Author ORCID Identifier
Date of Graduation
8-2019
Document Type
Thesis (MS)
Program Affiliation
Cancer Biology
Degree Name
Masters of Science (MS)
Advisor/Committee Chair
Anil K. Sood, M.D.
Committee Member
Menashe Bar Eli, Ph.D.
Committee Member
Wei Hu, M.D., Ph.D.
Committee Member
Kwong K. Wong, Ph.D.
Committee Member
Prahlad T. Ram, Ph.D.
Abstract
Purpose: To identify combination strategies using Bromodomain and Extra-Terminal Domain (BET) inhibitors in a mechanism driven fashion to maximize anti-tumor activity, and to determine the efficacy of BET inhibitor combinations in pre-clinical ovarian cancer mouse models.
Experimental Design: We used a novel, previously uncharacterized pan-BET inhibitor, CN210, for all in vitro (MTT, apoptosis, protein expression, reverse phase protein array (RPPA) analysis and in vivo (orthotopic mouse model)) experiments. The poly (ADP-ribose) polymerase (PARP) inhibitor olaparib, as well as the mechanistic target of rapamycin (mTOR) inhibitors rapamycin and INK-128 were also used. Statistical analyses of in vitro and in vivo experiments were performed using either Student t test or Mann-Whitney test with a p value < 0.05 considered significant (sig). RPPA data was analyzed by Cluster 3.0 using median value and the heat maps were generated by Java Treeview.
Results: Eight ovarian cancer cell lines were screened using MTT assays with CN210 to determine the most and least sensitive. HeyA8 and OVCAR8 ip1 cells displayed the greatest degree of sensitivity whereas OVCAR4 and OVCAR5 cells were the most resistant. We next examined the previously reported synergistic combination of olaparib and BETi and found only modest effects using in vitro and in vivo applications. RPPA analysis was then performed on all four cell lines after CN210 treatment revealing that activation of the mTOR pathway appeared to be an early adaptive response to BET inhibition. Decreased cell viability and increased apoptosis were seen with the addition of an mTORC1 or dual mTORC1/2 inhibitor to CN210 compared with CN210 alone. Notably, we found that decreased Akt activation and increased Rictor activation after CN210 treatment served as a possible marker for the increased response seen with CN210 and INK-128 combination. Lastly, we demonstrated a significant reduction in tumor weight using CN210/INK-128 combination treatment in an orthotopic ovarian cancer mouse model.
Conclusions: RPPA analysis identified mTOR activation as an early adaptive change to BET inhibition in ovarian cancer cells. Combination of BET and dual mTORC1/2 inhibitors led to robust anti-tumor responses, supporting the inclusion of these drugs into future clinical trials.
Keywords
BET inhibitors, PARP inhibitors, mTOR inhibitors, clinical trials, ovarian cancer, targeted therapeutics