Abstract 6719: GULP1 knockout promotes carcinogen-induced bladder cancer by regulating the NRF2-KEAP1 signaling axis

Abstract Urothelial carcinoma of bladder (UCB) is the ninth most commonly diagnosed cancer worldwide. Some key genetic and epigenetic alterations are hallmarks on the initiation and progression of UCB. The nuclear factor E2-related factor 2 (NRF2)-Kelch-like ECH-associated protein-1 (KEAP1) signaling pathway, essential for detoxifying reactive oxygen species, is normally regulated by the interaction of NRF2 with KEAP1and the actin cytoskeleton in the cytoplasm. However, genetic and epigenetic alterations can lead to a constitutive activation of NRF2 through the loss of KEAP1 function. This dysregulation results in NRF2 translocation to the nucleus, triggering the expression of numerous genes. We recently reported that phosphotyrosine binding (PTB) domain-containing engulfment adaptor protein 1 (GULP1) is frequently silenced in UCB by promoter hypermethylation, and interacts with the KEAP1-actin cytoskeleton complex to regulate NRF2. Furthermore, the loss of GULP1 expression led to uncontrolled NRF2 signaling, driving cancer progression and resistance to chemotherapy. Here, to strengthen our previous findings that GULP1 acts as a tumor suppressor gene (TSG) in UCB, we exposed GULP1 knockout (KO) mice to a most commonly used carcinogen for generating UCB model in mice. To this end, we exposed 4-week-old GULP1 KO and wild type (WT) C57BL/6 mice to N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) for 6 to 20 weeks. We then analyzed the resulting bladder tumors using various methods to investigate the NRF2-KEAP1 axis, tumor subtype, and several tumor immune microenvironment (TIME) factors. Our data demonstrates that GULP1 KO exhibits a higher frequency of carcinogen-induced UCB and muscle invasion compared to WT counterparts via regulating the NRF2-KEAP1 axis. Furthermore, GULP1 KO carcinogen-induced tumors appear at earlier time point compared to WT mice. Interestingly, GULP1 KO mice exhibited a characteristic subtype and TIME dynamics that differed from those of WT. In conclusion, our pilot in vivo data suggested that GULP1 may regulate the NRF2-KEAP1 signaling pathway in UCB. Further experiments are ongoing to understand the comprehensive mechanisms of GULP1 involvement in UCB and its association with chemo and immunotherapy response. Citation Format: Chonji Fukumoto, Naveen C. Pandey, Sylvia S. Sanchez, Pritam Sadhukhan, Christopher M. Koenig, Susmitha R. Pagadala, Fenna C. Sillé, Mohammad O. Hoque. GULP1 knockout promotes carcinogen-induced bladder cancer by regulating the NRF2-KEAP1 signaling axis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6719.