Pharmacological Effect and Underlying Molecular Insights of a Phytoagent Deoxyelephantopin and its Derivative against BRAF Mutant Melanoma

• Main Authors: Jia-Hua Feng (Kyaw Minn Hsan), 馮家華
• Other Authors: 徐麗芬
• Format: Others
• Language: en_US
• Published: 2016
• Online Access: http://ndltd.ncl.edu.tw/handle/51417790548152793499

博士 === 國立臺灣大學 === 植物科學研究所 === 104 === Melanoma is the most lethal form of skin cancer and cases are increasing year by year. BRAFV600E mutation is found approximately in 50% of melanoma patients that drives constitutive activation of MAP kinase (RAF/RAS/MEK/ERK) signaling for cancer progression. Vemurafenib, an orally available small molecule that specifically targets BRAFV600E mutation, received FDA approval for treatment of late-stage melanoma in 2011. Unfortunately, patients treated with vemurafenib commonly developed drug resistance and diverse cutaneous side effects mainly through paradoxical activation of MAP kinase signaling pathway. Recent combination therapy such as MEK inhibitor in combination with vemurafenib shows improvement in major clinical end points but percentage of patients with adverse toxic events are higher compared to vemurafenib monotherapy, and most patients relapse ultimately. It is therefore an urgent need to develop new anti-melanoma drug and/or adjuvant agent for vemurafenib therapy. Previously, our laboratory isolated a bioactive phytochompound deoxyelephantopin (DET) from Elephantopus scaber L. and showed that DET possessed anti-inflammation, hepatoprotective and anti-mammary tumor activities. We considered DET as a novel lead compound for further modification or optimization of its structure to create a more potent drug lead for cancer management. Therefore, we created a total of sixty semi-organically modified derivatives of DET (DETDs). In this thesis study, I showed that DETD-35 suppressed both parental human BRAFV600E mutant melanoma (A375) and vemurafenib resistance melanoma (A375-R) cell proliferation in vitro. Moreover, DETD-35 showed synergism with vemurafenib in suppressing the proliferation, colony formation and inducing apoptosis of A375 melanoma cell. In A375 xenograft study, DETD-35 suppressed tumor growth with 87.1% TGI and reduced tumor mass by 70.5%, as effective as vemurafenib (87.4% TGI and 71.9% reduction in tumor mass), compared to tumor control group. Moreover, compound-drug combinational treatment with alternate administration of DETD-35 and vemurafenib in which the treatment frequency was only half of the single compound administration frequency also showed similar tumor growth inhibition efficacy (91.0% TGI and 72.3% reduction in tumor mass), suggesting in vivo synergistic or additive effect of DETD-35 and vemurafenib. In A375-R xenograft study, vemurafenib treatment showed little or no anti-tumor activity with similar tumor growth rate and sizes relative to the tumor control group, whereas DETD-35 suppressed A375-R tumor growth with 47.1% TGI and reduced tumor mass by 46.3%. Notably, the combination of DETD-35 and vemurafenib treatment exhibited the most significant effect with 76.5% TGI and 65.2% reduction in tumor mass. Mechanistic studies revealed the reactivation MAP Kinase, Src-STAT3 and Akt signaling pathways in the acquired vemurafenib resistance A375-R cells, and DETD-35 suppressed those reactivated multiple signaling pathways. In addition, DETD-35 triggered ROS-induced apoptotic cell death in both A375 and A375-R cells. Furthermore, through a DMBA/TPA-induced mouse skin carcinogenesis model mimicking cutaneous side effect caused by vemurafenib, I showed that DETD-35 delayed vemurafenib-induced acceleration of skin papillomas formation and also decreased total papillomas number in mice. Overall, results from my thesis study offers strong evidence that DETD-35 inhibits BRAFV600E mutant melanoma growth, overcomes acquired vemurafenib resistance and reduces cutaneous side effect of the BRAF inhibitor in mice. It is therefore suggested that DETD-35 might have great therapeutic or adjuvant potential in management of melanoma patients with BRAFV600E mutation.