Abstract 84: Identification of HNRNPLL as a novel metastasis suppressor of colorectal cancer

Abstract

Abstract Despite the advances in surgery and chemotherapy for colorectal cancer patients, those with distant metastasis still suffer poor prognosis. Metastasis is a complex process regulated by a number of biomolecules, which can be conceptually divided into metastasis promoters and suppressors. To identify novel targets for prevention and/or therapy of colorectal cancer metastasis, we performed an in vivo shRNA library screen for colorectal cancer metastasis suppressor genes using an orthotopic transplantation model. CMT93 cells, a murine colorectal cancer cell line with poor metastasizing activity, were transduced with lentiviral shRNA library, and then transplanted into the rectum of C57/BL6 mice. Genomic DNA was collected from metastatic lesions, and the integrated shRNAs were retrieved by PCR for sequencing, followed by identification of the candidate genes targeted by the shRNAs. Among 47 candidate genes identified, we focused on Hnrnpll (heterogeneous nuclear ribonucleoprotein L-like) gene encoding a pre-mRNA binding protein involved in pre-mRNA splicing. Hnrnpll knockdown in CMT93 cells significantly increased their matrigel invasion in vitro and lung metastasis in vivo. An RNA-immunoprecipitation analysis identified Cd44 as a candidate target of Hnrnpll. The mRNA level of Cd44 variant 6 (Cd44v6), a splicing variant of Cd44 that had been shown to demarcate colon cancer stem cells driving colon cancer metastasis, was increased by knocking down Hnrnpll in colon cancer cells. Treatment with neutralizing anti-CD44v6 monoclonal antibodies suppressed matrigel invasion of colon cancer cells induced by Hnrnpll knockdown, suggesting that Hnrnpll may suppress invasion and metastasis, at least in part, by modulating the alternative splicing of Cd44. HNRNPLL expression level in colorectal cancer cells was drastically decreased upon induction of epithelial-mesenchymal transition (EMT), accompanied by increased expression of CD44v6. The link between HNRNPLL and EMT was further suggested by immunostaining analysis of clinical samples; colorectal cancer cells with low E-cadherin expression at the invasion front exhibited decreased HNRNPLL expression as compared with those in the tumor center. These results suggest that HNRNPLL is a novel metastasis suppressor in colorectal cancer. Possible mechanisms for regulation of HNRNPLL expression during EMT will also be discussed. Citation Format: Masahiro Aoki, Eiichi Sasaki, Kenya Kimura, Koji Komori, Yasuhiro Shimizu, Yasushi Yatabe, Keiichiro Sakuma. Identification of HNRNPLL as a novel metastasis suppressor of colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 84.