Abstract 2447: TLR4 expression as a determinant of EMT and stress response gene expression in UV exposed human keratinocytes

Abstract

Abstract Toll-like receptor 4 (TLR4) is gaining attention for its role in several cancer-associated signaling pathways, including the regulation of inflammation, DNA repair and epithelial-mesenchymal transition (EMT). There is evidence that TLR4 contributes to ultraviolet (UV) radiation-induced stress signaling in skin cells, including keratinocytes. Blockade of TLR4 activity using the pharmacological antagonist resatorvid (TAK-242) inhibits not only acute UV-induced signaling in vitro and in vivo, but blocks UV-induced skin carcinogenesis in mouse models. In order to genetically explore the role of TLR4 in keratinocyte responses to UV-induced stress, we recently utilized CRISPR/Cas9 techniques to delete TLR4 from the genome of human HaCaT keratinocyte cells in culture. First, using differential gene expression analysis (employing the EMTRT2 Profiler PCR array technology) it was observed that TLR4 KO status is associated with pronounced downregulation of EMT-related gene expression (MMP9, VIM, FN1, SNAI1, TWIST1) confirmed by independent RT-qPCR analysis. TLR4 was also a determinant of UV-induced inflammatory enzyme expression including PTGS2. Phenotypically, TLR4 KO cells displayed an attenuation of Matrigel transwell migration. TLR4 KO cells demonstrated increased sensitivity to UV-induced apoptosis as detected by flow cytometry of annexin V/propidium iodide stained cells, and by immunodetection of increased PARP cleavage. Moreover, NanoString nCounter expression analysis of murine skin chronically exposed to tumorigenic UV indicated that topical resatorvid treatment was associated with MMP9 downregulation. Future studies will employ this novel TLR4 KO keratinocyte cell line as a mechanistic tool to dissect the role of TLR4 in inflammatory signaling and EMT response in the context of skin photodamage and carcinogenesis. Supported by NIH grants ES029579, R03CA230830, P01CA229112. Citation Format: Viktoria Kirschnerova, Prajakta Vaishampayan, Maria Khawam, Clara Curiel-Lewandrowski, Georg T. Wondrak, Sally E. Dickinson. TLR4 expression as a determinant of EMT and stress response gene expression in UV exposed human keratinocytes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2447.