Abstract 10: A small molecule derivative effectively inhibits proliferation of pancreatic cancer cells by targeting Sp1 and survivin

Abstract

Abstract Pancreatic cancer is one of the most fatal malignancies due to its poor prognosis. With current standard treatment options, the overall 5 year survival rate is about 16% and these treatment plans often cause high toxicity. Therefore, there is an urgent need for identifying more effective and less toxic agents for the treatment of this malignancy. Tolfenamic acid (TA) is a generic drug used to treat migraine headaches but has been demonstrated to have anti-cancer activity in pre-clinical studies. It is known to downregulate the transcription factor Specificity protein 1 (Sp1). Sp1 regulates several genes involved in cell proliferation and apoptosis, including survivin, an inhibitor of apoptosis protein. Interestingly, a recent discovery proposed that copper(II) complex of TA can result in higher therapeutic response; however its efficacy was not tested in gastro-intestinal cancers. In this study, we assessed the therapeutic efficacy of a Cu(II)- containing complex of TA (Cu-TA) using human pancreatic cancer cell lines. MIA PaCa-2 and Panc-1 cells were treated with increasing concentrations of vehicle (DMSO), equimolar CuCl2 (negative control), TA or Cu-TA and the cell viability was measured at 24 and 48 h post-treatment using CellTiter-Glo kit. CuTA was further tested for its effect on Sp1 and survivin expression in MIA PACa-2 cells by Western blot (protein) and quantitative PCR (mRNA). The activation of apoptosis was determined by measuring the activity of effector caspases using the Caspase3/7-Glo kit and the apoptotic cell population through flow cytometric analysis using Annexin-V staining. Cell cycle arrest was assessed by flow cytometry with propidium iodide staining. While both TA and Cu-TA caused dose/time-dependent response to inhibit pancreatic cancer cell growth, Cu-TA showed higher efficacy when compared to TA. Cu-TA was highly effective in inhibiting Sp1 and survivin protein expression and showed similar trend for inducing apoptotic markers and causing cell cycle arrest in early phase (G0/G1). The results of qPCR demonstrated that the expression of survivin mRNA was significantly lower following both Cu-TA and TA treatment; however, the mRNA expression of Sp1 remained unchanged. This indicates that TA and Cu-TA could be working in similar mechanism by effecting Sp1 post-translationally, perhaps through proteasome-dependent degradation. These results demonstrate that Cu-TA is more effective than TA and potentially useful for pancreatic cancer treatment after clinical testing. Studies to precisely understand the underlying mechanisms of Cu-TA are currently under investigation through molecular profiling analysis. Citation Format: Myrna Hurtado, Umesh Sankpal, Aboubacar Kaba, Shahela Mahammad, Alvin Holder, Jamboor Vishwanatha, Riyaz Basha. A small molecule derivative effectively inhibits proliferation of pancreatic cancer cells by targeting Sp1 and survivin [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 10. doi:10.1158/1538-7445.AM2017-10