Abstract 4617: Targeting mTORC1 by overexpression of PRAS40(T246A) in basal keratinocytes suppresses proliferation and migration of keratinocyte stem cells and skin inflammation during tumor promotion

Abstract

Abstract Considerable evidence indicates a crucial role for Akt and mTORC1 signaling in cancer. Previously, we discovered that the Akt and mTORC1 signaling pathways play an important role in epithelial multistage carcinogenesis using the mouse skin model, especially during the tumor promotion stage. The proline-rich Akt substrate of 40 kDa (PRAS40), a subunit of mTORC1 was originally identified as a novel Akt substrate through Akt-mediated phosphorylation at Thr246 and this phosphorylation is critical for mTORC1 activity. Moreover, PRAS40 has been identified to negatively regulate mTORC1 signaling. In this study, transgenic mice were generated that overexpress a mutant form of PRAS40 [PRAS40(T246A)] using the bovine keratin 5 (BK5) promoter to further examine the importance of mTORC1 signaling in epithelial carcinogenesis. BK5.PRAS40(T246A) transgenic mice did not have a discernable gross skin phenotype. However, these mice were significantly less sensitive to TPA-induced epidermal hyperproliferation and hyperplasia. Overexpression of PRAS40(T246A) in the epidermis led to significant suppression of skin tumor development and significant reductions in tumor weight and tumor size compared to wild-type mice. Immunoprecipitation analysis of epidermal protein lysates from BK5.PRAS40(T246A) mice showed that PRAS40(T246A) remained bound to raptor even after TPA treatment. BK5.PRAS40(T246A) mice also displayed reduced mTORC1 signaling, reduced levels of cell cycle proteins and increased autophagy signaling as well as the skin inflammatory response in the epidermis following treatment with TPA. Notably, TPA-induced proliferation and migration of bulge-region label retaining cells were also significantly inhibited in BK5.PRAS40(T246A) mice. In addition, expression of PRAS40(T246A) in basal keratinocytes significantly inhibited keratinocyte migration both in vivo and in vitro and led to delayed wound healing. Decreased migration and impaired wound healing may be attributed to altered expression of EMT markers as identified in PRAS40(T246A) epidermal cells. The current data using this novel mouse model provides further evidence that mTORC1 specific signaling in keratinocytes contributes significantly to the process of skin tumor promotion by regulating proliferation and migration of keratinocyte stem cells. In addition, the data demonstrate an important role for mTORC1 specific signaling in keratinocytes in regulating skin inflammation. Collectively, using this novel mouse model where mTORC1 signaling is selectively suppressed in keratinocytes has provided further evidence that this signaling pathway regulates several important mechanistic events involved in the process of skin tumor promotion. Research supported by NIH Grants CA129409, CA037111, and American Cancer Research Center and Foundation Citation Format: Okkyung Rho, Jaya Srivastava, Jiyoon Cho, John DiGiovanni. Targeting mTORC1 by overexpression of PRAS40(T246A) in basal keratinocytes suppresses proliferation and migration of keratinocyte stem cells and skin inflammation during tumor promotion. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4617.