Abstract 1995: HuR dependent inhibition of PARG enhances PARP inhibitor therapy for DNA repair proficient and deficient pancreatic cancer cells
Authors: Chand Saswati N. , Zarei Mahsa , Kamath Akshay R. , Schiewer Matthew J. , Romeo Carmella , Cozzitorto Joseph A. , Meisner-Kober Nicole , Londin Eric , Rigoutsos Isidore , Knudsen Karen , Pascal John M. , Yeo Charles J. , Winter Jordan M. , Brody Jonathan R. 
Published: July 2016

Introduction: Despite our deep understanding of genetic drivers of the disease, pancreatic ductal adenocarcinoma (PDA) continues to be associated with dismal survival rates. Targeting the DNA repair machinery has emerged as a promising therapeutic strategy to treat pancreatic cancer patients carrying DNA damage repair (DDR) mutations. Such mutations promote tumorigenesis, but also paradoxically render tumor cells particularly susceptible to platinum-based agents and PARP inhibitors (PARPi). However, despite promising preclinical and clinical results, early data demonstrate that eventually most tumors, regardless of DDR status, become resistant to PARPi-therapies.

The mRNA-binding protein HuR, predominantly expressed in the nucleus, translocates to the cytoplasm upon tumor-associated stress where it post-transcriptionally regulates select mRNA cargo, resulting in resistance to DNA damaging agents in a harsh tumor microenvironment. Here, we sought to evaluate the role of HuR in regulating PARPi efficacy.

Results: In response to cellular stress induced by IC50 dosages of a panel of PARPis such as Veliparib, Olaparib, Rucaparib, Talazoparib and Niraparib, nuclear localized HuR undergoes cytoplasmic translocation. Silencing of HuR via siRNA, CRISPR and a DOX-inducible system resulted in significant decrease in long and short- term PDA cell survival, irrespective of DDR status. To complement and validate in vitro findings, we employed a heterotropic mouse xenograft model using Mia.sh290 cells wherein DOX induction significantly reduced HuR expression. Olaparib mediated PARP inhibition (50mg/kg, 5 days a week) combined with DOX-induced HuR silencing resulted in significant reduction in tumor volumes, compared to Olaparib alone or DOX alone.

Mechanistically, we demonstrate that the pro-survival protein HuR facilitates PDA cells to recover from PARPi insult by, in part, regulating poly ADP ribose glycohydrolase (PARG), the major enzyme responsible for hydrolyzing poly-ADP ribose (PAR) polymers, on chromatin and associated proteins. HuR binds to two 41- 43bp long sites in the 3’ untranslated region (3’UTR) of PARG, increasing its mRNA stability and protein expression. Increased PARG activity, further validated via exogenous overexpression, promotes DNA repair efficiency and increases PDA cell survival. Functional analysis indicate that such inhibition of HuR and/or PARG significantly enhances PARPi sensitivity in PDA cells, via increased accumulation of DNA damage γH2AX foci, preventing efficient removal of PAR polymers, and enhancing detrimental trapping of PARP1 on chromatin.

Conclusions: Taken together, our results indicate that HuR- mediated upregulation of PARG acts as a universal pro-survival mechanism and HuR inhibition could significantly potentiate PARPi therapy in PDA, irrespective of DNA repair status.

Citation Format: Saswati N. Chand, Mahsa Zarei, Akshay R. Kamath, Matthew J. Schiewer, Carmella Romeo, Joseph A. Cozzitorto, Nicole Meisner- Kober, Eric Londin, Isidore Rigoutsos, Karen Knudsen, John M. Pascal, Charles J. Yeo, Jordan M. Winter, Jonathan R. Brody. HuR dependent inhibition of PARG enhances PARP inhibitor therapy for DNA repair proficient and deficient pancreatic cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1995.