Compounds for use in the treatment of cancers that overexpress tspan1

Abstract

It relates to compounds of formula (I) for use in the treatment of cancers that overexpress TSPAN1 protein. It also relates to pharmaceutical compositions comprising these compounds, and to their therapeutical indication in the treatment of different types of cancer, in particular those resistant to conventional methods of treatment.

Description

[0001] Compounds for use in the treatment of cancers that overexpress TSPAN1

[0002] This application claims the benefit of European Patent Application EP23383269 filed 11 December 2023.

[0003] The invention refers to compounds for use in the treatment of cancers that overexpress TSPAN1 protein. It also relates to pharmaceutical compositions comprising these compounds, and to their therapeutical indication in the treatment of different types of cancer, in particular those resistant to conventional methods of treatment.

[0004] BACKGROUND ART

[0005] Cancer is an heterogeneous disease characterized by the accumulation of tumor cells, which can result in death in both animals and humans. Conventional methods of cancer treatment include surgical treatments, radiotherapy, administration of chemotherapeutic agents, and more recently immune-based treatments. However, so far, such treatments have had limited success in certain types of cancer.

[0006] Chemotherapy, despite all its limitations, is still today one of the most widespread methods for the treatment of different types of cancer. The inability of chemical agents to distinguish between normal, rapidly dividing cells and tumor cells can lead to the depression of the patient's immune system. This has been considered one of the main problems associated with chemotherapy, as well as the resistance mechanisms developed by cancer cells.

[0007] Drug discovery for use in cancer treatment is of vital importance for several reasons, in particular, to overcome the growing resistance of cancer cells to currently available treatments. One of the possible approaches is to combine new drugs with conventional treatments, thus enhancing effecti vity by synergy or by targeting different characteristics of tumor cells, especially in heterogenous tumors. This approach would also help reducing adverse effects associated to conventional treatments, such as chemotherapy and radiotherapy, given that doses would be reduced, improving tolerance and adherence to treatment. Moreover, the development of new drugs may also enable a more selective and targeted approach, by focusing on specific signaling pathways or genetic mutations present in resistant cancers. As a whole, new drugs for use in cancer treatment are needed in order to provide alternatives to conventional treatments, improve clinical results and provide hope, as well as better quality of life, to patients suffering with resistant cancers. Sensibilization of chemo-resistant tumor cells represents hence an important challenge for current cancer treatments. Tetraspanin-1 (TSPAN1) is a member of the family of proteins tetraspanins (TSPAN), whose principal characteristic is the capacity of forming aggregates between them or with several other transmembrane receptors, to become TSPAN-enriched microdomains.

[0008] These are essential for basic biological activities, such as cell adhesion, proliferation, and cell motility. Recently, TSPANTs overexpression has been associated to different types of cancers and to poor prognosis.

[0009] TSPANTs oncogenic role in cancer has been evidenced, mainly in digestives malignancies as pancreatic, gastric, colon cancer, and esophageal cancers, but also in hepatocellular carcinoma (Wang, G. L., et al., Oncol Rep 27, (2012), 1944-1952), skin squamous cell carcinoma, prostate cancer, osteosarcoma (Duan, J., et al., (2017), 5568- 5574) and non-small cell lung cancer (Wang, L., et al., Cell Death Dis 8, (2017), e2746).

[0010] In addition, the overexpression of TSPAN1 at mRNA and / or protein level in human tumor samples versus adjacent noncancerous tissues has been widely documented in cholangiocarcinoma, skin squamous cell carcinoma, esophageal carcinoma (Gu, T., et al., Oncology letters 14, (2017), 6815-6822), ovarian carcinomas (Scholz, C.J., et al., Cancer Lett 275 (2009), 198-203), prostate cancer (Munkley, J., et al., Sci Rep 7, (2017), 5249), pancreatic cancer and gastric carcinoma.

[0011] TSPAN1 is overexpressed (at mRNA and protein level) in colon cancer tissues. Its expression in tumor tissue is significantly associated with histological grade, proliferating cell nuclear antigen (PCNA), lymph node metastasis, and distant tumor node metastasis (TNM staging), according to Chen, L., et al., in “TSPAN1 protein expression: a significant prognostic indicator for patients with colorectal adenocarcinoma”, World J. Gastroenterol. 15 (2009), 2270. High TSPAN 1 expression is linked to poor survival and serves as an independent prognostic factor for colon cancer.

[0012] Zhang, G.-L., et al., in “The transcriptome difference between colorectal tumor and normal tissues revealed by single-cell sequencing”, J. Cancer 10 (2019), 5883-5890, disclose that gastric carcinoma, a highly aggressive digestive tract cancer, exhibits TSPAN 1 expression negatively associated with carcinoma differentiation and positively associated with tumor infiltration and lymph node status.

[0013] According to Hou, F.Q., et. al., in “Tetraspanin 1 is involved in survival, proliferation and carcinogenesis of pancreatic cancer”, Oncol Rep 34 (2015), 3068-3076, pancreatic cancer, known for its poor prognosis, identifies TSPAN 1 as an independent poor prognostic factor. TSPAN 1 has also been included in a diagnostic model for predicting pancreatic carcinoma. In head and neck squamous cell carcinoma (HNSCC), high TSPAN1 expression is related to poorly differentiated tumors.

[0014] S. Holters, et al., in “Tetraspanin 1 promotes invasiveness of cervical cancer cells”, Int. J. Oncol. 43 (2013), 503-512, disclose that, in high-grade cervical intraepithelial neoplasia, TSPAN1 overexpression has been shown to stimulate the invasive potential of cervical cancer cells. In ovarian serous borderline tumors and high-grade serous ovarian cancers, TSPAN1 is associated with worse overall survival. In endometriosis cell lines, frequently evolving into endometrial cancer, TSPAN1 overexpression triggers AMP-activated protein kinase (AMPK), ultimately increasing cell growth and invasion, suggesting TSPAN1 as a potential marker for these lesions. Overall, research suggests that TSPANI may serve as a new biomarker in various cancer models, with its overexpression generally linked to worse clinical prognoses.

[0015] Wang Y., et al., in “Tetraspanin 1 promotes epithelial-to-mesenchymal transition and metastasis of cholangiocarcinoma via PI3K / AKT signaling”, Journal of Experimental & Clinical Cancer Research, 2018, vol. 37, Article number 300, describe that TSPAN1 promotes epithelial-mesenchymal transition (EMT) and metastasis, acting through the p- ERK and p-AKT pathways, thus having a direct effect on the progression of the cancer. They furthermore report that TSPAN1 accelerates the progression, especially, of digestive malignancies, such as hepatocellular carcinoma (HCC), pancreatic, gastric, colorectal, and esophageal cancers, and some non-digestive cancers such as osteosarcoma and cervical cancer.

[0016] Huang R., et al., in “The role of tetraspanins pan-cancer”, iScience, 2022, vol. 25(8), 104777, describe a significantly high expression of TSPAN1 observed in breast, colon, kidney, pancreatic, and womb cancers. Moreover, they associate the overexpression of this protein to treatment resistance, proliferation speed and metastasis. Thus, it has been reported, that inhibition of TSPAN1 halts migration and invasion of cancer cells, as well as inducing their apoptosis in pancreatic cancer cells.

[0017] Garcia-Mayea Y., et al., in “TSPAN1 : A Novel Protein Involved in Head and Neck Squamous Cell Carcinoma Chemoresistance”, Cancers 2020, vol. 12, Article number 3269, describe that an increase in TSPAN1 expression, when compared to healthy tissue, can be observed in cancer cells resistant to cisplatin (CDDP-R), in cancer stem cells (CSC) and in biopsies obtained from patients with HNSCC. In the context of HNSCC, TSPAN1 has been identified as an oncogenic protein that contributes to the acquisition of chemoresistance of cancerous cells through the activation of its main target, p-Src. TSPAN1 emerges as a promising target for HNSCC therapy, given that its inhibition decreases size and proliferation of parental and resistant tumors, reduces metastasis, induces apoptosis, and sensitizes tumor cells to chemotherapeutic agents.

[0018] HNSCC is a form of cancer that affects the squamous cells lining the inner surfaces of the head and neck. This type of cancer can affect areas such as the oral cavity, pharynx, larynx, tonsils and sinuses. HNSCC accounts for approximately 90% of all head and neck cancers. It is a prevalent form of cancer with a significant burden of disease and mortality worldwide, mainly due to the lack of effective treatments and the resistance to conventional treatments, which in most cases leads to metastasis and relapse in patients, the latter occurring in more than 50% of HNSCC patients in the first 3 years after treatment.

[0019] Current available treatment for HNSCC include surgery, radiotherapy, and chemotherapy, depending on the stage of the cancer, the location and the general health of the patient. In some advanced cases, especially in metastatic patients, a combination of different therapeutic approaches is often necessary. Immunotherapy has entered the HNSCC treatment market in recent years, although only about 5% of all HNSCC patients can benefit from it. Immune checkpoint inhibitors, such as pembrolizumab and nivolumab, have been approved for the treatment of advanced or recurrent cases of HNSCC. However, the availability and approval of these therapies vary between countries and depend on several factors. Given that, unfortunately, this type of cancer is usually diagnosed in advanced stages, treatment options are limited, and metastases and relapses are frequent, reducing thus survival rates.

[0020] It is therefore imperative to find more effective and targeted therapeutic approaches to fight cancers in which TSPAN1, associated to poor prognosis, is overexpressed, to increase both life expectancy and life quality of patients, in particular, that of patients suffering from HNSCC.

[0021] SUMMARY OF THE INVENTION

[0022] Inventors have found certain compounds that are inhibitors of tetraspanin 1 (TSPAN1), and, therefore, have antitumor capacity against several cancer cell lines. Thus, as illustrated in the examples, the compounds of the invention inhibit TSPAN1 expression and, as a result, not only do they halt tumor proliferation, but moreover block activation of chemotherapy resistance and metastasis signaling pathways.

[0023] Furthermore, the compounds of the invention have effect against cancer cells but are not toxic for non-cancerous cells. Therefore, these compounds are advantageous because they represent a new treatment opportunity, in particular for cancers involving metastasis or chemotherapy-resistant cells, and provide more effective and targeted treatments for these challenging cancers.

[0024] Accordingly, a first aspect of the present invention relates to a compound of formula (I) or its pharmaceutically acceptable salts, for use in the treatment of a cancer expressing

[0025] TSPAN1 , in a mammal, including a human,

[0026] Formula (I) where: Ri is a radical selected from (Ci-C4)-alkyl, (C3-C6)-cycloalkyl, phenyl, and chlorophenyl; A is a biradical selected from S, N and C; Cy is a biradical selected from: on a single or double bond, m is an integer from 0 to 2; n is an integer from 0 to 2; o is an integer from 0 to 1 ; p is an integer from 0 to 1 ; q is an integer from 0 to 1 ; X is selected from O, S, N, and NH; Y is selected from C and N; Z is selected from C and N; R2, R3, R4, and Rs are selected from H, halogen, OH, (Ci-Ce)-alkyl, and O-(Ci-Ce)-alkyl; Re is selected from H and (Ci-C4)-alkyl-phenyl; R7-R20 is selected from H, halogen, OH, (Ci-Ce)-alkyl, and O-(Ci-Ce)-alkyl; with the proviso that: when (a) is a double bond then (b) is a single bond, and when (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond; X is selected from O, S, and NH; when (b) is a double bond, p is 0; when (b) is a single bond, p is 1 ; when Y is N, q is 0; when Z is N, o is 0; and when Z is C, o is 1.

[0027] Inventors have also found that the compounds of formula (I) as defined above in combination with cisplatin (CDDP) have synergistic effect, thus in a particular aspect of the invention, the combination of compounds of formula (I) and CDDP is for use in the treatment of cancers overexpressing TSPAN1.

[0028] Some of these compounds of formula (I) are new . and thus, a second aspect of the present invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof of formula (I),

[0029] Formula (I) wherein: R1 is a radical selected from (Ci-C4)-alkyl, and (C3-C6)-cycloalkyl;

[0030] A is S; Cy is a biradical selected from: a single or double bond; m is 1 ; n is an integer from 0 to 2; o is an integer from 0 to 1 ; p is an integer from 0 to 1 ; q is an integer from 0 to 1 ; X is selected from O, S, N; Y is selected from CH and N; Z is selected from C and N; R2 is selected from H, and halogen; Ra is selected from H, halogen, and OH; R4 is H; Rs is selected from H, halogen, and OH; Rs is selected from H and (Ci-C4)-alkyl-phenyl; and R7-R20 is selected from H, halogen, OH; with the proviso that: when (a) is a double bond then (b) is a single bond, and when

[0031] (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond; X is selected from O, S, and NH; when (b) is a double bond, p is 0; when

[0032] (b) is a single bond, p is 1 ; when Y is N, q is 0; when Z is N, o is 0; and when Z is C, o is 1 ; and with the proviso that: when Cy is phenyl and n is 0, R1 is other than methyl; when Cy is phenyl, n is 0 and X is O, R1 is other than ethyl or cyclopentyl; when Cy is phenyl, n is 0 and X is NH, R1 is other than isopropyl or 2-methylpropyl; and when Cy is phenyl, n is 0 and X is S, R1 is other than tert-butyl or cyclopentyl;

[0033] A third aspect of the present invention relates to a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein: R1 is a radical selected from (Ci-C4)-alkyl, and (Ca-Ce)- cycloalkyl; A is S; Cy has the same meaning as in formula (I), being a biradical selected from: formula (i), formula (ii), formula (iii), formula (iv), formula (v), and formula (vi); is a single or double bond; m is 1 ; n is an integer from 0 to 2; o is an integer from 0 to 1 ; p is an integer from 0 to 1 ; q is an integer from 0 to 1 ; X is selected from O, S, N; Y is selected from CH and N; Z is selected from C and N; R2 is selected from H, and halogen; Ra is selected from H, halogen, and OH; R4 is H; Rs is selected from H, halogen, and OH; Rs is selected from H and (Ci-C4)-alkyl-phenyl; and R7-R20 is selected from H, halogen, OH; with the proviso that: when (a) is a double bond then (b) is a single bond, and when

[0034] (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond; X is selected from O, S, and NH; when (b) is a double bond, p is 0; when

[0035] (b) is a single bond, p is 1 ; when Y is N, q is 0; when Z is N, o is 0; and when Z is C, o is

[0036] 1 , with the proviso that: when Cy is phenyl and n is 0, R1 is other than methyl; when Cy is phenyl, n is 0 and X is O, R1 is other than ethyl or cyclopentyl; when Cy is phenyl, n is 0 and X is NH, R1 is other than isopropyl or 2-methylpropyl; and when Cy is phenyl, n is 0 and X is S, R1 is other than tert-butyl or cyclopentyl; together with appropriate amounts of one or more pharmaceutical excipients or carriers.

[0037] BRIEF DESCRIPTION OF DRAWINGS

[0038] FIG. 1 shows expression of TSPAN1 in 4 cell line models of HNSCC (JHLI029, HTB-43, CCL-138 and SCC-25) by Western Blot (WB) (FIG. 1A) and the overexpression of TSPAN1 in CDDP-resistant cells (JHLI029, HTB-43 and CCL-138), as well as in CSC (generation 2 and 3 of HTB-43). p-actin and Vinculin were used as loading controls (FIG. 1 B).

[0039] FIG. 2 shows that depletion of TSPAN1 affects different signal cascades with p-Src as a central node as it is a commonly de-regulated protein.

[0040] FIG. 3 shows results of an in vivo study in mice, comparing growth of tumors formed by cells transfected with siRNA against TSPAN1 (siTSPANI) versus control cells, until day 41.

[0041] FIG. 4 shows reduction in tumor size, both in JHLI029 and CDDP resistant (R) cells, under TSPAN1 depletion (NC, negative control).

[0042] FIG. 5 shows a graphical representation (logarithmic scale) of a cytotoxic assay to calculate the respective IC50 values of each compound (If) for the lines JHLI029 and JHU029-R.

[0043] FIG. 6 shows the signaling pathway modulated for compound (If) (WB image) on HNSCC cell lines JHLI029, HTB-43 and CCL-138, both Parental and CDDP-resistant cells, at 8h and 24h after exposure to compound (If).

[0044] FIG. 7 shows morphological and proliferation changes on JHU029-R cells after 24hours of exposure to compound (If) at different concentrations.

[0045] FIG. 8 shows IC50 results of compound (If) on fibroblasts IMR90 and a cell line from HNSCC that doesn’t express TSPAN1 (line RPMI).

[0046] FIG. 9 shows a Volume vs Time graph comparing tumor growth of control group to that of the experimental group, treated with compound (If).

[0047] FIG. 10 shows tumor size comparison (at end point) between tumor samples of control group and tumor samples of experimental group.

[0048] FIG. 11 shows a graph representation of the mean tumor weight of the control group and that of the experimental group.

[0049] FIG. 12 shows anti-tumoral effect on JHLI029 cell line of CDDP, compound (If) and the combination of compound (If) and CDDP.

[0050] FIG. 13 shows a WB image of the effects of compound (If), CDDP and the combination of both on cell lines JHLI029 and JHU029-R, at 24h of exposition.

[0051] FIG. 14 shows microphotographies of the effect on the proliferation of JHU029-R cells of different treatment options, each option at two different concentrations.

[0052] FIG.15 shows a WB image of the effects of compounds (If), (Ir) and (Is) on cells lines JHU029-R and CCL-138-R at 24h and 48h.

[0053] FIG. 16 shows in vitro effectivity studies of compound (Is) in a graphic representation of tumor volume (mm3) vs time (days).

[0054] FIG. 17 shows the proliferation effect in number of cells at different times in JHLI029 and JHU029-R with compound (Is) versus control.

[0055] FIG. 18 shows differences in tumor weight (mean of each group) between control group and group treated with compound (Is) by the end of the treatment.

[0056] FIG. 19 shows the differences in tumor size by the end of the treatment with compound (Is).

[0057] FIG. 20 shows the expression in the proliferation marker Ki67 and the hematoxilin and eosin (H&E) staining in treated tumors with compound (If) and (Is).

[0058] FIG. 21 shows the cytotoxic effect of 11 compounds of the invention, at 100pM, in the HNSCC cell lines JHU029 and JHU029-R.

[0059] FIG. 22 shows IC50 values (mean ± SD) of compound (Is) at 72 hours when tested against cancer cells lines of different types of cancers (Hep27 for hepatocellular carcinoma, MDA-MD-23 for breast cancer, MNK-45 for gastric adenocarcinoma, SK-OV-3 for ovarian adenocarcinoma, and LI-2-OS and MG63 for osteosarcoma), where (%) is the survival rate of the given cells.

[0060] FIG. 23 shows WB test for TSPAN1 downstream signaling pathways of cancer cell lines of different types of cancers when treated for 24 hours with the corresponding IC50 concentration of compound (Is) obtained in FIG. 22, where NC DMSO is the “control” cell line, that is, without treatment, and asterisks (*) below the cell line indicate differences with respect to the control.

[0061] DETAILED DESCRIPTION OF THE INVENTION

[0062] All terms as used herein in this application, unless otherwise stated, shall be understood in their ordinary meaning as known in the art. Other more specific definitions for certain terms as used in the present application are as set forth below and are intended to apply throughout the description and claims.

[0063] As used herein, the indefinite articles “a” and “an” are synonymous with “at least one” or “one or more.” Unless indicated otherwise, definite articles used herein, such as “the,” also include the plural of the noun.

[0064] The word “comprise” for the purposes of the present invention encompasses the case of “consisting essentially of” and “consisting of”.

[0065] The term “treatment” is meant to include alleviating or eradicating a disorder, disease, or condition, or one or more of the symptoms associated with the disorder, disease or condition, or alleviating or eradicating the cause(s) of the disorder, disease, or condition itself.

[0066] The term “halogen” is meant to include the chemically related elements fluorine (F), chlorine (Cl), bromine (Br), and iodine (I).

[0067] As mentioned above, an aspect of the present invention relates to compounds of formula (I) as defined above, or their pharmaceutically acceptable salts, for use in the treatment of a cancer expressing TSPAN1 , in a mammal, including a human, where: Ri is a radical selected from (Ci-C4)-alkyl, (C3-C6)-cycloalkyl, phenyl, and chlorophenyl; A is a biradical selected from S, N and C; Cy is a biradical selected from: formula (i). formula (ii), formula (iii), formula (iv), formula (v), and formula (vi) as defined above; is a single or double bond, m is an integer from 0 to 2; n is an integer from 0 to 2; o is an integer from 0 to 1 ; p is an integer from 0 to 1 ; q is an integer from 0 to 1 ; X is selected from O, S, N, and NH; Y is selected from C and N; Z is selected from C and N; R2, R3, R4, and Rs are selected from H, halogen, OH, (Ci-Cs)-alkyl, and O-(Ci-Cs)-alkyl; Rs is selected from H and (C1-C4)- alkyl-phenyl; R7-R20 is selected from H, halogen, OH, (Ci-Cs)-alkyl, and O-(Ci-Cs)-alkyl; with the proviso that: when (a) is a double bond then (b) is a single bond, and when (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond; X is selected from O, S, and NH; when (b) is a double bond, p is 0; when (b) is a single bond, p is 1 ; when Y is N, q is 0; when Z is N, o is 0; and when Z is C, o is 1. This aspect can also be formulated as the use of compounds of formula (I) as defined above for the preparation of a medicament for the treatment and / or prevention of a cancer expressing TSPAN1 in a mammal, including a human.

[0068] The invention also relates to a method of treatment of a mammal, including a human, suffering from or being susceptible of suffering from cancer expressing TSPAN1 , in particular, to one of the cancers mentioned above, said method comprising the administration to said patient of a therapeutically effective amount of a compound of formula (I) as defined above, together with pharmaceutically acceptable excipients or carriers.

[0069] In a particular embodiment, the compounds of formula (I) as defined above, or their pharmaceutically acceptable salts, for use as defined above, are those where: A is a biradical selected from S and C, and m is an integer from 1 to 2.

[0070] In another particular embodiment, the compounds of formula (I) as defined above, or their pharmaceutically acceptable salts, for use as defined above, are those where: A is a biradical selected from S and C, m is an integer from 1 to 2, Cy is a biradical selected from formulas (i), (ii), (iii), (iv) and (v), and Re is selected from H and benzyl.

[0071] The compounds of formula (I) or their salts can exist in solvated, as well as unsolvated forms, including hydrated forms. Thus, they can contain in its structure stoichiometric amounts of solvent in the case of solvates, or of water in the case of hydrates. It is to be understood that the invention encompasses all such solvated, as well as unsolvated forms.

[0072] The pharmaceutically acceptable salts of the compounds of formula (I) include acid addition salts such as the hydrochloride, but also any other pharmaceutically acceptable salts of other acids such as hydrobromic, hydrofluoric, sulphuric, phosphoric, acetic, citric, fumaric, gluconic, lactic, maleic, succinic or tartaric acid.

[0073] The compounds of the present invention may be prepared by the process illustrated in Schemes I, II, III, IV and V. All radicals in the formulas of Schemes I, II, III, IV and V have the same meaning as the corresponding radicals in formula (I), as defined above. Cy in Schemes I, II, III, IV and V has the same meaning as the corresponding Cy in formula (I), as defined above. A, o, p, q, m, and n in Schemes I, III, IV and V have the same meaning as the corresponding A, o, p, q, m, and n in formula (I), as defined above. X in Schemes I, III, IV and V has the same meaning as the corresponding X in formula (I), and is thus selected from O, S, and NH, as defined above, with the proviso that in Scheme I X is selected from S or O.

[0074] Scheme I: Cyclisation when X is selected from O or S Compound (II) can be prepared by reacting (III) and (IV) in the presence of an acid, such as polyphosphoric acid (PPA). The reaction may be carried out at a temperature comprised in a range from 200 to 220°C. The mixture may be later cooled, neutralized with a base, such as NaOH, extracted with a solvent, such as EtOAc, dried over an anhydrous reagent, such as Na2SC>4, filtered, and concentrated under pressure to give cyclized product (II).

[0075] Scheme II: Cyclisation when X is N

[0076] Compound (IVi) can be prepared by reacting (VI) with (VII) ((Re)NH2) in the presence of a base, such as CS2CO3, for 2 hours, obtaining intermediated Vlh, followed by a hydrogenation reaction, using, for example, H2 and a palladium catalyst, for 3 hours. Both reactions may be carried out at room temperature and in an organic solvent, such as dioxane, and methanol. Compound (VIII) can be prepared through a cyclation reaction mediated by a coupling reagent, such as CDI, to obtain intermediate VII h, followed by a chlorination reaction, using, for example, a mixture of POCI3 and PCI5. Both reactions may be carried out in an organic solvent, such as THF. The cyclation reaction may be carried out at room temperature for 12 hours and the chlorination reaction may be carried out at a temperature comprised in a range from 90 to 110°C for 6 hours.

[0077] Compound of (II) can be prepared by reacting compound of (VIII) with compound of (III) boronic acid in the presence of a base, such as CS2CO3. Suzuki coupling reaction may be carried with a palladium catalyst, such as Pd(PPhs)4, in an organic solvent, such as DME, and with a reflux system.

[0078] Scheme III: Amide coupling

[0079] Compounds of formula (I) can be prepared by reacting (II) with (V) in the presence of a solvent, such as DMF, coupling reagents, such as EDC.HOBt, and HATLI, and an organic base, such as DI PEA, and triethylamine. The reaction may be stirred at room temperature, diluted with a solvent, such as EtOAc, extracted with a solvent, such as EtOAc, washed, dried over an anhydrous reagent, such as Na2SC>4, filtered, and concentrated under pressure. The product may be later purified to give (I).

[0080] Compounds of formula (V) can be prepared by using, for instance, mercaptoacetic acid (thioglycolic acid) and a halogenated compound, such as methyl iodide or propylbromide; or, for instance, by using mercaptoacetic ester instead of thioglycolic acid, and then a saponification process.

[0081] Compounds of formula (I) can be also prepared following Scheme III, by reacting (II) with compounds of formula (Vi), as defined below, in the presence of a solvent, such as DCM and an organic base, such as DI PEA or TEA. The reaction may be left stirring overnight at room temperature, washed, dried over an anhydrous reagent, such as Na2SC>4, filtered, and concentrated under pressure. The product may be later purified to give (I).

[0082] Scheme IV: Amide coupling when A is S

[0083] Compounds of formula (I) can be prepared in two steps following Scheme IV. In a first step, compound (lh) is obtained by reacting (II) with (V2.1) in the presence of a solvent, such as DCM, and an organic base, such as TEA. The reaction may be left stirring overnight at room temperature. In a second step, the resulting compound (lh) reacts with compound (V2.2), in the presence of a solvent, such as MeOH and an organic base, such as MeONa. The reaction may be left stirring at room temperature. Scheme V: Urea formation

[0084] Compounds of formula (I) can be prepared by reacting (II) with (V3) in the presence of a solvent, such as toluene. The reaction may be left stirring overnight at a temperature comprised in a range from 65 to 110°C to give the urea compounds of formula (I).

[0085] The preparation of pharmaceutically acceptable salts of the compounds of formula (I) can be carried out by methods known in the art. For instance, they can be prepared from the parent compound, which contains a basic or acidic moiety, by conventional chemical methods. Generally, such salts are, for example, prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate pharmaceutically acceptable base in water or in an organic solvent or in a mixture of them.

[0086] TSPAN1 is frequently overexpressed in several cancers and is correlated with advanced tumor stage and poor prognosis. The inventors have found, surprisingly, that the compounds of the present invention efficiently inhibit TSPAN1 overexpression, having a direct impact on proliferation and metastasis signaling pathways, as well as considerably reducing tumor size and weight, as can be seen in the examples and figures herein disclosed. This represents a major advance in cancer therapy, given that the compounds of the invention target a protein that is associated to several cancers, and have proved to be efficient against it.

[0087] In a particular embodiment, the compounds of the present invention for use as defined above are those where the cancer expressing TSPAN1 is selected from the group consisting of pancreatic cancer, colon cancer, gastric cancer, colorectal cancer, esophageal cancer, ovarian cancers, endometrial cancer, hepatocellular carcinoma, prostate cancer, osteosarcoma, non-small cell lung cancer, skin squamous cell carcinoma, cholangiocarcinoma, cervical intraepithelial neoplasia, and head and neck squamous cell carcinoma. In another particular embodiment, the compounds of the present invention for use as defined above are those where the cancer expressing TSPAN1 is selected from head and neck squamous cell carcinoma, hepatocellular carcinoma, breast cancer, gastric and ovarian adenocarcinoma, and osteosarcoma. In a more particular embodiment, the compounds of the present invention for use as defined above are those where the cancer expressing TSPAN1 is selected from HNSCC.

[0088] TSPAN1 activates several pathways involved in proliferation and apoptosis evasion, and even in metastasis. By inhibiting the overexpression of this protein, the compounds of the present invention halt these processes, therefore, ameliorating the patient’s prognosis, as well as their life expectancy and quality.

[0089] TSPAN1 is also involved in the activation of tumor cell mechanisms to develop resistance against currently approved cancer treatments, such as chemotherapy. The inventors have found that, unexpectedly, the compounds of the present invention are able to sensitize tumor cells to currently used chemotherapeutic agents, overcoming thus one of the major challenges in cancer therapy. Therefore, in a particular embodiment, the compounds of the present invention for use as defined above are those where the cancer is resistant to chemotherapeutic agents. In a more particular embodiment, the compounds of the present invention for use as defined above are those where the chemotherapeutic agent to which tumor cells are resistant is cisplatin (CDDP).

[0090] TSPAN1 is overexpressed in cancer stem cells (CSCs) and resistant HNSCC cells and in their CDDP-resistant derivatives JHU029-R, HTB-43-R and CCL-138-R. Thus, in a particular embodiment, the compounds of the present invention for use as defined above show antitumoral activity in a cancer cell line selected from JHLI029, JHU029-R, HTB-43, HTB-43-R, CCL-138, and CCL-138-R. In a more particular embodiment, the compounds of the present invention show antitumoral activity in the cancer cell line JHU029-R.

[0091] In another particular embodiment, the compounds of formula (I) for use as defined above are those where each one of the radicals R4, Rs, and R7-20 is H.

[0092] In another particular embodiment, the compounds of formula (I) for use as defined above are those where each one of the radicals R2 and R3 is selected from H and halogen.

[0093] In another particular embodiment, the compounds of formula (I) for use as defined above are those where the biradical A is S.

[0094] In another particular embodiment, compounds of formula (I) are those R1 is a radical selected from (Ci-C4)-alkyl, (Cs-Csj-cycloalkyl, phenyl, and chlorophenyl. In a more particular embodiment, compounds of formula (I) are those where R1 is a radical selected from (Ci-C4)-alkyl and (C3-C6)-cycloalkyl.

[0095] In another particular embodiment, compounds of formula (I) for use as defined above are those where m is 1. In another particular embodiment, compounds of formula (I) for use as defined above are those where X is O. In another particular embodiment, compounds of formula (I) for use as defined above are those where Y is N. In another particular embodiment, compounds of formula (I) for use as defined above are those where Z is C. In another particular embodiment, compounds of formula (I) for use as defined above are those where the biradical Cy is a phenyl.

[0096] In another particular embodiment, the compounds of formula (I) for use as defined above are selected from the following list: a) / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(phenylthio)acetamide (la) b) / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)butyramide (lb) c) / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)hexanamide (Ic) d) 1-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)-3-propylurea (Id) e) / \ / -(3-(benzo[c(]oxazol-2-yl)phenyl)-2-(ethylthio)acetamide (le)

[0097] In another particular embodiment, the compound of formula (I) for use as defined above is that where: f) when Ri is a chlorophenyl radical; n is 0; m is 1 ; Cy is a phenyl biradical; (a) is a single bond and (b) is a double bond; p is 0; o is 0; q is 0; A is S; Z is C; X is O; Y is N; and each one of R2 and R4 is H, its IIIPAC name is 2-((4-chlorophenyl)thio)- / V-(3- (oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide having formula (If) below,

[0098] In another particular embodiment, the compounds of formula (I) for use as defined above are selected from the following list: g) N-(5-(benzo[d]oxazol-2-yl)-2-chlorophenyl)-2-(ethylthio)acetamide (lea), h) N-(2-chloro-5-(oxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Iva), i) 2-(ethylthio)-N-(3-(7-hydroxybenzo[d]oxazol-2-yl)phenyl)acetamide, (leb), j) 2-(ethylthio)-N-(3-(4-hydroxybenzo[d]oxazol-2-yl)phenyl)acetamide (lec), and k) N-(3-(benzo[d]oxazol-2-yl)-4-hydroxyphenyl)-2-(ethylthio)acetamide (led).

[0099] The compounds of the present invention can be used in the same manner as other known chemotherapeutic agents, i.e., in combination with other treatments, either simultaneously or sequentially, depending on the condition to be treated. In another particular embodiment, the compounds of formula (I) for use in the treatment of cancers overexpressing TSPAN1 as defined above, are administered in combination with a chemotherapeutic agent. In a more particular embodiment, the chemotherapeutic agent is cisplatin. This is specially advantageous because of the fact that heterogenous cancers (more than a single population of cancer cells, i.e., bulk of cancer cells, CSCs, resistant cells [non-CSCs]), as is the case with HNSCC, tend to be resistant to certain treatments or develop mechanisms that enable cancer cells to keep proliferating, and by combining more than one strategy, it can be targeted different aspects of the cancer: In addition, the combination of both therapies have resulted in a synergistic effect, achieving a reduction of the tumor with a smaller dose and hence reduced side effects related to these treatments. In a particular embodiment, the compounds of the invention are administered simultaneously with the chemotherapeutic agent. In another particular embodiment, the compounds of the invention and the chemotherapeutic agent are administered separately, in any order, within a therapeutically effective interval. In another particular embodiment, the compounds of the invention and the chemotherapeutic agent are administered simultaneously or sequentially. In a more particular embodiment, the compounds of the invention are administered in combination with a chemotherapeutic agent at 50 to 800pM and 0,5 to 30pM, respectively.

[0100] The compositions of the present invention may be administered in parenteral form suitable for injection such as intravenous bolus injections, intravenous infusion, implantation into the body, oral, intratecal, or intranasal. Intratumoral administration may also be suitable for HNSCC tumors, due to their accessibility.

[0101] In a particular embodiment, the compounds for use in the treatment of a cancer expressing TSPAN1 , in a mammal, including a human, according to the present invention, are compounds of formula (11), or a pharmaceutically acceptable salt thereof, as described above.

[0102] As mentioned above, compounds of formula (I) or a pharmaceutically acceptable salt thereof, wherein: Ri is a radical selected from (Ci-C4)-alkyl, and (Cs-Cej-cycloalkyl;

[0103] A is S; Cy is a biradical selected from: a single or double bond; m is 1 ; n is an integer from 0 to 2; o is an integer from 0 to 1 ; p is an integer from 0 to 1 ; q is an integer from 0 to 1 ; X is selected from O, S, N; Y is selected from CH and N; Z is selected from C and N; R2 is selected from H, and halogen; R3is selected from H, halogen, and OH; R4 is H; Rs is selected from H, halogen, and OH; Rs is selected from H and (Ci-C4)-alkyl-phenyl; and R7-R20 is selected from H, halogen, OH; with the proviso that: when (a) is a double bond then (b) is a single bond, and when

[0104] (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond; X is selected from O, S, and NH; when (b) is a double bond, p is 0; when

[0105] (b) is a single bond, p is 1 ; when Y is N, q is 0; when Z is N, o is 0; and when Z is C, o is 1 , and with the proviso that: when Cy is phenyl and n is 0, R1is other than methyl; when Cy is phenyl, n is 0 and X is O, R1 is other than ethyl or cyclopentyl; when Cy is phenyl, n is 0 and X is NH, R1 is other than isopropyl or 2-methylpropyl; and when Cy is phenyl, n is 0 and X is S, R1 is other than tert-butyl or cyclopentyl, are also part of the invention.

[0106] In a particular embodiment the previous compounds of formula (I) are those where q is 1 and Rs is OH.

[0107] In another particular embodiment, the previous compounds of formula (I) are those where R3is OH.

[0108] In another particular embodiment, the previous compounds of formula (I) are those where R9 is Cl.

[0109] In another particular embodiment, the previous compounds of formula (I) are those where R7is OH. In another particular embodiment, the compound of formula (I) is a compoundof formula (h), or a pharmaceutically acceptable salt thereof,

[0110] Formula (h) where: Ri is a radical selected from (Ci-C4)-alkyl and (C3-C6)-cycloalkyl; Cy has the same meaning as in formula (I), being a biradical selected from: formula (i), formula (ii), formula (iii), formula (iv), formula (v), and formula (vi); a single or double bond; n is an integer from 0 to 2; o is an integer from 0 to 1 ; p is an integer from 0 to 1 ; X is selected from O, S and N; Y is selected from CH and N; Z is selected from C and N; R2 and R3are selected from H and halogen; Re is selected from H and (Ci-C4)-alkyl-phenyl; R7-R20 are H; with the proviso that: when (a) is a double bond then (b) is a single bond, and when (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond, X is selected from O, S, and NH; when (b) is a double bond, p is 0; when (b) is a single bond, p is 1 ; when Z is N, o is 0; and when Z is C, o is 1 and with the proviso that: when Cy is phenyl and n is 0, R1 is other than methyl; when Cy is phenyl, n is 0 and X is O, R1 is other than ethyl or cyclopentyl; when Cy is phenyl, n is 0 and X is NH, R1 is other than isopropyl or 2-methylpropyl; and when Cy is phenyl, n is 0 and X is S, R1 is other than tert-butyl or cyclopentyl; are also part of the present invention.

[0111] In a particular embodiment, compounds of formula (h) are those where, when Cy is phenyl and n is 0, R1 is other than methyl; when Cy is phenyl, n is 0 and X is O, R1 is other than ethyl or cyclopentyl; when Cy is phenyl, n is 0 and X is NH, R1 is other than isopropyl or 2-methylpropyl; and when Cy is phenyl, n is 0 and X is S, R1 is other than tert-butyl or cyclopentyl. The following compounds are commercially available, however, as far as the inventors know, no synthesis nor activity has been described: a) N-[3-(2-benzothiazolyl)phenyl]-2-(methylthio)acetamide; b) N-[3-(2-benzoxazolyl)phenyl]-2-(methylthio)acetamide; c) N-[3-(2-benzoxazolyl)phenyl]-2-(ethylthio)acetamide; d) N-[3-(1 H-benzimidazol-2-yl)phenyl]-2-(methylthio)acetamide; e) N-[3-(2-benzothiazolyl)phenyl]-2-[(1 , 1 -dimethylethyl)thio]acetamide; f) N-[3-(1 H-benzimidazol-2-yl)phenyl]-2-[(1-methylethyl)thio]acetamide; g) N-[3-(2-benzothiazolyl)phenyl]-2-(cyclopentylthio)-acetamide; h) N-[3-(2-benzoxazolyl)phenyl]-2-(cyclopentylthio)-acetamide; and i) N-[3-(1 H-benzimidazol-2-yl)phenyl]-2-[(2-methylpropyl)thio]acetamide.

[0112] In another particular embodiment, the compounds of formula (h) are those where n is an integer from 0 to 2; o is 1 ; p is 0; (a) is a single bond; (b) is a double bond; X is O; Y is N; Z is C; Cy is phenyl; Ri is a radical selected from (Ci-C4)-alkyl and (C3-C6)-cycloalkyl; R2 and R3are selected from H and halogen.

[0113] In a more particular embodiment, the compounds of formula (h) are those where n is an integer from 0 to 2; o is 0; p is 0; (a) is a single bond; (b) is a double bond; X is O; Y is N; Z is C; Cy is phenyl; R1 is a radical selected from (Ci-C4)-alkyl and (C3-C6)-cycloalkyl; R2 is H.

[0114] In another particular embodiment, the compounds of formula (h) as defined above are those selected from the following list: a) / V-(3-(3-benzyl-3 / 7-imidazo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Ig); b) 2-(ethylthio)- / V-(3-(oxazolo[5,4-c]pyridin-2-yl)phenyl)acetamide (Ih); c) 2-(ethylthio)- / V-(5-(oxazolo[4,5-b]pyridin-2-yl)oxazol-2-yl)acetamide (li); d) 2-(ethylthio)- / V-(5-(oxazolo[4,5-b]pyridin-2-yl)thiazol-2-yl)acetamide (Ij); e) / V-(5-(benzo[d]thiazol-2-yl)pyridin-3-yl)-2-(ethylthio)acetamide (Ik); f) / \ / -(5-(1 / 7-benzo[c]imidazol-2-yl)pyridin-3-yl)-2-(ethylthio)acetamide (Im); g) 2-(ethylthio)- / V-(2-(oxazolo[4,5-b]pyridin-2-yl)pyridin-4-yl)acetamide (In); h) 2-(ethylthio)- / V-(2-(oxazolo[4,5-b]pyridin-2-yl)pyrimidin-4-yl)acetamide(lo); i) / V-(3-(7-bromooxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Ip); j) / V-(3-(6-chlorooxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Iq); k) 2-(isopropylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Ir); l) 2-(ethylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Is); m) 2-(cyclopropylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (It); n) 2-(cyclopentylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (lu); o) 2-(methylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Iv); p) / V-(3-(6-chlorooxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(cyclopropylthio)acetamide (Iw); q) / \ / -(3-(benzo[c(]oxazol-2-yl)phenyl)-2-(cyclopropylthio)acetamide (lx); r) 2-(cyclopropylthio)-N-(3-(oxazolo[5,4-c]pyridin-2-yl)phenyl)acetamide (ly); s) / V-(3-(3-benzyl-3 / 7-imidazo[4,5-b]pyridin-2-yl)phenyl)-2-(cyclopropylthio)acetamide (Iz); t) / \ / -(5-(benzo[c(]oxazol-2-yl)pyridin-3-yl)-2-(ethylthio)acetamide (laa), and u) / \ / -(5-(benzo[c(]oxazol-2-yl)pyridin-3-yl)-2-(cyclopropylthio)acetamide (Ibb). In a more particular particular embodiment, the compounds of formula (h) as defined above are selected from the following list: a) / V-(3-(7-bromooxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Ip); b) / V-(3-(6-chlorooxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Iq); c) 2-(isopropylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Ir); d) 2-(ethylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Is); e) 2-(cyclopropylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (It); f) 2-(cyclopentylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (lu).

[0115] In a more particular particular embodiment, the compounds of formula (h) as defined above are selected from the following list: a) 2-(isopropylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Ir); b) 2-(ethylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Is); c) 2-(cyclopropylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (It); d) 2-(cyclopentylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (lu).

[0116] In a particular embodiment of the compounds of formula (I), the compound of formula (I) is selected from the following list:

[0117] - N-(5-(benzo[d]oxazol-2-yl)-2-chlorophenyl)-2-(ethylthio)acetamide (lea),

[0118] - N-(2-chloro-5-(oxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Iva),

[0119] - 2-(ethylthio)-N-(3-(7-hydroxybenzo[d]oxazol-2-yl)phenyl)acetamide, (leb),

[0120] - 2-(ethylthio)-N-(3-(4-hydroxybenzo[d]oxazol-2-yl)phenyl)acetamide (lec),

[0121] - N-(3-(benzo[d]oxazol-2-yl)-4-hydroxyphenyl)-2-(ethylthio)acetamide (led); and a pharmaceutically acceptable salt thereof; of any of them; is also part of the invention.

[0122] As mentioned above it is also part of the invention a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein: Ri is a radical selected from (C1-C4)- alkyl, and (C3-C6)-cycloalkyl; A is S; Cy has the same meaning as in formula (I), being a biradical selected from: formula (i), formula (ii), formula (iii), formula (iv), formula (v), and formula (vi); is a single or double bond; m is 1 ; n is an integer from 0 to 2; o is an integer from 0 to 1 ; p is an integer from 0 to 1 ; q is an integer from 0 to 1 ; X is selected from O, S, N; Y is selected from CH and N; Z is selected from C and N; R2 is selected from H, and halogen; Ra is selected from H, halogen, and OH; R4 is H; Rs is selected from H, halogen, and OH; Rs is selected from H and (Ci-C4)-alkyl-phenyl; and R7-R20 is selected from H, halogen, OH; with the proviso that: when (a) is a double bond then (b) is a single bond, and when (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond; X is selected from O, S, and NH; when (b) is a double bond, p is 0; when (b) is a single bond, p is 1; when Y is N, q is 0; when Z is N, o is 0; and when Z is C, o is 1, and with the proviso that: when Cy is phenyl and n is 0, Ri is other than methyl; when Cy is phenyl, n is 0 and X is O, Ri is other than ethyl or cyclopentyl; when Cy is phenyl, n is 0 and X is NH, Ri is other than isopropyl or 2-methylpropyl; and when Cy is phenyl, n is 0 and X is S, Ri is other than tert-butyl or cyclopentyltogether with appropriate amounts of one or more pharmaceutical excipients or carriers.

[0123] In a particular embodiment, the pharmaceutical composition according to the invention comprises a therapeutically effective amount of the compound of formula (h), or a pharmaceutically acceptable salt thereof as defined above, including the provisos mentioned above, together with appropriate amounts of one or more pharmaceutical excipients or carriers.

[0124] In another particular embodiment, the pharmaceutical composition according to the invention comprises a therapeutically effective amount of a compound of formula (I) which is selected from the group consisting of:

[0125] - N-(5-(benzo[d]oxazol-2-yl)-2-chlorophenyl)-2-(ethylthio)acetamide (lea),

[0126] - N-(2-chloro-5-(oxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Iva),

[0127] - 2-(ethylthio)-N-(3-(7-hydroxybenzo[d]oxazol-2-yl)phenyl)acetamide, (leb),

[0128] - 2-(ethylthio)-N-(3-(4-hydroxybenzo[d]oxazol-2-yl)phenyl)acetamide (lec),

[0129] - N-(3-(benzo[d]oxazol-2-yl)-4-hydroxyphenyl)-2-(ethylthio)acetamide (led), and a pharmaceutically acceptable salt thereof; of any of them; together with appropriate amounts of one or more pharmaceutical excipients or carriers.

[0130] The term “therapeutically effective amount” as used herein, refers to the amount of a compound that, when administered, is sufficient to prevent development of, or alleviate to some extent, one or more of the symptoms of the disease to be treated. The particular dose of compound administered according to this invention will of course be determined by the particular circumstances surrounding the case, including the compound administered, the route of administration, the particular condition being treated, and the similar considerations.

[0131] The term “pharmaceutical composition” refers to a mixture of a compound disclosed herein with other chemical components, such as diluents or carriers. The pharmaceutical composition facilitates administration of the compound to an organism.

[0132] The terms “pharmaceutically acceptable excipients or carriers” refer to pharmaceutically acceptable material, composition or vehicle, such as liquid or solid filler, diluent, excipient, solvent, or encapsulating material. Each component must be pharmaceutically acceptable in the sense of being compatible with the other ingredients of the pharmaceutical composition. It must also be suitable for use in contact with the tissue or organ of humans and animals without excessive toxicity, irritation, allergic response, immunogenicity or other problems or complications commensurate with a reasonable benefit / risk ratio.

[0133] Throughout the description and claims the word "comprise" and variations of the word, are not intended to exclude other technical features, additives, components, or steps. Furthermore, the word “comprise” encompasses the case of “consisting of” and of “consisting essentially of”. Additional objects, advantages and features of the invention will become apparent to those skilled in the art upon examination of the description or may be learned by practice of the invention. The following examples and drawings are provided by way of illustration, and they are not intended to be limiting of the present invention. Reference signs related to drawings and placed in parentheses in a claim, are solely for attempting to increase the intelligibility of the claim and shall not be construed as limiting the scope of the claim. Furthermore, the present invention covers all possible combinations of particular and preferred embodiments described herein.

[0134] EXAMPLES

[0135] Abbreviations: 3-(oxazolo[4,5-b]pyridin-2-yl)aniline (Ila); 3-(Benzo[d]oxazol-2-yl)aniline (lib); 5-(oxazolo[4,5-b]pyridin-2-yl)thiazol-2-amine (He); 3-(oxazolo[5,4-c]pyridin-2- yl)aniline (lid); 3-(7-bromooxazolo[4,5-b]pyridin-2-yl)aniline (He); 3-(6-chlorooxazolo[4,5- b]pyridin-2-yl)aniline (Ilf); 3-(3-benzyl-3H-imidazo[4,5-b]pyridin-2-yl)aniline (I Ig), 5- (benzo[d]thiazol-2-yl)pyridin-3-amine (Hh); 5-(Benzo[c(]oxazol-2-yl)pyridin-3-amine (Hi); 2- ((4-chlorophenyl)thio)acetic acid (Va); 2-(phenylthio)acetic acid (Vb); 2-(ethylthio)acetic acid (Vc); 2-(isopropylthio)acetic acid (Vd); 2-(cyclopropylthio)acetic acid (Ve); 2- (methylthio)acetic acid (Vf) ; 2-amino-4-bromopyridin-3-ol, bromine (IVa), and 2-Bromo-N- (3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Br1).

[0136] Abs (Absorbance); ACN (Acetonitrile); AcOH (Acetic acid); br (Broad); CV (Column Volume) ; ESI (Electrospray Ionisation) ; EtOAc (Ethyl Acetate); EtOH (Ethanol); FIA (Flux Injected Analysis); d (Doublet); dd (Doublet of Doublets); ddd (Doublet of Doublet of Doublets); dddd (Doublet of Doublet of Doublet of Doublets); DAD (Diode Array Detector); DCM (Dichloromethane); DIPEA (Diisopropylamine); DME (Dimethoxiethane); DMF (N,N- Dimethylformamide); DMSO (Dimethylsulfoxide); dt (Doublet of triplet); ee (enantiomeric excess); EDC (1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide); HOBt (hydroxybenzotriazole); ESI (Electrospray Ionization); h (Hours); H NMR (Proton Nuclear Magnetic Resonance); HNSCC (Head and Neck Squamous cell carcinoma), H&E (hematoxilin and eosin); P cells (parenteral cells); HPLC (High-performance Liquid Chromatography); IPA (Isopropanol); IUPAC (International Union of Pure and Applied Chemistry), LAH (Lithium aluminium hydride); m (Multiplet); MeOH (Methanol); mg (Miligrams); mL (Mililiter); pL (Microliter); MS (Mass Spectroscopy); MHz (Mega Hertz); min (Minutes); mm (Millimetre); mmol (Milimol); m / z (Mass to Charge Ratio); NC (Negative Control), NMR (Nuclear Magnetic Resonance Spectrometry); nm (Nanometre); ppm (Parts Per Million); q (Quartet); r.t. (room temperature); RT (Retention Time); s (Singlet); t (Triplet); THF (Tetrahydrofuran); UV (Ultraviolet); SD (standard deviation), and WB (Western Blot).

[0137] Materials and methods

[0138] All the Chemicals and solvents are from commercial suppliers and used without purification, except the anhydrous solvents such as DMF which were treated previously through a system of solvent purification (PureSolv), degasified with inert gases and dried over alumina or molecular sieves. Reactions were monitored by thin layer chromatography (60 F, 0.2 mm, Macherey-Nagel) by visualisation under 254 and / or 365 nm lamp. Purification was made by Flash column chromatography by using Merck Silica Gel 60, 40-63 microns RE or by Phase Reverse with an Isolera-Biotage equipment (SNAP KP-C18-HS; A: Water / Formic acid (0.05%), B: ACN / Formic Acid (0.05%): 5%B 3CV, 5%B-100%B 18CV, 100%B 5CV). NMR were performed in a Brucker 400 MHz. Chemical shifts 5 are reported in parts per million (ppm). HPLC 2795 Alliance Waters Aquity coupled to Detector DAD Agilent 1100 and Detector MS Waters ESI triple quadrupole Quattro micro, 10 pL of sample in ACN was injected, using a ZORBAX Extend-C18 3.5 pm 2.1x50mm (Agilent) column. The mobile phase used was a mixture of A = water + 0.05 formic acid and B = ACN + 0.05 formic acid with method described as follows: flow 0.5 mL / min; 5% B for 0.5 min; 5% to 100% B in 5 min, 100% B for 2 min.

[0139] Any of the compounds of formulas (I) and (h) may be prepared analogously, following the teachings of the document and the General Methods herein disclosed, using the corresponding starting products, as shown in the examples of the preparation process of some of the compounds of the invention.

[0140] General Method A: Cyclisation when X is selected from O or S

[0141] Following Scheme I, compound (IV) (1 equiv.) and compound (III) (1-1.2 equiv.) were added to polyphosphoric acid (c=1.4 M). The mixture was heated to 200°C. After 6 h, the mixture was poured onto cold water, neutralized with 5 M NaOH, extracted with EtOAc three times, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to give the cyclized product, compound (II), which was used in the following step without purification. General Method B: Cyclisation when X is N

[0142] Following Scheme II, to a solution of compound (VI) (1 equiv.) and CS2CO3 (2 equiv.) in dioxane (c=0.3 M) was added compound (VII) (1 equiv.) and heated at 100 °C for 2 h. The mixture was cooled to room temperature, filtered, and evaporated to give the substituted product, compound (Vh), which was used in the following step without purification, (stepl)

[0143] To a suspension of the resulting compound (Vh) (1 equiv.), and Pd / C (0.2 equiv.) in MeOH (c=0.3 M) was submitted a hydrogen atmosphere (1atm) for 3 h. Then, the mixture was filtered through Celite and evaporated to give the aniline of compound (IV1), which was used without further purification.

[0144] A solution of compound (IV1) (1 equiv.) and GDI (2 equiv.) in anhydrous THF was stirred under inert atmosphere for 12 h. Then, the product was partitioned between water and EtOAc. The organic layer was washed with brine, dried over anhydrous Na2SO4, and concentrated to give a cyclized product, compound (VII ), which was used in the following step without purification.

[0145] A solution of the resulting product (1 equiv.) in POCh (25 equiv.) was refluxed at 100°C for 10 min at which time PCI5 (1 equiv.) was added to the refluxing mixture and heated at 90°C for an additional 6 h. The mixture was cooled to room temperature and evaporated. The crude mixture was slowly added to ice-water and then basified with an aqueous 6M NaOH solution. The product was extracted with EtOAc. The organic layer was dried (Na2SO4) and purified by column chromatography to give the chlorinated imidazol compound.

[0146] The resulting compound (1 equiv.) and Pd(PPh3)4 (0.05 equiv.) were dissolved in DME (c=0.35 M) and stirred at room temperature for 10 min. The boronic acid of compound (III) (1.15 equiv.) was then added, followed by Na2COs (4 equiv.). The mixture was diluted with water, extracted with DCM, dried and evaporated. The crude residue was purified by flash column chromatography to give the aniline of compound (II).

[0147] General Method C: Amide coupling with EDC / HOBt

[0148] Following Scheme III, compound (II) (1 equiv.) and the required acid (1 equiv.) of compound (V) were in a solution with DMF (c = 0.2 M). EDC (1.5 equiv.), HOBt (1 .5 equiv.), and DI PEA (1.5 equiv.) were added to the solution. The mixture was stirred at room temperature overnight, then diluted with EtOAc, washed twice with brine, dried over anhydrous Na2SC>4, filtered, and concentrated under reduced pressure. The crude product was purified by flash column chromatography to give the amide product.

[0149] General Method D: Amide coupling with HATU

[0150] Following Scheme III, compound (II) (1 equiv.) and the required acid (1.5 equiv.) of compound (V) were in a solution with DMF (c = 0.1 M). HATU (2 equiv.) and triethylamine (3 equiv.) were added to the solution at 0 °C. The mixture was stirred at room temperature until completion, then extracted with EtOAc, dried over anhydrous Na2SC>4, filtered, and concentrated under reduced pressure. The crude product was purified by flash column chromatography to give the amide product.

[0151] General Method E: Amide coupling with the acid chloride

[0152] To a solution of compound (II) (1 equiv.) and triethylamine (2 equiv.) in DCM (c = 0.1 M), the required acid chloride (1.5 equiv.) was added, and the solution was stirred at room temperature overnight. Then, the mixture was evaporated and purified by reverse phase to give the amide product.

[0153] General Method F: Preparation of aromatic thioacetic acid derivatives

[0154] To a solution of the required benzenethiol (1 equiv.) in THF / water (1 :2, c = 0.7 M) at 0°C were added tetrabutylammonium bromide (0.01 equiv.), 2-bromoacetic acid (1 equiv.) and NaOH (2 equiv.). The mixture was stirred at room temperature overnight, then acidified with 1 M HCI, extracted with 3xEt2O, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to give the thioacetic product.

[0155] General Method G: Preparation of aliphatic thioacetic acid derivatives

[0156] To a solution of 2-mercaptoacetic acid (1 equiv.) in MeOH (c = 0.2 M) were added NaOH (2 equiv.) and the required aliphatic halide (1.02 equiv.). The solution was stirred at room temperature or at 40°C for 2 - 48 h, then concentrated under reduced pressure. The resulting solid was redissolved in water, acidified with 1 M HCI, extracted with Et20 three times, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to give the thioacetic product.

[0157] General Method H: Preparation of acid chloride from acid derivatives

[0158] To a solution of carboxylic acid (1 equiv.) in thionyl chloride (c = 0.1 M) were stirred at refluxed 1 h, then concentrated under reduced pressure. The resulting oil (acid chloride) was used without further purification.

[0159] Example 1 : Preparation process of compound (Ila)

[0160] Compound (Ila), with IIIPAC name 3-(oxazolo[4,5-b]pyridin-2-yl)aniline, was prepared from 2-aminopyridin-3-ol (500 mg, 4.54 mmol) and 3-aminobenzoic acid (623 mg, 4.54 mmol) following General Method A; yield 79% (757 mg, 3.58 mmol), yellow solid.1H NMR (400 MHz, DMSO-cfe) 6 8.53 (dd, J = 4.9, 1.4 Hz, 1 H), 8.21 (dd, J = 8.0, 1 .4 Hz, 1 H), 7.48 (t, J = 2.0 Hz, 1 H), 7.44 (dd, J = 8.0, 4.9 Hz, 1 H), 7.38 (dt, J = 8.0, 1.2 Hz, 1 H), 7.27 (t, J = 8.0 Hz, 1 H), 6.84 (ddd, J = 8.0, 2.0, 1 .2 Hz, 1 H), 5.54 (s, 2H); m / z (ESI+) 212.4 (MH+, 100%).

[0161] Example 2: Preparation process of compound (lib)

[0162] Compound (lib), with IIIPAC name 3-(Benzo[c(]oxazol-2-yl)aniline, was prepared from 2- aminophenol (796 mg, 7.29 mmol) and 3-aminobenzoic acid (1.00 g, 7.29 mmol) following General Method A; yield 31% (470 mg, 2.24 mmol), yellow solid.1H NMR (400 MHz, CD3OD) 5 7.74-7.69 (m, 1 H), 7.68-7.63 (m, 1 H), 7.55 (t, J = 1.8 Hz, 1 H), 7.52 (dt, J = 8.0, 1.2 Hz, 1 H), 7.46-7.34 (m, 2H), 7.28 (t, J = 8.0 Hz, 1 H), 6.92 (ddd, J = 8.0, 1.8, 1.2 Hz, 1 H).

[0163] Example 3: Preparation process of compound (lie)

[0164] Compound (He), with IIIPAC name 5-(oxazolo[4,5-b]pyridin-2-yl)thiazol-2-amine, was prepared from 2-aminopyridin-3-ol (764 mg, 6.94 mmol) and 2-aminothiazole-5-carboxylic acid (1.00 g, 6.94 mmol) following General Method A; yield 2% (35 mg, 0.16 mmol), yellow solid.1H NMR (400 MHz, DMSO-cfe) 6 8.42 (dd, J = 4.9, 1.4 Hz, 1 H), 8.10 (s, 2H), 8.08 (dd, J = 8.1 , 1.4 Hz, 1 H), 7.99 (s, 1 H), 7.34 (dd, J = 8.1 , 4.9 Hz, 1 H); m / z (ESI+) 219.3 (MH+, 100%).

[0165] Example 4: Preparation process of compound (lid)

[0166] Compound (lib), with IIIPAC name, 3-(oxazolo[5,4-c]pyridin-2-yl)aniline, was prepared from 4-aminopyridin-3-ol (1.00 g, 9.08 mmol) and 3-aminobenzoic acid (1.24 g, 9.08 mmol) following General Method A; yield 18% (351 mg, 1.66 mmol), white solid.1H NMR (400 MHz, DMSO-cfe) 6 9.11 (d, J = 1.0 Hz, 1 H), 8.55 (d, J = 5.3 Hz, 1 H), 7.84 (dd, J = 5.3, 1.0 Hz, 1 H), 7.49 (t, J = 2.0 Hz, 1 H), 7.40 (dt, J = 7.8, 1.2 Hz, 1 H), 7.27 (t, J = 7.8 Hz, 1 H), 6.86 (ddd, J = 7.8, 2.0, 1.2 Hz, 1 H), 5.56 (s, 2H); m / z (ESI+) 212.3 (MH+, 100%). Example 5: Preparation process of compound (IVa)

[0167] To prepare compound (IVa), with IIIPAC name 2-amino-4-bromopyridin-3-ol, bromine (1.17 mL, 22.7 mmol) was added dropwise to a stirred suspension of 2-aminopyridin-3-ol (2.08 g, 19.9 mmol) in acetic acid (31.5 mL) at 0°C. Then, the mixture was brought to room temperature. The reaction mixture was heated at 120°C for 12 h. After the reaction was completed, the reaction mixture was concentrated under reduced pressure and the residue was triturated with Et20 three times, filtered and dried to give 5.19 g of a black solid identified as compound (IVa), as a bromohydrate, with minoritary isomers 2-amino-5- bromopyridin-3-ol (IVb) and 2-amino-6-bromopyridin-3-ol (IVc), both bromohydrates.1H NMR (400 MHz, DMSO-86) 5 8.06 (s, 1 H), 7.42 (d, J = 6.9 Hz, 1 H), 7.04 (d, J = 6.9 Hz, 1 H), 4.13 (Brs, 2H).

[0168] Example 6: Preparation process of compound (He)

[0169] Compound (lie), with IIIPAC name 3-(7-bromooxazolo[4,5-b]pyridin-2-yl)aniline, was prepared from compound (IVa) bromohydrate (1.00 g, 3.70 mmol) and 3-aminobenzoic acid (508 mg, 3.70 mmol) following General Method A; yield 16% (173 mg, 0.596 mmol), pale yellow solid.1H NMR (400 MHz, DMSO-cfe) 6 8.40 (d, J = 5.4 Hz, 1 H), 7.74 (d, J = 5.4 Hz, 1 H), 7.48 (t, J = 2.0 Hz, 1 H), 7.40 (dt, J = 7.8, 1.2 Hz, 1 H), 7.29 (t, J = 7.8 Hz, 1 H), 6.87 (ddd, J = 7.8, 2.0, 1.2 Hz, 1 H), 5.61 (s, 2H). ; m / z (ESI+) 290.2 (MH+, 95%).

[0170] Example 7: Preparation process of compound (Ilf)

[0171] Compound (Ilf), with IIIPAC name 3-(6-chlorooxazolo[4,5-b]pyridin-2-yl)aniline, was prepared from 2-amino-5-chloropyridin-3-ol (1.00 g, 6.92 mmol) and 3-aminobenzoic acid (949 mg, 6.92 mmol) following General Method A, followed by trituration in DCM / MeOH instead of column chromatography; yield 13% (216 mg, 0.879 mmol), beige solid.1H NMR (400 MHz, DMSO-cfe) 61H NMR (400 MHz, DMSO-cfe) 6 8.57 (d, J = 2.2 Hz, 1 H), 8.52 (d, J = 2.2 Hz, 1 H), 7.46 (t, J = 2.0 Hz, 1 H), 7.36 (dt, J = 8.0, 1.2 Hz, 1 H), 7.27 (t, J = 8.0 Hz, 1 H), 6.85 (ddd, J = 8.0, 2.0, 1.2 Hz, 1 H), 5.57 (s, 2H) ; m / z (ESI+) 246.3 (MH+, 100%).

[0172] Example 8: Preparation process of compound (llg)

[0173] Compound (llg), with IIIPAC name 3-(3-benzyl-3H-imidazo[4,5-b]pyridin-2-yl)aniline, was prepared following General Method B.

[0174] Stepl : Compound (Vhg), with IIIPAC name / V-benzyl-3-nitropyridin-2-amine, was prepared from 2-chloro-3-nitropyridine (3.2 g, 20.2 mmol) and phenylmethanamine (2.2 mL, 20 mmol). Yield 81% (4.34 g, 16.3 mmol), yellow solid.1H NMR (400 MHz, CDCh) 5 8.52 (s, 1H), 8.45 (q, J = 1.8 Hz, 1 H), 8.43 (s, 1 H), 7.44-7.28 (m, 5H), 6.84-6.44 (m, 1H), 4.86 (d, J = 5.7 Hz, 2H); m / z (ESI+) 230.3 (MH+, 100%).

[0175] Step2: compound (I Vig), with IIIPAC name / V-benzylpyridine-2,3-diamine, was prepared from compound (Vhg) (4.34 g, 16.3 mmol). Yield (3.92 g, 86%) as a brown solid.1H NMR (400 MHz, CDCh) 6 7.79 (dd, J = 5.1, 1.5 Hz, 1 H), 7.44-7.38 (m, 2H), 7.36-7.27 (m, 5H), 6.88 (dd, J = 7.4, 1.5 Hz, 1H), 6.57 (dd, J = 7.4, 5.1 Hz, 1 H), 4.63 (d, J = 5.0 Hz, 2H), 4.39 (Brs, 1H); m / z (ESI+) 200.3 (MH+, 100%).

[0176] Step3: Compound (Vlllig), with IIIPAC name 3-benzyl-1 / 7-imidazo[4,5-b]pyridin-2(3 / 7)-one was prepared from compound (I Vig) (3.92 g, 16.3 mmol). Yield (3.52 g, 94%) as a brown semi-solid.1H NMR (400 MHz, CDCh) 6 8.06 (dd, J = 5.2, 1.4 Hz, 1H), 7.73 (s, 1H), 7.52- 7.44 (m, 2H), 7.39-7.24 (m, 3H), 7.22 (dd, J = 7.6, 1.4 Hz, 1H), 6.98 (dd, J = 7.6, 5.2 Hz, 1 H), 5.17 (s, 2H); m / z (ESI+) 226.3 (MH+, 100%).

[0177] Step4: Compound (VII Ig), with IIIPAC name 3 imidazopyridine 3-benzyl-2-chloro-3 / 7- imidazo[4,5-b]pyridine was prepared from compound (Vlllig) (1.03 g, 4.57 mmol). Yield (1.11 mg, 0.76 mmol, 99%) as a brown oil.1H NMR (400 MHz, CDCh) 6 8.31 (dd, J = 4.9, 1.4 Hz, 1 H), 7.89 (dd, J = 8.0, 1.4 Hz, 1H), 7.30-7.14 (m, 6H), 5.43 (s, 2H); m / z (ESI+) 244.3 (MH+, 100%).

[0178] Step5: Compound (I Ig), with IIIPAC name 3-(3-benzyl-3 / 7-imidazo[4,5-b]pyridin-2- yl)aniline was prepared from compound (VI I Ig) (800 mg, 3.28 mmol) and 3- aminophenylboronic acid (517mg, 3.78 mmol) and purified by column cromatography (DCM:MeOH 2%). Yield (390 mg, 40%) as a solid.1H-NMR (400 MHz, CDCI3) 5 8.41 (dd, J = 4.8, 1.4 Hz, 1H), 8.10 (dd, J = 8.0, 1.4 Hz, 1 H), 7.32-7.24 (m, 4H), 7.22 (t, J = 8.0 Hz, 1 H), 7.16-7.06 (m, 2H), 7.04-6.97 (m, 2H), 6.78 (dd, =8.0, 1.2 Hz, 1H ), 5.62 (s, 2H), 3.74 (Brs, 2H); m / z (ESI+) 301.5 (MH+, 100%).

[0179] Example 9: Preparation process of compound (I I h)

[0180] Compound (llh), with IIIPAC name 5-(benzo[c(]thiazol-2-yl)pyridin-3-amine, was prepared from 2-aminobenzenethiol (930 pL, 8.69 mmol) and 5-aminonicotinic acid (1.20 g, 8.69 mmol) following General Method A; yield quant. (2.18 g), green solid.1H NMR (400 MHz, DMSO-86) 5 8.41 (d, J = 2.0, 1 H), 8.17 (ddd, J = 8.4, 1.2, 0.8 Hz, 1 H), 8.09 (d, J = 2.4 Hz, 1 H), 8.08 (ddd, J = 8.4, 1.2, 0.8 Hz, 1H), 7.61 (dd, J = 2.4, 2.0 Hz, 1H), 7.57 (ddd, J = 8.4, 7.2, 1.2 Hz, 1H), 7.49 (ddd, J = 8.4, 7.2, 1.2 Hz, 1 H), 5.73 (s, 2H); m / z (ESI+) 228.4 (MH+, 100%).

[0181] Compound (Hi), with IIIPAC name 5-(Benzo[d]oxazol-2-yl)pyridin-3-amine, was prepared from 2-aminophenol (1.58 g, 14.5 mmol) and 5-aminonicotinic acid (2.00 g, 14.5 mmol) following General Method A; yield 6% (180 mg, 0.852 mmol), off-white solid. .1H NMR (400 MHz, DMSO-86) 5 8.41 (d, J = 2.0 Hz, 1 H), 8.17 (d, J = 2.8 Hz, 1 H), 7.83 (td, J = 7.6,

[0182] 2.4 Hz, 1 H), 7.82 (td, J = 7.6, 2.4 Hz, 1 H), 7.60 (dd, J = 2.8, 2.0 Hz, 1 H ), 7.59 - 7.53 (m, 1 H), 7.46 (td, J = 7.6, 2.4 Hz, 1 H), 7.43 (td, J = 7.6, 2.4 Hz, 1 H), 5.72 (s, 2H); m / z (ESI+)

[0183] 228.4 (MH+, 100%); m / z (ESI+) 212.3 (MH+, 100%).

[0184] Compound (llj), with IIIPAC name 4-amino-2-(benzo[d]oxazol-2-yl)phenol, was prepared from 2-aminophenol (726 mg, 7.29 mmol) and 5-amino-2-hydroxybenzoic acid (1.00 g, 7.29 mmol) following General Method A; yield 28% (415 mg, 1.83 mmol), yellow solid.1H NMR (400 MHz, DMSO-86) 5 10.40 (s, 1 H), 7.87-7.81 (m, 2H), 7.50-7.42 (m, 2H), 7.26 (d, J = 2.6 Hz, 1 H), 6.87 (d, J = 8.7 Hz, 1 H), 6.82 (dd, J = 8.7, 2.6 Hz, 1 H), 4.94 (s, 2H).

[0185] Compound (Ilk), with IIIPAC name 2-chloro-5-(oxazolo[4,5-b]pyridin-2-yl)aniline, was prepared from 2-aminopyridin-3-ol (661 mg, 6.00 mmol) and 3-amino-4-chlorobenzoic acid (1.03 g, 6.00 mmol) following General Method A; yield 43% (630 mg, 2.56 mmol), grey solid.1H NMR (400 MHz, DMSO-86) 5 8.55 (dd, J = 4.8, 1.6 Hz, 1 H), 8.23 (dd, J = 8.1 , 1.6 Hz, 1 H), 7.72 (d, J = 2.0 Hz, 1 H), 7.48-7.45 (m, 2H), 7.40 (dd, J = 8.3, 2.0 Hz, 1 H), 5.84 (s, 2H).

[0186] Compound (I Im), with IIIPAC name 5-(benzo[d]oxazol-2-yl)-2-chloroaniline, was prepared from 2-Aminophenol (782 mg, 7.17 mmol) and 3-Amino-4-chlorobenzoic acid (1.23 g, 7.17 mmol) following General Method A; yield 98% (1.72 g, 7.03 mmol), grey solid.1H NMR (400 MHz, DMSO-86) 5 7.81-7.77 (m, 2H), 7.68 (d, J = 2.0 Hz, 1 H), 7.46-7.39 (m, 3H), 7.36 (dd, J = 8.3, 2.0 Hz, 1 H), 5.78 (s, 2H). Compound (I In), with IIIPAC name 2-(3-aminophenyl)benzo[d]oxazol-7-ol, was prepared from of 3-aminobenzene-1 ,2-diol hydrochloride (234 mg, 1.45 mmol) and 3-Aminobenzoic acid (199 mg, 1.45 mmol) following General Method A; yield 6.8% (22 mg, 0.10 mmol), white solid.1H NMR (400 MHz, DMSO-86) 5 10.30 (s, 1 H), 7.34 (t, J = 2.0 Hz, 1 H), 7.23 (dt, J = 8.0, 1.6 Hz, 1H), 7.14 (t, J = 8.0 Hz, 1H), 7.11-7.04 (m, 2H), 6.75 (dd, J = 7.2, 1.6 Hz, 1H), 6.69 (ddd, J = 8.0, 2.0, 0.8 Hz, 1H), 5.39 (s, 2H).

[0187] Example 15: Preparation process of compound (Ho)

[0188] Compound (Ho), with IIIPAC name 2-(3-aminophenyl)benzo[d]oxazol-4-ol, was prepared from 2-aminobenzene-1,3-diol (2.00 g, 15.98 mmol) and 3-aminobenzoic acid (2.19 g, 15.98 mmol) following General Method A; yield 78% (2.82 g, 12.47 mmol), grey solid.1H NMR (400 MHz, DMSO-86) 5 10.35 (Brs, 1 H), 7.41 (t, J = 2.0 Hz, 1 H), 7.31 (dt, J = 7.6, 1.6 Hz, 1 H), 7.22 (t, J = 7.6 Hz, 1 H), 7.21-7.13 (m, 2H), 6.83-6.74 (m, 2H), 5.47 (Brs, 2H).

[0189] Example 16: Preparation process of compound (Va)

[0190] Compound (Va), with IIIPAC name 2-((4-chlorophenyl)thio)acetic acid, was prepared from of 4-chlorobenzenethiol (500 mg, 3.46 mmol) following General Method F; yield 94% (659 mg, 3.25 mmol), pink solid.1H NMR (400 MHz, CDCI3) 5 7.36 (d, J = 8.6 Hz, 2H), 7.29 (d, J = 8.6 Hz, 2H), 3.65 (s, 2H).

[0191] Example 17: Preparation process of compound (Vb)

[0192] Compound (Vb), with IIIPAC name 2-(phenylthio)acetic acid, was prepared from benzenethiol (500 pL, 4.90 mmol) following General Method G; yield quant. (833 mg, 4.95 mmol), pink solid.1H NMR (400 MHz, DMSO-cfe) 6 12.74 (s, 1 H), 7.39-7.26 (m, 4H), 7.20 (m, 1 H), 3.79 (s, 2H).

[0193] Example 18: Preparation process of compound (Vc)

[0194] Compound (Vc), with IIIPAC name 2-(ethylthio)acetic acid, was prepared from 2- mercaptoacetic acid (500 pL, 7.16 mmol) and iodoethane (590 pL, 7.31 mmol) following General Method G at room temperature for 48 h; yield 85% (732 mg, 6.09 mmol), light yellow oil.1H NMR (400 MHz, CDCI3) 5 3.27 (s, 2H), 2.69 (q, J = 7.4 Hz, 2H), 1.29 (t, J = 7.4 Hz, 3H).

[0195] Example 19: Preparation process of compound (Vd) Compound (Vd), with IIIPAC name 2-(isopropylthio)acetic acid, was prepared from 2- mercaptoacetic acid (500 pL, 7.16 mmol) and 2-bromopropane (710 pL, 7.52 mmol) following General Method F at room temperature for 2 h; yield 92% (881 mg, 6.56 mmol), light yellow oil.1H NMR (400 MHz, CDCI3) 5 3.29 (s, 2H), 3.10 (hept, J = 6.7 Hz, 1 H), 1.30 (dt, J = 6.7 Hz, 6H).

[0196] Example 20: Preparation process of compound (Ve)

[0197] Stepl: To a solution of ethyl 2-mercaptoacetate (0.91 ml, 8.32 mmol) in DMSO (8.32 ml) were added potassium tert-butoxide (1.17 g, 10.40 mmol), and bromocyclopropane (0.67 ml, 8.32 mmol). The resulting mixture were heated at 80 °C overnight. The resulting mixture was cooled at room temperature, and added saturated NaHCO3, extracted with EtOAc, three times, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give compound (Vei) with IIIPAC name ethyl 2- (cyclopropylthio)acetate, as a light yellow oil (721 mg, 4.50 mmol, 54%).1H NMR (400 MHz, CDCI3) 6 4.23 (q, J = 7.1 Hz, 2H), 3.31 (s, 2H), 2.11-2.03 (m, 1H), 1.32 (t, J = 7.1 Hz, 3H), 0.93-0.86 (m, 2H), 0.65-0.58 (m, 2H).

[0198] Step2: Ethyl 2-(cyclopropylthio)acetate (520 mg, 3.25 mmol) was dissolved in a 1 :1 mixture of THF / MeOH (32.5 mL) and then aqueous sodium hydroxide (2.5 M, 6.5 ml, 16 mmol) was added and reacted at room temperature overnight. The mixture was neutralised with 1 M hydrochloric acid until acidic pH and extracted with EtOAc three times. The organic layers were washed with brine, dried, and concentrated in vacuo to compound (Ve) as a yellow oil (480 mg, 3.63 mmol, quant.).1H NMR (400 MHz, CDCI3) 6 3.33 (s, 2H), 2.12-2.03 (m, 1H), 0.98-0.82 (m, 2H), 0.70-0.57 (m, 2H).

[0199] Example 21 : Preparation process of compound (Vf)

[0200] To a solution of 2-mercaptoacetic acid (500 pL, 7.16 mmol) in MeOH (42 mL) were added methyl iodide (460 pL, 7.31 mmol) and NaOH (573 mg, 14.3 mmol). The mixture was stirred at room temperature for 48 h, then concentrated. The residue was redissolved in water, acidified with 1 M HCI, extracted with Et20 three times, dried over anhydrous Na2SC>4, filtered, and concentrated under reduced pressure to give compound (Vf), with IIIPAC name 2-(methylthio)acetic acid, as a light yellow oil (609 mg, 5.74 mmol, 80%).1H NMR (400 MHz, CDCI3) 6 3.23 (s, 2H), 2.25 (s, 3H). The NMR signals are in agreement with the reported values.

[0201] Example 22: Preparation process of compound (Vea) Compound (Vea), with IIIPAC name 2-(ethylthio)acetyl chloride, was prepared from compound Vc (2.00 g, 16.6 mmol) following General Method H; yield: quantitative. (2.31 g, 16.6 mmol), light brown oil.

[0202] Example 23: Preparation process of compound (la)

[0203] Compound (la), with IIIPAC name / \ / -(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)-2- (phenylthio)acetamide, was prepared from compound (Ila) (450 mg, 2.13 mmol) and compound (Vb) (358 mg, 2.13 mmol) following General Method C; yield 52% (399 mg, 1.10 mmol), white solid.1H NMR (400 MHz, DMSO-cfe) 6 10.55 (s, 1 H), 8.62 (s, 1 H), 8.55 (d, J = 4.8 Hz, 1 H), 8.25 (d, J = 8.0 Hz, 1 H), 7.94 (d, J = 8.0 Hz, 1 H), 7.77 (d, J = 8.0 Hz, 1 H), 7.58 (t, J = 8.0 Hz, 1 H), 7.47 (dd, J = 8.0, 4.8 Hz, 1 H), 7.44 (d, J = 7.6 Hz, 2H), 7.34 (t, J = 7.6 Hz, 2H), 7.21 (t, J = 7.6 Hz, 1H), 3.92 (s, 2H); m / z (ESI+) 362.4 (MH+, 100%).

[0204] Example 24: Preparation process of compound (lb)

[0205] Compound (lb), with IIIPAC name / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)butyramide, was prepared from compund (Ila) (43 mg, 0.20 mmol) and Butyric acid (420 pL, 0.302 mmol following General Method C; The crude product was purified by reverse phase yielding compound (lb), 49% (28 mg, 0.098 mmol) as a white solid.1H NMR (400 MHz, CD3OD) 5 8.56 (t, J = 1.9 Hz, 1H), 8.50 (dd, J = 5.0, 1.4 Hz, 1H), 8.12 (dd, J = 8.2, 1.4 Hz, 1 H), 8.04 - 7.97 (m, 1 H), 7.83 (ddd, J = 8.2, 2.2, 1.1 Hz, 1 H), 7.53 (t, J = 8.0 Hz, 1H), 7.46 (dd, J = 8.2, 5.0 Hz, 1H), 2.40 (t, J = 7.4 Hz, 2H), 1.76 (h, J = 7.4 Hz, 2H), 1.03 (t, J = 7.4 Hz, 3H); m / z (ESI+) 282.4 (MH+, 100%).

[0206] Example 25 Preparation process of compound (Ic)

[0207] Compound (Ic), with IIIPAC name / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)hexanamide, was prepared from compound (Ila) (43 mg, 0.20 mmol) and hexanoyl chloride (40 pL, 0.30 mmol) following General Method E; yield 32% (20 mg, 0.065 mmol), white solid.1H NMR (400 MHz, CD3OD) 5 8.53 (t, J = 2.0 Hz, 1 H), 8.49 (dd, J = 5.0, 1.2 Hz, 1 H), 8.10 (dd, J = 8.0, 1.2 Hz, 1 H), 7.98 (ddd, J = 8.0, 2.0, 1.2 Hz, 1H), 7.81 (ddd, J = 8.0, 2.0, 1.2 Hz, 1 H), 7.51 (t, J = 8.0 Hz, 1 H), 7.44 (dd, J = 8.0, 5.0 Hz, 1H), 2.41 (t, J = 7.8 Hz, 2H), 1.80-1.65 (m, 2H), 1.44-1.33 (m, 4H), 0.94 (t, J = 7.2 Hz, 3H); m / z (ESI+) 310.4 (MH+, 100%).

[0208] Example 26: Preparation process of compound (Id)

[0209] 1-lsocyanatopropane (31 pL, 0.33 mmol) and compound (Ila) (47 mg, 0.22 mmol) were dissolved in dry toluene (2.2 mL) and stirred at room temperature for 30 min. At this point, more 1-isocyanatopropane (31 pL, 0.33 mmol) was added and continued stirring at 0°C for overnight. The reaction mixture was concentrated under vacuum and purified with reverse phase to give compound (Id), with IIIPAC name 1-(3-(oxazolo[4,5-b]pyridin-2- yl)phenyl)-3-propylurea, as a white solid (19 mg, 0.065 mmol, 29%).1H NMR (400 MHz, CD3OD) 58.50 (dd, J= 5.6, 1.6 Hz, 1H), 8.37 (t, J= 1.6 Hz, 1H), 8.12 (dd, J= 8.0, 1.6 Hz, 1H), 7.90 (ddd, J= 8.0, 2.0, 1.6 Hz, 1H), 7.64 (ddd, J= 8.0, 2.0, 1.6 Hz, 1H), 7.47 (t, J = 8.0 Hz, 1H ) 7.44 (dd, J= 8.0, 5.6 Hz, 1H), 3.19 (t, J= 7.4 Hz, 2H), 1.58 (h, J= 7.4 Hz, 2H), 0.98 (t, J= 7.4 Hz, 3H); m / z (ESI+) 297.4 (MH+, 100%). e

[0210] Compound (le), with IIIPAC name / \ / -(3-(benzo[c(]oxazol-2-yl)phenyl)-2- (ethylthio)acetamide, was prepared from compound (lib) (100 mg, 0.476 mmol) and compound (Vc) (82 mg, 0.68 mmol) following General Method C; yield 19% (28 mg, 0.088 mmol).1H NMR (400 MHz, CD3OD) 58.53 (t, J= 1.8 Hz, 1H), 7.99 (ddd, J= 8.0, 1.8, 0.8 Hz, 1H), 7.79 (ddd, J= 8.0, 2.0, 0.8 Hz, 1H), 7.77-7.72 (m, 1H), 7.71-7.67 (m, 1H), 7.54 (t, J= 8.0 Hz, 1H), 7.46-7.39 (m, 3H), 3.39 (s, 2H), 2.72 (q, J= 7.4 Hz, 2H), 1.31 (t, J = 7.4 Hz, 3H).; m / z (ESI+) 313.3 (MH+, 100%).

[0211] Compound (If), with IIIPAC name 2-((4-chlorophenyl)thio)- / V-(3-(oxazolo[4,5-b]pyridin-2- yl)phenyl)acetamide, was prepared from compound (Ila) (50 mg, 0.24 mmol) and compound (Va) (48 mg, 0.24 mmol) following General Method C; yield 70% (66 mg, 0.16 mmol), off-white solid.1H NMR (400 MHz, DMSO-cfe) 610.56 (s, 1H), 8.62 (s, 1H), 8.55 (d, J= 4.8 Hz, 1H), 8.26 (d, J= 8.0 Hz, 1H), 7.95 (d, J= 7.6 Hz, 1H), 7.76 (d, J= 7.6 Hz, 1H), 7.59 (t, J= 8.0 Hz, 1H), 7.47 (dd, J= 8.0, 4.8 Hz, 1H), 7.44 (d, J= 8.4 Hz, 2H), 7.34 (d, J= 7.6 Hz, 2H), 7.33 (d, J= 7.6 Hz, 2H), 7.21 (t, J= 7.6 Hz, 1H), 3.94 (s, 2H); m / z (ESI+) 396.3 (MH+, 100%).

[0212] Example 29: Preparation process of compound (Ig)

[0213] Compound (Ig), with IIIPAC name / V-(3-(3-benzyl-3 / 7-imidazo[4,5-b]pyridin-2-yl)phenyl)-2- (ethylthio)acetamide, was prepared from compound (I Ig) (90 mg, 0.30 mmol) and compound (Vc) (71 mg, 0.59 mmol) following General Method D. ; yield 4% (5 mg, 0.02 mmol).1H NMR (400 MHz, CD3OD) 58.43 (dd, J= 4.8, 1.4 Hz, 1H), 8.14 (dd, J= 8.0, 1.6 Hz, 1 H), 8.01 (dd, J = 2.0, 1.6 Hz, 1 H), 7.72 (ddd, J = 8.0, 2.0, 1.2 Hz, 1 H), 7.46 (t, J = 8.0 Hz, 1H), 7.42 (dd, J= 8.0, 4.8 Hz, 1H), 7.37 (ddd, J= 8.0, 2.0, 1.2 Hz, 1H), 7.23-7.18 (m, 3H), 7.01-6.96 (m, 2H), 5.70 (s, 2H), 3.35 (s, 2H), 2.67 (q, J= 7.4 Hz, 2H), 1.29 (t, J= 7.4 Hz, 3H). m / z (ESI+) 403.4 (MH+, 100%).

[0214] Compound (Ih), with IIIPAC name 2-(ethylthio)- / V-(3-(oxazolo[5,4-c]pyridin-2- yl)phenyl)acetamide, was prepared from compound (lid) (150 mg, 0.710 mmol) and compound (Vc) (102 mg, 0.852 mmol) following General Method C; yield 37% (82 mg, 0.26 mmol), white solid.1H NMR (400 MHz, CD3OD) 5 8.99 (s, 1H), 8.64-8.61 (m, 1H), 8.52 (d, J = 5.2 Hz, 1 H), 8.04 (d, J = 8.0 Hz, 1 H , 1 H), 7.85-7.76 (m, 2H), 7.56 (t, J = 8.0 Hz, 1 H , 1 H ), 3.39 (s, 2H), 2.72 (q, J = 7.4 Hz, 2H), 1.31 (t, J = 7.4 Hz, 3H); m / z (ESI+) 314.3 (MH+, 100%).

[0215] Compound (Ij), with IIIPAC name 2-(ethylthio)- / V-(5-(oxazolo[4,5-b]pyridin-2-yl)thiazol-2- yl)acetamide, was prepared from compound (He) (35 mg, 0.16 mmol) and compound (Vc) (23 mg, 0.19 mmol) following General Method C; yield 10% (5 mg, 0.016 mmol).1H NMR (400 MHz, CD3OD) 5 8.48 (dd, J = 5.0, 1.4 Hz, 1 H), 8.40 (s, 1 H), 8.09 (dd, J = 8.2, 1.4 Hz, 1 H), 7.44 (dd, J = 8.2, 5.0 Hz, 1H), 3.48 (s, 2H), 2.70 (q, J = 7.4 Hz, 2H), 1.30 (t, J = 7.4 Hz, 3H); m / z (ESI+) 321.2 (MH+, 100%).

[0216] Compound (Ik), with IIIPAC name / \ / -(5-(benzo[c(]thiazol-2-yl)pyridin-3-yl)-2- (ethylthio)acetamide, was prepared from compound (I Ih) (90 mg, 0.40 mmol) and compound (Vc) (54 mg, 0.45 mmol) following General Method D; yield 8% (9 mg, 0.02 mmol). 1 H NMR (400 MHz, CD3OD) 5 8.99 (d, J = 2.2 Hz, 1 H), 8.89 (t, J = 2.2 Hz, 1 H), 8.85 (d, J = 2.2 Hz, 1 H), 8.08 (apparently t, J = 8.6 Hz, 2H), 7.58 (ddd, J = 8.4, 7.2, 1.2 Hz, 1 H), 7.49 58 (ddd, J = 8.4, 7.2, 1.2 Hz, 1 H), 3.42 (s, 2H), 2.73 (q, J = 7.4 Hz, 2H), 1.31 (t, J = 7.4 Hz, 3H); m / z (ESI+) 330.3 (MH+, 100%).

[0217] Compound (Ip), with IIIPAC name / V-(3-(7-bromooxazolo[4,5-b]pyridin-2-yl)phenyl)-2- (ethylthio)acetamide, was prepared from compound (He) (98 mg, 0.34 mmol) and compound (Vc) (49 mg, 0.40 mmol) following General Method C; yield 27% (35 mg, 0.090 mmol).1H NMR (400 MHz, CDCh) 6 9.03 (s, 1H), 8.38 (d, J = 5.2 Hz, 1 H), 8.35 (t, J = 2.0 Hz, 1 H), 8.08 (apparently dt, J = 8.0, 1.2 Hz, 1 H), 8.03 (ddd, J = 8.0, 2.0, 1.2 Hz, 1 H ), 7.53 (t, J = 8.0 Hz, 1 H), 7.46 (d, J = 5.2 Hz, 1H), 3.43 (s, 2H), 2.67 (q, J = 7.4 Hz, 2H), 1.32 (t, J = 7.4 Hz, 3H); m / z (ESI+) 392.2 (MH+, 85%).

[0218] Compound (Iq), with IIIPAC name / V-(3-(6-Chlorooxazolo[4,5-b]pyridin-2-yl)phenyl)-2- (ethylthio)acetamide, was prepared from compound (Ilf) (109 mg, 0.444 mmol) and compound (Vd) (64 mg, 0.53 mmol) following General Method C; yield 10% (13 mg, 0.046 mmol), white solid.1H NMR (400 MHz, CD3OD) 5 8.58 (t, J = 1.8 Hz, 1 H), 8.52 (d, J = 2.4 Hz, 1 H), 8.26 (d, J = 2.4 Hz, 1 H), 8.02 (apparently dt, J = 8.0, 1.4 Hz, 1 H), 7.83 (ddd, J = 8.0, 2.4, 1.2 Hz, 1H), 7.56 (t, J = 8.0 Hz, 1 H), 3.38 (s, 2H), 2.72 (q, J = 7.4 Hz, 2H), 1.31 (t, J = 7.4 Hz, 3H); m / z (ESI+) 348.3 (MH+, 100%).

[0219] Compound (Ir), with IIIPAC name 2-(lsopropylthio)-N-(3-(oxazolo[4,5-b]pyridin-2- yl)phenyl)acetamide, was prepared from compound (Ila) (450 mg, 2.13 mmol) and compound (Vd) (343 mg, 2.56 mmol) following General Method C; yield 22% (152 mg, 0.464 mmol), off-white solid.1H NMR (400 MHz, DMSO-cfe) 6 10.42 (s, 1 H), 8.66 (t, J = 2.2 Hz, 1 H), 8.56 (dd, J = 4.8, 1.6 Hz, 1H), 8.27 (dd, J = 8.0, 1.6 Hz, 1H), 7.95 (apparently dt, J = 8.0, 1.2 Hz, 1 H), 7.79 (ddd, J = 8.0, 2.2, 1.2 Hz, 1 H), 7.59 (t, J = 8.0 Hz, 1 H), 7.48 (dd, J= 8.0, 4.8 Hz, 1H), 3.39 (s, 2H), 3.11 (hept, J = 6.7 Hz, 1 H), 1.26 (d, J = 6.7 Hz, 6H); m / z (ESI+) 328.3 (MH+, 100%).

[0220] Compound (Is), with IIIPAC name 2-(ethylthio)-N-(3-(oxazolo[4,5-b]pyridin-2- yl)phenyl)acetamide, was prepared from compound (Ila) (470 mg, 2.22 mmol) and compound (Vc) (321 mg, 2.67 mmol) following General Method C; yield 28% (196 mg, 0.625 mmol), off-white solid.1H NMR (400 MHz, DMSO-cfe) 6 10.42 (s, 1 H), 8.65 (t, J = 2.0 Hz, 1 H), 8.56 (dd, J = 4.8, 1.6 Hz, 1H), 8.27 (dd, J = 8.0, 1.6 Hz, 1H), 7.94 (ddd, J = 8.0, 2.0, 1.0 Hz, 1H), 7.79 (ddd, J = 8.0, 2.0, 1.0 Hz, 1 H), 7.59 (t, J = 8.0 Hz, 1 H), 7.48 (dd, J = 8.0, 4.8 Hz, 1H), 3.36 (s, 2H), 2.67 (q, J = 7.4 Hz, 2H), 1.23 (t, J = 7.4 Hz, 3H); m / z (ESI+) 314.5 (MH+, 100%).

[0221] Compound (It), with IIIPAC name 2-(cyclopropylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2- yl)phenyl)acetamide, was prepared from compound (Ila) (87 mg, 0.41 mmol) and compound (Ve) (82 mg, 0.62 mmol) following General Method C; yield 20% (28 mg, 0.09 mmol).1H NMR (400 MHz, CD3OD) 5 8.58 (t, J = 1.6 Hz, 1H), 8.51 (dd, J = 4.8, 1.2 Hz, 1 H), 8.13 (dd, J = 8.0, 1.2 Hz, 1H), 8.03 (ddd, J = 8.0, 1.6, 1.2 Hz, 1 H), 7.85 (ddd, J = 8.0, 1.6, 1.2 Hz, 1H), 7.55 (t, J = 8.0 Hz, 1 H), 7.46 (dd, J = 8.0, 4.8 Hz, 1H), 3.44 (s, 2H), 2.08 (tt, J = 7.2, 4.4 Hz, 1 H), 0.91 (dt, J = 7.2, 4.4 Hz, 2H), 0.58 (dt, J = 7.2, 4.4 Hz, 2H); m / z (ESI+) 326.3 (MH+, 100%).

[0222] Example 38: Preparation process of compound (lu)

[0223] 2-Bromo- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Br1) was prepared from compound (Ila) (1.07 g, 5.07 mmol) and 2-bromoacetyl chloride (640 pl, 7.60 mmol) following General Method E; yield 19% (316 mg, 0.951 mmol), pale yellow solid.1H NMR (400 MHz, CDCI3) 6 8.52 (dd, J = 4.8, 1.2 Hz, 1 H), 8.30 (ddd, J = 2.0 Hz, 1 H), 8.04-8.00 (m, 12H), 7.90 (dd, J = 8.0, 1.2 Hz, 1 H), 7.51 (t, t, J = 8.0 Hz, 1H), 7.33 (dd, J = 8.0, 4.8 Hz, 1H), 4.17 (s, 2H); m / z (ESI+) 332.1 (MH+, 100%).

[0224] To a suspension of cyclopentanethiol (58 pL, 0.54 mmol) in anhydrous MeOH (2.7 mL) was added sodium methoxide (30% in MeOH, 110 pL, 0.542 mmol) and the resulting mixture was stirred at room temperature for 30 min. The reaction mixture was then added to a suspension of bromide (Br1) (90 mg, 0.27 mmol) in MeOH (2.7 mL) and stirring at room temperature was continued for another 2 h. The reaction mixture was concentrated under reduced pressure and purified by reverse phase column chromatography to give compound (lu), with IIIPAC name 2-(cyclopentylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2- yl)phenyl)acetamide (45 mg, 0.13 mmol, 47%), as a white solid.1H NMR (400 MHz, CD3OD) 5 8.57 (s, 1 H), 8.50 (d, J = 4.8 Hz, 1 H), 8.12 (d, J = 8.0 Hz, 1 H), 8.02 (d, J = 8.0 Hz, 1 H), 7.83 (d, J = 8.0 Hz, 1H), 7.55 (t, J = 8.0 Hz, 1 H), 7.46 (dd, J = 8.0, 4.8 Hz, 1 H), 3.41 (s, 2H), 2.20-1.93 (m, 2H), 1.84 - 1.71 (m, 2H), 1.67 - 1.46 (m, 4H); m / z (ESI+) 354.4 (MH+, 100%).

[0225] Example 39: Preparation process of compound (Iv)

[0226] Compound (Iv), with IIIPAC name 2-(methylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2- yl)phenyl)acetamide, was prepared from compound (Ila) (450 mg, 2.13 mmol) and compound (Vf) (271 mg, 2.56 mmol) following General Method C; yield 29% (184 mg, 0.62 mmol), off-white solid.1H NMR (400 MHz, DMSO-cfe) 6 10.40 (s, 1 H), 8.65 (t, J = 2.0 Hz, 1 H), 8.55 (dd, J = 4.8, 1.6 Hz, 1 H), 8.26 (dd, J = 8.0, 1.6 Hz, 1 H), 7.94 (aparently dt, J = 8.0, 1.2 Hz, 1 H), 7.80 (ddd, J = 8.0, 2.0, 1.2 Hz, 1 H), 7.59 (t, J = 8.0 Hz, 1H), 7.47 (dd, J = 8.0, 4.8 Hz, 1H), 3.32 (s, 2H), 2.20 (s, 3H); m / z (ESI+) 300.3 (MH+, 100%).

[0227] Example 40 Preparation process of compound (Iw) Compound (Iw), with IIIPAC name / V-(3-(6-chlorooxazolo[4,5-b]pyridin-2-yl)phenyl)-2- (cyclopropylthio)acetamide, was prepared from compound (Ilf) (65 mg, 0.26 mmol) and compound (Ve) (52 mg, 0.40 mmol) following General Method C; yield 10% (9 mg, 0.03 mmol), beige solid.1H NMR (400 MHz, CD3OD) 5 8.61 (t, J = 2.0 Hz, 1 H), 8.54 (d, J = 2.2 Hz, 1 H), 8.28 (d, J = 2.2 Hz, 1 H), 8.07 (ddd, J = 8.0, 2.0, 1.2 Hz, 1 H), 7.86 (ddd, J = 8.0, 2.0, 1.1 Hz, 1H), 7.58 (t, J = 8.0 Hz, 1 H), 3.44 (s, 2H), 2.08 (tt, J = 7.2, 4.4 Hz, 1H), 0.95- 0.91 (m, 1 H ), 0.62-0.58 (m, 1H); m / z (ESI+) 360.3 (MH+, 100%).

[0228] Compound (lx), with IIIPAC name / \ / -(3-(benzo[c(]oxazol-2-yl)phenyl)-2- (cyclopropylthio)acetamide, was prepared from compound (lib) (100 mg, 0.476 mmol) and compound (Vd) (82 mg, 0.62 mmol) following General Method C; yield 21% (28 mg, 0.09 mmol), beige solid. 1 H NMR (400 MHz, CD3OD) 5 8.52 (t, J = 2.0 Hz, 1 H), 7.97 (dt, J = 8.0, 1.2 Hz, 1 H), 7.79 (ddd, J = 8.2, 2.0, 1.2 Hz, 1 H), 7.75-7.72 (m, 1H), 7.69-7.64 (m, 1 H), 7.53 (t, J = 8.0 Hz, 1 H), 7.44-7.37 (m, 2H), 3.44 (s, 2H), 2.08 (tt, J = 7.4, 4.3 Hz, 1H), 1.01-0.86 (m, 2H), 0.65-0.52 (m, 2H); m / z (ESI+) 325.3 (MH+, 100%).

[0229] Compound (ly), with IIIPAC name 2-(cyclopropylthio)- / V-(3-(oxazolo[5,4-c]pyridin-2- yl)phenyl)acetamide, was prepared from compound (lid) (61 mg, 0.29 mmol) and compound (Ve) (57 mg, 0.43 mmol) following General Method C; yield 11% (10 mg, 0.03 mmol). 1 H NMR (400 MHz, CD3OD) 5 9.02 (brs, 1 H), 8.67 (t, J = 2.0 Hz, 1 H), 8.54 (d, J = 5.4 Hz, 1 H), 7.98 (ddd, J = 8.0, 2.0, 1.2 Hz, 1H), 7.75 (ddd, J = 8.0, 2.0, 1.2 Hz, 1H), 7.73 (dd, J = 5.4, 0.8 Hz, 1H), 7.59 (t, J = 8.0 Hz, 1 H), 3.44 (s, 2H), 2.08 (tt, J = 7.4, 4.4 Hz, 1 H), 0.97-0.82 (m, 2H), 0.66-0.49 (m, 2H); m / z (ESI+) 326.3 (MH+, 100%). of compound (lz'

[0230] Compound (lz), with IIIPAC name / V-(3-(3-benzyl-3 / 7-imidazo[4,5-b]pyridin-2-yl)phenyl)-2- (cyclopropylthio)acetamide, was prepared from compound (llg) (90 mg, 0.30 mmol) and compound (Ve) (59 mg, 0.45 mmol) following General Method C; yield 9% (11 mg, 0.03 mmol).1H NMR (400 MHz, CD3OD) 5 8.43 (dd, J = 4.8, 1.2 Hz, 1H), 8.13 (dd, J = 8.0, 1.2 Hz, 1 H), 8.02 (t, J = 2.0 Hz, 1 H), 7.73 (ddd, J = 8.0, 2.0, 1.2 Hz, 1 H), 7.46 (t, J = 8.0 Hz, 1 H), 7.41 (dd, J = 8.0, 4.8 Hz, 1H), 7.37 (dt, J = 8.0, 1.2 Hz, 1 H), 7.21-7.18 (m, 3H), 6.99- 6.96 (m, 2H), 5.69 (s, 2H), 3.39 (s, 2H), 2.02 (tt, J = 7.4, 4.4 Hz, 1H), 0.93-0.81 (m, 2H), 0.58-0.51 (m, 2H); m / z (ESI+) 415.4 (MH+, 100%).

[0231] Compound (laa), with IIIPAC name / V-(5-(benzo[d]oxazol-2-yl)pyridin-3-yl)-2- (ethylthio)acetamide, was prepared from compound (Hi) (75 mg, 0.36 mmol) and compound (Vc) (43 mg, 0.36 mmol) following General Method D; yield 8% (9 mg, 0.03 mmol). 1 H NMR (400 MHz, CD3OD) 5 9.09 (d, J = 2.0 Hz, 1 H), 8.97 (t, J = 2.0 Hz, 1 H), 8.88 (d, J = 2.0 Hz, 1 H), 7.80-7.77 (m, 1 H), 7.73-7.71 (m, 1 H), 7.47 (td, J = 7.2, 1.6 Hz, 1 H), 7.43 (td, , J = 7.2, 1.6 Hz, 1H), 3.42 (s, 2H), 2.73 (q, J = 7.4 Hz, 2H), 1.31 (t, J = 7.4 Hz, 3H); m / z (ESI+) 314.3 (MH+, 100%).

[0232] Compound (Ibb), with IIIPAC name / \ / -(5-(benzo[c(]oxazol-2-yl)pyridin-3-yl)-2- (cyclopropylthio)acetamide, was prepared from compound (Hi) (75 mg, 0.36 mmol) and compound (Ve) (47 mg, 0.36 mmol) following General Method D; yield 11% (13 mg, 0.04 mmol).1H NMR (400 MHz, CD3OD) 5 9.10 (d, J = 2.0 Hz, 1H), 8.99 (t, J = 2.0 Hz, 1 H), 8.90 (d, J = 2.0 Hz, 1 H), 7.79-7.77 (m, 1 H), 7.73-7.70 (m, 1 H), 7.47 (td, J = 7.4, 1.4 Hz, 1 H), 7.47 (td, J = 7.4, 1.2 Hz, 1H), 3.47 (s, 2H), 2.09 (td, J = 7.2, 4.4 Hz, 1 H), 0.98-0.89 (m, 2H), 0.62-0.54 (m, 2H); m / z (ESI+) 326.3 (MH+, 100%).

[0233] Compound (Icc), with IIIPAC name / V-(3-(6-Bromooxazolo[4,5-b]pyridin-2-yl)phenyl)-2- (ethylthio)acetamide, was prepared analogously to compound (Ip), following General Method C, but using isomer 2-amino-5-bromopyridin-3-ol (IVb), as disclosed in Example 5, as starting product.1H NMR (400 MHz, CD3OD) 5 8.61 (d, J = 2.0 Hz, 1 H), 8.58 (t, J = 1.8 Hz, 1 H), 8.39 (d, J = 2.0 Hz, 1 H), 8.07 - 7.99 (m, 1 H), 7.87 - 7.79 (m, 1 H), 7.56 (t, J = 8.0 Hz, 1 H), 3.38 (s, 2H), 2.72 (q, J = 7.4 Hz, 2H), 1.31 (t, J = 7.4 Hz, 3H); m / z (ESI+) 392.2 (MH+, 85%).

[0234] Compound (lea), with IIIPAC name / \ / -(5-(benzo[d]oxazol-2-yl)-2-chlorophenyl)-2- (ethylthio)acetamide, was prepared from compound (Hm) (32 mg, 0.13 mmol) and compound (Vea) (27 mg, 0.20 mmol) following General Method E; yield 23% (10 mg, 0.03 mmol).1H NMR (400 MHz, CDCI3) 5 9.59 (brs, 1 H), 9.29 (d, J = 2.0 Hz, 1H), 8.02 (dd, J = 8.4, 2.0 Hz, 1 H), 7.82-7.77 (m, 1H), 7.63-7.61 (m, 1H), 7.57 (d, J = 8.4 Hz, 1 H), 7.42-7.35 (m, 2H), 3.51 (s, 2H), 2.72 (q, J = 7.4 Hz, 2H), 1.36 (t, J = 7.4 Hz, 3H).

[0235] Compound (Iva), with IIIPAC name / \ / -(2-chloro-5-(oxazolo[4,5-b]pyridin-2-yl)phenyl)-2- (ethylthio)acetamide, was prepared from compound (Ilk) (177 mg, 0.72 mmol) and compound (Vea) (144 mg, 1.04 mmol) following General Method E; yield 24% (57 mg, 0.16 mmol).1H NMR (400 MHz, CDCI3) 5 9.45 (s, 1H), 9.15 (d, J = 2.0 Hz, 1 H), 8.44 (dd, J = 5.0, 1.6 Hz, 1 H), 7.93 (dd, J = 8.4, 2.0 Hz, 1 H), 7.74 (dd, J = 8.1 , 1.6 Hz, 1 H), 7.42 (d, J = 8.4 Hz, 1H), 7.17 (dd, J = 8.1, 5.0 Hz, 1H), 3.34 (s, 2H), 2.54 (q, J = 7.4 Hz, 2H), 1.18 (t, J = 7.4 Hz, 3H).

[0236] Compound (leb), with IIIPAC name 2-(ethylthio)- / V-(3-(7-hydroxybenzo[d]oxazol-2- yl)phenyl)acetamide, was prepared from compound (I In) in two steps:

[0237] Step 1 : Compound (lin) (70 mg, 0.31 mmol) and compound (Vc) (64 mg, 0.46 mmol) following General Method C; yield 35% (46 mg, 0.11 mmol) of 2-(3-(2- (ethylthio)acetamido)phenyl)benzo[d]oxazol-7-yl 2-(ethylthio)acetate.

[0238] 1H NMR (400 MHz, CDCI3) 6 8.87 (s, 1 H), 8.16 (t, J = 1.0 Hz, 1H), 7.93 (dd, J = 8.0, 1.0 Hz, 1H), 7.90 (dt, J = 7.6, 1.0 Hz, 1H), 7.59 (dd, J = 8.0, 1.0 Hz, 1H), 7.43 (t, J = 8.0 Hz, 1 H), 7.28 (t, J = 8.0 Hz, 1H), 7.09 (dd, J = 8.0, 1.0 Hz, 1 H), 3.53 (s, 2H), 3.34 (s, 2H), 2.81 (q, J = 7.4 Hz, 2H), 2.58 (q, J = 7.4 Hz, 2H), 1.33 (t, J = 7.4 Hz, 3H), 1.25 (t, J = 7.4 Hz, 3H).

[0239] Step 2: To a solution of 2-(3-(2-(ethylthio)acetamido)phenyl)benzo[d]oxazol-7-yl 2- (ethylthio)acetate (40 mg, 0.09 mmol) in THF (1 mL), LiOH 1 N (186 .L, 0.18 mmol) was added and stirred at r.t. for 6 h. Then THF was evaporated and the aqueous solution was purified by biotage, reverse phase, yield 15% (5 mg, 0.14 mmol)

[0240] 1H NMR (400 MHz, CDCI3) 6 8.86 (s, 1 H), 8.17 (t, J = 2.0 Hz, 1 H), 7.85 (dt, J = 8.0, 1.2 Hz, 1H), 7.76 (ddd, J = 8.0, 2.0, 1.0 Hz, 1 H), 7.33 (t, J = 8.0 Hz, 1 H), 7.18 (dd, J = 8.0, 1.0 Hz, 1H), 7.06 (t, J = 8.0 Hz, 1 H), 6.79 (d, J = 8.0 Hz, 1 H), 3.30 (s, 2H), 2.53 (q, J = 7.4 Hz, 2H), 1.18 (t, J = 7.4 Hz, 3H).

[0241] Compound (lec), with IIIPAC name 2-(ethylthio)-N-(3-(4-hydroxybenzo[d]oxazol-2- yl)phenyl)acetamide, was prepared from compound (Ho) (100 mg, 0.44 mmol) and compound (Vc) (92 mg, 0.66 mmol) following General Method C; yield 41% (61 mg, 0.18 mmol).

[0242] 1H NMR (400 MHz, CDCI3) 6 7.48 (ddd, J = 7.8, 1.6, 0.8 Hz, 1H), 7.42 (dd, J = 1.6, 0.8 Hz, 1 H), 7.34 (dd, J = 8.0, 0.8 Hz, 1 H), 7.20 (t, J = 8.0 Hz, 1H), 7.15 (t, J = 8.0 Hz, 1 H), 7.00 (dd, J = 8.0, 0.8 Hz, 1 H), 6.71 (ddd, J = 8.0, 2.0, 0.8 Hz, 1 H), 3.49 (s, 3H), 2.77 (q, J = 7.4 Hz, 2H), 1.26 (t, J = 7.4 Hz, 3H).

[0243] Example 51 : Preparation process of compound (led)

[0244] Compound (led), with IIIPAC name / \ / -(3-(benzo[d]oxazol-2-yl)-4-hydroxyphenyl)-2- (ethylthio)acetamide, was prepared from compound (llj) (100 mg, 0.44 mmol) and compound (Vc) (64 mg, 0.53 mmol) following General Method C; yield 56% (82 mg, 0.03 mmol).1H NMR (400 MHz, CDCI3) 5 8.70 (brs, 1 H), 8.25 (d, J = 2.8 Hz, 1 H), 7.68-7.62 (m, 1 H), 7.55-7.51 (m, 1 H), 7.45 (dd, J = 8.8, 2.8 Hz, 1 H), 7.34-7.27 (m, 2H), 7.03 (d, J = 8.8 Hz, 1 H), 3.35 (s, 2H), 2.60 (q, J = 7.4 Hz, 2H), 1.26 (t, J = 7.4 Hz, 3H).

[0245] Example 52: TSPAN1 inhibition sensitize chemotherapy resistant cancers and blocks tumoriqenic signaling pathways

[0246] Of the 4 HNSCC models available in the laboratory (JHLI029, HTB-43, CCL-138 and SCC-25), TSPAN1 is most expressed in the CCL-138 cell line (FIG. 1A). Furthermore, the overexpression of TSPAN1 can be corroborated in the resistant lines as well as in CSC (generation 2 and 3 of HTB-43) (FIG.1 B). Phosphorylation of Src kinase (p-Src) is a signaling pathway of chemotherapy resistance and it has been observed that, as a result of TSPAN1 inhibition, not only p-Src, but also the activation of other signaling pathways, such as protein kinase B (AKT) and mitogen-activated protein kinase (ERK), are blocked. FIG. 2 shows these results, hence confirming that inhibition of TSPAN1 affects different signal cascades with Src as a central node. Inhibition of chemoresistance pathways in cancerous cells represents an important advantage and progress in the management of cancer, given that this is one of the main obstacles in cancer treatment and would ameliorate prognosis, life expectancy and life quality of patients.

[0247] Example 53: In vivo studies showed that TSPAN1 inhibition significantly reduced tumor size

[0248] An immunosuppressed mouse model (NMRI-FOXn1 nu / nu strain) was used to demonstrate that TSPAN1 absence diminishes proliferation in vivo. JHLI029 and JHU029- R cells transduced with a small RNA (siRNA) fragment inhibiting TSPAN1 (siTSPANI) or siRNA negative controls (NC) were subcutaneously injected. A significant reduction in tumor size was observed in the TSPANI-inhibited groups: JHLI029 NC vs siTSPANI (days 10-28) and JHU029-R NC vs siTSPANI (days 12-41) (FIG. 3). Tumor size was also compared between groups at the end point, showing that in both cases (parental (P) cells and CDDP-R cells) tumor size was reduced when TSPAN1 was inhibited (FIG. 4). Phenotypic characteristics and proliferative potential of the tumors formed in mice were also observed, showing similarities to human HNSCC. Hence, the correlation between TSPAN1 inhibition and halt of tumor progression, which is part of the state of the art, is confirmed with these in vitro results obtained by the inventors, meaning that TSPAN1 is a valuable target for drug development in cancer treatment, in particular HNSCC.

[0249] Example 54: In vitro studies of compound (If)

[0250] In vitro studies were performed to evaluate the cytotoxic effects at 72h of the compounds of the invention. Three HNSCC cell lines (JHLI029, HTB-43 and CCL-138) were used, including both parental cells and CDDP-resistant cells (R), to determine the IC50 of compound (If) with the MTS method in p96 plates as described in Riss TL, et al. (in “Cell Viability Assays”, 2013 [Updated 2016 Jul 1], in: Markossian S., et al., “Assay Guidance Manual” [Internet], Bethesda (MD), Eli Lilly & Company and the National Center for Advancing Translational Sciences, 2004), (see Table 1).

[0251] Table 1: IC50 values of compound (If) in different HNSCC cell lines

[0252] As can be seen in Table 1 , results for IC50 of compound (If) in CCL-138 cells are higher, indicating a general resistance to treatment, probably given that this cell line comes from metastasis. Thus, the use of this compound represents a path for progress in the treatment of cancer, including resistant and metastatic cells.

[0253] Example 55: Western blot (WB) assays under compound (If) effect in HNSCC cell lines at three different times

[0254] WB assays were pursued on compound (If), showing effect over TSPAN1 in a short period of time after exposition (8 h). Thus, surprisingly, TSPAN1 inhibition by this compound seems a direct effect and not a collateral consequence of a possible unspecific action of this compound. The fact that, not only does this compound show activity against TSPAN1 , but moreover, does not require long periods of exposure to achieve the desired effect, represents a great advantage in terms of reducing possible side effects related to drug administration. This will have an impact on quality of life of patients suffering from cancer, in comparison to current methods of treatment. FIG. 5 is a graphical representation of IC50 results of compound (If) against TSPAN1 for the cell lines JHLI029 and JHU029-R at 72h by MTS assay, as described in Riss TL, et al. (in “Cell Viability Assays”, 2013 [Updated 2016 Jul 1], in: Markossian S., et al., “Assay Guidance Manual” [Internet], Bethesda (MD), Eli Lilly & Company and the National Center for Advancing Translational Sciences, 2004). WB analysis mentioned above shows that TSPAN1 inhibition, as well as inhibition of active form of Src and ERK (p-Src y p-ERK), is achieved with compound (If) on HNSCC cell lines JHU029, HTB-43 and CCL-138, both parental and resistant to CDDP (R), at 8h and 24h after exposure to compound (If) at 2 times the IC50. Moreover, apoptosis marker PARP1 is also cleaved at 8 and 24 h in HTB- 43-P cells (FIG. 6). These effects were concentration dependent, and this was reflected in the morphology and proliferation of cells after 24hours of exposure (see FIG. 7).

[0255] Example 56: In vitro toxicological studies on non-tumorous and other HNSCC cell lines

[0256] In vitro toxicological studies and WB tests were performed on non-tumorous cell lines that express TSPAN1 IMR90 and a cell line from HNSCC that doesn’t express TSPAN1 (RPMI). The aim of these studies was to characterize the effect of the compounds of the invention on these non-cancerous cell lines and in absence of TSPAN1. Compound (If) showed no toxicologic effect on the cell line coming from non-cancerous cells for the concentration range studied. The fact that the compounds of the invention have effect against cancer cells but are not toxic for non-cancerous cells is an advantage in view of considering their use for human cancer treatment (see FIG. 8). It is also shown that in the absence of TSPAN1 the IC50 increases, so it is not as cytotoxic without its target, suggesting target specificity.

[0257] Example 57: In vivo toxicological studies

[0258] In vivo toxicological studies with the compounds of the invention were conducted on mice, at different concentrations of drug (20, 40 and 80mg / Kg), based on IC50 results obtained. The vehicle used in all treatments was PBS + 12,5% chromophore + 5% DMSO and the treatment were administrated via intraperitoneal injection, 3 times a week for a total of 4 weeks. Possible toxicological effects were evaluated by means of visual examination of pain and weight loss in the mice. Samples of serum, spleen, liver, lungs, and kidneys were extracted from the dead animals (sacrificed under an overdose of 5% deep anesthesia) to evaluate possible toxicity by means of histological technigues and molecular biology. No differences in animal behavior, general weight, nor relative weight of organs were remarked when the experimental group was compared to the control group (treated solely with the vehicle). No damage was observed in the liver, spleen, brain, kidneys in histological samples (H&E staining) and neither at the level of biochemical parameters in mice serum samples. These results hence confirm that the compounds of the invention, surprisingly, not only show activity against cancerous cells expressing TSPAN1 , including resistant and metastatic cells, but moreover they are safe to use for treatment and do not incur in side effects.

[0259] Example 58: In vivo experiments to show antitumoral efficacy on JHU029-R cells

[0260] To show antitumoral efficacy of the compounds of the invention, an intermediate concentration of compound (If) was used. 1x106cancerous CDDP resistant cells (JHU029-R) that overexpress TSPAN1 were injected subcutaneously on the side of the mouse, allowing tumor growth for approximately 7-9 days before starting the treatment (40mg / kg). Necropsies of the animals were performed once treatment had finished, extracting samples of the spleen, kidneys, liver, lungs, and serum, as well as extracting the tumor. FIG. 9 shows that tumor volume was significantly reduced in the experimental group after 2-3 weeks of treatment, when compared to the control group. Moreover, by the end of the study, total size (FIG. 10) and weight (FIG. 11) of the tumor were also significantly reduced in the experimental group, in comparison to the control group.

[0261] Hence, these in vivo results confirm that the compounds of the invention not only show the desired activity, but moreover, the short period of time needed to achieve desired results also represents a significant advantage.

[0262] Example 59: Drug levels in tumor, liver and serum detected by HPLC technique

[0263] HPLC technique was used to detect drug levels at end point in tumor, liver, and serum in 3 mice of each group (control and experimental). A final dose of treatment was injected in these animals 2h before euthanasia was conducted. The presence of compound (If) in serum and tumor indicates that it is distributed correctly and that it effectively reaches the tumor. In this way we can conclude that the reduction in size and weight is due to the presence of the drug and not to indirect / unspecific effects of the compound (If), mentioned in Example 47, in the experimental group. Drug levels were also reported in liver tissue, as expected, given that the drug is injected in peritoneal zone and, furthermore, the liver has metabolic functions. The variability is due to the heterogeneity of the in vivo model and the quantity is not as important as the mere presence of the compound in the tumor and serum samples.

[0264] Table 2: Drug concentration at end point in tumor, liver and serum detected by HPLC

[0265] Example 60: Combination of the compound of the invention with CDDP

[0266] Synergistic effect of compound (If) with CDDP was evaluated by conducting several in vitro drug combination studies, including the study of the combinatorial index (Cl) in constant and non-constant ratios. Subsequently, expression of several markers, including TSPAN1 , was evaluated by WB test at the established concentrations.

[0267] Cell line JHLI029 and JHU029-R were exposed to increasing concentrations of one of the drugs (compound (If), for example), together with the other drug (CDDP) at fixed IC50 concentration, and vice versa. Results obtained were analyzed using isobole method, and synergy was calculated using the multi-drug effect equation and were quantified by means of combination index (Cl) (calculated using CompuSy (ComboSyn Inc., NJ) software).

[0268] Tables 3 and 4 show Cl results for non-constant ratio combinations of compound (If) at fixed IC50 concentration and CDDP at different concentrations, and vice versa, respectively in JHU029-R. As can be read in the tables, synergy (Cl<1) of the combination of drugs was confirmed by the Cl results, meaning that treatment could be improved by such combination.

[0269] FIG. 12 shows a graphic representation of the effects of CDDP (0,5|JM, 1 ,75|JM and 2,6pM), compound (If) (26pM, 88|JM and198|jM), and the combination of both on the cell line JHU029-R, showing synergistic effect of the combination. Finally, a WB assay was performed with the obtained ratio (1 ,8:444 pM CDDP: compound (If)) and the most adequate concentration (one time the ratio and two times), in order to evaluate the expression of aforementioned markers (p-ERK, p-Src and TSPAN1), as well as other markers that will indicate the effect of CDDP on cells. FIG. 13 shows the corresponding WB assay, where inhibition of TSPAN1 , p-Src and p-ERK both in cell lines JHLI029 and JHU029-R (resistant to CDDP) is increased by the combination of compound (If) and CDDP when compared to CDDP alone. In addition, CD44 (a classic CSC marker in HNSCC and other types of cancers) also significantly reduced its expression with both compound (If) concentrations and with their respective combinations with CDDP.

[0270] FIG. 14 shows microphotographies that represent the effect of different treatment conditions: control (DMSO), CDDP alone, compound (If) alone and combination of compound (If) and CDDP, on the proliferation of JHU029-R cells. All three treatment options were evaluated at two different concentrations and, as can be seen in the microphotographies, proliferation is proportionally reduced with increasing concentrations, but most importantly, it is significantly reduced with compound (If) in comparison to CDDP (probably due to this cell line’s resistance to CDDP) and even more reduced with the combination of both drugs, as suggested by the Cl synergistic results from Tables 3 and 4. The fact that both drugs are used in combination represents an advantage, since when using each drug individually the concentrations are higher (than when using them in combination with each other) and therefore the side effects associated with each drug individually are reduced (this is very significant in the case of CDDP).

[0271] Table 3: Cl Data for non-constant combination of compound (If) at IC50 concentration and CDDP on cell line JHU029-R

[0272] Table 4: Cl Data for non-constant combination of CDDP at IC50 concentration and compound (If) on cell line JHU029-R

[0273] Example 61 : In vitro tests of compounds (Ir) and (Is) of the invention

[0274] Compounds (Ir) and (Is), which are structurally similar to compound (If), were developed and synthesized. Both (Ir) and (Is) showed adequate results when compared to compound (If) at 350|JM regarding TSPAN1 and p-Src inhibition, in both cell lines JHU029-R and CCL-138-R (FIG. 15). In addition, compound (Is) shows a proliferation inhibition when compared with non-treated (control, DMSO or NC) cells at 150|JM (FIG. 16).

[0275] Example 62: In vivo toxicological and effectivity studies of compounds (Ir) and (Is) of the invention

[0276] Toxicological studies for compounds (Ir) and (Is) were performed in the same manner as in Example 46, this time at two concentrations (40 and 80mg / kg). No precipitation of the drug nor inflammatory tissue around it was observed when histological samples were analyzed, even at the highest concentration. Histological and molecular markers did not indicate any possible toxicological effect produced by these two compounds of the invention.

[0277] Results for effectivity studies showed that compound (Is) produced a similar therapeutic effect to compound (If) over time (FIG. 17), due to their structural similarity. Weight (FIG. 18) and size (FIG. 19) of the respective tumors at end point of treatment were compared and conclusions drawn from FIG. 17 were confirmed. It is also shown the H&E staining and the proliferation marker Ki67, that is reduce in both treatments compare to the control (FIG. 20).

[0278] Both in vitro and in vivo results with compound (Is), structurally similar to compound (If), show that the compounds of the present invention, by virtue of their chemical structure and hence interaction with TSPAN1, represent a path for progress in the treatment of cancers that overexpress TSPAN1. They not only show desired activity, but have moreover demonstrated to be safe, given that no toxicity nor side effects were reported.

[0279] Example 63 Cytotoxic effect of 11 compounds of the invention, at 100uM, in the HNSCC cell lines JHU029 and JHU029-R

[0280] Figure 21 shows two graphs, in which 11 compounds of the invention (namely (lb), (Ic), (Id), (le), (Ig), (Ih), (Ik), (lu), (lx), (Iz), and (Icc)) were tested against HNSCC cell lines JHLI029 and JHU029-R. The survival effect was measured at 72h after treatment withlOOpM of each compound by MTS assay, as described in Riss TL, et al. (in “Cell Viability Assays”, 2013 [Updated 2016 Jul 1], in: Markossian S., et al., “Assay Guidance Manual” [Internet], Bethesda (MD), Eli Lilly & Company and the National Center for Advancing Translational Sciences, 2004). As can be seen, these compounds of the invention show cytotoxic effect in these cell lines, like (If) and (Is) which have been disclosed in previous examples.

[0281] Example 64: In vitro toxicological studies and WB tests performed in cancer cell lines of hepatocellular carcinoma, breast cancer, gastric and ovarian adenocarcinoma, and osteosarcoma.

[0282] In vitro toxicological studies and WB test were performed in the following cancer cell lines: Hep27 (hepatocellular carcinoma), MDA-MD-231 (breast cancer), MKN-45 (gastric adenocarcinoma), SK-OV-3 (ovarian adenocarcinoma), U-2-OS and MG-63 (osteosarcoma). The cell lines were cultured in DM EM F-12 medium (Gibco, Thermo Fisher Scientific, Waltham, MA, USA) with 10% FBS (Biowest, Nuaille-France), 1% L- glutamine (200 mM, Gibco, Thermo Fisher Scientific, Waltham, MA, USA), and 1% penicillin / streptomycin (Pen 20 U / mL and Strep 20 pg / mL, Gibco, Thermo Fisher Scientific, Waltham, MA, USA).

[0283] These cancer models have been described, as mentioned above, to have a relation with TSPAN1 , and thus the inventors have pursued these studies to further characterize the effect of compound (Is) in these cancers. The IC50 value was determined at 72h by the MTS assay in the six above-cited cancer cell lines and a efficacy of the compound against them was observed in all models. Upon treating the cells with the determined IC50 for 24 hours, a decrease in TSPAN1 expression and in the active form of Src was advantageously noted. These results, shown in FIG. 22 and 23, support that the use of the compounds of the invention may be extended to the treatment of further cancer models where TSPAN1 plays a significant role. Citation List

[0284] Non-Patent Literature

[0285] Chen, L., et al., “Clinicopathological significance of overexpression of TSPAN1 , Ki67 and CD34 in gastric carcinoma”, Tumori 94 (2008), 531-538.

[0286] Chen, L., et al., “TSPAN1 protein expression: a significant prognostic indicator for patients with colorectal adenocarcinoma”, World J. Gastroenterol, 15 (2009), 2270.

[0287] Duan, J., et al., “miR-491-3p suppresses the growth and invasion of osteosarcoma cells by targeting TSPAN1”, Molecular medicine reports 16 (2017), 5568-5574.

[0288] Garcia-Mayea Y., et al., “TSPAN1 : A Novel Protein Involved in Head and Neck Squamous Cell Carcinoma Chemoresistance”, Cancers, vol. 12 (2020), Article number 3269.

[0289] Gu, T., et al., “Expression and function of tetraspanin 1 in esophageal carcinoma”, Oncology letters 14 (2017), 6815-6822.

[0290] Holters, S., et al., “Tetraspanin 1 promotes invasiveness of cervical cancer cells”, Int. J. Oncol. 43 (2013), 503-512.

[0291] Hou, F.Q., et. al., “Tetraspanin 1 is involved in survival, proliferation and carcinogenesis of pancreatic cancer”, Oncol Rep 34 (2015), 3068-3076S.

[0292] Huang R., et al., “The role of tetraspanins pan-cancer”, iScience, vol. 25(8), (2022), 104777.

[0293] Ji, Z.J., Wang, et al., “Inhibition of skin squamous cell carcinoma proliferation and promote apoptosis by dual silencing of NET-1 and surviving”, Oncol Rep 34 (2015), 811-822.

[0294] Munkley, J., et al., “The cancer-associated cell migration protein TSPAN1 is under control of androgens and its upregulation increases prostate cancer cell migration”, Sci Rep 7 (2017), 5249.

[0295] Riss TL, et al. “Cell Viability Assays”, 2013 [Updated 2016 Jul 1], in: Markossian S, et al., “Assay Guidance Manual” [Internet], Bethesda (MD), Eli Lilly & Company and the National Center for Advancing Translational Sciences, 2004. Available from: https: / / www.ncbi.nlm.nih.gov / books / NBK144065 / Scholz, C.J., et al., “Tspan-1 is a tetraspanin preferentially expressed by mucinous and endometrioid subtypes of human ovarian carcinomas”, Cancer Lett 275 (2009), 198-203.

[0296] Subrungruanga, I., et al., “Gene expression profiling of intrahepatic cholangiocarcinoma. Asian Pacific journal of cancer prevention”, APJCP 14 (2013), 557-563. Tian, J., et al., “Silencing Tspanl inhibits migration and invasion, and induces the apoptosis of human pancreatic cancer cells”, Molecular medicine reports 18, (2018), 3280-3288.

[0297] Wang, G.L., et al., “The effect of NET-1 on the proliferation, migration and endocytosis of the SMMC-7721 HCC cell line”, Oncol Rep 27, (2012), 1944-1952. Wang, L., et al., “Cisplatin-enriching cancer stem cells confer multidrug resistance in nonsmall cell lung cancer via enhancing TRIB1 / HDAC activity”, Cell Death Dis 8 (2017), e2746.

[0298] Wang Y., et al., “Tetraspanin 1 promotes epithelial-to-mesenchymal transition and metastasis of cholangiocarcinoma via PI3K / AKT signaling”, Journal of Experimental & Clinical Cancer Research, vol. 37, (2018), Article number 300.

[0299] Zhang, G.-L., et al., “The transcriptome difference between colorectal tumor and normal tissues revealed by single-cell sequencing”, J. Cancer 10 (2019), 5883-5890.

Claims

Claims1 . A compound of formula (I) or a pharmaceutically acceptable salt thereof, for use in the treatment of a cancer expressing TSPAN1 , in a mammal, including a human,Formula (I) wherein:Ri is a radical selected from (Ci-C4)-alkyl, (C3-C6)-cycloalkyl, phenyl, and chlorophenyl;A is a biradical selected from S, N and C;Cy is a biradical selected from:m is an integer from 0 to 2; n is an integer from 0 to 2; o is an integer from 0 to 1 ; p is an integer from 0 to 1 ; q is an integer from 0 to 1 ;X is selected from O, S, N,and NH;Y is selected from C and N;Z is selected from C and N;R2, R3, R4, and Rs are selected from H, halogen, OH, (Ci-Cs)-alkyl, and O-(Ci-Cs)-alkyl;Re is selected from H and (Ci-C4)-alkyl-phenyl; andR7-R20 is selected from H, halogen, OH, (Ci-Ce)-alkyl, and O-(Ci-Ce)-alkyl, with the proviso that: when (a) is a double bond then (b) is a single bond, and when (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond; X is selected from O, S, and NH; when (b) is a double bond, p is 0; when (b) is a single bond, p is 1 ; when Y is N, q is 0; when Z is N, o is 0; and when Z is C, o is 1.

2. The compound for use according to claim 1 wherein the cancer expressing TSPAN1 is selected from the group consisting of pancreatic cancer, colon cancer, gastric cancer, colorectal cancer, esophageal cancer, ovarian cancers, endometrial cancer, hepatocellular carcinoma, prostate cancer, osteosarcoma, non-small cell lung cancer, skin squamous cell carcinoma, cholangiocarcinoma, cervical intraepithelial neoplasia and neck squamous cell carcinoma.

3. The compound for use according to claim 2, wherein the cancer expressing TSPAN1 is selected from head and neck squamous cell carcinoma, hepatocellular carcinoma, breast cancer, gastric and ovarian adenocarcinoma, and osteosarcoma.

4. The compound for use according to claims 1-3, wherein each one of the radicals R4, Rs, and R7-20 is H.

5. The compound for use according to claims 1-4, wherein each one of the radicals R2 and R3 is selected from H and halogen.

6. The compound for use according to claims 1-5, wherein R1 is a radical selected from (Ci-C4)-alkyl, (C3-Cs)-cycloalkyl, phenyl, and chlorophenyl.

7. The compound for use according to claims 1-6, wherein Y is N.

8. The compound for use according to claims 1-7, wherein Z is C.

9. The compound for use according to claim 1 , which is 2-((4-chlorophenyl)thio)- / V-(3- (oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide.

10. The compound for use according to claims 1-8, which is administered in combination with a chemotherapeutic agent.

11. A compound of formula (I) or a pharmaceutically acceptable salt thereof,Formula (I) wherein:Ri is a radical selected from (Ci-C4)-alkyl, and (C3-C6)-cycloalkyl;A is S;Cy is a biradical selected from:m is 1 ; n is an integer from 0 to 2; o is an integer from 0 to 1 ;p is an integer from 0 to 1 ; q is an integer from 0 to 1 ;X is selected from O, S, N;Y is selected from CH and N;Z is selected from C and N;R2 is selected from H, and halogen;Ra is selected from H, halogen, and OH;R4 is H;Rs is selected from H, halogen, and OH;Re is selected from H and (Ci-C4)-alkyl-phenyl; andR7-R20 is selected from H, halogen, OH; with the proviso that: when (a) is a double bond then (b) is a single bond, and when (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond; X is selected from O, S, and NH; when (b) is a double bond, p is 0; when (b) is a single bond, p is 1; when Y is N, q is 0; when Z is N, o is 0; and when Z is C, o is 1 ; and with the proviso that: when Cy is phenyl and n is 0, R1 is other than methyl; when Cy is phenyl, n is 0 and X is O, R1 is other than ethyl or cyclopentyl; when Cy is phenyl, n is 0 and X is NH, R1 is other than isopropyl or 2-methylpropyl; and when Cy is phenyl, n is 0 and X is S, R1 is other than tert-butyl or cyclopentyl.

12. The compound of formula (I) according to claim 11 which is a compound of formula(h) or a pharmaceutically acceptable salt thereof of formula (h),Formula (h) wherein:R1 is a radical selected from (Ci-C4)-alkyl and (C3-C6)-cycloalkyl;Cy is a biradical selected from:n is an integer from 0 to 2; o is an integer from 0 to 1 ; p is an integer from 0 to 1 ;X is selected from O, S and N;Y is selected from CH and N;Z is selected from C and N;R2 and R3are selected from H and halogen;Re is selected from H and (Ci-C4)-alkyl-phenyl; and R7-R20 are H, with the proviso that: when (a) is a double bond then (b) is a single bond, and when (a) is a single bond then (b) is a double bond; when (a) is a double bond, X is N; when (a) is a single bond, X is selected from O, S, and NH; when (b) is a double bond, p is 0; when (b) is a single bond, p is 1 ; when Z is N, o is 0; and when Z is C, o is 1 , and with the proviso that: when Cy is phenyl and n is 0, R1 is other than methyl; when Cy is phenyl, n is 0 and X is O, R1 is other than ethyl or cyclopentyl;when Cy is phenyl, n is 0 and X is NH, R1 is other than isopropyl or 2-methylpropyl; and when Cy is phenyl, n is 0 and X is S, R1 is other than tert-butyl or cyclopentyl.

13. The compound according to claim 12, wherein: n is an integer from 0 to 2; o is 1 ;P is 0;(a) is a single bond;(b) is a double bond;X is O;Y is N;Z is C;Cy is phenyl;Ri is a radical selected from (Ci-C4)-alkyl and (C3-C6)-cycloalkyl; andR2 and R3are selected from H and halogen.

14. The compound according to claim 12, wherein: n is an integer from 0 to 2; o is 0;P is 0;(a) is a single bond;(b) is a double bond;X is O;Y is N;Z is C;Cy is phenyl;Ri is a radical selected from (Ci-C4)-alkyl and (C3-C6)-cycloalkyl; and R2is H.

15. The compound according to claim 12, which is selected from the following list: a) / V-(3-(3-benzyl-3 / 7-imidazo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Ig); b) 2-(ethylthio)- / V-(3-(oxazolo[5,4-c]pyridin-2-yl)phenyl)acetamide (Ih); c) 2-(ethylthio)- / V-(5-(oxazolo[4,5-b]pyridin-2-yl)oxazol-2-yl)acetamide (li); d) 2-(ethylthio)- / V-(5-(oxazolo[4,5-b]pyridin-2-yl)thiazol-2-yl)acetamide (Ij); e) / V-(5-(benzo[d]thiazol-2-yl)pyridin-3-yl)-2-(ethylthio)acetamide (Ik); f) / V-(5-(1 / 7-benzo[d]imidazol-2-yl)pyridin-3-yl)-2-(ethylthio)acetamide (Im); g) 2-(ethylthio)- / V-(2-(oxazolo[4,5-b]pyridin-2-yl)pyridin-4-yl)acetamide (In); h) 2-(ethylthio)- / V-(2-(oxazolo[4,5-b]pyridin-2-yl)pyrimidin-4-yl)acetamide(lo);i) / V-(3-(7-bromooxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Ip); j) / V-(3-(6-chlorooxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Iq); k) 2-(isopropylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Ir); l) 2-(ethylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Is); m) 2-(cyclopropylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (It); n) 2-(cyclopentylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (lu), and o) 2-(methylthio)- / V-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide (Iv), p) / V-(3-(6-chlorooxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(cyclopropylthio)acetamide (Iw); q) / \ / -(3-(benzo[c(]oxazol-2-yl)phenyl)-2-(cyclopropylthio)acetamide (lx); r) 2-(cyclopropylthio)-N-(3-(oxazolo[5,4-c]pyridin-2-yl)phenyl)acetamide (ly); s) / V-(3-(3-benzyl-3 / 7-imidazo[4,5-b]pyridin-2-yl)phenyl)-2-(cyclopropylthio)acetamide (Iz); t) / \ / -(5-(benzo[c(]oxazol-2-yl)pyridin-3-yl)-2-(ethylthio)acetamide (laa), and u) / \ / -(5-(benzo[c(]oxazol-2-yl)pyridin-3-yl)-2-(cyclopropylthio)acetamide (Ibb).

16. The compound of formula (I) according to claim 11, which is selected from the following list:- N-(5-(benzo[d]oxazol-2-yl)-2-chlorophenyl)-2-(ethylthio)acetamide (lea),- N-(2-chloro-5-(oxazolo[4,5-b]pyridin-2-yl)phenyl)-2-(ethylthio)acetamide (Iva),- 2-(ethylthio)-N-(3-(7-hydroxybenzo[d]oxazol-2-yl)phenyl)acetamide, (leb),- 2-(ethylthio)-N-(3-(4-hydroxybenzo[d]oxazol-2-yl)phenyl)acetamide (lec), and- N-(3-(benzo[d]oxazol-2-yl)-4-hydroxyphenyl)-2-(ethylthio)acetamide (led).

17. A pharmaceutical composition comprising a therapeutically effective amount of the compound of formula (I) as defined in any of the claims 11-16, or a pharmaceutically acceptable salt thereof, together with appropriate amounts of one or more pharmaceutical excipients or carriers.

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