Elucidating the epigenetic landscape of non-muscle invasive bladder cancer.

e16580 Background: Non-muscle invasive bladder cancer (NMIBC) presents a significant clinical challenge due to its high recurrence rate. Recently, a deeper understanding of tumor epigenetics has provided new opportunities for cancer detection and treatment. While significant efforts have been made to understand NMIBC biology, the molecular and epigenetic mechanisms driving its progression remain elusive. Methods: We used multiomics analysis including expression, DNA methylation, and cell-free DNA chromatin immunoprecipitation sequencing (cfChIP-seq) assays to uncover the molecular and epigenetic landscape of NMIBC from human tissue samples and urine. Results: High-throughput sequencing revealed more than 6,000 differentially methylated regions (DMRs) within regulatory elements, many of which were linked to cancer-related pathways and exhibited distinct signatures in cancer tissues compared to healthy controls. Next, we distinguished high and low-grade tumors based on unique DNA methylation profiles. These patterns correlated with transcription level of genes associated with cell cycle regulators, consistent with the higher proliferative capacity of high-grade tumors. Moreover, Among the DMRs, we also identified genes involved in T cell regulation. Supporting this, we demonstrated reduced CD8+ T cells infiltration in high-grade tumors, further highlighting immune dysregulation in the more aggressive disease. We extended our investigation to urine samples, performing cfChIP-seq on samples from healthy donors and NMIBC patients. By using the H3K4me3 promoter histone mark on cfDNA we could detect cell origin. Utilizing this assay, we were able to identify changes in the urine of NMIBC patients, owing to cf-nucleosome contributions from both the tumor and immune cells. Additionally, we stratified the cancer cohort by tumor grade and found enrichment in the number of differentially marked genes in high-grade tumors. Conclusions: Our detection of unique epigenetic signatures offer opportunities for biomarker development, enabling early detection through non-invasive urine based diagnostic approach. This could grade tumor more accurately and tailor personalized patient care and monitoring.