Ponatinib Effectively Inhibits C-KIT Mutations In Patient Derived Tumor Model Of Mucosal Melanoma Of Head And Neck

PURPOSE: Mocosal melanoma is a malignant tumor developed from melanocyte with five years survival is merely 20%.Mocosal melanoma is more common in estern Asia area while is rare in western population.Mocosal melanoma compose 22.6% of all melanoma in China.It is reported that 9%-21% of patient with Mocosal melanoma harboring c-kit mutation,which provide a vital therapeutic target for clinical management.Recent three clinical trials demonstrated that approximately 20% of patients benefit from the treatment of Imatinib.Therefore,it is an urgent problem to overcome Imatinib resistant.The aim of the study is to establish patient derived tumor cell model(PDC)and patient derived tumor xenograft model(PDX)as well as utlize these models to screen and determine sensitive inhibitor for clinical treatment.METHODS: 1.Tumor specimens of mocosal melanoma of Head and Neck were collected and subcutaneously implanted into the immune-compromised nude mice.Xenografted tumor tissue was surgically excised and divided into small fragments and were re-implanted to the second generation nude mice.Histological features between the xenograft tumor and the parental tumor were compared by H-E staining.2.HNMM PDXs harboring c-kit V560 D and K642 E mutation were treated with Imatinib.The volume of xenograft tumor were observed to predict the effect of Imatinib for PDXs harboring c-kit V560 D and K642 E mutation.3.Tumor cell were isolated from xenografted tumor tissue and were dedicated to screen sensitive inhibitors for c-kit V560 D,K642E and D816 V mutation.Effects of c-kit signaling pathway were detected by western blotting after PDC were treated by inhibitors.4.PDXs were selected to validate and determine the sensitivity of inhibitor screened by PDC model.Immunohistochemistry staining of Ki67 and TUNEL apopotosis staining were conducted to investigate the mechanism of the sensitive inhibitors.RESULTS: 1.PDX of HNMM were successfully established and could be re-transplanted in serial generation.The established PDX models retain the complex histopathological features of HNMM.Imatinib showed no significant inhibition on the growth of xengraft tumor harboring c-kit V560 D and K642 E mutation(P>0.05)2.Tumor cell were isolated from xenografted tumor tissue and validated as melanoma cell by Immunofluorescence staining and subcutaneous tumor in nude mice and immunochemistry staining.V560 D,K642E and D816 V mutation were more sensitive to Ponatinib,Dasatinib and Axitinib than other inhibitors.3.Ponatinib could significantly inhibit the growth of xengraft tumor harboring V560 D,K642E and D816 V mutation by inhibit tumor cell proliferation and induce cell apopotosis.CONCLUSIONS: Patient derived tumor xenograft model and Patient derived tumor cell model could be established in mucosal melanoma of Head and Neck and could be a validated preclinical model for drug screening.Ponatinib could significantly inhibit the growth of xengrafted tumor harboring V560 D,K642E and D816 V mutation than Imatinib,which may provide potential therapeutic strategy for HNMM with c-kit mutation.