Abstract 953: MEK/ERK induces the EMT transcription factor ZEB1 that is regulated by USP10 and promotes colorectal cancer metastasis that can be inhibited by BRAFV600E-directed therapy

Abstract

Abstract Background: Constitutive MEK-ERK activation induced by BRAFV600E or RAS mutation promotes tumor metastasis in CRC that is mediated by transcription factors regulating epithelial-to-mesenchymal transition (EMT). We determined the role of ZEB1, an EMT master transcription factor, and its potential regulation by the ubiquitin specific peptidase 10 (USP10), in tumor metastasis mediated by MEK-ERK. Materials and Methods: Isogenic RKO [BRAFV600E or wild-type (wt)] and Colo320 (BRAF/KRAS wt/wt) human CRC cells were utilized. Colo320 cells were transfected with mutant KRASG12D. In addition to parental cells, we generated USP10 shRNA expressing RKO and Colo320 cells. Cells were treated with the MEK1/2 inhibitor cobimetinib or BRAF inhibitor encorafenib. Cell migration was analyzed by wound healing and transwell assays. Protein/protein interaction was analyzed by immunoprecipitation. Protein ubiquitination level was determined by ubiquitination assay. In an orthotopic model, lung metastases in mice were generated by tail vein injection of RKO cells. Results: We demonstrate MEK-ERK activation, induced by BRAFV600E, promotes CRC metastasis through inhibition of the USP10-ZEB1 axis. USP10 is shown to interact with ZEB1 and to remove its K27 linkage ubiquitination chain, thus increasing K48 ubiquitination of ZEB1 protein that results in proteosome-mediated degradation of ZEB1. MEK-ERK induced the phosphorylation of USP10 that is shown to attenuate the USP10-ZEB1 interaction and results in ZEB1 stabilization in RKO and Colo320 cell lines. Increased ZEB1 expression promotes tumor cell migration in vitro and metastasis in a murine xenograft model that could be suppressed by cobimetinib or BRAFV600E-directed therapy with encorafenib and cetuximab. Conclusions: Targeting RAF/RAS-driven MEK/ERK signaling can suppress ZEB1 stabilization to inhibit tumor metastasis. Suppression of metastasis occurs by a novel mechanism of ZEB1 stabilization that is regulated by an atypical USP10 deubiquitinase activity. Citation Format: Bo Qin, Lei Sun, Frank A. Sinicrope. MEK/ERK induces the EMT transcription factor ZEB1 that is regulated by USP10 and promotes colorectal cancer metastasis that can be inhibited by BRAFV600E-directed therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 953.