Combination therapy for treatment of pancreatic cancer

Abstract

To provide a combination pharmaceutical for treating liver metastasis in a subject in need of treatment thereof.SOLUTION: A combination pharmaceutical includes a therapeutically effective amount of a combination of active agents. The combination includes cobimetinib or a pharmaceutically acceptable salt thereof and ponatinib or a pharmaceutically acceptable salt thereof. The cobimetinib or a pharmaceutically acceptable salt thereof is present in an amount of 45-75 mg. The ponatinib or a pharmaceutically acceptable salt thereof is present in an amount of 30-60 mg.SELECTED DRAWING: Figure 20A

Description

[Technical field]

[0001] The problem described herein is the combination of MEK inhibitors with RTK, S6, and JAK / S The treatment of pancreatic cancer in combination with multi-targeted agents targeting TAT.

[0002] [Reference to sequence listing submitted as a text file via EFS-Web] The official copy of the sequence listing was created on August 10, 2017 and has a size of 471 bytes. The file named 501159SEQLIST.TXT is in ASCII format. and submitted electronically via EFS-Web as a sequence listing of the present application. The sequence listing contained in this ASCII document is incorporated herein by reference in its entirety. and is incorporated herein by reference. [Background technology]

[0003] Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers, and the pancreas has been studied in culture models. Numerous therapeutic approaches have been investigated based on the outcome of drug responses from cancer cells in PDAC tumors. The majority of these diseases are caused by KRAS mutation-driven activation of MAPK signaling. (Jones S, Zhang X, Parsons DW, Lin JC, Lea ry RJ,Angenendt P,Core signaling pathway s in human pancreatic cancers revealed b y global genomic analyses,Science,2008;3 21(5897):1801-6,Epub 2008 / 09 / 06, Biankin AV, Waddell N, Kassahn KS, Gingras MC, Muthu swamy LB,Johns AL,Pancreatic cancer geno mes reveal aberrations in axon guidance pathway genes,Nature,2012;491(7424):399- 405, Epub 2012 / 10 / 30, Cid-Arregui A, Juarez V,Perspectives in the treatment of panc reatic adenocarcinoma,World Journal of G astroenterology,2015;21(31):9297-316,Epu b 2015 / 08 / 27), thereby supporting the idea that MEK is a potential therapeutic intervention in PDAC patients. However, preclinical and clinical studies have not yet been carried out. Resistance to MAPK inhibitors through various compensatory mechanisms rapidly develops, which This may limit the efficacy of inhibitors and lead to the development of drug-resistant tumors. We found that MAPK pathway inhibition alone was ineffective. (Baines AT, Xu D, Der CJ, Inhibition of Ras for cancer treatment:the search continues,Future M edicinal Chemistry,2011;3(14):1787-808,E pub 2011 / 10 / 19, McCubrey JA, Steelman LS,C happell WH, Abrams SL, Franklin RA, Montalt o G,Ras / Raf / MEK / ERK and PI3K / PTEN / Akt / mT OR cascade inhibitors:how mutations can result in therapeutic resistance and how to overcome resistance,Oncotarget,2012;3(10 ):1068-111, Epub 2012 / 10 / 23, Junttila MR,D Evasthali V, Cheng JH, Castillo J, Metcalfe C,Clermont AC,Modeling targeted inhibit ion of MEK and PI3 kinase in human pancr edic cancer,Molecular Cancer Therapeuti cs,2015;14(1):40-7,Epub 2014 / 11 / 08).

[0004] To improve our understanding of the mechanisms underlying sensitivity and resistance to MEK inhibitors We have identified inhibitors that can be used in combination with MEK inhibitors to target pancreatic cancer cells. It can be determined. Summary of the Invention

[0005] In one aspect, the problem described herein is a subject in need of treatment for pancreatic cancer. A method for treating pancreatic cancer in a subject, comprising administering to said subject a therapeutically effective amount of a combination of active agents. and administering to said patient a combination of a MEK inhibitor and PDGFRα, S6, and and a multikinase agent that targets STAT3.

[0006] In another aspect, the subject matter described herein relates to a method for the treatment of pancreatic cancer, comprising: a) administering to said patient a therapeutically effective amount of a medicament for the treatment of pancreatic cancer; a) a MEK inhibitor or a pharma- ceutical acceptable salt thereof; and b) a PDGFRα, S6, and ST Combination with a multikinase inhibitor targeting AT3 or a pharma- ceutical acceptable salt thereof Regarding.

[0007] In another aspect, the subject matter described herein is a method for treating a pulmonary artery disease by administering an effective amount of a MEK inhibitor or The present invention relates to a compound that targets PDGFRα, S6, and STAT3, and a pharmacopoeia thereof. A combination of a rutilated kinase agent or a pharma- ceutically acceptable salt thereof and a pharma- ceutically acceptable and an excipient, wherein the composition is for treating pancreatic cancer. do.

[0008] In another aspect, the subject matter described herein relates to a method for the treatment of pancreatic cancer, comprising: a) administering to said patient a therapeutically effective amount of a medicament for the treatment of pancreatic cancer; MEK inhibitors or pharma- ceutically acceptable salts thereof, and PDGFRα, S6, and S a multikinase inhibitor targeting TAT3 or a pharma- ceutical acceptable salt thereof; and b) a pancreatic A kit for treating pancreatic cancer, including a package insert or label indicating the treatment of cancer. .

[0009] These and other aspects are described more fully below. [Brief description of the drawings]

[0010] [Figure 1] Figure 2. Differential response to MEK inhibition by KP4 PDAC cells in 2D monolayer and 3D spheroid cultures. Small molecule inhibitor screening of KP4 cancer cells in 2D and 3D culture. [Diagram 2] FIG. 14 shows the differential response to MEK inhibition by KP4 PDAC cells in 2D monolayer and 3D spheroid cultures. Phosphokinase array layout with darkened positions of differentially activated proteins in panel C. [Diagram 3]Figure 2 shows the differential response to MEK inhibition by KP4 PDAC cells in 2D monolayer and 3D spheroid cultures. Phosphokinase arrays of KP4 cells in 2D and 3D. [Figure 4] Figure 1. Differential response to MEK inhibition by KP4 PDAC cells in 2D monolayer and 3D spheroid cultures. Protein kinase array by RT-PCR of KP4 cells cultured in 2D or 3D and treated with cobimetinib for 24 hours. Samples were normalized to DMSO-treated KP4 2D controls. [Diagram 5] Differential response to MEK inhibition by KP4 PDAC cells in 2D monolayer and 3D spheroid cultures. Luciferase reporter assay of KP4 cells cultured in 2D or 3D and treated with cobimetinib for 24 hours. Transcription factors with significant differences in fold change in luciferase activity are shown in red. [Figure 6A-B] We show that MEK inhibition induces activation of PDGFRα, S6, and STAT3. (Figure 6A) Immunoblot verifying activation of PDGFRα, S6, and STAT3 upon cobimetinib treatment in KP4 cells. (Figure 6B) Western blot verifying increase in p-STAT3 upon cobimetinib treatment in PDAC cell lines and KPP-GEMM-derived cell lines. [Figure 7] We show that MEK inhibition induces activation of PDGFRα, S6, and STAT3.IL-6 secretion levels were measured in 2D culture supernatants of PDAC cell lines treated with cobimetinib for 24 hours. [Figure 8] Figure 2 shows that MEK inhibition induces activation of PDGFRα, S6, and STAT3.CTG assay measuring the effect of Dox-induced STAT3 knockdown in combination with cobimetinib on the viability of KP4 and MIA-PACA2 cells. [Figure 9] Figure 2 shows that MEK inhibition induces activation of PDGFRα, S6, and STAT3. Western blot confirming efficient knockdown of STAT3 after 72 h treatment with Dox (1 μg / ml). [Figure 10]Showing that MEK inhibition induces activation of PDGFRα, S6, and STAT3 Quantification of the invasive potential of pancreatic cancer cells cultured in 2D and 3D in response to 10% FBS in a Matrigel invasion assay. [Figure 11] We show that inhibition of S6 and STAT3 in combination with cobimetinib enhances the killing of pancreatic cancer cells.The effect of the multi-RTK inhibitor ponatinib in combination with cobimetinib on the activation of signaling receptors and downstream targets was examined by Western blot. [Figure 12] We show that inhibition of S6 and STAT3 in combination with cobimetinib enhances pancreatic cancer cell killing. The effects of cobimetinib + / - ponatinib or JAK inhibitors on STAT3 activation and induction of cleaved PARP were examined by Western blot. [Figure 13] Showing that inhibition of S6 and STAT3 in combination with cobimetinib increases pancreatic cancer cell killing. PathScan Multiplex Western blot analysis of KP4 cells pretreated with cobimetinib + / - ponatinib in 2D and 3D culture. [Figure 14] We show that inhibition of S6 and STAT3 in combination with cobimetinib enhances pancreatic cancer cell killing.The effect of cobimetinib + / - ponatinib / S6 / PDGFRα inhibitors on S6 activation and induction of cleaved PARP was examined by Western blot. [Figure 15] We show that inhibition of S6 and STAT3 in combination with cobimetinib increases pancreatic cancer cell killing.The effect of the PDGFR inhibitor crenolanib in combination with cobimetinib and ruxolitinib on the viability of KP4 cells was measured using a CTG assay. [Figure 16] We show that inhibition of S6 and STAT3 in combination with cobimetinib increases pancreatic cancer cell killing.The synergy between cobimetinib and ponatinib was analyzed in pancreatic cancer cell lines using the Bliss analysis. [Figure 17]We show that inhibition of S6 and STAT3 in combination with cobimetinib increases pancreatic cancer cell killing.The synergy between cobimetinib and ponatinib was analyzed in pancreatic cancer cell lines using the Bliss analysis. [Figure 18] Showing that inhibition of S6 and STAT3 in combination with cobimetinib increases pancreatic cancer cell killing. Bliss analysis of synergy between cobimetinib and the S6 inhibitor rapamycin in KP4 cells. [Figure 19] Showing that inhibition of S6 and STAT3 in combination with cobimetinib increases pancreatic cancer cell killing. BLISS analysis of synergy between cobimetinib and GDC-0980 in KP4 cells. [Figure 20A-B] Inhibition of PDGFRα / S6 / STAT3 and MEK inhibits tumor growth and reduces serum PDGFα (Figure 20A).Tumor growth curves (mean ± SEM) of KP4 xenograft models treated with cobimetinib (5 mg / kg, PO, QD) and / or ponatinib (30 mg / kg, PO, QD) every 24 hours (n=10 for each cohort), (Figure 20B).Tumor growth curves (mean ± SEM) of KPP xenograft models treated as in (Figure 20A). [Figure 21] Figure 1 shows that inhibition of PDGFRα / S6 / STAT3 and MEK inhibits tumor growth and reduces serum PDGFα. Representative tumor sections from KP4 xenografts were stained for p-STAT3, p-Erk, and cleaved caspase 3 by IHC or co-stained with p-STAT3 and antibodies against F4 / 80 or p-PDGFRα by IF. Scale bars: 20 μm for IHC slides and 50 μm for IF slides. [Figure 22] Showing that inhibition of PDGFRα / S6 / STAT3 and MEK inhibits tumor growth and reduces serum PDGFα. Immunoblots validating activation of PDGFRα, S6, and STAT3 upon cobimetinib / ponatinib treatment of KP4 xenograft tumors. [Diagram 23]Figure 1 shows that inhibition of PDGFRα / S6 / STAT3 and MEK inhibits tumor growth and reduces serum PDGFα. Luminex assay for growth factors and cytokines in plasma samples from KP4 xenografted mice. [Figure 24] 1 shows an analysis of p-Erk and p-STAT3 in pancreatic tumors and metastases from PDAC patient samples. Distribution of p-Erk and p-STAT3 positive samples at different stages of malignancy in primary tumors and metastases from 82 PDAC patients. [Diagram 25] Analysis of p-Erk and p-STAT3 in pancreatic tumors and metastases from PDAC patient samples. Representative primary tumor and liver sections stained for p-Erk and p-STAT3 in PDAC patient samples. Scale bar: 20 μm. [Figure 26] 4 shows analysis of p-Erk and p-STAT3 in pancreatic tumors and metastases from PDAC patient samples.Microarray analysis of PDGFR and IL-6R expression in PDAC patient tumor samples. [Figure 27A-B] We show that KP4-derived 3D spheroids with histopathological features of micrometastases exhibit differential sensitivity to small molecule inhibitors.KP4 cells were cultured for 3 days in RPMI medium containing 10% normal FBS (for 2D culture) or 10% pre-boiled FBS (for 3D culture), and IHC was performed on 2D cell pellets or 3D spheroids for Ki67, p-STAT3, and cleaved caspase-3. [Figure 28] We show that KP4-derived 3D spheroids with histopathological features of micrometastases display differential sensitivity to small molecule inhibitors. Spheroids formed using 10% pre-boiled FBS display a similar drug sensitivity profile as seen with other conventional 3D culture methods. [Figure 29] Figure 1. KP4-derived 3D spheroids with histopathological features of micrometastases exhibit differential sensitivity to small molecule inhibitors. Cell viability and colony count assays of spheroids formed from different 3D culture methods. [Diagram 30]Figure 1. Cobimetinib / ponatinib co-treatment inhibits tumor growth and reduces tumor-associated macrophage infiltration. Representative tumor sections from KPP xenografts probed with p-Erk and p-STAT3 antibodies by IHC or co-stained with anti-p-STAT3 and anti-F4 / 80 by immunofluorescence. Scale bars: 20 μm for IHC slides and 50 μm for IF slides. [Fig. 31A-B] Figure 2. Cobimetinib / ponatinib co-treatment inhibits tumor growth and reduces tumor-associated macrophage infiltration. Body weight change in cobimetinib + / - ponatinib-treated mice in KP4 and KPP xenograft models. [Diagram 32] Figure 1. Luminex assay of growth factors in plasma samples from KP4 xenografted mice. [Diagram 33] Showing that cobimetinib / ponatinib cotreatment inhibits tumor growth and reduces tumor-associated macrophage infiltration. Immunoblots validating activation of PDGFRα, S6, and STAT3 upon cobimetinib / ponatinib treatment of KPP xenograft tumors.

[0011] Data in Figures 6-23 are expressed as mean ± SEM. *P<0.05, Student's T t test. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0012] Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal human diseases with few available drug treatments. Most pancreatic cancers remain largely refractory to KRAS-driven M The EK pathway is activated by mutations and is an important therapeutic target. Inadequate targeting of known carcinogenic drivers and activation of compensatory feedback loops, In addition, the inability to prevent metastatic spread contributes to the poor prognosis of this disease.

[0013] Activation of transmembrane receptor tyrosine kinases (RTKs) results in the regulation of cell fate. This activation triggers numerous biochemical cascades that activate the cells. Growth factors, extracellular matrix (ECM) proteins, and adhesion molecules present on the surfaces of neighboring cells The cell's response to these signals is regulated by the molecules that adsorb to the cell's surface. Many growth factors (GFs) bind to specific receptors present in the Binds to and activates transmembrane glycoproteins of the tyrosine kinase (RTK) family do.

[0014] RTKs consist of an extracellular ligand-binding domain, a single transmembrane domain, and a tyrosine kinase. It contains an endonuclease domain, as well as several regulatory tyrosine-containing intracellular components. K can phosphorylate and activate cytoplasmic STAT family transcription factors. Activated STATs translocate to the nucleus. Once inside the nucleus, STATs mediate the expression of genes involved in cell proliferation. Increases gene transcription.

[0015] Platelet-derived growth factor receptor (PDGFR) is a member of the platelet-derived growth factor (PDGF) family. PDGFRα and P are cell surface tyrosine kinase receptors for members of the PDGFR family. DGFRβ isoforms are factors that regulate cell proliferation, cell differentiation, cell growth and development. Upon binding, the two isoforms dimerize and confer kinase activity and PDG Activates phosphorylation of FR. Tyrosine phosphorylation sites of growth factor receptors activate kinases. Downstream signaling that controls the levels and is often a substrate for kinases as well These are binding sites for downstream signaling molecules such as the S6 ribosomal protein. This signaling cascade is involved in many diseases, including cancer.

[0016] As described herein, two-dimensional monolayer culture systems and three-dimensional spheroids can be used. By using a PD-L1 culture system, we were able to confirm the efficacy of MEK inhibitors or their pharma- ceutical acceptable salts in the treatment of PD-L1. A multi-targeting agent targeting GFRα, S6, and STAT3 or a pharma- ceutical acceptable salt thereof Salt combination treatment targets pancreatic cancer cells in both monolayers and spheroids Without wishing to be bound by theory, it has been found that this combination We report that STAT3 and S6-mediated compensatory feedback loop activation in cancer cells. It effectively blocks signaling through PDGFRα and MEK kinase while also preventing activation. Furthermore, as described herein, the present invention can be used to block the expression of IL-1 in xenograft models. In this study, co-treatment with MEK inhibitors and ponatinib caused tumor regression and concomitantly suppressed pancreatic tumor microenvironment function. Preventing PDAC tumor progression by targeting tumor-associated macrophages in the microenvironment Inhibiting myeloid populations that promote tumor regression through a dual mechanism PDAC patient samples show increased expression of STAT3 activation in tumors and Erk expression in liver metastases. revealed increased activation of STAT3 and Erk in PDAC tumors and liver, respectively These results suggest that IL-16 is an important driver of metastasis. Thus, these results suggest a combination drug therapy strategy for the effective treatment of pancreatic cancer. Furthermore, active agents administered alone may not be effective against pancreatic cancer. do.

[0017] The survival pathways identified as PDGFRβ, MEK, S6, and STAT3 were associated with various Lesina M. (Lesina M. Lesina M.) ,Kurkowski MU,Ludes K,Rose-John S,Treibe r M,Kloppel G,Stat3 / Socs3 activation by IL-6 transsignaling promotes progression of pancreatic intraepithelial neoplasia and development of pancreatic cancer,Ca ncer Cell,2011;19(4):456-69,Epub 2011 / 04 / 13、Yen TW,Aardal NP,Bronner MP,Thorning DR,Savard CE,Lee SP,Myofibroblasts are responsible for the desmoplastic reaction n surrounding human pancreatic carcinoma s,Surgery,2002;131(2):129-34,Epub 2002 / 0 2 / 21, Weissmueller S, Stained E, Saborowsk in M,Morris JPt,Wagenblast E,Davis CA,et al.Mutant p53 drives pancreatic cancer m etastasis through cell-autonomous PDGF r receptor beta signaling,Cell,2014;157(2): 382-94,Epub 04 / 15 / 2014;Corcoran RB,Rothe nberg SM,Hata AN,Faber AC,Piris A,Nazari and RM,TORC1 deletion predicts response iveness to RAF and MEK inhibition in BRA F-mutant melanoma,Science Translational Medicine,2013;5(196):196ra98,Epub 2013 / 0 8 / 02、Corcoran RB,Cheng KA,Hata AN,Faber AC,Ebi H,Coffee EM,Synthetic lethal inte raction of combined BCL-XL and MEK inhib ition promotes tumor regressions in KRAS mutant cancer models,Cancer Cell,2013;2 3(1):121-8,Epub 2012 / 12 / 19、Deng J,Liu Y, Lee H,Herrmann A,Zhang W,Zhang C,S1PR1-S TAT3 signaling is crucial for myeloid ce ll colonization at future metastatic sit es,Cancer Cell,2012;21(5):642-54,Epub 20 12 / 05 / 26、Corcoran RB,Contino G,Deshpande V,Tzatsos A,Conrad C,Benes CH,STAT3 pla ys a critical role in KRAS-induced pancr eatic tumorigenesis,Cancer Research,2011 ;71(14):5020-9,Epub 2011 / 05 / 19) MEK inhibition in PDGFRα inhibits alternative signaling through PDGFRα, S6, and STAT3 We now find that it induces activation of the pathway that contributes to resistance to single-agent MEK inhibition. In particular, individual agents that individually showed no effect on cell viability were found to be significantly reduced in combination. It has been shown to occasionally inhibit cell proliferation in both 2D and 3D cultures. Thus, inhibition of PDGFRα, S6, and STAT3 activation in combination with MEK inhibition for the MEK inhibitor cobimetinib or its pharmacologic acceptable salt in pancreatic tumors. In one embodiment, the multi-RTK and JAK2 inhibitor ponacin was administered to prevent induction of resistance. or a pharma- ceutical acceptable salt thereof with cobimetinib or a pharma- ceutical acceptable salt thereof Combination treatment, as mentioned above, is one of the drug treatment regimens currently being explored in the clinic. This has been highly effective in targeting pancreatic tumors, which are generally refractory to chemotherapy.

[0018] Pancreatic ductal adenocarcinoma (PDAC) is prone to aggressive tumor growth, metastasis, immunosuppression, and drug resistance. This is associated with a marked involvement of various stromal elements that contribute to the progression of the disease. (Cid-Arregui A,e t al.,(2015), Hidalgo M.Pancreatic cancer ,The New England Journal of Medicine,201 0;362(17):1605-17,Epub 2010 / 04 / 30, Korc M .Pancreatic cancer-associated stroma pro duction,American Journal of Surgery,2007 ;194(4 Suppl):S84-6,Epub 2007 / 12 / 06, Inma n KS,Francis AA,Murray NR.Complex role f or the immune system in initiation and p rogression of pancreatic cancer,World Jo urnal of Gastroenterology,2014;20(32):11 160-81,Epub 2014 / 08 / 30、Wormann SM,Diakop oulos KN,Lesina M,Algul H,The immune net work in pancreatic cancer development an d progression,Oncogene,2014;33(23):2956- 67,Epub 2013 / 07 / 16、Moses AG,Maingay J,Sa ngster K,Fearon KC,Ross JA,Pro-inflammat ory cytokine release by peripheral blood mononuclear cells from patients with ad vanced pancreatic cancer:relationship to acute phase response and survival,Oncol ogy Reports,2009;21(4):1091-5,Epub 2009 / 03 / 17、Tjomsland V,Niklasson L,Sandstrom P,Borch K,Druid H,Bratthall C,The desmop lastic stroma plays an essential role in the accumulation and modulation of infi ltrated immune cells in pancreatic adeno carcinoma,Clinical & Developmental Immun ology,2011;2011:212810,Epub 2011 / 12 / 23,S ilvestris N, Gnoni A, Brunetti AE, Vincenti L,Santini D,Tonini G,et al.Target thera pies in pancreatic carcinoma,Current Med icinal Chemistry,2014;21(8):948-65,Epub 2013 / 09 / 03, Vaccaro V,Sperduti I,Vari S,B ria E,Melisi D,Garufi C,Metastatic pancr eatic cancer:Is there a light at the end of the tunnel? World Journal of Gastroe terology,2015;21(16):4788-801.Epub 2015 / 05 / 07, Tuveson DA, Neotolemos JP, Underst anding metastasis in pancreatic cancer:a call for new clinical approaches,Cell,2 012;148(1-2):21-3,Epub 2012 / 01 / 24). Clophages are the most common stromal cell type that infiltrates pancreatic tumors and correlates with poor patient prognosis. (Vonderheide RH, Bayne LJ, Inflammat ory networks and immune surveillance of pancreatic carcinoma,Current Opinion in Immunology,2013;25(2):200-5,Epub 2013 / 02 / 21、Di Caro G,Cortese N,Castino GF,Grizz i F,Gavazzi F,Ridolfi C,Dual prognostic significance of tumor-associated macrop hages in human pancreatic adenocarcinoma treated or untreated with chemotherapy, Gut,2015.Epub 2015 / 07 / 15; B, Zeng LJ, Lian GD, Li JJ, Qian CC, Distribu tion and clinical significance of tumor -associated macrophages in pancreatic do ctal adenocarcinoma:a retrospective anal analysis in China,Curr.Oncol.2015;22(1):e11- 9,Epub 2015 / 02 / 17). The TAM's Effects of Noy R,Pollard JW.Tu mor-associated macrophages:from mechanisms ms to therapy,Immunity,2014;41(1):49-61, Epub 2014 / 07 / 19、Melero I,Gaudernack G,Ge Ritsen W,Huber C,Parmiani G,Scholl S,Th erapeutic vaccines for cancer:an overvie w of clinical trials,Nature Reviews Clin ical Oncology,2014;11(9):509-24,Epub 201 4 / 07 / 09、Drake CG.Combination immunothera py approaches,Annals of oncology:officia l journal of the European Society for Me dical Oncology / ESMO,2012;23 Suppl 8:vi ii41-6,Epub 2012 / 08 / 29、Weden S,Klemp M,G ladhaug IP,Moller M,Eriksen JA,Gaudernac k G,Long-term follow-up of patients with resected pancreatic cancer following va ccination against mutant K-ras,Internati onal Journal of Cancer,Journal Internati onal du Cancer,2011;128(5):1120-8,Epub 2 010 / 05 / 18、Bronte V,Murray PJ.Understandi ng local macrophage phenotypes in diseas e:modulating macrophage function to trea t cancer,Nature Medicine,2015;21(2):117- 9, Epub 2015 / 02 / 06). Our findings indicate that the IL-16 expression of IL-16 was responsive to MEK inhibition. This suggests the role of the STAT3 signaling axis in both tumor cells and TAMs in PDAC. Our results further demonstrate that MEK inhibitors, such as cobimetinib / PDGFR Co-treatment of α, S6 with a multikinase inhibitor that targets STAT3, such as ponatinib The therapy can target both myeloid and cancer cells within the primary tumor, promoting cell death and tumor suppression. This suggests that it may cause increased tumor regression.

[0019] The problems disclosed herein are explained more fully hereinafter. Many modifications and other embodiments of the subject matter of this disclosure described herein are possible. It will be readily apparent to one skilled in the art to which the disclosed subject matter pertains having the benefit of the teachings presented in the preceding description. Therefore, it is to be understood that the scope of the present disclosure should not be limited to the specific embodiments disclosed. However, modifications and other embodiments are intended to be within the scope of the appended claims. In other words, the problem described herein is solved by all alternatives. The present invention includes all forms, modifications, and equivalents of the present invention. One or more of the following may be included in the specification, including but not limited to defined terms, usage of terms, and described techniques. In the event that any provision of this application differs or conflicts with the provisions of this application, this application will take precedence. Unless otherwise specified, technical and scientific terms used herein are understood to be of ordinary skill in the art. All publications, patents, and other references referred to herein have the same meaning as those referred to herein. Applications, patents, and other references are incorporated herein by reference in their entireties. do.

[0020] I. Definition The term "pancreatic cancer" refers to neoplasms originating from the pancreas. Pancreatic cancer includes exocrine and endocrine cancers. Most pancreatic cancers are exocrine tumors. Pancreatic endocrine tumors are classified as islet cell tumors. Unfortunately, the prognosis for pancreatic cancer is currently poor, even when detected early. Pancreatic cancers that can be treated with the methods described herein include exocrine Exocrine pancreatic cancer includes, but is not limited to, pancreatic cancer and endocrine pancreatic cancer. , adenocarcinoma, acinic cell carcinoma, adenosquamous carcinoma, mucinous carcinoma, and undifferentiated with osteoclast-like giant cells. Hepatoid adenocarcinoma, intraductal papillary mucinous neoplasm, mucinous cystic neoplasm, pancreatoblastoma, serous cystadenoma, These include, but are not limited to, ring cell carcinoma, solid pseudopapillary tumor, pancreatic ductal carcinoma, and undifferentiated carcinoma. Not limited thereto. In some embodiments, the pancreatic exocrine cancer is pancreatic ductal carcinoma. Cancers include, but are not limited to, insulinoma and glucagonoma.

[0021] In some embodiments, the pancreatic cancer is early stage pancreatic cancer, non-metastatic pancreatic cancer, primary pancreatic cancer. , resected pancreatic cancer, advanced pancreatic cancer, locally advanced pancreatic cancer, metastatic pancreatic cancer, unresectable pancreatic cancer, remission pancreatic cancer in the adjuvant setting, or in the neoadjuvant setting In some embodiments, the pancreatic cancer is locally advanced pancreatic cancer, unresectable pancreatic cancer, Or metastatic pancreatic ductal carcinoma. In some embodiments, the pancreatic cancer is treated with gemcitabine-based chemotherapy. In some embodiments, the pancreatic cancer is resistant to gemcitabine-based therapy. It is refractory to treatment.

[0022] The methods described herein may be used for any one or more of the following purposes: It can: alleviate one or more symptoms of pancreatic cancer, delay the progression of pancreatic cancer, reduce the size of pancreatic cancer tumors reduction in size, destruction of the pancreatic cancer stroma (e.g., destruction), inhibition of pancreatic tumor growth, and increased overall survival. Extending disease-free survival, extending time to pancreatic cancer disease progression, and preventing or delaying pancreatic cancer tumor metastasis extension, reduction (e.g. eradication) of metastasis of existing pancreatic cancer tumors, occurrence or burden of metastasis of existing pancreatic cancer tumors reducing the severity of pancreatic cancer, preventing recurrence of pancreatic cancer, and / or improving clinical benefit for patients with pancreatic cancer.

[0023] The pancreatic cancer treatment methods described herein include those for primary and metastatic tumors. A therapeutically effective amount of the compound to inhibit, slow, or reverse the growth, development, or spread of pancreatic cancer. MEK inhibitor or a pharma- ceutical acceptable salt thereof and PDGFRα, S6, and STAT Combination with a multikinase inhibitor targeting 3 or a pharma- ceutical acceptable salt thereof The method includes administering the compound to a subject (human or animal) in need of treatment. The administration may include administering to one or more kinases and / or kinases that are inhibited by one of the administered active agents. or a method for the treatment or prevention of a disease mediated by a survival pathway. "Administration" or "administering" of the compounds disclosed herein is intended to mean the same as, but not limited to, the administration of The active agent, or its prodrug, may be administered in any suitable formulation or route of administration as described above. The present invention includes delivery of a MEK inhibitor or other pharma- ceutically acceptable derivative thereof to a recipient. and multikinase inhibitors, in either order, sequentially, sequentially, or both. Combinations may be administered.

[0024] The term "subject" refers to any animal, including primates (e.g., humans), cows, sheep, goats, horses, dogs, cats, It refers to animals such as mammals, including but not limited to rabbits, rats, mice, and the like. In certain embodiments, the subject is a human.

[0025] The terms "prevention" or "prophylactic" refer to the same or similar effects that would be expected if the combination were not administered. It refers to the continued absence of symptoms of a presumed disease or condition.

[0026] As used herein, the term "synergistic" refers to an effect greater than the additive effect of two or more single agents. This also refers to an effective therapeutic combination that targets PDGFRα, S6, and STAT3. The determination of synergistic interactions between a multi-kinase inhibitor and a MEK inhibitor is described herein. For example, the present invention can be based on the results obtained from the assays described herein. The in vivo or in vitro methods disclosed herein are based on the results of these assays. To obtain the index, we use the combined method of Chou and Talalay and CalcuSyn The software can be used to perform dose-effect analysis (Chou and T alalay, 1984, Adv. Enzyme Regul. 22:27-55). proposal The combinations provided can be evaluated in one or more assay systems and data collected. The data set is a standard program for quantifying synergy, additivity, and antagonism between anticancer drugs. An exemplary program is Chou and Tala. New Avenues in Developmental Canlay cer Chemotherapy,Academic Press,1987,Cha pter 2. A combination index value of less than 0.8 indicates synergy, and a combination index value of 1. Values ​​above 2 indicate antagonism, while values ​​between 0.8 and 1.2 indicate additive effects. Drugs provide a "synergistic effect" and are "synergistic," i.e., active agents used together enhance the compound's Demonstrate the effect achieved when the effect is greater than the sum of the effects resulting from each use separately A synergistic effect can be achieved when the active agents are: (1) in combination (2) coformulated and administered or delivered simultaneously in a single-site dosage formulation; delivered in alternation therapy, either in parallel or in parallel with each other, or (3) in different dosing regimens. When the compounds are administered or delivered sequentially, for example in separate syringes, Synergistic effects may be achieved by administration of the same or similar drugs. Generally, during alternation therapy, effective doses of each active agent are administered. Sequentially, i.e., consecutively, in combination therapy, effective amounts of the two The above active agents are administered together. The combination effect is evaluated in the Bliss independent model and the best single agent (HSA) model. Both models can be used to assess lar Systems Biology, 3:80(2007)). Bliss scores are simply The degree of potentiation from the agent was quantified, and a positive Bliss score (greater than 0) indicates simple additive potentiation. A cumulative positive Bliss score of more than 250 indicates a greater than normal toxicity in the range of concentrations tested. A strong synergistic effect observed within the range of 0 was considered. This suggests a combination effect that is greater than the maximum response of the single agents at a given concentration.

[0027] "Spheroids" are organisms in which biological cells grow or grow in all three dimensions. These are known in the art as three-dimensional (3D) cell cultures in artificial environments that interact with their surroundings. Unlike a 2D environment such as a Petri dish, 3D cell culture allows It allows cells to grow in all directions in vitro, similar to how they grow in vivo. The industrial environment includes a culture medium that provides specific nutrients and other factors that allow cells to grow. The media described herein provide an environment for rapid cell culture. " refers to a period of time that is shorter than the period required for identical cell growth in a comparison medium.

[0028] The term "contact" or "contacting" refers to the process by which an active agent binds to a target and inhibits its activity. Target enzymes or survival pathways so that they can be activated, decreased, or reduced. This means that the active agent is placed in close proximity to the surface of the substrate.

[0029] As used herein, the term "mammal" includes humans and all other mammals. As used herein, the term "mammal" refers to a "subject" or "patient." " and refers to warm-blooded animals. Guinea pigs, dogs, cats, rats, mice, horses, Animals, cattle, sheep, zoo animals, livestock, primates, and humans all fall within the meaning of this term. are understood to be examples of animals within the scope of the invention.

[0030] As used herein, a "mammal in need thereof" refers to a subject being treated. The specific condition for which the treatment is intended, such as being diagnosed with pancreatic cancer or a particular type of pancreatic cancer, This is the subject of the study.

[0031] The terms "treat" and "treatment" include both therapeutic treatment and prophylactic treatment. refers to both preventative and therapeutic measures, the purpose of which is to prevent the onset or spread of cancer. The aim of the study is to prevent or slow down (reduce) undesirable physiological changes or disorders such as hypertension. For purposes of this disclosure, beneficial or desired clinical results include alleviation of symptoms, reduction in severity of disease, reduction in the severity of the disease, stabilization of the disease state (i.e., not worsening), delay or slowing of disease progression, Remission or improvement of disease, and remission (partial or complete) whether detectable or undetectable "Treatment" also refers to the condition that would be expected if no treatment was given. It can also mean an increased survival time compared to the survival time expected in a patient with a disease that is not in need of treatment. Those who have a condition or disorder and those who are prone to having a condition or disorder or to prevent a condition or disorder.

[0032] The term "inhibitor" is used herein to refer to a molecule that inhibits the activity of a target enzyme. As used herein, "inhibit" refers to the reduction in the activity of an enzyme in the absence of an inhibitor. In some embodiments, the activity of a target enzyme is reduced relative to the activity of the target enzyme. In the present context, the term "inhibit" means to inhibit by at least about 5%, at least about 10%, at least about 20%, or %, at least about 25%, at least about 50%, at least about 60%, at least about 70 %, at least about 80%, at least about 90%, or at least about 95% MEK activity In another embodiment, inhibition of a particular survival pathway in cancer cells is referred to as a reduction in the ability of the cancer cells to survive. The harm is about 5% to about 25%, about 25% to about 50%, about 50% to about 75%, or about 75% to about In some embodiments, inhibition is about 95% to 100%. 00% decrease in activity, e.g., 95%, 96%, 97%, 98%, 99%, or 100% decrease in activity. % decrease in activity. Such a decrease would be recognizable by one of ordinary skill in the art. This can be measured using a variety of techniques, including in vitro kinase assays.

[0033] As used herein, a "MEK inhibitor" refers to an inhibitor of MEK (MEK1 and / or Reduce, inhibit, or otherwise decrease one or more biological activities of MEK2 The activity is, for example, at least about 5%, 10%, 15%, or more relative to a suitable control. %, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65 Statistically significant including %, 70%, 75%, 80%, 85%, 95% or 100% reduction The amount of MEK activity can be reduced.

[0034] As used herein, a "multikinase inhibitor" refers to an inhibitor of PDGFRα, S6, and and molecules that reduce, inhibit, or otherwise diminish the biological activity of STAT3. A suitable multi-kinase inhibitor would exhibit activity against each of these three targets, and may have activity against additional targets. MEK inhibitors also act on tumor-associated macrophages associated with pancreatic tumors. The ability of a multikinase inhibitor to act on each of these targets when combined with pancreatic The activity may be, for example, at least Also about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% , 55%, 60%, 65%, 70%, 75%, 80%, 85%, 95% or 100% The activity of the target may be decreased by a statistically significant amount, including a decrease.

[0035] The compounds of the present disclosure may or may not be specific inhibitors. Reduce, inhibit, or enhance the activity of a defined target relative to the activity of an unrelated target. For example, a MEK-specific antagonist is an agent that inhibits or reduces the expression of MEK. At least one biological activity can be determined by the addition of any other protein (e.g., other serine / threonine The inhibitory effect of antagonists on the kinase (kinase) is reduced by a significantly greater amount than that of antagonists. In some embodiments, the IC50 of an antagonist for a target is greater than the IC50 of an antagonist for a non-target. Approximately 90%, 80%, 70%, 60%, 50%, 40%, 30% of the IC50 of the antagonist. 20%, 10%, 5%, 1%, 0.1%, 0.01%, 0.001% or less Specific MEK inhibitors block the biological activity of one or more MEK family members, if desired. Antagonists against other proteins of interest (e.g., other serine / threonine kinases) In some of these embodiments, the inhibitory effect of The IC50 of MEK inhibitors against RAF is higher than that of other serine / threonine kinases, other ME M against K family members or other types of kinases (e.g., tyrosine kinases) Approximately 90%, 80%, 70%, 60%, 50%, 40%, 30% of the IC50 of the EK inhibitor 20%, 10%, 0.1%, 0.01%, 0.001% or less.

[0036] The term "target" or "targeting" refers to a specific MEK enzyme or a specific targeting associated with a survival pathway. This refers to the ability of an active agent to act specifically on a particular enzyme. can be determined through routine experimentation as described herein, or by other known methods. It can be determined using

[0037] As used herein, a "survival pathway" refers to a pathway that promotes the survival and proliferation of cancer cells. As used herein, the term refers to an adaptive mechanism or ability to promote cell proliferation and become resistant to active agents. When used in this study, the survival pathway involves PDGFRα, S6, and STAT3. do.

[0038] The phrase "therapeutically effective amount" refers to a compound that (i) treats or prevents pancreatic cancer, (ii) inhibits one or more of the pancreatic cancer or (iii) reduce, ameliorate, or eliminate one or more symptoms of pancreatic cancer. A therapeutically effective amount of a drug is an amount of an active agent that prevents or delays the onset of cancer. It can reduce tumor size and inhibit cancer cell invasion into peripheral organs (i.e. slowing to some extent and preferably halting tumor metastasis (i.e., slowing to some extent, and preferably halting, tumor growth, inhibiting to some extent, and / or or relieve to some extent one or more of the symptoms associated with cancer. It is cytostatic and / or capable of killing existing cancer cells. In the case of cancer treatment, efficacy can be measured, for example, by the time to disease progression (T This can be measured by assessing response rate (TP) and / or determining response rate (RR). This can be done.

[0039] The term "pharmaceutically acceptable salt" refers to a salt that is not biologically or otherwise undesirable. Pharmaceutically acceptable salts include both acid addition salts and base addition salts. "Pharmaceutically acceptable" means that a substance or composition is compatible with other ingredients and / or It must be chemically and / or toxicologically compatible with the mammal being treated with it. As used herein, the phrase "pharmaceutically acceptable salt" refers to a molecule that is Exemplary salts include sulfate, citrate, and the like. Acid salts, acetates, oxalates, chlorides, bromides, iodides, nitrates, bisulfates, phosphates, Acid phosphates, isonicotinates, lactates, salicylates, acid citrates, tartrates, Oleate, tannate, pantothenate, bitartrate, ascorbate, succinate Salt, maleate, gentisate, fumarate, gluconate, glucuronate, saccharate Carboxylate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate phosphate, benzenesulfonate, p-toluenesulfonate, and pamoate (i.e. This includes 1,1'-methylene-bis-(2-hydroxy-3-naphthoate) salts. Pharmaceutically acceptable salts include, but are not limited to, acetate ions, succinate ions, or other salts. The counterion may include another molecule, such as a counterion of the formula: It may be any organic or inorganic moiety. Furthermore, a pharma- ceutically acceptable salt may have one or more amines in its structure. Multiple charged atoms may be part of a pharma- ceutically acceptable salt. In some cases, there may be more than one counter ion. Thus, a pharma- ceutically acceptable salt is: It can have one or more charged atoms and / or one or more counter ions. include cobimetinib and ponatinib HCl fumarate or hemifumarate salts But not limited to these.

[0040] Additional definitions are provided below as needed.

[0041] II. In Vivo Methods MEK inhibitor and multikinase inhibitor targeting PDGFRα, S6, and STAT3 Combinations of the present disclosure with cytotoxic agents are useful in the treatment of pancreatic cancer.

[0042] In an embodiment, the present invention provides a method of treating pancreatic cancer in a subject in need thereof. The method comprises administering to the subject a therapeutically effective amount of a combination of active agents, The combination comprises a MEK inhibitor or a pharma- ceutical acceptable salt thereof and PDGFRα, S6, and and a multikinase agent targeting STAT3 or a pharma- ceutical acceptable salt thereof. A method is disclosed herein for inhibiting the proliferation of tumor associated macrophages. It may further include inhibiting.

[0043] Examples of MEK inhibitors that may be used include cobimetinib, GDC-0623, trametinib, Nib, binimetinib, selumetinib, pimasertinib, refametinib, PD-0325 901 and BI-847325, or pharma- ceutically acceptable salts thereof. In an embodiment, the MEK inhibitor is cobimetinib or a pharma- ceutically acceptable salt thereof. Cobimetinib has the following structure: [ka]

[0044] Cobimetinib is marketed as Cotellic® and is marketed under the trademark Hemihumerus. It is found in commercial formulations as the phosphate salt.

[0045] Ponaty is a useful multikinase agent that targets PDGFRα, S6, and STAT3. Ponatinib has the following structure: [ka]

[0046] Ponatinib is sold as Iclusig® and is available commercially as the hydrochloride salt. It is found in formulations of.

[0047] Pancreatic cancer includes endocrine and exocrine cancers. In embodiments, pancreatic cancer is Adenocarcinoma, acinic cell carcinoma, adenosquamous carcinoma, mucinous carcinoma, undifferentiated carcinoma with osteoclast-like giant cells , hepatoid adenocarcinoma, intraductal papillary mucinous neoplasm, mucinous cystic neoplasm, pancreatoblastoma, serous cystadenoma, signet ring tumor The present invention relates to a method for treating pancreatic cancer, the method comprising administering to the patient a therapeutically effective amount of ... In one embodiment, the pancreatic cancer is pancreatic ductal adenocarcinoma.

[0048] In an embodiment, a MEK inhibitor or a pharma- ceutically acceptable salt thereof and PDGFRα, A multikinase inhibitor targeting S6 and STAT3 or a pharma- ceutical acceptable derivative thereof The combination is synergistic with the salts of the corresponding drugs, i.e., the combination produces a greater than additive effect of both agents. In embodiments, one or both active agents alone provide efficacy against pancreatic cancer. Cobimetinib or a pharma- ceutical acceptable salt thereof and ponatinib do not show the desired efficacy. In the case of rifabutin or its pharmacologic acceptable salt, neither agent alone is effective against pancreatic cancer. It is not known to exist.

[0049] In an embodiment, a MEK inhibitor or a pharma- ceutical acceptable salt thereof and the multikinase The enzyme inhibitor or a pharma- ceutically acceptable salt thereof is administered as a combination formulation. In certain embodiments, the combination therapy may be administered as a simultaneous or sequential regimen. When administered sequentially, the combination may be administered in two or more administrations. Administration may be by co-administration using separate formulations or a single pharmaceutical formulation, and by administration of both (or all) In any order, there is preferably a period during which the active agents simultaneously exert their biological activity. This includes continuous administration of

[0050] Methods for treating pancreatic cancer include reducing or inhibiting the activity of one or more targeted enzymes or survival pathways. Tumor-associated macrophages associated with pancreatic tumors are also It can also be targeted by in vitro kinase assays, using antibodies specific for phosphorylated targets. MEK activity or PDG, including immunoblots or measurements of downstream biological effects of activity To measure the levels of FRα, S6, STAT3 and tumor-associated macrophage activation Any method known in the art can be used to determine the activity of the combination.

[0051] Pancreatic cancer may be at an early or late stage and may have metastasized. The combinations described herein are intended to treat any stage of cancer, including cancer that has metastasized. can be used for.

[0052] Effective doses of MEK inhibitors and multi-kinases targeting PDGFRα, S6, and STAT3 Treatment of a subject in combination with an enzyme inhibitor can include monotherapy or can be a single therapy. The treatment may include a series of

[0053] Useful amounts of active agent are as described elsewhere herein. In embodiments, the MEK inhibitor is administered in an amount of from about 5 mg to about 100 mg, or from about 45 mg to about 75 mg. g, and the multikinase inhibitor is administered in an amount of about 5 mg to about 100 mg, or about 3 In an embodiment, the MEK inhibitor is administered in an amount of about 60 mg to about 60 mg. In one embodiment, the multikinase inhibitor is administered in an amount of about 45 mg. In an embodiment, the administration of the combination therapy is oral once daily. the K inhibitor is cobimetinib or a pharma- ceutical acceptable salt thereof, is preferably ponatinib or a pharma- ceutically acceptable salt thereof. In this embodiment, each active agent is administered at a dose of about 0.001 μg / kg to about 1000 mg / kg. The subject is administered about 0.001 μg / kg, 0.01 μg / kg, 0.05 μg / kg, g / kg, 0.1μg / kg, 0.5μg / kg, 1μg / kg, 10μg / kg, 25 μg / kg, 50μg / kg, 100μg / kg, 250μg / kg, 500μg / kg , 1mg / kg, 5mg / kg, 10mg / kg, 25mg / kg, 30mg / kg, 4 Includes 0mg / kg, 50mg / kg, 100mg / kg, and 200mg / kg. Each active agent present in the combination composition to be administered may be, but is not limited to, The amount is about 1 mg to about 1000 mg, or about 5 mg to about 100 mg, or about 10 mg. about 80 mg, or about 45 mg to about 75 mg, or about 30 mg to about 60 mg. The appropriate dose of the active compound will depend on a number of factors within the knowledge of one of ordinary skill in the art or veterinarian. It will be understood that the dose of the active agent will depend, for example, on the age, weight, and overall health of the subject. The condition, sex, and diet of the subject, the time of administration, the route of administration, the excretion rate, and any drug combinations The effective dose used for treatment will vary depending on the course of the particular treatment. It will also be understood that the dosage may be increased or decreased over the course of a diagnostic assay. It can arise from and be evident in the results.

[0054] In some embodiments, the present invention relates to a method for treating pancreatic cancer in a subject in need thereof. The method comprises administering to the patient a MEK inhibitor or a pharma- ceutical acceptable salt thereof and a PDGFRα inhibitor. A multikinase inhibitor targeting STAT3, S6, and STAT3 or a pharma- ceutical acceptable salt thereof and administering to the subject a combination of the compound and a salt thereof, Methods described herein include, but are not limited to, radiation therapy, surgery (e.g., chemotherapy, chemotherapy of the ovarian cancer ... (e.g., lumpectomy and mastectomy), chemotherapy, gene therapy, DNA therapy, viral therapy immunotherapy, RNA therapy, immunotherapy, bone marrow transplantation, nanotherapy, monoclonal antibody therapy, or The additional therapy may be in the form of adjuvant or neoadjuvant therapy. In some embodiments, the additional therapy is administration of an anti-metastatic drug. In some embodiments, the additional therapy is a side effect limiting agent (e.g., an anti-nausea agent) that reduces the occurrence of side effects of the treatment. and / or administration of an agent intended to reduce the severity of the condition. In some embodiments, the additional therapy is radiation therapy. In some embodiments, the additional therapy is a combination of radiation therapy and surgery. In some embodiments, the additional therapy is gamma radiation.

[0055] In some embodiments, the treatment produces a sustained response in the subject after cessation of treatment. "Durable response" refers to a sustained effect on reducing tumor growth after treatment has stopped. For example, tumor size remains the same or increases compared to the size at the beginning of the dosing phase. In some embodiments, a sustained response may be at least as long as the duration of treatment. For the same period, at least 1.5, 2.0, 2.5, or 3.0 times longer than the treatment period It has a quality.

[0056] The therapeutic methods disclosed herein may result in a partial or complete response. As used herein, "complete response" or "CR" refers to the disappearance of all target lesions. A partial response (PR) was defined as a response based on the sum of the longest diameters (SLD) of the target lesions. A reduction of at least 30% from the line SLD is considered to be "stable disease" or " "SD" refers to the smallest SLD since treatment initiation that is sufficient to qualify for PR. As used herein, the term refers to a condition that shows neither a shrinkage of the lesions nor a sufficient increase in the lesions to qualify for PD. When the treatment is administered, the overall response rate (ORR) is the sum of the complete response (CR) rate and the partial response (PR) rate. Refers to...

[0057] The methods of treatment disclosed herein include preventing progression of disease in subjects administered the combination therapy. As used herein, "can result in increased survival and overall survival." "Progression-free survival" (PFS) is the period during which a disease (e.g., cancer) does not get worse during treatment or Progression-free survival refers to the length of time after treatment when a patient achieves a complete or partial response. These may include the length of time the patient has experienced stable disease, and the length of time the patient has experienced stable disease.

[0058] As used herein, "overall survival" refers to the survival after a particular period of time. The proportion of subjects in a group likely to be affected.

[0059] In some embodiments, the subject to which the combination is administered is a domestic animal (e.g., a domestic animals (e.g., sheep, cats, dogs, and horses), primates (e.g., humans and non-humans such as monkeys) mammals, such as primates, rabbits, and rodents (e.g., mice and rats); In some embodiments, the subject to be treated is a human.

[0060] Subjects who require treatment for pancreatic cancer are those who show symptoms of cancer, those who have been diagnosed with cancer, and those who have pancreatic cancer. Subjects who are in remission from cancer or who are at high risk of developing cancer (e.g., those with a genetic predisposition) , certain dietary or environmental exposures).

[0061] Many of the combinations described herein will be determined to be synergistic.

[0062] The in vitro method described herein involves adding boiling water to the medium for culture. Prepare spheroid cell cultures, including culturing cells in medium containing heated serum The medium can be modified to rapidly culture 3-D spheroids. The modified medium was supplemented with fetal bovine serum (FBS) prior to contact with the cells to be cultured. In one embodiment, the FBS is heated to 95-100° C. It is boiled for 10 minutes, at which point the serum turns yellow. The FBS was centrifuged at 8000 × g for 20 min and filtered through a filter with a pore size of 0.22 μm. Filter through a sterile Corning vacuum filter unit. Useful medium is approximately 5% to 10%. The medium for spheroid culture contains approximately 20% (v / v) fetal bovine serum. RP supplemented with glutamine, 100 units ml-1 of penicillin and streptomycin Prepare MI medium by adding 10% (v / v) boiled FBS. The Lloyd medium must be freshly prepared before use. Boiled FBS can be stored at 4°C for 2-3 months. The medium for monolayer culture is 2 mM L-glutamine, 100 units ml-1 penicillin, RPMI medium supplemented with phosphate and streptomycin, 10% (v / v) normal FBS The medium for migration assay is prepared by adding 2 mM L-glutamine, RPMI medium supplemented with 100 units ml-1 penicillin and streptomycin Prepare by adding 0% (v / v) normal FBS.

[0063] III. Combination Pharmaceutical Compositions In an embodiment, the subject matter described herein is the prevention or treatment of pancreatic cancer. a) a MEK inhibitor or a pharma- ceutical acceptable salt thereof, and b) a PDGFR A multikinase inhibitor targeting α, S6, and STAT3 or its pharma- ceutical acceptable salt The present invention relates to combinations with salts that are

[0064] In an embodiment, the subject matter described herein is a method for the treatment of a disease comprising administering to a patient a therapeutically effective amount of a MEK inhibitor or a pharmaceutical composition thereof. A multikinase inhibitor targeting PDGFRα, S6, and STAT3 with a commercially available commercially acceptable salt. and a combination of the agent or a pharma- ceutical acceptable salt thereof, and a pharma- ceutical acceptable excipient. In certain embodiments, the pharmaceutical composition is for treating pancreatic cancer. It is useful.

[0065] In embodiments, the composition comprises cobimetinib, GDC-0623, trametinib, bimetinib, Metinib, selumetinib, pimasertinib, refametinib, PD-0325901 and and BI-847325, or a pharma- ceutically acceptable salt thereof. In one embodiment, the MEK inhibitor is cobimetinib or a pharmaceutical In certain embodiments, the MEK inhibitor is cobime hemifumarate. It is tinib.

[0066] In embodiments, the multikinase agent is ponatinib or a pharma- ceutically acceptable salt thereof. In one embodiment, the multikinase inhibitor is ponatinib hydrochloride.

[0067] In an embodiment, the composition comprises cobimetinib or a pharma- ceutical acceptable salt thereof and ponamylase. In an embodiment, the composition comprises hemifumab or a pharma- ceutical acceptable salt thereof. This includes cobimetinib hydrochloride and ponatinib hydrochloride.

[0068] The amount of each active agent present in the combination may range from about 1 mg to about 1000 mg, or from about 5 mg to about 100 mg, or about 10 mg to about 80 mg, or about 45 mg to about 75 mg, Or, the MEK inhibitor may be about 30 mg to about 60 mg. In an embodiment, the MEK inhibitor is about 45 mg. g to about 75 mg, and the multikinase inhibitor is present in an amount of about 30 mg to about 60 mg. It exists in.

[0069] A combination therapy provides a "synergistic effect" and is "synergistic," i.e., has activity The effect achieved when the active agents are used together is different from that achieved when these active agents are used separately. A synergistic effect can be achieved when the active agents: (1) Coformulated and administered or delivered simultaneously in a combined unit dosage formulation. (2) delivered as separate formulations, either alternating or in parallel; or (3) delivered in combination with other When delivered in alternation therapy, when active agents are administered or delivered sequentially For example, different injections in separate syringes, separate pills or capsules, or separate injections. Synergistic effects may be achieved by the inclusion of multiple active agents. Generally, during alternation therapy, effective doses of each active agent are administered. Sequentially, i.e., consecutively, in combination therapy, effective amounts of the two The above active agents are administered together.

[0070] MEK inhibitors and / or multikinase inhibitors are each referred to herein as active agents. and can be formulated as a pharmaceutical composition in accordance with standard pharmaceutical practice. Ponatinib, a potent multikinase inhibitor, is available in strengths of 15 mg, 30 mg, and 45 mg. It is available as a tablet for oral administration. An exemplary MEK inhibitor, cobimetinib, is It is available in a strength of 20 mg as a tablet for administration. However, according to this embodiment , a MEK inhibitor or a multikinase inhibitor, or both, in a carrier or diluent, etc. Other pharmaceutical compositions are provided that include, in combination with a pharma- ceutically acceptable excipient.

[0071] A typical formulation is prepared by mixing an active agent with one or more excipients. Suitable excipients are well known to those skilled in the art and include carbohydrates, waxes, water-soluble and / or swellable polymers, and the like. These include materials such as mers, hydrophilic or hydrophobic substances, gelatin, oils, solvents, and water. The particular excipient used will depend on the means and purpose for which the active agent is being applied. is generally a solvent recognized by those skilled in the art as safe for administration to mammals (G Generally, safe solvents are non-toxic aqueous solvents such as water and and other non-toxic solvents that are soluble or miscible in water. Suitable aqueous solvents include water, ethanol, and the like. propylene glycol, polyethylene glycol (e.g., PEG400, PEG3 00), and mixtures thereof. The formulation may also contain one or more buffers, stabilizing agents, agents, surfactants, wetting agents, lubricants, emulsifiers, suspending agents, preservatives, antioxidants, opacifying agents, glidants Provides a sophisticated presentation of processing aids, colorants, sweeteners, fragrances, flavorings, and active agents. or other known additives to aid in the manufacture of a pharmaceutical product (i.e., a drug). It may include.

[0072] The formulations can be prepared using conventional dissolution and mixing procedures. The drug (i.e., the active agent or a stabilized form of the active agent (e.g., a cyclodextrin derivative or or other known complexing agents) in the presence of one or more of the above-mentioned excipients in a suitable solvent. The active agent typically provides an easily controllable dosage of the drug and is easily administered. The therapeutic agent is formulated into a pharmaceutical dosage form to enable patient compliance with the prescribed dosing regimen.

[0073] The pharmaceutical composition (or formulation) for application depends on the method used to administer the drug. In general, articles for distribution contain pharmaceutical products in an appropriate form. Suitable containers include containers for holding the agent therein. Suitable containers are well known to those skilled in the art and include bottles (plastic and glass), sachets, ampoules, plastic bags, metal cylinders, etc. The containers also have tamper-evident features to prevent inadvertent access to the package contents. Additionally, the container may have a label thereon that describes the contents of the container. The label may also include appropriate warnings. In one embodiment, the container is It's a star pack.

[0074] Pharmaceutical formulations can be prepared for a variety of routes and forms of administration. The active agent having the desired degree of purity is optionally combined with a pharma- ceutically acceptable excipient in a lyophilized form. It can be mixed as a powder, crushed powder, or in an aqueous solution (Remington's rmaceutical Sciences(1980)16th edition,O sol, A.Ed., Mack Publishing Co., Easton, PA) The formulation is carried out at ambient temperature, at the appropriate pH, at the desired purity, in a physiologically acceptable carrier, i.e. by mixing with a carrier which is nontoxic to the recipient at the dosage and concentration employed. The pH of the formulation depends primarily on the particular application and the concentration of the compound, but may be about 3 to 50%. It may range from about 8. Formulation in acetate buffer at pH 5 is a preferred embodiment.

[0075] The active agent can be sterile. In particular, formulations to be used for in vivo administration must be sterile. Such sterilization is readily accomplished by filtration through sterile filtration membranes.

[0076] The active agent may generally be stored as a solid composition, a lyophilized formulation, or an aqueous solution.

[0077] The pharmaceutical composition containing the active agent may be formulated in a manner, i.e., amount, concentration, schedule, course of treatment, biology, Formulate, administer, and administer the drug in a vehicle and route of administration consistent with good medical practice. Factors to consider in this regard include the particular injury being treated, the specific the mammal, the clinical condition of the individual patient, the cause of the injury, the site of delivery of the agent, the method of administration, the administration schedule, and other factors known to those skilled in the art. A "therapeutically effective amount" of a compound to be administered is Acceptable excipients will be governed by such considerations. and concentrations that are non-toxic to the recipient and contain phosphate, citrate, and other Buffers such as organic acids; antioxidants including ascorbic acid and methionine; preservatives (such as chlorinated Benzalkonium chloride;Cutadecyldimethylbenzylammonium chloride;Hexamethonium chloride benzethonium chloride; phenol, butyl, or benzyl alcohol; methyl or Propylparaben and other alkylparabens; catechol; resorcinol; cyclohexyl 3-pentanol; and m-cresol; low molecular weight (less than about 10 residues) poly Peptides; proteins such as serum albumin, gelatin, or immunoglobulins; polyvinyl chloride Hydrophilic polymers such as ruthenium pyrrolidone; glycine, glutamine, asparagine, histidine , arginine, lysine, and other amino acids; monosaccharides, disaccharides, and glucose, mannose, or other carbohydrates, including dextrin; chelating agents such as EDTA; sucrose, mannitol, etc. a sugar such as tallow, trehalose, or sorbitol; a salt-forming counterion such as sodium; gold complexes (e.g., Zn-protein complexes); and / or polysorbates (e.g., T WEEN™), poloxamers (e.g., PLURONICS™), or The active agent may include a non-ionic surfactant such as polyethylene glycol (PEG). For example, by coacervation techniques or by interfacial polymerization, Hydroxymethylcellulose or gelatin microcapsules and poly(methyl Colloids in microcapsules prepared by methacrylate microcapsules Drug delivery systems (e.g., liposomes, albumin microspheres, in chromium emulsions, nanoparticles and nanocapsules) or in macroemulsions Such techniques are described in Remington's Pharmacy, The results are published in the journal Cal Sciences.

[0078] Sustained release formulations of the active agent can be prepared. Suitable examples of sustained release formulations include The matrix may be in the form of a shaped article, For example, in the form of a film or a microcapsule. are polyesters, hydrogels (e.g., poly(2-hydroxyethyl-methacrylate) ), or poly(vinyl alcohol)), polylactide (US3773919), L-glucose Copolymer of glutamic acid and gamma-ethyl-L-glutamate, non-degradable ethylene-acetate Injectable microspheres composed of vinyl, lactic acid-glycolic acid copolymer and leuprolide acetate Degradable lactic acid-glycolic acid copolymers such as Fair (LUPRON DEPOT™) ) and poly-D-(-)-3-hydroxybutyric acid.

[0079] Formulations include those suitable for routes of administration as detailed herein. and may be prepared by any of the methods well known in the art of pharmacy. The techniques and formulations are generally based on Remington's Pharmaceuticals Such methods involve mixing the active agent with one or more accessory components. Generally, the formulation involves bringing the active agent into association with a liquid carrier or finely divided uniformly and intimately bringing into association the product with a solid carrier, or both, and then, if necessary, shaping the product. Manufactured by.

[0080] Formulations of the active agent suitable for oral administration may be in the form of pills, capsules, or capsules, each containing a predetermined amount of the active agent. It may be prepared as discrete units such as capsules or tablets.

[0081] Compressed tablets may optionally contain binders, lubricants, inert diluents, preservatives, surface active agents or The active ingredient in a free-flowing form such as a powder or granules mixed with a dispersing agent is compressed in a suitable machine. Molded tablets may be prepared by mixing the powdered active ingredient moistened with an inert liquid diluent. The mixture can be made by molding in a suitable machine. Tablets can be made by optionally Can be coated or scored, optionally for slow or controlled release of active agent therefrom. The compound is formulated to provide release.

[0082] Tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions capsules, hard or soft capsules, e.g. gelatin capsules, syrups or elixirs are administered intravenously The formulation of the active agent intended for oral use may be prepared according to the method of preparation of the pharmaceutical composition. The composition may be prepared according to any method known in the art for the manufacture of such compositions. Such compositions may contain sweetening, flavoring, coloring and preservative agents to provide a palatable preparation. The composition may contain one or more agents, including preservatives. Tablets containing the active agent mixed with an excipient are acceptable. These excipients include, for example, calcium carbonate. or sodium carbonate, lactose, calcium phosphate or sodium phosphate, etc. Inert diluents, granulating and disintegrating agents such as corn starch or alginic acid, de Binders such as starch, gelatin, or acacia, and magnesium stearate, sucrose, Lubricants such as stearic acid, talc, etc. Tablets may be uncoated or may be coated with Delays disintegration and absorption in the intestinal tract, thereby providing a sustained effect over a longer period The particles may be coated by known techniques, including microencapsulation for example. A time delay material such as glyceryl monostearate or glyceryl distearate alone It may be used alone or with a wax.

[0083] Aqueous suspensions contain the active agent in admixture with excipients suitable for the manufacture of aqueous suspensions. Such excipients include sodium carboxymethylcellulose, croscarmellose, and povidone. Methylcellulose, Hydroxypropylmethylcellulose, Sodium alginate, Suspending agents such as polyvinylpyrrolidone, gum tragacanth and gum acacia, and natural Phosphatides (e.g. lecithin), condensation products of alkylene oxides with fatty acids (e.g. , polyoxyethylene stearate), condensation of ethylene oxide with long-chain aliphatic alcohol Synthesis products (e.g., heptadecaethyleneoxycetanol), ethylene oxide and fatty acids and condensation products with partial esters derived from hexitol anhydrides (e.g., polyoxyethylene Aqueous suspensions may contain dispersing or wetting agents such as sodium sorbitan monooleate. , ethyl p-hydroxybenzoate or n-propyl p-hydroxybenzoate one or more preservatives, one or more coloring agents, one or more flavoring agents, and sucrose or saccharin. The composition may also contain one or more sweetening agents such as glycerin.

[0084] The pharmaceutical compositions of the active agents may be in the form of a sterile injectable preparation, such as a sterile injectable aqueous or oleaginous suspension. This suspension can be formulated using suitable dispersing or wetting agents and suspending agents, as mentioned above. It can be formulated according to known techniques. Sterile injectable formulations can be prepared by adding 1,3-butanediol to the formulation. A sterile injectable solution or suspension in a non-toxic parenterally acceptable diluent or solvent, such as The sterile injectable preparation may also be prepared as a lyophilized powder. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution. For this purpose, sterile, fixed oils may be conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil may be used including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid can likewise be used in the preparation of injectables. do.

[0085] The amount of active agent which can be combined with a carrier material to produce a single dosage form will vary depending on the therapeutic These may vary depending on the host and the particular method of administration. For example, The sustained release formulation may range from about 5 to about 95% (weight:weight) of the total composition. The pharmaceutical composition may contain from about 1 to 1000 mg of active agent combined with a convenient amount of carrier material. The compositions can be prepared to provide easily measurable amounts for administration. For example, Aqueous solutions intended for intravenous infusion can be administered in suitable volumes at a rate of approximately 30 mL / hour. For example, each milliliter of solution may contain about 3 to 500 μg of active agent.

[0086] Formulations suitable for parenteral administration may include additives such as antioxidants, buffers, bacteriostats and agents that are suitable for the intended recipient of the formulation. Aqueous and non-aqueous sterile injection solutions, which may contain solutes that render them isotonic with the blood of the patient, as well as suspensions and and thickening agents.

[0087] Formulations for rectal administration may be prepared with a suitable base comprising, for example, cocoa butter or a salicylate. The pharmaceutical composition may be provided as a suppository.

[0088] Formulations suitable for pulmonary or nasal administration include, for example, formulations having a particle size in the range of 0.1 to 500 microns. Diameter (0.1 micron, increasing in micron increments such as 0.5, 1, 30 microns, 35 microns, etc.) The particles have a diameter ranging from 100 to 500 microns, which allows them to reach the alveolar sacs. It is administered by rapid inhalation through the nasal passages or by inhalation through the mouth. Formulations suitable for aerosol or dry powder administration can be prepared by conventional methods. and can be used in the treatment or prophylaxis of disorders as described below. The therapeutic agent may be delivered along with other therapeutic agents, such as compounds that have previously been used in

[0089] Formulations suitable for vaginal administration include those known in the art that are suitable in addition to the active agent. Pessaries, tampons, creams, gels, pastes, and foams containing such carriers Alternatively, it may be provided as a spray formulation.

[0090] The formulations may be packaged in unit-dose or multi-dose containers, such as sealed ampoules and vials. and just prior to use, for injection, only the addition of a sterile liquid carrier, e.g., water, may be required. It can be stored in a lyophilized (freeze-dried) state, requiring no preparation. Suspensions may be prepared from sterile powders, granules and tablets of the kind previously described. The agent may be administered in a daily dose or unit daily sub-dose as described herein above, or thereabouts. The active agent may include an appropriate portion of the active agent.

[0091] The subject matter concerns a veterinary drug comprising at least one active agent as defined above together with a veterinary carrier. The compositions are further provided. A veterinary carrier is a material useful for administering the composition, otherwise inert or acceptable in the veterinary arts and compatible with the active agent. These veterinary compositions may be administered parenterally or in the form of a solid, liquid, or gaseous substance. The composition may be administered orally, or by any other desired route.

[0092] IV.Manufactured products In another aspect, described herein are articles of manufacture, e.g., methods for treating pancreatic cancer. The kit contains useful substances such as MEK inhibitors for the treatment of pancreatic cancer. or a pharma- ceutical acceptable salt thereof, and a compound that targets PDGFRα, S6, and STAT3. A container containing a combination of a multikinase inhibitor or a pharma- ceutical acceptable salt thereof. The kit may further comprise a label or package insert on or associated with the container. "Package insert" refers to the indications, directions for use, dosage, administration, and / or other information regarding the use of such therapeutic product. Information normally included in commercial packaging of therapeutic products, including information on contraindications and / or warnings Suitable containers include, for example, bottles, vials, Containers include cartridges, blister packs, etc. Containers can be made of a variety of materials such as glass or plastic. The container can be formed from a material that is effective in treating the condition. a pharma- ceutical acceptable salt thereof and a multikinase inhibitor or a pharma- ceutical acceptable salt thereof and may have a sterile access port (e.g., the container may be The intravenous solution bag or vial may have a stopper that can be penetrated by a needle. The label or package insert indicates that the composition is used for treating pancreatic cancer. Alternatively or in addition, the product may be a bacteriostatic solution for injection (BWFI), phosphate buffered saline, a second container containing a pharma- ceutically acceptable buffer, such as dextrose solution, It may further include other buffers, diluents, filters, needles, and syringes. The invention may include other materials desirable from a commercial and user standpoint, including, for example,

[0093] The kit comprises a MEK inhibitor or a pharma- ceutical acceptable salt thereof, and a separately formulated If present, instructions for administration of a multikinase inhibitor or a pharma- ceutical acceptable salt formulation thereof For example, the kit may further include a MEK inhibitor or a pharma- ceutically acceptable salt thereof. a first composition comprising a salt, and a multikinase inhibitor or a pharma- ceutically acceptable salt thereof; When the kit includes a second pharmaceutical formulation containing the first and second It may further include instructions for the simultaneous, sequential or separate administration of the pharmaceutical compositions.

[0094] In another embodiment, the kit comprises a combination of a MEK inhibitor or a pharma- ceutical acceptable salt thereof. and a combination of a cyclophosphamide, ... and a cytochrome P450 inhibitor or a pharma- ceutical acceptable salt thereof. Such a kit preferably includes a plurality of unit dosages. The kit may include a card having the doses arranged in the order of their intended use. An example of such a kit is a "blister pack". Blister packs are a common feature of the packaging industry. are well known and widely used in packaging pharmaceutical unit dosage forms. In the form of letters, letters, or other markings, or a treatment schedule by which dosages may be administered. A memory aid can be provided with a calendar insert designating the days of the week.

[0095] According to one embodiment, the kit comprises: (a) a MEK inhibitor or a pharma- ceutically acceptable salt thereof; and optionally (b) a multikinase inhibitor or a pharma- ceutical acceptable salt thereof. and a second container containing the salt formulation to be administered. bacteriostatic fluid for immunosuppression (BWFI), phosphate-buffered saline, Ringer's solution, and dextrose solution The method may further include a third container containing a pharma- ceutically acceptable buffer, such as a buffer solution. from a commercial and user perspective, including buffers, diluents, filters, needles, and syringes The composition may contain other materials as desired.

[0096] The kit comprises a MEK inhibitor or a pharma- ceutical acceptable salt thereof and a multikinase inhibitor or In other specific embodiments, including compositions with pharma- ceutically acceptable salts thereof, the kit comprises Containers containing separate compositions, such as separate bottles or individually separated foil packets, Although it is possible to separate the compositions, the separate compositions may be contained within a single, undivided container. Typically, the kit comprises directions for the administration of the separate components. Preferably, when administered in different dosage forms (e.g., oral and parenteral), they are administered at different dosage intervals. or when titration of the individual components of the combination is desired by the prescribing physician. This is particularly advantageous.

[0097] In yet another embodiment, the problem described herein is solved by using a pre-heated boiling water. Culture media that use serum that has been boiled for a period of time before use in the medium. This medium is superior to medium using non-scalded normal serum in terms of the number of spun spores. In an embodiment, the medium for culturing the cells is about 1 0% boiled fetal bovine serum, which is diluted with 95-100% water before being added to the medium. Heat in boiling water at 37°C for 10 minutes.

[0098] The subject matter described herein includes the following specific embodiments.

[0099] 1. A method for treating pancreatic cancer in a subject in need thereof, comprising administering an effective amount of administering to said subject a combination of active agents, said combination comprising an active ingredient selected from the group consisting of a MEK inhibitor and a combination of active ingredients selected from the group consisting of a medicament for treating inflammatory bowel disease. and a pharma- ceutical or pharma- ceutical acceptable salt thereof, targeting PDGFRα, S6, and STAT3. or a pharma- ceutically acceptable salt thereof.

[0100] 2. The above MEK inhibitors are cobimetinib, GDC-0623, trametinib, and binimetinib. Nib, selumetinib, pimasertinib, refametinib, PD-0325901 and B I-847325, or a pharma- ceutically acceptable salt thereof. The method according to embodiment 1.

[0101] 3. The MEK inhibitor is cobimetinib, trametinib, or their pharmaceutical equivalents. 3. The method of any of the preceding embodiments, wherein the compound is selected from the group consisting of a salt thereof that is acceptable for

[0102] 4. The method according to any one of the preceding claims, wherein the MEK inhibitor is cobimetinib or a pharma- ceutical acceptable salt thereof. 3. The method according to any one of the preceding embodiments.

[0103] 5. The method according to claim 1, wherein the multikinase agent is ponatinib or a pharma- ceutically acceptable salt thereof. 20. The method according to any of the preceding embodiments.

[0104] 6. The method of any of the above embodiments, wherein said pancreatic cancer is an endocrine cancer.

[0105] 7. The method of any of the above embodiments, wherein said pancreatic cancer is an exocrine cancer.

[0106] 8. The above pancreatic cancer is adenocarcinoma, acinar cell carcinoma, adenosquamous carcinoma, mucinous carcinoma, osteoclast-like giant cell carcinoma, Undifferentiated carcinoma with cells, hepatoid adenocarcinoma, intraductal papillary mucinous neoplasm, mucinous cystic neoplasm, pancreatoblastoma, Serous cystadenoma, signet ring cell carcinoma, solid pseudopapillary tumor, pancreatic ductal carcinoma, and undifferentiated carcinoma 4. The method according to any of the preceding embodiments.

[0107] 9. The method of any of the above embodiments, wherein said pancreatic cancer is pancreatic ductal adenocarcinoma.

[0108] 10. The method of any of the preceding embodiments, wherein said combination is synergistic.

[0109] 11. The method according to any of the previous embodiments, wherein the active agents are administered sequentially.

[0110] 12. The method according to any of the previous embodiments, wherein said active agents are administered simultaneously.

[0111] 13. The above MEK inhibitor and the above multikinase inhibitor are administered as a combination drug. 4. The method according to any of the preceding embodiments,

[0112] 14. The MEK inhibitor is administered in an amount of about 45 mg to about 75 mg, and the multikinase In any of the above embodiments, the enzyme inhibitor is administered in an amount of about 30 mg to about 60 mg. How to.

[0113] 15. The method according to any of the previous embodiments, wherein said amount is administered once a day.

[0114] 16. The above MEK inhibitor is cobimetinib or its equivalent such as cobimetinib hemifumarate. A pharma- ceutical acceptable salt, wherein the multikinase inhibitor is ponatinib or The method according to any of the above embodiments, wherein the compound is a pharma- ceutical acceptable salt thereof, such as HCl. Law.

[0115] 17. a) MEK inhibitor or its pharmaceutical composition for preventive or therapeutic treatment of pancreatic cancer and b) a multi-kinase inhibitor targeting PDGFRα, S6, and STAT3. Combination with an enzyme inhibitor or a pharma- ceutically acceptable salt thereof.

[0116] 18. A method for treating an MSC inhibitor or a pharma- ceutical agent comprising administering to a patient a therapeutically effective amount of a MEK inhibitor or a pharma- ceutical agent having a PDGFRα, S 6, and a multikinase agent targeting STAT3 or a pharma- ceutical acceptable salt thereof. and a pharma- ceutical acceptable excipient, The composition is for treating pancreatic cancer.

[0117] 19. The above MEK inhibitors are cobimetinib, GDC-0623, trametinib, and binimetinib. tinib, selumetinib, pimasertinib, refametinib, PD-0325901 and BI-847325, or a pharma- ceutically acceptable salt thereof; The composition of any of the preceding embodiments.

[0118] 20. The MEK inhibitor is cobimetinib, trametinib, or a pharmaceutical combination thereof. The composition according to any of the above embodiments, wherein the compound is selected from the group consisting of a physiologically acceptable salt thereof. .

[0119] 21. The above-mentioned MEK inhibitor is cobimetinib or a pharma- ceutical acceptable salt thereof. The composition of any of the preceding embodiments.

[0120] 22. The multikinase agent is ponatinib or a pharma- ceutically acceptable salt thereof. The composition of any of the preceding embodiments.

[0121] 23. Cobimetinib or a pharma- ceutical acceptable salt thereof and ponatinib or a pharma- ceutical acceptable salt thereof and an acceptable salt thereof.

[0122] 24. The MEK inhibitor or a pharma- ceutical acceptable salt thereof is about 45 mg to about 75 mg. and the multikinase inhibitor or a pharma- ceutical acceptable salt thereof is present in an amount of about 30 mg to about 50 mg. The composition of any of the preceding embodiments, wherein the composition is present in an amount of about 60 mg.

[0123] 25. A method for treating a patient with a MEK inhibitor or a pharma- ceutical acceptable salt thereof, and a combination of PDGFRα, S6, and S A multikinase agent or a pharma- ceutical acceptable salt thereof that targets TAT3, a container, and and a supporting document or label.

[0124] The following examples are offered by way of illustration and not by way of limitation. EXAMPLES

[0125] Materials and Methods Cell lines and reagents: PA-TU-8988T cells derived from the DSMZ and SUIT-2 cells derived from the JCRB All cell lines were from ATCC, except for . All cell lines were obtained from ATCC with the identity of the cell line confirmed. Genentech Cell Line Core routinely performs SNP and STR analysis to identify All cell lines were maintained in a 10% FBS, 2 mM L-glutamine, 100% CO. RPMI medium (Gibco) supplemented with penicillin and streptomycin at 1 ml Rapamycin and ponatinib were obtained from Selleckchem (Hepatitis C). Crenolanib was obtained from Arog Pharmaceuticals, and Covi Metinib, GDC-0941, and GDC-0980 were synthesized at Genentech. The human phosphokinase array kit was purchased from R&D Systems (ARY003B). , and Cignal-45-Pathway reporter assay was purchased from Qiagen (CCA-9 01L), the human tyrosine kinase RT2 Profiler PCR array is gen (PAHS-161Z), and the MDSC isolation kit was from Miltenyi Biotech. All assays using these kits were performed according to the manufacturer's instructions. provided.

[0126] Western blotting and cell viability assays: Cells were seeded in 10 cm dishes and treated with 1 μM small molecule inhibitors for 24 h. Lysates were purified by ELISA using a protease inhibitor cocktail (Thermo Scientific) The cells were prepared in RIPA lysis buffer (Thermo Scientific) containing DS-PAGE was performed and proteins were transferred to a nitrocellulose membrane. Imaging was performed using standard methods. Protein analysis was performed using ImageJ software. The bands were quantified. Primary antibodies: p-Erk, p-STAT3, p-S6, total S6, pP DGFRα, p-PDGFRβ, p-RSK3, cleaved PARP, Bcl-xL, Mcl, Survivin, Rab11, p-Akt, β-actin, GAPDH (Cell Sign aling), total STAT3, total PDGFRα (Santa Cruz Biotech nology), p-EphA7 (Gene Tex), and p-EphA2 / 3, phA2 / 5(MyBioscience).

[0127] Cell viability assays involve injecting cells into a dose titration of various pharmacological inhibitors (0.001–10 μg / mL). M) or a fixed dose (1 μM) for 72 hours and then analyzed by CellTiter Glo (Pro The survival rate was measured using a Bliss score as previously described. (Borisy AA, Elliott PJ, Hurst NW, Lee MS, Lehar J, Price ER, Systematic d discovery of multicomponent therapeutics, Proceedings of the National Academy of S. sciences of the United States of America, 2003;100(13):7977-82,Epub 2003 / 06 / 12).

[0128] Lentiviral infection: Lentiviral particles against shSTAT3 were generated as previously described. (Moffat J, Grueneberg DA, Yang X, Kim SY, Kl oepfer AM,Hinkle G,A lentiviral RNAi lib rary for human and mouse genes applied o an arrayed viral high-content screen,C ell,2006;124(6):1283-98.Epub 2006 / 03 / 28) shSTAT3 in the Dox-inducible system was obtained from LakePharma Inc. The target sequence used for shSTAT3 was AATCTTAGCAGGAAGGTGC KP4 and MIA-PACA2 cells were infected with lentivirus at optimized titers. The cells were transduced with 1 μg / ml puromycin and selected.

[0129] Luminex Assays and ELISA: Culture supernatant from cancer cells cultured for 72 hours or plasma samples collected at the end point from animal studies Luminex assays for the samples were multiplexed according to the manufacturer's instructions. ELISA for PDGFα was performed using a kit (Bio-Rad). The assay was performed using a kit from systems.

[0130] Invasion assay: Cells in RPMI containing 2% FBS were inoculated onto BD BioCoat Matrigel invasion tubes. The cells were seeded onto inverted inserts in the lower chamber (BD Biosciences). The plates were filled with RPMI supplemented with 10% FBS as a chemoattractant and incubated for 18 h. After incubation, the number of cells that had invaded the lower chamber was counted using a SpectraMax M5 (Mo Measurements were performed using a vitreous endoscopic device.

[0131] Animal testing: All animal experiments were conducted in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals. All dosing regimens were well tolerated in the xenografts. We used: human PDAC-derived cell line KP4 and KPP-GEMM mouse isolated cell lines. Cells derived from immunocompetent tumors were cultured at 5 × 106 cells without any matrigel. All mice (Charles River Laboratories) were injected subcutaneously into the right flank. After tumors reached approximately 200 mm3, mice were randomized and treated with vehicle (MC T+25mM citrate buffer, pH 2.75), cobimetinib (5mg / kg)+ / - Ponatinib (30 mg / kg) was administered PO, QD at the indicated intervals. Tumor volumes were measured using Cal IV calipers. Repeated tumor volumes from the same animals were To adequately analyze the return measurements over time, a mixed modeling approach was used. ose Pinheiro DB,Saikat DebRoy,Deepayan S arkar,R Core team.Linear and Nonlinear m ixed effects models.Package 'nlme'2008). This approach accounts for repeated measures and for all non-treatment-related deaths of animals before the end of the study. The time course of log2 tumor volume at each dose level was A cubic regression spline was used to fit the largely nonlinear profile. Thus, These nonlinear profiles were related to dose in a mixed model. Tumor growth inhibition (% TGI) as an index was calculated as 1 / 1 ratio relative to vehicle using the following formula: Percentage of area under the fitted curve (AUC) for each dose group per day It was calculated as: %TGI=100×(1−AUC dose / AUC vehicle)

[0132] The fitted curve and fitted covariance matrix were used to determine the uncertainty interval (UI) of the %TGI. A random sample was generated as an approximation to the distribution of %TGI using the It consisted of 1,000 simulated realizations of the mixed model, with the %TGI recalculated for each realization. The reported UI is the recurrence rate of %TGI 95% of the time given the fitted model. The calculated value was within this range: 2.5 percentile of the simulated distribution and 97. The 5th percentile was used as the upper and lower UI. >60% tumor growth inhibition was The results were deemed significant. (Wong H, Choo EF, Alicke B, Ding X ,La H,McNamara E,Antitumor activity of t Argeted and cytotoxic agents in murine s ubcutaneous tumor models correlates with clinical response,Clinical Cancer Resea rch:an official journal of the American Association for Cancer Research,2012;18( 14):3846-55.Epub 2012 / 06 / 01).

[0133] Plots were generated using R version 2.8.1 (R Development Core Te am 2008;R Foundation for Statistical Com putting; Vienna, Austria) and Excel version 12.0. 1 (Microsoft Corporation). The data were analyzed using R, version 2.8.1, and mixed models were analyzed using the nlme package, Fits were generated in R using version 3.1-89 ( 41 ).

[0134] Immunohistochemistry and immunofluorescence: Immunohistochemistry and immunofluorescence were performed on formalin-fixed, paraffin-embedded, cell pellets, whole Tissue sections and 4μ sections cut from TMA blocks (US Biomax, Inc.) Slides were deparaffinized and antigen retrieval was performed using the Target Retrieval Solution (Dako) was used to obtain the PT Module (Th Endogenous peroxidase activity was assayed using 3% H2 Blocking with O2 and 3% BSA was used to block endogenous immunoglobulins. p-Erk and Primary antibody against p-STAT3 (Cell Signaling) at 1 μg / ml; p - PDGFRα (Cell Signaling) at 0.4μg / ml, F4 / 80 (Serotec) at 10 μg / ml, Gr-1 (BD Pharmingen) at 1 μg / ml, and cleaved caspase 3 (Cell Signaling) at 0.6 μg / ml. CD8α (Genentech) was used at 5 μg / ml. Antibody binding was For IHC staining, Vecstatin ABC Elite Horseradish Pepper Peroxidase (Vector Laboratories) or PowerVisi on poly-HRP (Leica Biosystems) and metal-enhanced 3,3'-diazide For immunofluorescence staining, we used the TSA kit (Li Fluorescent Technologies) and PowerVision Poly-HRP (Leufe Technologies) The IHC slides were counterstained using a Ma The analysis was performed using Yer's hematoxylin (Rowley Biochemical).

[0135] Histology: All slide images were scanned using the Nanozoomer XR automated slide scanning platform. Hamamatsu, Hamamatsu City, Shizuoka Prefecture ef., Japan) at a final magnification of 200x.

[0136] Microarray and RNA sequencing analysis: Gene expression in normal and PDAC human tissues was analyzed using the Gene Logic Aff Gene expression data was measured using the ymetrix HGU133P array. Normalization was performed by the Random Number Average (RMA) method. Expression probe set 203131_ at to human PDGFRα, 202273_at to human PDGFRβ, 205945_ at was used for human IL6-R and 204171_at for PRS6KB1.

[0137] statistical analysis All data are expressed as the mean ± standard error of the mean (SEM). A two-tailed t-test was used to compare the two groups and was performed using Prism or Excel. P values ​​were calculated using the Kaplan-Meier method. P<0.05 was considered significant. Survival curves were analyzed using the Kaplan-Meier method. was used to plot.

[0138] Example 1: Sensitivity to MEK inhibition in pancreatic cancer cell spheroid cultures Pancreatic cancer cells grown in two-dimensional (2D) and three-dimensional (3D) cultures can be used to treat a variety of cancers To compare how KRAS-mutant PDAC cancer cells respond to The cell line KP4 was then screened against a panel of small molecule inhibitors of 203 cancer-relevant targets and known chemotherapeutic drugs. and treated with (Table 1). [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5]

[0139] With respect to 3D culture, described herein is a relatively rapid "high-throughput" analysis. This is a method to promote precipitation. Before adding to the medium, pre-boil the solution at 95-100℃ for 10 minutes. Cultivating cancer cells in medium containing heated 10% fetal bovine serum consistently results in 3D “staining” It was observed that "spheroids" formed rapidly. Lloyd noted predominant localization of Ki67 staining in the peripheral layer and cleavage debris up to 72 hours. Low-level necrotic / apoptotic centers with apoptosis indicated by MAP-3 The cells exhibited all the typical characteristics of spheroids previously described, including proliferation of bells (Figure 2 7) Their drug sensitivity patterns were also similar to those of previously described spheroids. (Figures 28 and 29).

[0140] Treatment of KP4 cells in 2D and 3D culture conditions with a panel of 203 compounds These results revealed different sensitivities to several drugs (Figure 1). KP4 spheroids are more resistant to chemotherapy, and KP4 spheroids are more resistant to chemotherapy. were indeed significantly more resistant than 2D cultures to a subset of 18 inhibitors from the Surprisingly, the inventors also found that 3D culture was more effective than 2D culture. We identified 11 inhibitors that inhibited multiple signaling pathways (Figure 1). The inhibitors consisted of various molecules that have been reported to have activity targeting these Four of them (cobimetinib, GDC-0623, AZD6244, and PD901) target the MEK pathway. and three target the PI3K pathway (GDC-0941, GDC-0980, G 38390), and one that targets the MET pathway (G45203).

[0141] Example 2: Multiple differentially regulated 2D vs. 3D culture of pancreatic cancer cells Signal transduction pathways> Scaffolding potentially contributes to the observed differential treatment sensitivity of pancreatic cancer cells in 2D vs. 3D. To identify signal transduction pathways, we analyzed KP4 cells under 2D and 3D conditions. Phosphokinase arrays, gene expression arrays, and luciferase expression reports were performed in cells. A transduction assay was performed and the changes in the expression levels of signaling proteins were compared. Phosphokinase arrays in KP4 spheroids compared to KP4 cells cultured in 2D In this study, p38, ERK, EGFR, MSK, Akt, TOR, CREB, and HSP-27 were , STAT2, STAT5α, Hck, Chk-2, c-Jun, RSK1 / 2 / 3, e Tyrosine depletion in several signaling proteins, including NOS, p27, and PLC-γ1 Showed increased phosphorylation, suggesting a substantial "rewiring" of signaling pathways in 3D conditions In KP4 monolayer cultures, MEK inhibition inhibited the activation of ERK, RSK, and and down-regulates PLC-γ1 phosphorylation, and up-regulates STAT family proteins and In contrast, KP4 spheroids showed increased Akt activation upon MEK inhibition. In addition to ERK, RSK, and PLC-γ1, Akt and STAT family proteins MEK inhibition reduced c-Jun activation in KP4 spheroids. Only activation was induced.

[0142] mRNAs corresponding to protein kinases in KP4 monolayer and spheroid cultures qRT-PCR analysis of the expression of A. cerevisiae revealed a number of RTKs, particularly FGFR3, EP Increased expression of HA1, EPHB6, and INSR in KP4 spheroids We demonstrated that MEK inhibition suppressed these cell deaths in both KP4 monolayers and spheroids (Figure 4). However, PDGFRα and PDGF Rβ expression was significantly upregulated in KP4 monolayer cultures upon cobimetinib treatment. In contrast, expression of EPHA1 and EPHB1 was upregulated in KP4 spheroids upon MEK inhibition. The increase in the IL-10 signal was selectively observed in the IL-10 signal-receptor region (Figure 4). PDAC-derived leukocytes may contribute to the different sensitivity to drug treatment observed compared with those Reveal drug response profiles specific to 3D culture conditions for cells.

[0143] Transcription upon cobimetinib treatment using a large panel of luciferase reporter assays Further evaluation of changes in factor pathway activation demonstrated that KP4 monolayer cultures upregulate STAT3, c-Myc, We found that the expression of GRE, KLF4, ISRE, and GAS transcription factors increased activation. In contrast, KP4 spheroids expressed similar levels of these transcription factors. These results demonstrated no activation of the MEK pathway in monolayer versus spheroid conditions. Multiple pathways that may contribute to the activation of alternative pro-survival signaling pathways in response to inhibition This implies differential regulation of transcription factors.

[0144] The results of phosphokinase array and gene expression array were compared with those of cobimetinib treatment of KP4 cells. p-PDGFRα, p-STAT3, p-Erk, p-S6, and p-RSK during treatment The results were verified by Western blot analysis of p-PDGFRα, p-STAT3, and Increased activation levels of p-STAT3 and p-S6 were observed (Fig. 6A). was reproducibly detected upon cobimetinib treatment in multiple PDAC human cancer cell lines (Figure 6B). Importantly, IL-6 ligands can promote STAT3 activation. Increased secretion of STAT3 was also observed in multiple PDAC cell lines (Figure 7). Consistent with a requirement for activation, STAT3 RNase activity in KP4 and MIA-PACA2 cells was significantly increased in Ai knockdown induced cell death upon cobimetinib treatment (Figures 8 and 9). In vitro cell lines using multiple PDAC cell lines and KPP-GEMM tumor-derived cell lines Invasion assays demonstrated a decrease in the invasive ability of cancer cells upon MEK inhibition (Figure 10). Our results suggest that MEK inhibition as a single agent is not sufficient to cause primary tumor regression. , suggesting that it may inhibit the invasive ability of cancer cells. showed that the signaling pathway mediated by PDGFRα, S6, and STAT3 regulates pancreatic These results suggest that MEK inhibition in cancer cells may enable cancer cell survival.

[0145] Example 3: Inhibition of PDGFRα, S6, and STAT3 with MEK inhibition enhances PDA Effectively promote apoptosis in C cells> Previous studies have shown that MEK inhibition upregulates and activates multiple RTKs as well as “bypasses” "We report that the IL6 / JAK / STAT pathway may be a survival mechanism. There are (Duncan JS, Whittle MC, Nakamura K, Abell AN,Midland AA,Zawistowski JS,Dynamic re programming of the kinome in response to targeted MEK inhibition in triple-negat ive breast cancer,Cell,2012;149(2):307-2 1.Epub 2012 / 04 / 17, Lee HJ, Zhuang G, Cao Y, Du P, Kim HJ, Settleman J, Drug resistance via feedback activation of Stat3 in onco gene-addicted cancer cells,Cancer Cell,2 014;26(2):207-21,Epub 2014 / 07 / 30, Dai B,M eng J, Peyton M, Girard L, Bornmann WG, Ji L ,STAT3 mediates resistance to MEK inhibition tor through microRNA miR-17, Cancer Resea rch,2011;71(10):3658-68,Epub 2011 / 03 / 30) However, pancreatic cancer was not involved. S6 and the JAK2 / STAT3 cascade Ponatinib, a multi-RTK inhibitor that targets both RA and RA alone or in combination with cobimetinib, The efficacy of cobimetinib was tested in tumor cell proliferation assays. In mice, it promoted the activation of STAT3 and S6 (Figure 6A), and cobimetinib / ponatinib Combined treatment with PDGFRα, STAT3, and S in 2D and 3D cultures 6. Inhibits activation of ERK1 / 2 and Akt, and suppresses the expression of pro-survival proteins Bcl-XL and Mcl -1, reduced survivin expression and increased cleaved PARP levels (Figure 11, 12 and 13). Ponatinib potentially inhibits downstream signaling pathways in PDAC cells. It has been previously reported that β-lactamase inhibits the kinase activity of ephrin receptors, which can be regulated by However, in PDAC models, the combination treatment with cobimetinib / ponatinib has It affects the activity of phospho-receptors (EphA2 / 3, EphA2 / 5, and EphA7). No (Figure 11).

[0146] Ponatinib inhibits not only S6 and JAK2 / STAT3 signaling, but also FGFR, In addition to inhibiting VEGFR and ephrin receptors, PDGFRα is also inhibited, and the inventors of the present application have Examine the effects of other PDGFRα, S6, and JAK2 inhibitors in combination with metinib The selective activity of ponatinib was further investigated by comparing it with the EGFR inhibitor erlotinib or In contrast to the effects of PDGFR inhibitors, cobimetinib and S6 inhibitors, rapamycin, or downregulation of S6 activation by combination treatment with GDC-0980 in KP4 cells The PDGFRα inhibitor, Cleno, promoted cell death in the rats (Figures 14, 18, and 19). A triple combination of ranib, a JAK2 inhibitor, ruxolitinib, and cobimetinib Concomitant downregulation of PDGFRα and STAT3 activation after treatment also suppressed KP4 cell proliferation. The results showed that the IL-11 expression of IL-11 inhibited the expression of IL-11 and caused significant cell death (Figure 15). , co-targeting RTK, S6, and JAK / STAT and inhibiting proliferation of PDAC cells The strongest synergy was observed with cobimetinib / ponatinib treatment, which effectively inhibited 11, 12, 13, 14, 16, and 17). Cobimetinib and / or ponatinib Treatment of PDAC cell lines with 2 were synergistic in inhibiting cell proliferation in combination in both D and 3D cultures, and PDA To clarify the combination of cobimetinib / ponatinib as a potentially effective treatment strategy for C. This was made clear (Figures 12, 16 and 17).

[0147] Example 4: Cobimetinib / ponatinib co-treatment induces tumor regression in mouse models > To test the efficacy of the cobimetinib / ponatinib combination in vivo, The inventors performed xenograft studies using KP4 and KPP-GEMM-derived tumor cells Both cobimetinib and ponatinib were ineffective as single agents in KP4 xenografts. However, cotreatment with cobimetinib and ponatinib resulted in significant inhibition of tumor growth and the cleavage of This resulted in increased cell death accompanied by an increase in caspase 3 (Fig. 20 (KP4 cells), 21 and 22). Similar results were obtained with KPP-GEMM-derived cell line xenograft tumors, We observed a significant delay in tumor growth, as opposed to predominantly tumor regression (Figure 20). Cobimetinib / ponatinib combination treatment also inhibited IFN-γ expression in KP4 xenografts. 103% TGI (% tumor growth inhibition) in guinea pigs and 71% TGI in KPP xenografts The combination of cobimetinib / ponatinib treatment was shown to be effective in the KP4 xenograft study. In mice, it caused some associated weight loss, but not due to KPP-GEMM. In the KP4 xenograft study, the incidence was >20%. Two mice had to be withdrawn early from the study due to weight loss. During the course of the combination treatment, significant changes above the acceptable threshold were observed in the remaining mice. No significant weight loss was observed (Figure 31A). Histopathological analysis of the transplanted tumors, IH C staining and immunoblotting showed that p-PDGFRα and p- We confirmed the expected induction of STAT3 and pS6, which was consistent with cobimetinib and ponatinib. was significantly down-regulated following co-treatment with (Figures 21, 22, 30, and 33). High serum levels of PDGFα, PDGFβ, and increased levels of IL-6 were also associated with This was observed in bimetinib-treated mice (Figure 23). Treatment reduced serum PDGFα and PDGFβ levels but not VEGF or FGF levels. ponatinib did not reduce PDGFRα expression in PDAC cancer models, but rather specifically inhibited the PDGFRα pathway in The association was highlighted (Figures 23 and 32).

[0148] Analysis of the myeloid cell compartment within PDAC tumors revealed that F4 / 80+ tumor-associated macrophages (TA M) was found to be the predominant myeloid population (Figure 21). Co-treatment with tinib significantly reduced the number of TAMs in tumors and upregulated the macrophage inhibitory factor (GCS). Induces serum levels of F, and macrophage-activating factors GM-CSF and MCSF These findings suggest that cobimetinib / ponatinib reduced the levels of Co-treatment of PDGFRα inhibited downstream MEK, S6, and STAT3 survival pathways, as well as PDGFRα Targeting the pathway is effective in neutralizing both tumor and myeloid cell populations. Furthermore, the findings suggest that this combination treatment may promote tumor regression. This suggests that: [Table 2]

[0149] Example 5: Differential activation of Erk and STAT3 in liver metastases of PDAC patients To explore the potential clinical relevance of these findings, we investigated p-Erk and IHC of 76 human PDAC tissue samples and 4 liver metastases for p-STAT3 and p-STAT3 Analysis was performed. Erk was primarily detected in patients with stage I and II malignancies. , was activated in 13% of PDAC tissue samples (10 / 76), and STAT3 activation was was observed in 26% of the samples (20 / 76), and 6.5% of the samples (5 / 76) were primarily In stage I malignancies, both Erk and STAT3 were activated (Figure 2 4 and 25). Furthermore, 50% of liver metastases showed Erk activation (2 / 4), and S TAT3 activation was seen in 25% of samples (1 / 4) (Figures 24 and 25). Furthermore, gene expression analysis demonstrated that PDAC tumors from human patients exhibited increased expressing comparable levels of the RTKs PDGFRα, PDGFRβ, and IL6-R. This was further clarified (Figure 26, Table 3). [Table 3-1] [Table 3-2] [Table 3-3] [Table 3-4] [Table 3-5]

[0150] Data in Table 3 demonstrate that PDAC tumors express PDGFR, IL-6, and STAT3 signaling. and that these pathways limit the response to MEK pathway inhibition. These findings also suggest that the MEK pathway may be a key regulator of pancreatic cancer in human patients. Survival of tumor cells in liver metastases without switching to a STAT3-mediated survival cascade upon migration This shows that it can play an important role in

[0151] All technical and scientific terms used herein have the same meaning. Although efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperatures, etc.), , some experimental error and deviation should be considered.

[0152] Throughout this specification and the claims, the words "comprise", "including The words "comprises" and "including" are used except as otherwise required by the context. Except where otherwise noted, the terms are used in a non-exclusive sense. It is understood to include embodiments "consisting of" and / or "consisting essentially of".

[0153] As used herein, the term "about" when referring to values ​​means any In some embodiments, ±50%, in some embodiments, ±20%, in some embodiments, In some embodiments, ±10%, in some embodiments, ±5%, in some embodiments In some embodiments, the range is ±1%, in some embodiments ±0.5%, and in some embodiments ±1% from the specified amount. In embodiments, the present invention is intended to encompass variations from the specified amounts of ±0.1%. Such variations may be appropriate for carrying out the disclosed methods or using the disclosed compositions. This is because.

[0154] When a range of values ​​is given, the upper and lower limits of the range are used, unless the context clearly dictates otherwise. Between the lower limit and any other stated or intermediate value within that stated range , each intermediate value is inclusive to one-tenth of the unit of the lower limit. The upper and lower limits of these smaller ranges that may be included in the ranges also include any are included subject to any specifically excluded limitations. If one or both of the bounds are included, one or both of those included limits are excluded. Ranges are also included.

[0155] Many modifications and other embodiments described herein are within the scope of the foregoing description and related teachings. BRIEF DESCRIPTION OF THE DRAWINGS It will be readily apparent to one skilled in the art to which this subject pertains having the benefit of the teachings presented in the accompanying drawings. It will be appreciated, therefore, that the subject matter should not be limited to the particular embodiments disclosed, but rather should be modified as necessary. It is understood that these and other embodiments are intended to be within the scope of the appended claims. Although specific terms are used herein, they are intended only in a generic and descriptive sense. Used for taste only and not for limitation purposes.

Claims

1. 1. A method of treating pancreatic cancer in a subject in need thereof, comprising administering a therapeutically effective amount of Administering to the subject a combination of active agents, the combination comprising a MEK inhibitor and a multikinase agent that targets PDGFRα, S6, and STAT3. ,method.

2. The MEK inhibitor is cobimetinib, GDC-0623, trametinib, or binimetinib. , selumetinib, pimasertinib, refametinib, PD-0325901 and BI- 847325, or a pharma- ceutically acceptable salt thereof.

1. The method according to claim 1.

3. The MEK inhibitor is selected from the group consisting of cobimetinib, trametinib, and pharma- ceuticals thereof.

3. The method of claim 2, wherein the compound is selected from the group consisting of an acceptable salt thereof.

4. 3. The method of claim 2, wherein the MEK inhibitor is cobimetinib or a pharma- ceutically acceptable salt thereof. The method according to

5. The method of claim 1, wherein the multikinase agent is ponatinib or a pharma- ceutically acceptable salt thereof.

1. The method according to claim 1.

6. 2. The method of claim 1, wherein the pancreatic cancer is an endocrine cancer.

7. 2. The method of claim 1, wherein the pancreatic cancer is an exocrine pancreatic cancer.

8. The pancreatic cancer is selected from the group consisting of adenocarcinoma, acinar cell carcinoma, adenosquamous carcinoma, mucinous carcinoma, osteoclast-like giant cell carcinoma, and pancreatic cancer. undifferentiated carcinoma with cystic fibrosis, hepatoid adenocarcinoma, intraductal papillary mucinous neoplasm, mucinous cystic neoplasm, pancreatoblastoma, serous cystadenoma, signet ring cell carcinoma, solid pseudopapillary tumor, pancreatic ductal carcinoma, and undifferentiated carcinoma. The method of claim 7.

9. The method of claim 8, wherein the pancreatic cancer is pancreatic ductal adenocarcinoma.

10. The method of claim 1 , wherein the combination is synergistic.

11. The method of claim 1 , wherein the active agents are administered sequentially.

12. The method of claim 1 , wherein the active agents are administered simultaneously.

13. The MEK inhibitor and the multikinase inhibitor are administered as a combined formulation The method according to claim 1 .

14. the MEK inhibitor is administered in an amount of about 45 mg to about 75 mg; and the multikinase inhibitor 10. The method of claim 1, wherein the agent is administered in an amount of about 30 mg to about 60 mg.

15. 15. The method of claim 14, wherein the amount is administered once a day.

16. The MEK inhibitor is cobimetinib or a pharma- ceutical acceptable salt thereof, 2. The method of claim 1, wherein the kinase inhibitor is ponatinib or a pharma- ceutically acceptable salt thereof. method.

17. A combination of a) a MEK inhibitor or a pharma- ceutical acceptable salt thereof and b) a P Multikinase inhibitor targeting DGFRα, S6, and STAT3 or pharmaceutical composition thereof In combination with a physiologically acceptable salt.

18. An effective amount of a MEK inhibitor or a pharma- ceutical acceptable salt thereof and PDGFRα, S6, and and a combination of a multikinase agent targeting STAT3 or a pharma- ceutical acceptable salt thereof. and a pharma- ceutical composition comprising a combination of the above and a pharma- ceutical acceptable excipient, wherein the composition is A composition for the treatment of cancer.

19. The MEK inhibitor is cobimetinib, GDC-0623, trametinib, or binimetinib. , selumetinib, pimasertinib, refametinib, PD-0325901 and BI- 847325, or a pharma- ceutically acceptable salt thereof.

19. The composition described in 18.

20. The MEK inhibitor is selected from the group consisting of cobimetinib, trametinib, and pharma- ceuticals thereof.

20. The composition of claim 19, wherein the compound is selected from the group consisting of an acceptable salt thereof.

21. 2. The method of claim 1, wherein the MEK inhibitor is cobimetinib or a pharma- ceutically acceptable salt thereof.

10. The composition according to claim 0.

22. The method of claim 1, wherein the multikinase agent is ponatinib or a pharma- ceutically acceptable salt thereof.

19. The composition described in 18.

23. Cobimetinib or a pharma- ceutical acceptable salt thereof and ponatinib or a pharma- ceutical acceptable salt thereof and a salt thereof.

24. The MEK inhibitor or a pharma- ceutically acceptable salt thereof is present in an amount of about 45 mg to about 75 mg. and the multikinase inhibitor or a pharma- ceutical acceptable salt thereof is present in an amount of from about 30 mg to about 60 mg.

20. The composition of claim 18, wherein the composition is present in an amount of 100 mg.

25. MEK inhibitor or a pharma- ceutically acceptable salt thereof and PDGFRα, S6, and STA A multikinase agent targeting T3 or a pharma- ceutical acceptable salt thereof, a container, and an attached document. A kit comprising:

26. the MEK inhibitor is administered at a dose of about 1 mg / kg to about 50 mg / kg; The method of claim 1, wherein the kinase inhibitor is administered at a dose of about 1 mg / kg to about 50 mg / kg. Method of posting.

27. the MEK inhibitor is administered at a dose of about 1 mg / kg to about 10 mg / kg; 27. The method of claim 26, wherein the kinase inhibitor is administered at a dose of about 10 mg / kg to about 40 mg / kg. The method according to

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