Project Details
Projekt
Targeting Rpa3 for cancer treatment
Applicants
Dr. Daniel Dauch, Ph.D.; Professor Dr. Stefan Laufer
Subject Area
Hematology, Oncology
Term
from 2015 to 2019
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 267467939
The Replication protein A heterotrimeric complex is composed of Rpa1, 2 and 3 and plays an essential role in proliferation. As the major single-stranded DNA binding protein it is required for DNA repair, DNA recombination and induction of DNA damage checkpoints. During DNA replication Rpa protects the ssDNA in the replication forks and prohibits replication catastrophe upon replicative stress. Global exhaustion of Rpa in the replication forks is prevented by the Atr-Chk1 pathway by confining the amount of replication origins. By applying a pooled negative selection in vivo RNAi screen we identified Rpa3 as a promising therapeutic target for the treatment of therapy resistant liver carcinomas. Based on the rationale that essential processes in cancer cells might represent vulnerabilities that can be exploited therapeutically, we thought to determine the potential and the therapeutic window of targeting Rpa in therapy resistant solid tumors. By applying knockdown of Rpa3 in different cancer cells as well as transposon-based mouse models in vivo we could show that liver cancer- and pancreatic cancer growth can be efficiently decreased by Rpa3 suppression. In vitro experiments furthermore showed that a striking antitumor activity was observed when Rpa3 knockdown was combined with Chk1 inhibition, resulting in rapid induction of cell death specifically in cancer but not in normal cells. By applying a shRpa3 transgenic mouse strain, we could show that a nine-day knockdown of Rpa3 is tolerated by mice. Together, these data indicate that a therapeutic window for targeting Rpa3 may exist and we started to develop pharmacological inhibitors of Rpa3, for which on target activity and in vitro therapeutic efficacy could already be shown. In this proposal, we aim to characterize the cellular stress response of tumor cells upon Rpa3 inhibition with or without simultaneous Chk1 inhibition. We further plan to characterize why the stress- and the therapy response differ between cancerous and non-cancerous cells. To determine the potential of targeting Rpa3 in a cancer therapy, we aim to perform preclinical treatment studies in transposon-based mouse models for liver- and pancreatic cancer which are combined with shRpa3 transgenic mice. These studies will be conducted based on established tolerable doses of Rpa3 knockdown alone or in combination with Chk1 inhibition. In addition, we are planning to perform in vivo negative selection RNAi screening in a transposon-based liver cancer mouse model to either identify other gene products involved in DNA replication or DNA damage that can be exploited therapeutically or to identify candidate genes whose knockdown may also synergize with Rpa3 inhibition. Finally, we aim to further develop our pharmacological Rpa3 inhibitors to obtain a tool compound for in vivo studies. New Rpa3 inhibitors will be analyzed by detailed ADME and toxicity profiling and applied in an anti-cancer therapy of liver- and pancreatic cancer in vivo.
DFG Programme
Research Units
