The role of SERPINB3 in cervical cancer therapeutic resistance

PROJECT SUMMARY High SERPINB3 is a biomarker of radioresistance, and a potential therapeutic target for cervical cancer. Our recent work demonstrated that CRISPR-Cas9 knock out of SERPINB3, an inhibitor of lysosomal proteases, sensitized cervical tumor cells to radiation therapy (RT)-induced cell death. In vivo targeting of SERPINB3 with

siRNA not only altered the tumor microenvironment, but significantly sensitized SERPINB3-high tumor models to RT. Importantly, we established that tumor cells died via lysoptosis, a newly described and evolutionarily conserved mode of cell death that is dependent upon lysosomal membrane permeabilization (LMP) and leakage

of potent proteases into the cytoplasm. SERPINB3-KO tumor cells underwent widespread lysoptosis when treated with a variety of cytotoxic agents and chemotherapies. This suggests that loss of SERPINB3 exposed lysoptosis as a default cell death mechanism in these cells. The molecular features of tumor cells that are

susceptible to RT-induced LMP and lysoptosis are unknown. Validation that lysoptosis leads directly to tumor control is required. Additionally, the mechanism through which RT induces LMP is unknown. New unpublished data suggest that SERPINB3-high tumors have a distinct molecular signature characterized by enrichment of

lysosome-related pathways. In these cells, LMP occurs soon after treatment with clinically relevant doses of RT, while lysosomal rupture occurs days later just prior to cell death. Finally, we have identified candidate small molecules that bind specifically to SERPINB3. The long-term goal of this work is to define new treatment

approaches for radioresistant SERPINB3-high tumors. This proposal tests the hypothesis that targeting SERPINB3 in SERPINB3-high tumor cells renders them susceptible to RT-induced lysoptosis, and that LMP is the initiating event that results from unrepaired lysosomal damage. Three aims are proposed to directly address

this hypothesis: Aim 1: Determine the predominant cell death mechanism induced by RT in SERPINB3-high tumors; Aim 2: Determine the mechanism of therapy-induced LMP leading to lysoptosis; Aim 3: Identify SERPINB3-targeting strategies that specifically sensitize tumor cells and not normal cells to RT. To accomplish

these aims, we propose to use ex vivo organelle-based approaches, step-wise in vitro and in vivo analysis of cell death mechanisms leading to tumor control, and an innovative pre-clinical brachytherapy system that we have developed to mimic treatment delivered in the clinic. Success of this proposal will confirm that SERPINB3

is a predictive biomarker for lysoptosis-inducing CRT strategies, generate new knowledge of the critical events leading to RT-induced LMP and cell death, and provide innovative therapeutic strategies to target SERPINB3 and improve survival for patients with cervical cancer using a personalized treatment approach.