Oncogenic mutations in RAS provide a compelling yet intractable therapeutic target. Using co-immunoprecipitation mass spectrometry, we recently identified an interaction between RAS and Argonaute 2 (AGO2), the core component of the RNA silencing machinery exerting translational control of mRNA transcripts. RAS and AGO2 co-sediment and co-localize in intracellular endomembrane bound organelles. The AGO2 N-terminal domain directly binds the KRAS Switch II region, irrespective of GDP/GTP bound to RAS. Functionally, we observed that AGO2 was required for maximal oncogenic KRAS levels and AGO2 knock-down attenuates cell proliferation in mutant KRAS-dependent cells. Intriguingly, AGO2 mediated microRNA processing (RISC activity) was attenuated in cancer cells expressing mutant KRAS compared to those expressing wild type RAS. Yet, these initial investigations using cell line models do not reveal a clear role for AGO2 in KRAS driven oncogenesis. Therefore, we probed the role of AGO2 in the well-established pancreatic ductal adenocarcinoma (PDAC) mouse model driven by oncogenic KRAS.
Towards this end, we generated LSL-KrasG12D; Pft1a Cre; AGO2flox/flox (KCA) mice. Preliminary analysis (8-10 weeks of age) suggests that compared to LSL-KRASG12D (KC) mice, homozygous loss of AGO2 results in increased KRAS driven Acinar to Ductal Metaplasia (ADM) phenotype considered as precursors of PanIN (pancreatic intraepithelial neoplasia) and PDAC. This mirrors the phenotype of loss of Dicer, another microRNA processing enzyme, in the same model further reinforcing a role for microRNAs in restraining KRAS oncogenic programs. Yet unlike Dicer, loss of AGO2 alone did not cause any gross or histological changes in the development of the mouse pancreata. In an effort to understand the underlying mechanisms that are regulated by AGO2, we also studied the activated levels of a variety of signaling molecules in pancreata obtained from different genotypes. Using both Western blot and Immunohistochemical analyses, we demonstrate a previously unknown and critical role for AGO2 in KRAS-MAPK signaling pathway.
Together our data from the in vivo mouse model suggests that AGO2 is an important regulator of KRAS signaling and oncogenic KRAS alters the translational machinery through its interaction with AGO2.
Citation Format: Sunita Shankar, Jean Tien, Vijaya L. Dommeti, Sylvia Zelenka-Wang, Arul M. Chinnaiyan. An in vivo model reveals a role for Argonaute 2 in oncogenic KRAS driven pancreatic cancer initiation. [abstract]. In: Proceedings of the AACR Special Conference on Translational Control of Cancer: A New Frontier in Cancer Biology and Therapy; 2016 Oct 27-30; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2017;77(6 Suppl):Abstract nr A05.