Characterization of T cell receptors diversity present in blood and urine: correlation between immune system activation with clinical-pathological parameters of superficial bladder cancer patients

Abstract

The most common type of bladder cancer is the non invasive form, not reaching muscular tissue (non muscle invasive bladder cancer, NMIBC), corresponding to 70-80% of cases. NMIBC treatment is cirurgical (transurethral resection of the bladder, TURB), followed by immunotherapy with BCG (Bacillus Calmette-Guerin) vaccine in cases with high or intermediate recurrence risk. The recurrence risk classification is based on clinical-pathological criteria, such as T stage (TNM classification), histological grade, presence/absence of carcinoma in situ, size and number of tumors. Around 30 to 40% of patients do not respond to BCG treatment and relapse, with 10 to 25% progressing to the invasive type (MIBC). The immune system is central in the control of tumor progression, which explains immunotherapies success, like BCG for NMIBC and immune checkpoint inhibitors for metastatic MIBC. As bladder cancers have a high mutational burden, they also have a greater chance of generating neoantigens. However, the antitumor immunological response and the immunotherapy success occur only if T lymphocytes recognize the tumor-presented neoantigens through TCRs (T cell receptors). TCRs are expressed in T lymphocytes after genomic sequences rearrangement and pre-mRNAs gene processing, resulting in unique TCR sequences, specific to each recognized antigen. After recognizing neoantigens, T-cells expand clonally, increasing the representation of specific TCR sequences (capable of recognizing those neoantigens) and, consequently, reducing diversity of the TCR repertoire of the patient's tumoral infiltrate, blood and urine - as demonstrated by previous studies. We believe that a lower TCR diversity (related to higher clonality) in blood and urine from NMIBC patients is an evidence of immune response activation against neoantigens, and it interferes directly in tumor evolution. For that reason, in this proposal, we aim at investigating if immune system activation, measured by TCR diversity, can be observed in blood and urine samples of NMIBC patients, and if it is correlated to clinical pathological parameters. This proposal integrates two other larger projects supervised by Dr. Masotti, which aims to access TCR diversity as a molecular biomarker for NMIBC prognosis and BCG response, using urine and blood samples. In the first phase of this project, we will explore the correlation between diversity and tumor growth, using computational pipelines and TCR-seq data from NMIBC tumoral infiltrates previously generated by our group. In the second phase, for which sample collecting has already begun, we will sequence the circulating TCR repertoire from blood and urine samples from approximately 20 NMIBC patients hospitalized for TURB at the Advanced Urology Nucleus from Sírio-Libanês Hospital. The data generated in the second phase will allow us to: (I) estimate TCRs diversity using bioinformatics, (II) compare diversity estimates between blood and urine samples, and (III) explore the correlation of diversity with clinical-pathological parameters. This study aims to contribute to the future development of molecular tests for relapse risk prediction and, consequently, to improve patient care through better therapeutic choice for NMIBC.(AU)