Abstract 5647: Investigating the role of AP-2α in melanomagenesis

Abstract

Abstract Introduction: Transcription factor AP-2α (encoded by the TFAP2A gene) is a protein involved in the early development of multiple tissues, particularly the neural crest. Melanocytes are derived from the neural crest and their development appears to rely on AP-2α and its paralogues, along with master regulator MITF. Prior studies have indicated that AP-2α is a crucial regulator of melanoma metastasis. However, it is unclear if AP-2α plays a role in melanomagenesis. We sought to understand the role of AP-2α in melanomagenesis, hypothesizing that knock out (KO) of TFAP2A would suppress malignant potential in vitro and downregulate genetic pathways linked to malignant transformation. Methods: TFAP2A KO was generated with a single-guide RNA CRISPR/Cas9 system targeting exon 3 of the gene in the human melanoma cell line A375. Tumorigenesis phenotype was assessed by evaluating contact-independent colony formation with the in vitro soft agar assay. To identify downstream regulatory programs controlled by TFAP2A, we then utilized RNA-seq and Gene Set Enrichment Analysis (GSEA, San Diego). RNA-seq was performed on four clones each for wildtype and TFAP2A KO samples and the resulting data was processed using GSEA to identify gene sets modified after TFAP2A KO. Results: Targeting of TFAP2A with CRISPR/Cas9 effectively knocked out AP-2α protein expression in the A375 melanoma cell line. Colony formation on soft agar assay was significantly decreased after TFAP2A KO (P < 0.01). RNA-seq revealed significant differential expression of 11,195 genes after TFAP2A KO. Gene sets with negative enrichment after TFAP2A KO included those overall higher in tumors versus normal tissue (FWER P = 0.003) and multiple signaling pathways related to malignant transformation such as TGF-β (FWER P = 0.003), MYC (FWER P = 0.009), KRAS (FWER P = 0.03), and Notch (FWER P = 0.043). Conclusions: Our data suggests that TFAP2A plays a role in cutaneous melanomagenesis. TFAP2A may represent a therapeutic target for melanoma, though further pre-clinical studies are needed. We plan to further test this hypothesis in a melanoma mouse model with conditional TFAP2A KO under the control of inducible Cre recombinase driven by the mouse tyrosinase gene promoter. Citation Format: Nicholas Bartschat, Jeremy Chang, Mohammed Suraju, Ryan Nagel, Zhijie Li, Jeffrey White, Colin Kenny, Ronald Weigel. Investigating the role of AP-2α in melanomagenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5647.