Abstract 1730: RET tyrosine kinase receptor represses Noxa transcription and prevents genotoxic or endoplasmic reticulum stress-induced apoptosis.
Authors: Bagheri-Yarmand Rozita , Gururaj Anupama E , Williams Michelle , Ahmed Zamal , Ladbury Jean E , Bogler Oliver , Huang Sue-chen , Cote Gilbert G , Gagel Robert F 
Published: August 2014

Abstract
One of the hallmarks of human cancers is the intrinsic or acquired resistance to apoptosis. Apoptosis resistance contributes to carcinogenesis, tumor progression, and treatment resistance. Patients with medullary thyroid carcinoma (MTC) have been shown to be resistant to standard chemotherapy and conventional radiotherapy. Dominant activating mutations in the RET protooncogene are the cause of hereditary MTC and are found in more than 40% of sporadic cases. Despite an ever-growing wealth of information about RET's role in cell survival/proliferation, a detailed understanding of how this tyrosine kinase receptor acts to prevent cell death in tumorigenesis remains elusive. We hypothesized that activated RET prevents apoptosis by repressing the transcription of the proapoptotic gene Noxa in the nucleus in response to genotoxic or endoplasmic reticulum (ER) stress.
Here we report that activated RET translocates to the nucleus in cultured MTC cells as well as MTC primary tumor tissues. RET is recruited to the Noxa promoter and is able to repress its transcription. The formation of the RET-Noxa promoter complex depends on RET expression and requires the tyrosine kinase activity of RET. Additionally, stable depletion of RET in MTC cells, using lentiviral shRNA, sensitizes MTC cells to genotoxic or endoplasmic reticulum stress. We specifically show that RET is recruited to the Activating Transcription Factor 4 (ATF4) binding site on the Noxa promoter. RET physically interacts with ATF4 and phosphorylates ATF4 leading to the blockage of its transcriptional activity. Moreover, RET knockdown is associated with increased ATF4-dependent expression of Noxa. Inhibition of RET kinase activity by sunitinib results in ATF4 activation, induction of Noxa expression and apoptosis. Conversely, silencing of ATF4 in MTC cells decreased sunitinib-mediated Noxa induction and apoptosis. ATF4 protein levels markedly decreased and/or localized in the cytoplasm of 60% of primary MTC (n=40) and associated with poor overall survival.
Our data suggest that RET activating mutations confers resistance to chemotherapeutic agents by preventing apoptosis through inhibition of ATF4 activity and repression of proapoptotic gene Noxa. These findings reveal the importance of nuclear localization of RET, as newly discovered modulator of nuclear gene expression. This newly identified interaction between RET kinase and ATF4 has important implication for MTC and other RET/ATF4-mediated cancer types. Since ATF4 transcription is induced by endoplasmic reticulum stress, and activation of ATF4 induces cell cycle arrest and apoptosis, the concept of promoting endoplasmic reticulum stress in combination with tyrosine kinase inhibitors could be considered as a therapeutic strategy for cancer.
Citation Format: Rozita Bagheri-Yarmand, Anupama E Gururaj, Michelle Williams, Zamal Ahmed, Jean E Ladbury, Oliver Bogler, Sue-chen Huang, Gilbert G Cote, Robert F Gagel. RET tyrosine kinase receptor represses Noxa transcription and prevents genotoxic or endoplasmic reticulum stress-induced apoptosis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1730. doi:10.1158/1538-7445.AM2013-1730