Pilot: Development of New Protacs and Light-Activated Inhibitors for Cytochrome P450 (2A6, 1B1) for Treatment of Bladder Cancer

The American Cancer Society’s estimates for bladder cancer in the United States for 2022 are about 81,180 new cases and about 17,100 deaths. The treatment of bladder cancer includes surgery, intravesical, chemo-, radiation, immuno- and targeted therapies. Over 70% of bladder cancer patients with non-muscle invasive bladder cancer (NMIBC) survive for more than 5 years postdiagnosis. However, the 5-year survival rate drops to 35% when invasive cancer is diagnosed. Drug resistance is observed in more than 30% of cases after chemotherapy. A cystectomy is recommended for these patients, which is accompanied by a significant reduction in the quality of life and high costs to the health care system. Cytochrome P450s (CYPs) are responsible for metabolic activation of carcinogens and overexpressed in several cancers, including bladder cancer.1, 2 Overexpression of CYP2A6 and 1B1 is associated with chemotherapy resistance, and silencing of CYP2A6 resulted in improved cell killing.3 Therefore, CYP1B1 and 2A proteins can be considered as therapeutic targets, and inhibition or degradation of these proteins could overcome the cisplatin resistance in chemotherapy of bladder cancer. The objective of this project is to develop new systems for inhibition and degradation of CYP proteins overexpressed in bladder cancer and associated with drug resistance. The strategy relies on two synthetic approaches: 1) PROTAC (PROteolysis TArgeting Chimera) and 2) photocaging. The selected approaches are independent, but connected through the targeting of CYP proteins. The two synthetic aims also were developed to work independently OR in combination. The approach is validated by literature precedent, as PROTAC molecules have been proven to degrade important proteins, such as Brd4, FKBP12, ER, and BTK.4 Indeed, the pharmaceutical company Arvinas has developed PROTACs targeting the androgen receptor and estrogen receptors that have progressed into a phase 2 clinical study. However, it has emerged that PROTAC approaches still have undesired effects, because systemic application can affect untargeted tissue, a disadvantage shared with traditional inhibitor-based therapeutics. The first Specific Aim is to develop the CYP-based PROTACs for ubiquitination and degradation of the CYP2A or 1B1 proteins by the proteasome. The second Specific Aim is to utilize photocaging approaches for development of Ru(II) based prodrugs that are activated with red light, and will possess a rapid photorelease of CYP-inhibitors or PROTACs. Finally, the third Specific Aim is to investigate the toxicity in vivo for new PROTACs and Ru-based prodrugs. In summary, this proposal seeks to develop new multifunction compounds that will target proteins associated with the invasive phenotype of bladder cancer and resistance to chemotherapy. These compounds will be synthesized with the ability for light independent (Aim 1) and light dependent (Aim 2) inactivation of CYPs, resulting in restoration of chemosensitivity of bladder cancer cells to chemotherapy.