrosine kinases can be categorized as either the receptor type tyrosine kinases, that have an extracellular, a transmembrane, and an intracellular portion, or the non-receptor type tyrosine kinases that are wholly intracellular.

The receptor-type tyrosine kinases are comprised of a large number of transmembrane receptors with diverse biological activity, such as EGFR, HER2, HER3, HER4, TGF-α, amphiregulin, HB- EGF, betacellulin and heregulin, INS-R, IGF-IR, and IR-R, PDGF-α and β receptors, CSFIR, c- kit and FIk-II. VEGFR-I (FIt-I), VEGFR-2 (KDR) and VEGFR-3 (Flt-3) (32). The non-receptor type of tyrosine kinases is also comprised of numerous subfamilies, including Src, Frk, Btk, Csk, AbI, Zap70, Fes/Fps, Fak, Jak, Ack, and LIMK. Each of these subfamilies is further sub-divided into varying receptors (33).

As mentioned above VEGF binds to both the VEGFR-I and VEGFR-2 tyrosine kinases. However, the biological responses appear to be quite different (14). For example, activation of VEGFR-2 leads to proliferation and migration of endothelial cells, whereas VEGFR-I is unable to transduce such signals when stimulated with VEGF (14). VEGF is the prototype of an enlarging family of growth peptides that includes four other structurally related members. These re- cently identified VEGF-related molecules are placenta growth factor (PlGF) (15), VEGF- B/VEGF-related factor (16, 17), VEGF-C (18, 19), and c-/os-induced growth factor (FIGF/VEGF-D) (20, 29, 30). They show a striking similarity in their primary sequences, especially in the platelet-derived growth factor-like domain containing eight conserved cysteine residues. PlGF is predominantly expressed in the placenta and binds to VEGFR-I, but not to VEGFR-2 (21, 22). VEGF-B has been identified as a weak mitogen for endothelial cells and a robust expression is particularly detected in skeletal and cardiac muscle tissues (16). Both PlGF and VEGF-B modulate VEGF activity via formation of heterodimers (16, 23). VEGF-C is a ligand for two receptors, VEGFR-2 and VEGFR-3 (Flt-4) (18, 24). The latter differs from VEGFR-I and VEGFR-2 by being predominantly expressed in lymphatic endothelial cells in adult tissues, but at low levels in most other vascular endothelial cells (25, 31). VEGF-C recently has been characterized as being a fairly selective growth factor for lymphatic vessels (26, 27). In addition, proteolytic processing is involved in the regulation of VEGF-C activity (24). FIGF/VEGF-D, which is drastically induced by c-fos activation (20), also binds to VEGFR-2 and VEGFR-3 (30). Although these factors are believed to stimulate endothelial cell growth in vitro, their in vivo angiogenic effects have not yet been fully characterized.

Tyrosine kinases are without doubt interesting targets for finding methods for preventing or cur- ing diseases that are developed due to over-expression of growth factors. Solid tumors can be treated by tyrosine kinase inhibitors since these tumors depend on angiogenesis for the formation of the blood vessels necessary to support their growth and, indeed, there are a number of tyrosine kinase inhibitors already in the clinic.

An important aspect of tyrosine kinase inhibitors is selectivity since, as mentioned herein above, many of the growth factors as well as the tyrosine kinases are very closely structurally related and also show many different biological responses. Many of the known kinase inhibitors of today are multi-targeting and show different kinase inhibiting profiles and thus different biological responses. Thus, a "good kinase profile" will be important when developing a new drug with improved selectivity.

Recent research has established that blocking VEGFR-I induces some unwanted side-effects such as bone marrow toxicity (37) and liver complications (38), which means that a selectivity between VEGFR-2 and VEGFR-I is important and would be an advantageous feature of a me- dicament aimed at the treatment of disorders involving essentially the VEGFR-2 compared to the VEGFR-I. Accordingly, one object of the invention is to provide a compound which specifically inhibits, regulates and/or modulates the signal transduction of tyrosine kinases, in particular VEGFR-2, for use as a medicament.

Quinoline derivatives have been described earlier in a biological context and also in a number of therapeutic areas. As an example, Bi et al (34) have described quino lines as extremely potent and selective PDE5 inhibitors, being potential agents for treatment of erectile dysfunction. Allais et al (44) describe 2-methyl/phenyl quinolines having anti-inflammatory activity. Hanifin et al (45) describe 4-anilino-3-quinolinecarboxylic acids and esters as diuretic and anti-depressant agents.

A number of quinoline derivatives also are known as tyrosine kinase inhibitors. As an example, Boscelli et al (35) have described quino line-3-carbonitriles as Src Kinase inhibitors. Wissner et al (36) have described quino line-3-carbonitrile as an inhibitor of epidermal growth factor recep- tor kinase. Quinoline-3-carbonitrile also has been described in Wissner et al (46) as not only EGFR inhibitor, but also as HER-2 inhibitor.

Recently Kim et al described quinoline-3-carboxylic acid esters as DYRKl inhibitors (47) for the treatment of learning and memory deficits of people with Down syndrome

Thomas et al (48) describe 3-fluoro/hydrogen quinolines as VEGF tyrosine kinase inhibitors. Tang et al (49) describe quinoline-3-carboxylic acid esters as VEGF tyrosine kinase inhibitors. Sapelkin et al describe quinoline-3-carboxylic acid esters as CK2 inhibitors for treatment of cancer (50).

Macular degeneration is a clinical term used to describe a group of disorders that are generally characterized by a progressive loss of central vision associated with abnormalities of Bruch's membrane, the choroid, the neural retina and/or the retinal pigment epithelium. These disorders include very common conditions that affect older individuals (age-related macular degeneration or AMD) as well as rarer, earlier-onset dystrophies that in some cases can be detected in the first decade of life. Other maculopathies include North Carolina macular dystrophy, Sorsby's fundus dystrophy, Stargardt's disease, pattern dystrophy, Best disease, and Malattia Leventinese.

Age-related macular degeneration (AMD), is the most prevalent form of macular degeneration. It is associated with progressive diminution of visual acuity in the central portion of the visual field, changes in color vision, and abnormal dark adaptation and sensitivity. Two principal clinical manifestations of AMD have been described as the dry, or atrophic, form, and the wet, or exudative, form. The most significant risk factor for the development of both forms are age and the deposition of drusen, abnormal extracellular deposits, behind the retinal pigment epithelium. Drusen causes a lateral stretching of the retinal pigment epithelium and physical displacement thereof from its immediate vascular supply, the choriocapillaris. This displacement creates a physical barrier that may impede normal metabolite and waste diffusion between the choriocapillaris and the retina.

Another form of macular degeneration is Malattia Leventinese (ML), also termed Doyne's honeycomb choroiditis or dominant drusen. This is a genetic, early onset form of macular degeneration characterized by numerous, often confluent drusen that may radiate peripherally from the macula. Advanced AMD, which is responsible for profound vision loss, has two forms: dry and wet. Central geographic atrophy, the dry form of advanced AMD, results from atrophy to the retinal pigment epithelial layer below the retina, which causes vision loss through loss of photoreceptors (rods and cones) in the central part of the eye.

Neovascular or exudative AMD, the wet form of advanced AMD, causes vision loss due to abnormal blood vessel growth in the choriocapillaries, through Bruch's membrane, ultimately leading to blood and protein leakage below the macula. Bleeding, leaking, and scarring from these blood vessels eventually cause irreversible damage to the photoreceptors and rapid vision loss if left untreated.

Until recently, no effective treatments were known for wet macular degeneration. Lately, new drugs, called anti-angiogenics or anti-VEGF (anti- Vascular Endothelial Growth Factor) agents, have been approved by the FDA for use in the treatment of AMD, namely Lucentis and Macugen, from Genenetech. These, however, must be administered by injection directly into the vitreous humor of the eye, which makes their use limited to a hospital or clinical setting. Additionally, the administration by injection into the eye, which frequently has to be repeated on a monthly or bi-monthly basis, may be associated with distress and pain to the subject being treated and thereby give rise to a redcued treatment compliance.

Summary of the invention

The present inventors now have found that quinoline-3-carboxylic acid esters with certain novel side chain pattern inhibit the VEGFR-2 and VEGFR-3 tyrosine kinase with good potency and high selectivity, while being capable also of inhibiting other tyrosine kinases as well.

Consequently, according to one aspect the present invention relates to a compound of formula (I)

wherein n is 0-5;

Z is O or S and R¹ is selected from -OH, C1-C6 alkylNH-, (C1-C6 alkyl)₂N- and C1-C6 alkyl-

S(O)₂NH-; or Z is N forming together with R¹ an unsaturated, aromatic or non-aromatic hetero- cycle optionally containing further heteroatoms, which heterocycle is optionally substituted with at least one C 1 -C6 alkyl;

R² is Cl-C6 alkyl;

R³ _m represents m radicals R³, independently selected from halogen, NC-(CH2)p-, R⁴-,

R⁴O-(CH₂)P-, R⁴S-(CH₂)P-, R⁴S(O)₂-(CH₂)p-, R⁴R⁵N-(CH₂)p-, R⁴R⁵NC(O)-(CH₂)p-,

(R⁴C(O))(R⁵)N-(CH₂)p-, R⁴C(O)-(CH₂)P-, R⁴OC(O)-(CH₂)p-, R⁴C(O)O-(CH₂)p-; and when m > 2, two R³ may together form -O-(CR⁶R⁷)_k-O-, m is 0-5; k is 1 or 2; p is 0-3;

R⁴ and R⁵ are independently selected from H and C1-C6 alkyl; R⁶ and R⁷ are independently selected from H and halogen; and any C1-C6 alkyl optionally is substituted by one or several substituents selected from halogen and OH; as well as pharmaceutically acceptable salts or prodrugs thereof.

According to a further aspect, the present invention provides compounds of formula (I) for use as a medicament, e.g. a medicament for the treatment of diseases such as cancer, diabetic retinopathy, macular degeneration-related disorders, e.g. age-related macular degeneration, chronic inflammation, stroke, ischemic myocardium, atherosclerosis, tumor growth and macular edema.

According to still a further aspect, the present invention provides the use of the compounds of formula (I) or pharmaceutically acceptable salts or prodrugs thereof for manufacturing a medicament for the treatment of diseases such as cancer, diabetic retinopathy, macular degeneration-related disorders, chronic inflammation, stroke, ischemic myocardium, atherosclerosis, tumor growth and macular edema.

According to still a further aspect, the present invention provides the compounds of formula (I) or pharmaceutically acceptable salts or prodrugs thereof for the treatment of diseases such as cancer, diabetic retinopathy, macular degeneration-related disorders, chronic inflammation, stroke, ischemic myocardium, atherosclerosis, tumor growth and macular edema. According to another aspect, the invention provides a method of treatment of a disorder selected from cancer, diabetic retinopathy, macular degeneration-related disorders, chronic inflammation, stroke, ischemic myocardium, atherosclerosis, tumor growth and macular edema by administration of a therapeutically effective amount of a compound of formula (I) to a mammal in need of such treatment.

According to one aspect, a method of preparing a compound of formula (I) according to the invention is provided by reacting an aniline derivative (a)

(a) with a malonic acid derivative (b)

(b) cyclizing the formed intermediate (c)

(C) so as to obtain quinoline-4-ol derivative (d)

reacting (d) with a halogenating agent so as to obtain (e),

(e) wherein X is halogen, and reacting (e) with an amine (f)

According to one aspect, a method of preparing a compound of formula (I) according to the invention, wherein Z is O, is provided, by reacting a p-halogenoaniline (g)

(9) wherein Y is halogen, with a malonic acid derivative (b)

(b) so as to obtain a compound (h)

(H) cyclizing and halogenating (h) by reacting it with a suitable halogenating agent, so as to obtain a compound of formula (i) wherein Y and X are independently selected halogens,

reacting (i) with an amine (f)

so as to obtain a compound (j)

U) and reacting (j) with R¹H in the presence of Herrmann's palladacycle (£rα/?s-Di(μ-acetato)-bis[o- (di-o-tolylphosphino)benzyl]-dipalladium(II)); [(J-Bu)₃PH]BF₄; Mo(CO)₆; and DBU (1,8- Diazabicyclo[5.4.0]undec-7-ene).

Further aspects and embodiments of the invention are as defined in the claims.

Brief description of the drawings

Fig. 1 contains photos showing actin reorganization upon stimulation with VEGF, examined by fluorescence microscopy using tetramethyl-rhodamine isothiocyanate-phalloidin staining (A-C): A. PAE/VEGFR-2 cells without the presence of VEGF; B. spindle-like-shaped PAE/VEGFR-2 cells, leading lamellae, lamellipodium extensions and actin reorganization in the presence of VEGF; and C. restitution of PAE/VEGFR-2 cells morphology in the presence of VEGF and inventive Compound 5. Fig. 2 contains light microscope images of PAE cells expressing VEGFR-2 (PAE/VEGFR-2; column 1) and PAE cells lacking the expression of VEGFR-2 (PAE without R; column 2). Row 1 shows the cells without adding VEGF. Row 2 shows the morphology of the cells in the presence of VEGF. Rows 3-5 show the cells in the presence of VEGF and Compounds 9, 5 and 11 of the invention, respectively. The pictures show that the inventive compounds act as VEGF inhibitors (column 1) without affecting the cells lacking the VEGFR-2 receptor (column 2).

Fig. 3 is a bar chart representing percentage of blockade of the cell migration in a Boyden chamber in VEGFR-2 and VEGFR-3 dependence in the presence of inventive Compounds 9, 5 and I r respectively.

Fig. 4 is a diagram showing growth rate of xenotransplanted mouse fibrosarcoma cells. Mice were treated with vehicle only (control), or inventive Compound 9, Compound 5 or Compound 11. Tumor growth inhibition at the last day of the experiment with Compound 9: 35%, Com- pound 5 : 86% and Compound 11 : 59%.

Fig. 5 is a photo of representative examples of mice from the different treatment groups treated with either vehicle only or different compounds of the invention. From the left: Control, Compound 9, Compound 5, Compound 11.

Fig. 6 is a photo of excised tumours from the animals in Fig. 5. From the left: Control, Compound 9, Compound 5, Compound 11.

Fig. 7 is a diagram showing growth rate of xenotransplanted human lung carcinoma cells in mice. Mice were treated with vehicle only (control) or inventive Compound 10. Tumor growth inhibition at the last day of the experiment was 67%.

Fig. 8 contains photos of A. representative examples of immunodeficient mice having received human lung carcinoma cells and treated with either vehicle only or inventive Compound 10; B. the same mice where the tumor area has been freed from skin; and C. the excised tumors from the same mice. Fig. 9 is a bar chart representing choroidal neovascularization area (CNV) induced in mice by laser photocoagulation, in mice treated with vehicle only (100%) and in mice treated with inventive Compound 10.

Detailed description of the invention

The present invention relates to novel quinoline-3-carboxylic acid ester derivatives which inhibit, regulate and/or modulate growth factor and tyrosine kinase signal transduction, compositions which contain these derivatives, and methods of using them to treat and/or prevent growth factor and tyrosine kinase-dependent diseases and conditions such as cancer, diabetic retinopa- thy, age-related macular degeneration, chronic inflammation, stroke, ischemic myocardium, atherosclerosis, tumor growth, macular edema, and the like in mammals.

Consequently, according to one aspect, the present invention provides a compound of formula (I)

as generally defined herein above.

Any alkyl group in a compound of formula (I) may generally be selected from cyclic, branched or linear alkyl groups, and may be selected from C1-C6 alkyl groups, e.g. from methyl, ethyl, n- propyl, iso-propyl, cyclopropyl, n-butyl, iso-butyl, tert-butyl, cyclobutyl, n-pentyl, 2- methylbutyl, 2,2-dimethylpropyl, cyclopentyl, hexyl, 2-methylpentyl, 3-methylpentyl, 2,2- dimethylbutyl, 2,3-dimethylbutyl, cyclohexyl etc.

Any halogen in a compound according to the invention may generally be selected from F, Cl, Br and I; in particular F, Cl and Br; or F and Cl.

In one embodiment of the invention, R² is selected from C1-C4 alkyl, or C1-C3 alkyl, e.g. methyl, ethyl and propyl, in particular ethyl. In one embodiment, n in formula (I) is 0-5, 0-4, 0-3, or 0-2, e.g. 0 or 1. In one particular embodiment, n in formula (I) is 0. In another embodiment, n is 1.

The phenyl ring in a compound of formula (I) is substituted with m radicals R³, whereby m is 0- 5, e.g. 0-4, 0-3, or 0-2, e.g. 1-2.

Thus, in one embodiment, the phenyl ring is substituted with one or several radials R , independently selected from halogen, NC-(CH₂)p-, R⁴-, R⁴O-(CH₂)p-, R⁴S-(CH₂)p-, R⁴S(O)₂-(CH₂)p-, R⁴R⁵N-(CH₂)P-, R⁴R⁵NC(O)-(CH₂)P-, (R⁴C(O))(R⁵)N-(CH₂)p-, R⁴C(O)-(CH₂)p-, R⁴OC(O)- (CH₂)P-, R⁴C(O)O-(CH₂)P-; and any two R³ may together form a moiety -O-(CR⁶R⁷)_k-O-; wherein k is 1 or 2, preferably 1; p is 0-3; or 0-2, or 1 or 0, preferably 0; R⁴ and R⁵ are independently selected from H and C1-C6 alkyl; e.g. from H and C1-C5 alkyl or from H and C1-C4 alkyl; and R⁶ and R⁷ are independently selected from H and halogen.

In one embodiment, the phenyl ring in a compound of formula (I) is substituted with one or several radials R³, independently selected from halogen, NC-(CH₂)p-, R⁴-, R⁴O-(CH₂)p-, R⁴R⁵N-(CH₂)P-, R⁴C(O)-(CH₂)P-, R⁴OC(O)-(CH₂)p-; and any two R³ may together form a moiety -O-(CR⁶R⁷)_k-O-; R⁴ and R⁵ are independently selected from H and C1-C6 alkyl; e.g. from H and C1-C5 alkyl or from H and C1-C4 alkyl; and R⁶ and R⁷ are independently selected from H and halogen.

In -O-(CR⁶R⁷)k-O-, each R⁶ and R⁷ is independently selected. For example, if k is 1, R⁶ may be H and R⁷ a halogen, e.g. F or Cl; or R⁶ both and R⁷ may be H or halogen, e.g F or Cl. Also, if k is 2, each R⁶ is independently selected and each R⁷ also is independently selected. Thus, for exam- pie, -0-(CR⁶R⁷VO- may be selected from , -0-CX₂-O-, -O-CHX-O-, -0-CH₂-O-, -O-CXY-O-, -O-(CX₂)₂-O-, -0-CHXCX₂-O-, -0-CH₂CX₂-O-, -0-(CXY)₂-O-, -0-CH₂CXY-O-, -0-(CHX)₂-O-, -O-CHX-CXY-O-, and -0-CX₂CXY-O-, wherein X and Y are different halogens, e.g. X is F and Y is Cl, or vice versa.

When two R³ taken together are -O-(CR⁶R⁷)k-O-, the oxygen atoms thereof suitably are bound to adjacent atoms of the phenyl ring.

In one embodiment, R³ is independently selected from C1-C6 alkyl, e.g. C1-C4 alkyl, i.e. methyl, ethyl, n-propyl, iso-propyl, cyclo-propyl, n-butyl, iso-butyl, tert-butyl, cyclo-butyl; Cl- C6 alkyloxy, e.g. C1-C4 alkyloxy, or C1-C3 alkyloxy, e.g. methoxy, ethoxy or propoxy, in particular methoxy; CF₃, OH, cyano, (C1-C6 alkyl)₂N-, e.g. (C1-C4 alkyl)₂N-, or (C1-C3 alkyl)₂N-, e.g. dimethylamino; C1-C6 alkylNH-, e.g. C1-C4 alkylNH-, or C1-C3 alkylNH-, C1-C6 alkyl- C(O)-, e.g. C1-C4 alkyl-C(O)-, such as C1-C3 alkyl-C(O)-, e.g. CH₃C(O)-; C1-C6 alkyl-OC(O)-, e.g. C1-C4 alkyl-OC(O)-, such as C1-C3 alkyl-OC(O)-, e.g. CH₃C(O)-; -C(O)OH, halogen, e.g. F or Cl; or two R³ may together form -O-(CR⁶R⁷)_k-O-, wherein k is 1 or 2, e.g. 1, and R⁶ and R⁷ are independently selected from H and halogen.

In one embodiment of the invention, any R³ is independently selected from methyl, ethyl, iso- propyl, tert-butyl, methoxy, CF₃, OH, cyano, F, Cl, OH, (CH₃)₂N-, -C(O)OH, CH₃C(O)-, CH₃CH₂OC(O)-, or two R³ may together form methylenedioxy or difluoromethylenedioxy.

Any radical R³ may be in ortho, meta or para position, in relation to the -NH-[CH]₂- linking the phenyl ring to the quinoline moiety and, in case more than one R is present on the phenyl ring, these may be in any position on the phenyl ring. E.g. in case m is 2, any two R may be in or- tho,ortho'; ortho,meta'; ortho,para; ortho,meta; meta,meta'; or meta,para position.

In one embodiment, at least one R³ is in para position, e.g. m is 1 and R³ is in para position, or m is 2 and one R³ is in para position while the other R³ is in any other position on the phenyl ring.

In one embodiment, the moiety

is selected from phenyl, metylphenyl, ethylphenyl, isopropylphenyl, tert-butylphenyl, hydroxy- phenyl, methoxyphenyl, dimethylaminophenyl, trifluoromethylphenyl, cyanophenyl, acetoxy- phenyl, chlorophenyl, fluorophenyl, difluorophenyl, dimethylphenyl, carboxyphenyl, difluoro- benzodioxolyl, ethoxycarbonylphenyl, (trifluoromethyl)(methyl)phenyl and benzodioxolyl. For example, the moiety may be selected from

wherein * denotes the point of attachment to the other moiety of the compound of formula (I).

In one embodiment of the invention, Z is O or S, whereby R¹ is selected from -OH, C1-C6 al- kylNH-, (C1-C6 alkyl)₂N-, and C1-C6 alkyl- S (O)₂NH-; wherein any C1-C6 alkyl is independently selected and e.g. is a C1-C4 alkyl, or C1-C3 alkyl, and wherein any alkyl carbon is optionally substituted with OH.

In one embodiment, R¹ is C 1 -C6 alkylNH- or (C 1 -C6 alkyl)₂N-.

In one embodiment, when R¹ is C1-C6 alkylNH- or (C1-C6 alkyl)₂N-, any C1-C6 alkyl is selected from methyl, ethyl, and propyl such as iso-propyl, optionally substituted with OH, such as hydroxyethyl.

In one embodiment, when R¹ is C1-C6 alkylNH- or (C1-C6 alkyl)₂N-, it is selected from CH₃NH-, (CHs)₂CHNH-, HO(CH₂CH₂)NH- and (CH₃)₂N-.

In another embodiment, R is -OH. In still another embodiment, R¹ is C1-C6 alkyl- S (O)₂NH-, wherein any C1-C6 alkyl is independently selected and e.g. is a C1-C4 alkyl, or C1-C3 alkyl.

In the embodiment where Z is O, formula (I) may be written as:

wherein R¹, R², R³, m and n are as defined herein above.

In one embodiment of the invention, in a compound of formula (I) Z is N forming together with

R¹ an unsaturated, aromatic or non-aromatic heterocycle, which heterocycle may optionally con- tain one or several further heteroatoms, e.g. 1-3 further heteroatoms, such as 1 or 2 further het- eroatoms, and which heterocycle may be substituted by one or several C1-C6 alkyl groups, e.g.

1-3 C1-C6 alkyl groups, such as 1 or 2 Cl-C6-alkyl groups. The further heteroatoms may be selected e.g. from N, O and S, or from N and O. The C1-C6 alkyl groups may be selected e.g. from C1-C4 alkyl, e.g. from C1-C3 alkyl, such as methyl or ethyl, e.g. methyl.

In one embodiment, the heterocycle contains 1 or 2 further heteroatoms, selected from N and O.

In one embodiment, the heterocyclic moiety formed by N and R¹ is 5-7-membered, e.g. 5-6- membered, or 5-membered. For example, the heterocyclic moiety may be selected from pyrrolyl, 2H-pyrrolyl, oxadiazolyl, thiadiazolyl, thiazolyl, oxazolyl, pyrazolyl, triazolyl, tetrazolyl, isoxa- zo IyI, isothiazolyl, imidazolyl, furazanyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, triazinyl, diazinyl, thiadiazinyl, oxadiazinyl, 2H-azepinyl, oxazepinyl, thiazepinyl etc.

In one embodiment, the heterocycle is 5-membered, unsaturated, aromatic or non-aromatic, con- tains 1 or 2 further heteroatoms selected from N and O and is optionally substituted with at least one C1-C6 alkyl, e.g. at least one C1-C4 alkyl, or at least one C1-C3 alkyl, such as at least one methyl. In one embodiment, the heterocycle is a 5-membered, unsaturated, aromatic or non-aromatic heterocycle containing 1 or 2 further heteroatoms selected from N and O and optionally is substituted with methyl.

In one embodiment, the heterocycle is aromatic.

In one embodiment, the heterocycle formed by N together with R may be represented by the formula (II)

(II) wherein X is O or S; the curbed line

linking X and N represents a 2-4-membered, saturated or unsaturated chain of covalently bound atoms selected from C and heteroatoms, e.g. N, O and S, e.g. N and O; and * denotes the point of attachment of the heterocycle to the other moiety of the compound of formula (I), which chain optionally is substituted by one or several C1-C6 alkyl radicals, e.g. C1-C4 alkyl radicals, or Cl- C3 alkyl radicals, e.g. methyl and ethyl.

In one embodiment, X in formula (II) is O.

In one embodiment, the curbed line linking X and N represents a 2-membered, saturated or unsaturated chain of covalently bound atoms selected from C and heteroatoms.

In one embodiment, the heterocycle formed by N together with R¹ is represented by the formula (III)

A-N (III) wherein X is O or S; A is CH₂, NH, CH or N; the curbed line linking X and A represents a 1-3-membered, saturated or unsaturated chain of covalently bound atoms selected from C and heteroatoms, e.g. N, O and S, or N and O, which chain optionally is substituted by one or several C1-C6 alkyl radicals, e.g. C1-C4 alkyl radicals, or C1-C3 alkyl radicals, e.g. methyl and ethyl; and * denotes the point of attachment to the other moiety of the compound of formula (I).

In one embodiment, X in formula (III) is O.

In one embodiment, A is selected from CH₂ and NH and the curbed line linking X and A repre- sents the moiety -CR⁸R⁹- , whereby formula (III) may be represented by

wherein R⁸ and R⁹ are independently selected from H and C1-C6 alkyl, e.g. C1-C4 alkyl, or Cl- C3 alkyl, e.g methyl and ethyl.

In another embodiment, A is selected from CH and N and the curbed line linking X and A represents the moiety =CR10-, whereby formula (III) may be represented by

wherein R¹⁰ is selected from H and C1-C6 alkyl, e.g. C1-C4 alkyl, or C1-C3 alkyl, e.g methyl and ethyl.

In one embodiment, any heteroatom in formula (III) is selected from N and O.

In one embodiment, the heterocycle of formula (II) or (III) is substituted by one or more C1-C6 alkyl groups.

In one embodiment, the heterocycle formed by N together with R¹ is a 5-membered, unsaturated, aromatic or non-aromatic heterocycle selected from

optionally substituted by at least one Cl-C6-alkyl, e.g. at least one C1-C4 alkyl, or C1-C3 alkyl, e.g. at least one methyl, wherein * denotes the point of attachment to the other moiety of the compound of formula (I).

In one embodiment, the 5-membered, unsaturated, aromatic or non-aromatic heterocycle is selected from

wherein R¹¹ is C1-C6 alkyl, e.g. C1-C4 alkyl, or C1-C3 alkyl, methyl or ethyl, in particular methyl; and * denotes the point of attachment to the other moiety of the compound of formula (I).

In one embodiment, the compound according to the invention is selected from 6-methylcarbamoyl-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-isopropylcarbamoyl-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-methanesulfonylaminocarbonyl-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-(2-hydroxy-ethylcarbamoyl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 4-p-tolylamino-quinoline-3,6-dicarboxylic acid 3-ethyl ester,

6-methylcarbamoyl-4-(4-methoxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-(4,5-dihydro-oxazol-2-yl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-(5-methyl-[l,3,4]oxadiazol-2-yl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(2,4-difluoro-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(2,4-dimethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(4-carboxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(3-acetyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(4-acetyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester,

6-methylcarbamoyl-4-(2,2-difluoro-benzo[l,3]dioxol-5-ylamino)-quinoline-3-carboxylic acid ethyl ester,

6-methylcarbamoyl-4-(2-methoxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(4-ethoxycarbonyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(4-dimethylamine-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(3 -trifluoromethyl-4-methyl-phenylamino)-quino line-3 -carboxylic acid ethyl ester,

6-methylcarbamoyl-4-(4-fluoro-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(4-chloro-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(4-hydroxy-phenylamino)-quino line-3 -carboxylic acid ethyl ester,

6-methylcarbamoyl-4-(4-trifluoromethyl-phenylamino)-quino line-3 -carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(4-nitril-phenylamino)-quinoline-3 -carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(benzo[l,3]dioxol-5-ylamino)-quinoline-3-carboxylic acid ethyl ester, 6-dimethylcarbamoyl-4-(4-methoxy-phenylamino)-quino line-3 -carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(benzylamino)-quino line-3 -carboxylic acid ethyl ester, 6-dimethylcarbamoyl-4-(benzylamino)-quino line-3 -carboxylic acid ethyl ester, 6-dimethylcarbamoyl-4-(4-ethyl-phenylamino)-quino line-3 -carboxylic acid ethyl ester, 6-dimethylcarbamoyl-4-(4-isopropyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-dimethylcarbamoyl-4-(4-tert.butyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-dimethylcarbamoyl-4-(2,3-dimethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester,

6-dimethylcarbamoyl-4-(2,6-dimethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester and 6-dimethylcarbamoyl-4-(2,5-dimethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester, or a pharmaceutically acceptable salt of any of these.

The preparation of the novel quino line-3 -carboxylic acid esters of the present invention lies well within the capability of the person skilled in the art.

As an example, a quino line-3 -carboxylic acid ester derivative of the invention, may be formed in a four step procedure wherein, first, a suitable aniline derivative (a) is reacted with a malonic acid derivative (b), the formed intermediate (c) is cyclized to give a quinoline-4-ol derivative (d), which is then converted to the corresponding halogen derivative (e) (wherein X is halogen), using a suitable halogenating agent, e.g. POCI₃, which derivative (e) is finally reacted with a suitable amine (f) to form the selected quino line-3 -carboxylic acid ester derivative of formula (I) according to the invention, as illustrated by the following Reaction Scheme 1 : Reaction Scheme 1

halogenating agent

(e) (I)

With regard to the above reaction sequence, it is well within the capability of the person skilled in the art to select suitable reaction components as well as reaction conditions. As an example, when X is Cl, a suitable halogenating agent is e.g. POCI3.

In addition to the general method the inventors of the present invention also developed a useful high throughput palladium mediated synthesis method which allows the synthesis of the inventive compounds of formula (I) wherein Z is O. In one advantageous embodiment of this latter method, at least one of the reaction steps is performed under microwave irradiation. In a preferred embodiment, all the reaction steps of the palladium mediated synthesis method are performed under under microwave irradiation.

In the first steps of the palladium mediated synthesis method, a p-halogenoaniline (g) (wherein Y is halogen), e.g. p-bromoaniline, is reacted with a malonic acid derivative (b), the formed intermediate (h) is cyclized and reacted with a suitable halogenating agent, such as e.g. POCI3, to give a 4-halogenoquinoline derivative (i) (i.e. X is halogen).

In the final reaction step of the method, compound (j) is reacted with a nucleophilic compound corresponding to the moiety R¹ (denoted as R¹H) in a suitable solvent in the presence of Herrmann's palladacycle (trans-Di(μ-acetato)-bis[o-(di-o-tolylphosphino)benzyl]- dipalladium(II)); [(J-Bu)₃PH]BF₄; Mo(CO)₆; and DBU (l,8-Diazabicyclo[5.4.0]undec-7-ene) to provide a compound of formula (I) according to the invention wherein Z is O.

Reaction Scheme 2

U) (I)

The compounds according to formula (I) will be useful for treating various diseases such as cancer, diabetic retinopathy, macular degeneration-related disorders, chronic inflammation, stroke, ischemic myocardium, atherosclerosis, tumor growth, macular edema. The treatment may be preventive, palliative or curative.

In one particular embodiment, the inventive compounds are for use in the treatment of a disorder selected from cancer and tumor growth.

Examples of cancers that may suitably be treated by administration of the inventive compounds are bladder cancer, breast cancer, colorectal cancer, gastric cancer, head and neck cancer, kidney cancer, liver cancer, lung cancer, ovarian cancer, pancreas/gall bladder cancer, prostate cancer, thyroid cancer, osteosarcoma, rhabdomyosarcoma, MFH/fϊbrosarcoma, melanoma, glioblastomas/astrocytomas, or mesothelioma. Thus, for example, the compounds of the invention are useful in the treatment of a variety of cancers, including, but not limited to, carcinoma, including that of the bladder, breast, colon, kidney, liver, lung, including small cell lung cancer, esophagus, gall bladder, ovary, pancreas, stomach, cervix, thyroid, prostate, and skin, including squamous cell carcinoma; hematopoietic tumors of lymphoid lineage, including leukemia, acute lymphocytic leukemia, acute lymphoblastic leukemia, B-cell lymphoma, T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkins lymphoma, hairy cell lymphoma and Burkett's lymphoma; hematopoietic tumors of myeloid lineage, including acute and chronic myelogenous leukemias, myelodysplastic syndrome and promyelo- cytic leukemia; tumors of mesenchymal origin, including fibrosarcoma and rhabdomyosarcoma; tumors of the central and peripheral nervous system, including astrocytoma, neuroblastoma, glioma and schwannomas; and other tumors, including melanoma, seminoma, teratocarcinoma, osteosarcoma, xenoderoma pigmentosum, keratoctanthoma, thyroid follicular cancer and Kaposi's sarcoma.

In another embodiment, the compounds are for use in a macular degeneration-related disorder. The term "macular degeneration-related disorder" refers to any of a number of conditions in which the retinal macula degenerates or becomes dysfunctional, e.g. age-related macular degeneration, North Carolina macular dystrophy, Sorsby's fundus dystrophy, Stargardt's disease, pattern dystrophy, Best disease, dominant drusen, and Malattia Leventinese (radial drusen). The term also encompasses extramacular changes that occur prior to, or following dysfunction and/or degeneration of the macula. Thus, the term "macular degeneration-related disorder" also broadly includes any condition which alters or damages the integrity or function of the macula (e.g., damage to the RPE or Bruch's membrane).

In one embodiment, the macular degeneration-related disorder is age-related macular degeneration.

Examples of pharmaceutically acceptable addition salts for use in the pharmaceutical compositions of the present invention include those derived from mineral acids, such as hydrochlorid, hydrobromic, phosphoric, metaphosphoric, nitric and sulphuric acids, and organic acids, such as tartaric, acetic, citric, malic, lactic, fumaric, benzoic, glycolic, gluconic, succinic, and arylsul- phonic acids. The pharmaceutically acceptable excipients described herein, for example, vehicles, adjuvants, carriers or diluents, are well-known to those who are skilled in the art and are readily available to the public. The pharmaceutically acceptable carrier may be one that is chemically inert to the active compounds and that has no detrimental side effects or toxicity under the conditions of use. Pharmaceutical formulations are found e.g. in Remington: The Science and Practice of Pharmacy. A. R. Gennaro, Editor. Lippincott, Williams and Wilkins, 20th edition (2000).

Prodrugs of the compounds of formula (I) may be prepared by modifying functional groups present on the compound in such a way that the modifications are cleaved, in vivo when such prodrug is administered to a mammalian subject. The modifications typically are achieved by synthesizing the parent compound with a prodrug substituent. Prodrugs include compounds of formula (I) wherein a hydroxy, amino, sulfhydryl, carboxy or carbonyl group in a compound of formula (I) is bonded to any group that may be cleaved in vivo to regenerate the free hydroxyl, amino, or sulfhydryl group, respectively. Examples of prodrugs include, but are not limited to, esters and carbamates of hydroxy functional groups, esters groups of carboxyl functional groups, N-acyl derivatives and N-Mannich bases. General information on prodrugs may be found e.g. in Bunde- gaard, H. "Design of Prodrugs" pl-92, Elesevier, New York-Oxford (1985).

The composition according to the invention may be prepared for any route of administration, e.g. oral, intravenous, cutaneous or subcutaneous, nasal, intramuscular, or intraperitoneal. The precise nature of the carrier or other material will depend on the route of administration. For a par- enteral administration, a parenterally acceptable aqueous solution is employed, which is pyrogen free and has requisite pH, isotonicity and stability. Those skilled in the art are well able to prepare suitable solutions and numerous methods are described in the literature. A brief review of methods of drug delivery is also found in e.g. (43).

The dose administered to a mammal, particularly a human, in the context of the present invention should be sufficient to effect a therapeutic response in the mammal over a reasonable time frame. One skilled in the art will recognize that dosage will depend upon a variety of factors including the potency of the specific compound, the age, condition and body weight of the patient, as well as the stage/severity of the disease. The dose will also be determined by the route (administration form) timing and frequency of administration. In the case of oral administration the dosage can vary from about 0.01 mg to about 1000 mg per day of a compound of formula (I) or the corresponding amount of a pharmaceutically acceptable salt thereof. The compounds of the present invention may be used or administered in combination with one or more additional pharmaceutically active ingredients or drugs, e.g. drugs useful in the treatment of hyperproliferative diseases, e.g. a cytostatic or cytotoxic agent (i.e. anti-tumor agents). The components may be in the same formulation or in separate formulations for administration si- multaneously or sequentially. The compounds of the present invention may also be used or administered in combination with other treatment such as irradiation for the treatment of cancer.

Examples of cytostatic or cytotoxic agents, are DNA interactive agents, such as cisplatin or doxorubicin; topoisomerase II inhibitors, such as etoposide; topoisomerase I inhibitors such as CPT-11 or topotecan; tubulin interacting agents, such as paclitaxel, docetaxel or the epothilones, hormonal agents, such as tamoxifen; thymidilate synthase inhibitors, such as 5-fluorouracil; and anti-metabolites, such as methotrexate, other tyrosine kinase inhibitors such as Iressa and OSI- 774; angiogenesis inhibitors; EGF inhibitors; VEGF inhibitors; CDK inhibitors; SRC inhibitors; c-Kit inhibitors; Her 1/2 inhibitors and monoclonal antibodies directed against growth factor re- ceptors such as erbitux (EGF) and herceptin (Her2).

The invention is further illustrated by the following, non-limiting examples.

Examples Example 1

Intermediary Compound 1. 2-[(4-Bromo-phenylamino)-methylene]-malonic acid diethyl ester

A 20 mL microwave vial was charged with 4-bromoaniline (6.881 g, 40.0 mmol), diethyl eth- oxymethylenemalonate (8.650 g, 40.0 mmol) and toluene (5 mL). The vial was capped and the mixture was microwave heated at 150⁰C for 30 min. After cooling, the solution was poured onto 50 ml of vigorously stirred isohexane. A thick, white precipitate formed and the suspension was stirred for another 15 min. The suspension was filtered and the product washed with 20 mL of isohexane. The product was dried under vacuum to give 11.678 g (85%) of the title compound. MS (ESI⁺) m/z 342, 344 (MH⁺) Example 2

Intermediary Compound 2. 6-Bromo-4-chloroquinoline-3-carboxylic acid ethyl ester

A 20 niL microwave vial was charged with 2-[(4-Bromo-phenylamino)-methylene]-malonic acid diethyl ester (1.711 g, 5.0 mmol) and phosphoryl chloride (10.0 mL, 16.8 g, 109 mmol. The vial was capped and the mixture was microwave heated stepwise up to 180⁰C (watching the pressure) over 5 min and then kept at 180⁰C for 30 min. Excess POCI3 was evaporated and the residue partitioned between CH₂Cl₂ (40 mL) and 2 M NaOH (aq) (40 mL). The aqueous layer was ex- tracted with CH₂Cl₂ (2x40 mL). The organic layers were combined, dried with Na₂CO₃ and evaporated. The residue was purified by silica flash chromatography using CH₂Cl₂ as eluent. Pure fractions were combined, evaporated and the residue dried under vacuum to give 0.821 g (52%) of the title compound. MS (ESI⁺) m/z 314, 316 (MH⁺)

Example 3

Intermediary Compound 3. 6-Bromo-4-p-tolyl-amino-quinoline-3-carboxylic acid ethyl ester

A 20 mL microwave vial was charged with 6-Bromo-4-chloro-quinoline-3-carboxylic acid ethyl ester (0.786 g, 2.50 mmol), p-toluidine (0.268 g, 2.50 mmol) and dry 1,4-dioxane (15 mL). The vial was capped and the mixture was microwave heated at 150⁰C for 30 min. After cooling, a yellow precipitate had formed. The suspension was poured onto 2 M NaOH (aq) (100 mL) and the aqueous layer was extracted with CH₂Cl₂ (3x80 ml). The organic layers were combined and washed with H₂O (100 mL), dried with MgSO₄ and evaporated. The residue was purified by a short silica column using isohexane/EtOAc (1 :1) as eluent. Pure fractions were combined, evaporated and the residue was dried under vacuum to give 0.748 g (78%) of the title compound. MS (ESI⁺) m/z 385, 387 (MH⁺) Example 4

Intermediary Compound 4. 6-Bromo-4-(4-methoxyphenylamino)-quinoline-3-carboxylic acid ethyl ester

A 20 niL microwave vial was charged with 6-Bromo-4-chloro-quinoline-3-carboxylic acid ethyl ester (0.157 g, 0.500 mmol), p-anisidine (61.6 mg, 0.500 mmol) and dry 1,4-dioxane (5 rnL). The vial was capped and the mixture was microwave heated at 150⁰C for 30 min. Triethylamine (0.10 mL) was added and the mixture evaporated, then slurried in CH₂C₂ and TEAHCl filtered off. The remainder was purified by silica flash chromatography using 1% MeOH in CH₂Cl₂ as eluent. Pure fractions were combined, evaporated and the residue dried under vacuum to give 0.153 g (76%) of the title compound. MS (ESI⁺) m/z 402 (MH⁺)

¹H NMR (400 MHz, CDCl₃): 10.49 (s, IH), 9.21 (s, IH), 7.80 (d, IH), 7.73 (s, IH), 7.64 (d, IH), 7.13, (d, 2H), 6.96 (d, 2H), 4.42 (q, 2H), 2.36 (s, 3H), 1.44 (t, 3H).

Compounds of the invention wherein, in formula (I), Z is O, also were synthesized by a microwave assisted process, the last step of which was carried out according to the procedure outlined in relation to Reaction Scheme 2 herein above, and as generally described herein below:

A microwave vial was charged with compound (10) (1 eq.), Herrmann's palladacycle (trans- Di(μ-acetato)-bis[o-(di-o-tolylphosphino)benzyl]dipalladium(II)) (0.05 eq.), [(J-Bu)₃PH]BF₄ (0.2 eq.), Mo(CO)₆ (2 eq.), a nucleophilic compound corresponding to R¹, (suitably in excess, e.g. 1.5-10 eq.) and dry THF. Finally, DBU (l,8-Diazabicyclo[5.4.0]undec-7-ene) (3 eq.) was added and the vial was immediately capped with a Teflon septum and irradiated with microwaves for suitable time, e.g. 5 min at 130⁰C. Volatiles are removed under reduced pressure and the product was purified.

Example 5

Compound 5. 6-Methylcarbamoyl-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester

A 2-mL microwave vial was charged with 6-Bromo-4-p-tolyl-amino-quinoline-3-carboxylic acid ethyl ester (38.5 mg, 0.100 mmol), Herrmann's palladacycle (tra/?s-Di(μ-acetato)-bis[o-(di-o- tolylphosphino)benzyl]dipalladium(II), 4.7 mg, 0.0050 mmol), [OBu)₃PH]BF₄ (5.9 mg, 0.020 mmol), Mo(CO)₆ (52.8 mg, 0.20 mmol), methylamine (0.150 mmol, 2 M in THF) and dry THF (1.0 mL). Finally, DBU (l,8-Diazabicyclo[5.4.0]undec-7-ene, 0.045 μL, 0.30 mmol) was added and the vial was immediately capped with a Teflon septum and irradiated with microwaves for 5 min at 130⁰C. Volatiles were removed under reduced pressure and the product was purified by silica flash chromatography using EtOAc as eluent. Yield: 70%

MS (ESI⁺) m/z 364 (MH⁺)

¹H NMR (400 MHz, CDCI₃/CD₃OD 1 :1): 9.12 (s, IH), 8.04 (s, IH), 7.93 (d, IH), 7.85 (d, IH), 7.15, (d, 2H), 7.00 (d, 2H), 4.39 (q, 2H), 2.79 (s, 3H), 2.34 (s, 3H), 1.41 (t, 3H).

Example 6

Compound 6. 6-Isopropylcarbamoyl-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester

The compound was synthesized in the same way as Compound 5, but using 5 equiv. of isopro- pylamine instead of methylamine. The product was purified by silica flash chromatography using EtOAc as eluent. Yield: 53%, MS (ESI⁺) m/z 392 (MH⁺) Example 7

Compound 7. 6-Methanesulfonylaminocarbonyl-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester

The compound was synthesized in the same way as Compound 5, but using 5 equiv. of methane- sulfonamide instead of methylamine. The product was purified by silica flash chromatography using 5% MeOH in CH₂Cl₂ as eluent.

Yield: 71%

MS (ESI⁺) m/z 428 (MH⁺)

Example 8

Compound 8. 6-(2-Hydroxy-ethylcarbamoyl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester

The compound was synthesized in the same way as Compound 5, but using 5 equiv. of etha- nolamine instead of methylamine. Product purified by silica flash chromatography using 2.5-5% methanol in CH₂Cl₂ as eluent.

Yield: 73%

MS (ESI⁺) m/z 394 (MH⁺)

Example 9

Compound 9. 4-p-Tolylamino-quinoline-3,6-dicarboxylic acid 3-ethyl ester

The compound was synthesized in the same way as Compound 5, but using 10 equiv. of water instead of methylamine. The product was purified by preparative RP-LCMS using 10-50% ace- tonitrile gradient in H₂O (0.05% HCOOH).

Yield: 23%

MS (ESI⁺) m/z 351 (MH⁺)

¹H NMR (400 MHz, CDCI₃/CD₃OD 1 :1): 9.15 (s, IH), 8.41 (s, IH), 8.14 (d, IH), 7.86 (d, IH),

7.12, (d, 2H), 7.00 (d, 2H), 4.40 (q, 2H), 2.31 (s, 3H), 1.41 (t, 3H).

Example 10

Compound 10. 6-Methylcarbamoyl-4-(4-methoxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester

A 2-mL microwave vial was charged with 6-Bromo-4-(4-methoxyphenylamino)-quinoline-3- carboxylic acid ethyl ester (40.1 mg, 0.100 mmol), Herrmann's palladacycle (trans-Di(μ- acetato)-bis[o-(di-o-tolylphosphino)benzyl]dipalladium(II), 4.7 mg, 0.0050 mmol), [(t- Bu)₃PH]BF₄ (5.9 mg, 0.020 mmol), Mo(CO)₆ (52.8 mg, 0.20 mmol), methylamine (0.150 mmol, 2 M in THF) and dry THF (1.0 mL). Finally, DBU (l,8-Diazabicyclo[5.4.0]undec-7-ene, 0.045 μL, 0.30 mmol) was added and the vial was immediately capped with a Teflon septum and irradiated with microwaves for 5 min at 130⁰C. Volatiles were removed under reduced pressure and the product was purified by silica flash chromatography using 2.5% methanol in CH₂Cl₂ as elu- ent. Yield: 83%, MS (ESI⁺) m/z 380 (MH⁺) Example 11

Compound 11. 6-(4,5-Dihydro-oxazol-2-yl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester

6-(2-Hydroxy-ethylcarbamoyl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester (10.0 mg, 0.0254 mmol), CHCl₃ (0.5 mL) and SOCl₂ (3.0 μL, 1.6 equiv.) were charged to a 2 mL microwave vial. The mixture was heated at 100⁰C for 10 min. The resulting suspension was dissolved in an additional 5 mL Of CHCl₃ and loaded onto a small silica column. The yellow product was eluted with 10% MeOH in CH₂Cl₂ to give 8.0 mg of 6-(2-Chloro-ethylcarbamoyl)-4-p- tolylamino-quinoline-3-carboxylic acid ethyl ester of which 1.2 mg was collected for biological testing. The remainder was dissolved in 2 mL Of CH₂Cl₂ and added to a 5 mL microwave vial together with 2 mL of saturated NaHCO₃ (aq) and the mixture was heated at 100⁰C for 90 min. The organic layer was dried and evaporated to give 4.7 mg (49%) of the title compound. MS (ESI⁺) m/z 376 (MH⁺) 1H NMR (400 MHz, CDC1₃/CD₃OD 1 :1): 9.21 (s, IH), 8.13 (s, IH), 8.09 (d, IH), 7.84 (d, IH), 7.13, (d, 2H), 6.99 (d, 2H), 4.39 (q, 2H), 4.26 (t, 2H), 3.93 (t, 2H), 2.33 (s, 3H), 1.41 (t, 3H).

Example 12

Compound 12. 6-(5-Methyl-[l,3,4]oxadiazol-2-yl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester

4-p-Tolylamino-quinoline-3,6-dicarboxylic acid 3-ethyl ester (4.0 mg, 0.011 mmol) and acetic hydrazide (4.0 mg, 0.054 mmol) was heated in 1 mL POCl₃ at 100⁰C for 7 h. Excess POCl₃ was evaporated and the residue was purified by silica flash chromatography using 1-3% MeOH in CH₂Cl₂ to give 3.4 mg of the title compound. Yield: 77% MS (ESI⁺) m/z 389 (MH⁺) The following compounds were produced according to the methods described above:

Compound 13. 6-Methylcarbamoyl-4-(2,4-difluoro-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 386.4 (MH⁺), Purity>95%

Compound 14. 6-Methylcarbamoyl-4-(2,4-dimethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 378.4 (MH⁺), Purity>95%

Compound 15. 6-Methylcarbamoyl-4-(4-carboxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 394.4 (MH⁺), Purity>95% Compound 16. 6-Methylcarbamoyl-4-(3-acetyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 392.4 (MH⁺), Purity>95%

Compound 17. 6-Methylcarbamoyl-4-(4-acetyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 392.4 (MH⁺), Purity>95%

Compound 18. 6-Methylcarbamoyl-4-(2,2-Difluoro-benzo[l,3]dioxol-5-ylamino)-quinoline- 3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 430.4 (MH⁺), Purity>95% Compound 19. 6-Methylcarbamoyl-4-(2-methoxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 380.4 (MH⁺), Purity>95%

Compound 20. 6-Methylcarbamoyl-4-(4-ethoxycarbonyl-phenylamino)-quinoline-3- carboxylic acid ethyl ester

MS (ESI⁺) m/z MlA (MH⁺), Purity>95%

Compound 21. 6-Methylcarbamoyl-4-(4-dimethylamine-phenylamino)-quinoline-3- carboxylic acid ethyl ester

MS (ESI⁺) m/z 393.4 (MH⁺), Purity>95% Compound 22. 6-Methylcarbamoyl-4-(3-trifluoromethyl-4-methyl-phenylamino)-quinoline- 3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 432.4 (MH⁺), Purity>95%

Compound 23. 6-Methylcarbamoyl-4-(4-fluoro-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 368.4 (MH⁺), Purity>95%

Compound 24. 6-Methylcarbamoyl-4-(4-chloro-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 384.7 (MH⁺), Purity>95%

Compound 25. 6-Methylcarbamoyl-4-(4-hydroxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 366.4(MH⁺), Purity>95%

Compound 26. 6-Methylcarbamoyl-4-(4-trifluoromethyl-phenylamino)-quinoline-3- carboxylic acid ethyl ester

MS (ESI⁺) m/z 418.4(MH⁺), Purity>95%

Compound 27. 6-Methylcarbamoyl-4-(4-cyano-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 375.4(MH⁺), Purity>95%

Compound 28. 6-Methylcarbamoyl-4-(benzo[l,3]dioxol-5-ylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 394.4(MH⁺), Purity>95% Compound 29. 6-Dimethylcarbamoyl-4-(4-methoxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 394.4(MH⁺), Purity>95%

Compound 30. 6-Methylcarbamoyl-4-(benzylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 364.4(MH⁺), Purity>95%

Compound 31. 6-Dimethylcarbamoyl-4-(benzylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 378.4(MH⁺), Purity>95% Compound 32. 6-Dimethylcarbamoyl-4-(4-ethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 378.1(MH⁺), Purity>95%

Compound 33. 6-Dimethylcarbamoyl-4-(4-isopropyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 392.1(MH⁺), Purity>95%

Compound 34. 6-Dimethylcarbamoyl-4-(4-tert.butyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 406.1 (MH⁺), Purity>95% Compound 35. 6-Dimethylcarbamoyl-4-(2,3-dimethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 378.1 (MH⁺), Purity>95%

Compound 36. 6-Dimethylcarbamoyl-4-(2,6-dimethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 378.1 (MH⁺), Purity>95%

Compound 37. 6-Dimethylcarbamoyl-4-(2,5-dimethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester

MS (ESI⁺) m/z 378.1 (MH⁺), Purity>95%

Biology Cells

Stable porcine aortic endothelial (PAE) cell lines expressing VEGFR-2 or VEGFR-3, respectively, were established as reported (14, 24) and maintained in Ham's F12 medium supplemented with glutamine, penicillin/streptomycin (PEST) and 10% fetal bovine serum (FBS).

For experiments on PAE cells, the wildtype porcine aortic endothelial cell line was cultivated. Mouse fibrosarcoma cell line T241 was used to prepare conditioned medium contained recombinant human VEGF (cm/VEGF). The T241 cells were grown in DMEM medium supplemented with 5% FBS for 72 hours.

Cell shape assay

PAE/VEGFR-2 cells were grown in 48-well plates to about 50% confluence in Ham's F12 medium supplemented with Glutamine, PEST, 10%FBS. The medium was removed and replaced with fresh Ham's F 12 medium containing 10% FBS and minimal concentration of cm/VEGF that activated cell shape change, with or without test compounds in a final concentration of 10 μM/well. After 12 h of incubation, cells were fixed with 3% paraformaldehyde in phosphate buffer (pH 7.5) for 30 min at room temperature and rinsed three times with PBS. The cells were examined under light microscope (Fig. 2).

Actin staining PAE/VEGFR-2 cells were grown on covers lips in 6-well plates to about 50-70% confluence in Ham's F12 medium supplemented with 10% FBS. The medium was removed and replaced with fresh Ham's F 12 medium containing 10% FBS and minimal concentration of cm/VEGF with or without test compounds. After 12 h of incubation, cells were fixed with 3% paraformaldehyde in phosphate buffer (pH 7.5) for 30 min at room temperature. After rinsing three times with PBS, the cells were permeabilized with 0.5% Triton X-100 for 30 min. The cells then were washed three times with PBS and stained for 30 min at room temperature with 1 mg/ml of rhodamine- phalloidin (Sigma) in PBS. After washing with PBS five times, the coverslips were mounted in a mixture of glycerol and PBS (90:10), and the cells were examined under light and fluorescence microscopes (Fig 1).

Chemotaxis assay

The motility response of PAE/VEGFR-2 cells and PAE/VEGFR-3 cells to VEGF and test compounds were assayed by a modified Boyden chamber technique (39) as described (14, 40), by using micropore polycarbonate filters (PVPF, 8 micron) coated with 1% gelatine solution. Cells were trypsinized and resuspended at a concentration of 6xlO⁵ cells/ml in serum- free medium. The cells (30,000 cells per well) were placed in the upper chamber with or without test compounds in a final concentration of 10 μM/well and serum- free medium with or without VEGF in the lower chamber. After 6 h at 37°C, the medium was removed and cells attached to the filter were fixed in methanol and stained with Giemsa solution. The cells migrating through the filter were counted and plotted as number of migrating cells per optic field (x32) or were detected by ImageJ software and plotted as integrated density of migrated cells (Fig. 3). All experiments were performed in triplicate.

Kinase binding assay

Testing for tyrosine kinase activity was performed using Cerep Kinase Profiling Service (www.cerep.com) . The kinases used were recombinant human proteins expressed in bacteria or in insect cells. They were assayed with appropriate biotinylated substrates in the presence of 0.05-20 μM ATP (0.3 to 3 times the Km of the individual kinase) at 22 ⁰C for 15 to 90 min, ac- cording to the kinase. Kinase activity was detected by an HTRF assay. HTRF (a trademark of CisBio International) is based on fluorescence transfer, using Europium (Eu³⁺) cryptate and XL665 as donor and acceptor, respectively. When the biotinylated (or tagged) substrate is phos- phorylated by the kinase, it cross-reacts with a cryptate-labeled phosphospecific antibody. Addition of Streptavidin-XL665, SA-XL665, (or anti-tag antibody XL665) causes the juxtaposition of the cryptate and XL665 fluorophore, resulting in FRET (fluorescence resonance energy transfer). FRET intensity depends on the amount of bounded cryptate antibody, which is proportional to the extent of substrate phosphorylation (41).

In-vivo analysis Experiments using T241 tumor fibrosarcoma cells in mice

T241 tumor fibrosarcoma cells

T241 tumor fibrosarcoma cells were kindly provided by Dr Yihai Cao, KI, Stockholm, Sweden. The cells were grown in Dulbecco 's modified essential medium supplemented with fetal calf serum, glutamine, and penicillin/streptomycin. The cells were grown in humidified air (95%) and CO₂ (5%) at 37 ⁰C. The medium was changed twice a week and confluent cultures were subcul- tivated after treatment with trypsin. For s.c. tumor injections, a single cell suspension was prepared, where cells were resuspended in medium and the viability and cell concentration were calculated after addition of trypan blue dye. The cell suspension was kept on ice during the injection procedure. The culture was shown to be free from mycoplasm.

Animals

Female C57B1/6J mice were used for xenografting at the age of 6-7 weeks (body weight 18-20 g). The mice were housed in an isolated room at 24 ⁰C with a 12-h light, 12-h dark cycle. They were fed ad libitum with water and food pellets. The animal weight and general appearance were recorded every day throughout the experiment. The experiment was approved by regional ethics committee for animal research.

Xenografting Tumor cells (1 x 10⁶ in 0.1 ml medium) were implanted s.c. in the hind leg of the animal. Animals were anesthetized with isofluoran supplemented with oxygen. Tumor volume measurements began when the tumor became palpable (~ 0.1 ml) and were repeated every day using a calliper. The tumor volume was calculated by the formula: 0.52 x length x width².

Administration of drug

Treatment started at the same time as tumor implantation and all animals received treatment for 20 days. Control animals were given vehicle only. The test compounds were given at a dose of 25 mg/kg/day s.c. in the neck.

Perfusion fixation and autopsy

The animals were anesthetized by an ip injection of 25 mg/kg of avertin. A cannula was inserted in the thoracic aorta, and the animal was perfused with PBS/heparin. The thoracic and abdominal viscera were examined for macroscopic metastases. The true tumor weight and volume were recorded at autopsy and correlated well to the calculated volume.

Tissue analyses

The dissected tumors were immersion- fixed in 4% formaldehyde for approximately 2 days before dehydration and paraffin embedding. Sections were cut at 3 μm and put on slides, dewaxed, rehydrated and stained immunohistochemically.

Immunohistochemistry

To quantify angiogenesis (vessel growth), Bandeira Simplicifolia-1 lectin histochemistry was used for highlighting endothelial cells.

Stereological quantification

A representative section from the geometrical center of each tumor was used. Structure was counted at x400 with an eyepiece grid. The grid was placed at random at the upper left-hand corner of the section and systematically advanced every 1 to 3 mm, depending on tumor size, in both directions by use of the gonimeter stage of microscope. Vascular parameters from 25 to 35 grids were quantified from each tumor.

Experiments using human lung carcinoma cell xenografts in mice 30 million human lung carcinoma cells were implanted s.c. in the hind leg of immunodefϊcient mice (SCID), using a 23G needle. Tumor measurements began when the tumor became palpable (~ 0.1 ml) and were repeated every day. When tumors reached a volume of 200 mm the treatment began. The mice received oral treatment (50 mg/kg/day) with inventive compound for 17 days. Controls received vehicle only. Analyses, e.g. of angiogenesis and tumor volume, were performed as described herein above.

Experiments using laser induced eye model for macula degeneration

Choroidal neovascularization was induced by laser photocoagulation with an Argon laser (50 μm spot size; 0.05 s duration; 250 mW). Three laser spots were placed in each eye close to the optic nerve. Treatment started at the same day as laser treatment with 50 mg/kg/day of inventive compound, administered orally. 8 days after laser treatment, all animals were perfused with 1 ml of PBS containing 50 mg/ml fluorescein-labeled dextran (FITC-dextran; average molecular mass, 2 xlO⁶; Sigma- Aldrich) and sacrificed. The eyes were harvested and fixed in 10% phosphate- buffered formalin, and retinal pigment epithelium (RPE)-choroid-scleral flat mounts were pre- pared, stained red. The experiment was repeated 3 times. Controls received vehicle only.

Results

It is known that the mature form of VEGF binds to the angiogenic receptor VEGFR-2 and mediates VEGF-stimulated biological responses including membrane ruffling, cell migration and pro- liferation (24). In the presence of VEGF the VEGFR-2 expressing cells acquire an elongated cell shape with distinct spindle-like processes and actin reorganization (Fig. 1). The cell shape change appeared to be independent of the addition of bovine serum, because a similar change was observed in the presence and absence of serum. These data suggested that other serum factors were not required for the morphological effect (42).

Cell shape assay

One of the used assays comprised a culture of PAE/VEGFR-2 cells. Morphological changes of the cells were recorded microscopically after addition of the VEGF, followed by the test compound at final concentration up to lOOμM. Growth inhibitions of the PAE/VEGFR-2 cells were detected in the presence of a number of compounds according to the invention at 10 μM. These compounds were retested in primary screening at lower concentration (1, 2.5 and 5μM) as well as higher concentration.

Chemotaxis assay

In addition to above mentioned effects on endothelial cell shape the chemotactic effect of the test compounds on PAE/VEGFR-2 cells in a modified Boyden chamber assay was studied. The migration of the receptor expressing PAE cells through micropore polycarbonate filter toward VEGF as chemo-attractant was scored in the absence of serum. As shown in Fig. 3 several compounds could block the cell migration in both a VEGFR-2 and VEGFR-3 dependence. The assay gives a quantitative measure of the activity of the antagonist.

In Table 1 data from both the cell shape assay and the chemotaxis assay are shown. Thus, under "PAE/VEGFR-2 with VEGF Antagonist cone. (μM)" the concentration of the indicated inven- tive compound that gave restitution of cell morphology in the cell shape assay is shown. Data under "Chemotaxis VEGFR-3 % inhibition of cell migration" show the percentage inhibition of PAE cells expressing VEGFR-3 in the presence of 10 μM of the indicated inventive compound.

In-vitro Kinase inhibition

The IC50 values for a number of compounds from the kinase inhibition assays are indicated in Table 2.

In Table 3, the kinase inhibition (in % of control) in the presence of 10 μM of different inventive compounds is shown.

nt=not tested, - =no effect at 10 μM.

Table 4 shows IC50 values of Compound 10 tested on a kinase panel screen of 32 different kinases. It should be noted that the differences in IC50 between Tables 2 and 4 are due to the differences in assay arrangement between the two different CRO companies that have run the studies.

Table 4. Cont.

T241 tumor fibrosarcoma cells in mice

S. c. treatment with compounds 5, 9 and 11, respectively, suppressed all the tumor growth in mice having received the T241 fibrosarcoma cell line. Tumor growth inhibition at the last day of the experiment using Compound 9: 35%; Compound 5: 86%; and Compound 11 : 59% (Fig. 4). Neither toxicity nor metastases were seen and the weight increased/stayed the same throughout the experiment. The tumors from animals having received the inventive compounds were all smaller, paler and harder than the control (Figs. 5 and 6).

Human lung carcinoma cell xenografts in mice

The mean tumor inhibition after 17 days of treatment was 67% (Fig. 7) and the angiogenesis was inhibited by 43%. Tumors of some representative animals after 17 days of treatment are shown in Fig. 8.

Laser induced eye model for macula degeneration

As shown in Fig. 9, the CNV area was reduced by about 50% in the animals receiving Compound 10. References

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Claims

1. A compound of formula (I)

(I) wherein n is 0-5;

Z is O or S and R¹ is selected from -OH, C1-C6 alkylNH-, (C1-C6 alkyl)₂N-, and C1-C6 alkyl-

R² is Cl-C6 alkyl;

R³ _m represents m radicals R³, independently selected from halogen, NC-(CH₂)p-, R⁴-,

(R⁴C(O))(R⁵)N-(CH₂)p-, R⁴C(O)-(CH₂)P-, R⁴OC(O)-(CH₂)p-, R⁴C(O)O-(CH₂)p-; and when m > 2, two R³ may together form -O-(CR⁶R⁷)_k-O-; m is 0-5; k is 1 or 2; p is 0-3;

2. The compound according to claim 1, wherein R² is selected from C1-C3 alkyl.

3. The compound according to claim 1 or 2, wherein R is ethyl.

4. The compound according to any one of the claims 1-3, wherein m is 0-3.

5. The compound according to claim 4, wherein m is 0-2.

6. The compound according to any one of the claims 1-5, wherein n is 0-2.

7. The compound according to any claim 6, wherein n is 0 or 1.

8. The compound according to any one of the claims 1-7, wherein k is 1.

9. The compound according to any one of the claims 1-8, wherein p is 0 or 1.

10. The compound according to claim 9, wherein p is 0.

11. The compound according to any one of the claims 1-10, wherein each R .3 i •s independently selected from halogen, NC-(CH2)p-, R⁴-, R⁴O-(CH2)p-, R⁴R⁵N-(CH2)p-, R⁴C(O)-(CH2)p-, R⁴OC(O) -(CH2)p-; and when m > 2, two R³ may together form -O-(CR⁶R⁷)_k-O-, wherein R⁴, R⁵, R⁶, R⁷, p, m and k are as defined in any of the claims 1-10.

12. The compound according to any one of the claims 1-11, wherein R⁴ and R⁵ are independently selected from H and C1-C4 alkyl.

13. The compound according to any one of the claims 1-12, wherein each R³ is independently selected from C1-C4 alkyl, C1-C4 alkyloxy, CF₃, OH, cyano, (C1-C4 alkyl)₂N-, C1-C4 al- kylNH-, (C1-C4) alkyl-C(O)-, (C1-C4) alkyl-OC(O)-, -C(O)OH, halogen, or two R³ may together form methylenedioxy, optionally substituted by 1-2 halogens.

14. The compound according to claim 13, wherein each R³ is independently selected from methyl, ethyl, iso-propyl, tert-butyl, methoxy, CF₃, OH, cyano, F, Cl, OH, (CH₃ )₂N-, -C(O)OH, CH₃C(O)-, CH₃CH₂OC(O)-, or two R³ taken together are methylenedioxy or di- fluoromethy lenedio xy .

15. The compound according to any one of the claims 1-14, wherein Z is O.

16. The compound according to claim 15, wherein R¹ is selected from -OH, C1-C6 alkylNH-, (C1-C6 alkyl)₂N-, and C1-C6 alkyl- S (O)₂NH-; and any alkyl is optionally substituted with OH.

17. The compound according to claim 16, wherein R¹ is selected from C1-C6 alkylNH- or (Cl- C6 alkyl)₂N- wherein any alkyl is optionally substituted with OH.

18. The compound according to any one of the claims 1-14, wherein Z is N forming together with R¹ an unsaturated, aromatic or non-aromatic heterocycle optionally containing further het- eroatoms, which heterocycle is optionally substituted with at least one C1-C6 alkyl.

19. The compound according to claim 18, wherein in the heterocycle each further heteroatom is independently selected from N, O and S.

20. The compound according to claim 18 or 19, wherein the heterocycle is 5-7 membered.

21. A compound according to claim 1, selected from 6-methylcarbamoyl-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-isopropylcarbamoyl-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-methanesulfonylaminocarbonyl-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-(2-hydroxy-ethylcarbamoyl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 4-p-tolylamino-quinoline-3,6-dicarboxylic acid 3-ethyl ester,

6-methylcarbamoyl-4-(4-methoxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-(4,5-dihydro-oxazol-2-yl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-(5-methyl-[l,3,4]oxadiazol-2-yl)-4-p-tolylamino-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(2,4-difluoro-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(2,4-dimethyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(4-carboxy-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(3-acetyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(4-acetyl-phenylamino)-quinoline-3-carboxylic acid ethyl ester, 6-methylcarbamoyl-4-(2,2-difluoro-benzo[l ,3]dioxol-5-ylamino)-quinoline-3-carboxylic acid ethyl ester,

22. A compound according to any one of the claims 1-21, or a pharmaceutically acceptable salt thereof, for use as a medicament.

23. A compound according to claim 22, for use as a medicament having a VEGFR-2 tyrosine kinase inhibiting activity.

24. A compound according to any one of the claims 1-23, for the treatment of a disorder selected from cancer, diabetic retinopathy, macular degeneration-related disorder, chronic inflammation, stroke, ischemic myocardium, atherosclerosis, tumor growth, and macular edema.

25. A compound according to claim 24, for the treatment of a macular degeneration-related disorder selected from age-related macular degeneration, North Carolina macular dystrophy, Sorsby's fundus dystrophy, Stargardt's disease, pattern dystrophy, Best disease, dominant drusen, and Malattia Leventinese (radial drusen).

26. A compound according to claim 25, for the treatment of age-related macular degeneration.

27. A compound according to claim 24, for the treatment of cancer.

28. A compound according to any one of the claims 1-27, for the treatment of a disorder related to VEGF overexpression.

29. A pharmaceutical composition comprising a therapeutically effective amount of a compound according to any one of the claims 1-28, or a pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable excipient.

30. A pharmaceutical composition according to claim 29, comprising at least one further, pharmaceutically active compound.

31. A pharmaceutical composition according to claim 27, wherein the further, pharmaceutically active compound is an anti-tumor agent.

32. Use of a compound according to any one of the claims 1-20, or a pharmaceutically acceptable salt thereof, for the manufacturing of a medicament for the treatment of a disorder selected from cancer, diabetic retinopathy, macular degeneration-related disorder, chronic inflammation, stroke, ischemic myocardium, atherosclerosis, tumor growth, and macular edema.

33. A method of preparing a compound of formula (I) according to any one of the claims 1-20, comprising reacting an aniline derivative (a)

(a) with a malonic acid derivative (b)

(b) cyclizing the formed intermediate (c)

(C) so as to obtain quinoline-4-ol derivative (d)

reacting (d) with a halogenating agent so as to obtain (e)

wherein X is halogen; and reacting (e) with an amine (f)

so as to obtain a compound of formula (I).

34. A method of preparing a compound of formula (I) according to any one of the claims 15-17, comprising reacting a p-halogenoaniline (g), wherein Y is halogen,

(9) with a malonic acid derivative (b)

R'

(b) so as to obtain a compound (h)

(h) ; cyclizing and halogenating (h) by reacting with a suitable halogenating agent, so as to obtain a compound of formula (i), wherein X is halogen,

reacting (i) with an amine (f)

so as to obtain a compound (j)

and reacting (j) with R¹H in the presence of Herrmann's palladacycle (£rα/?s-Di(μ-acetato)-bis[o- (di-o-tolylphosphino)benzyl]-dipalladium(II)); [(J-Bu)₃PH]BF₄; Mo(CO)₆; and DBU (1,8- Diazabicyclo[5.4.0]undec-7-ene); so as to obtain a compound of formula (I).

35. A method of therapeutic treatment of a mammal in need thereof comprising administering to said mammal a compound according to any one of the claims 1-28, or a pharmaceutically acceptable salt thereof.