Abstract 5831: Urinary exosome analysis as a marker of treatment response in bladder cancer patients
Authors: Valera Vladimir , Walter Beatriz , Merino Maria J. 
Published: June 2022

Abstract
Introduction: Intravesical Bacillus Calmette-Guerin (BCG) is an effective immunotherapy for non-muscle invasive bladder cancer (NMIBC). However, BCG treatment failure will lead to recurrence and tumor progression. In this study, urinary exosomes content (miRNA profile) was evaluated as a possible marker of BCG treatment response in NMIBC patients.
Methods: Urine samples from patients with bladder cancer were collected at the time of surgery and during patient follow up to 1 year. Urine from healthy volunteers were also included. Clinical and pathological information such as tumor grade (High Grade (HG), Low Grade (LG)), depth of invasion (Ta, T1, CIS), and response to BCG treatment was also obtained. Exosome Isolation and total RNA extraction including microRNAs from cell-free urine after centrifugation were obtained. Library preparation for miRNA expression was done (QIAseq® miRNA Library) for Next-Generation Sequencing (NGS) analysis in a NextSeq 2000 single read platform, 75 bp with 15-20 million reads per sample. Reads were then queried against miRDeep2 software for identification. Only miRNAs having at least 20 counts considering all samples were included. After normalization, significantly and deferentially expressed microRNAs (&gt;2-Fold) were selected for analysis. Bioinformatic analysis including sequence alignment was performed under the STAR-based approach. Identified microRNAs were then used to classify/predict the response to treatment and its relationship with other clinicopathologic variables.
Results: A total of 56 urine samples from 13 patients were available/used for analysis including 10 High Grade Ta and 3 High Grade T1 patients. Urine from normal healthy donors (N=3) was also included. Clinicopathological features were patients with HGTa=10, HGT1=3 and 3 control samples. Regarding treatment response 9 patients were BCG responders and 4 BCG unresponsive. When compared to BCG unresponsive patients, BCG responders showed 45 differentially expressed miRNAs. Statistically significant differentially expressed miRNAs (Fold-change &gt;2, p value &lt;0.05) were 12 miRNAs, upregulated were miR132-3p (p=0.042); miR-187-3p (p=0.021); miR-409-3p (p=0.043) and miR1301-3p (p=0.048). Downregulated miRNAs were miR-let7-5p (p=0.007), miR-3605-3p (p=0.047), miR-140-5p (p=0.031), miR-500a-5p (p=0.051), miR-629-5p (p=0.039), miR-454-3p (p=0.05), miR-2110 (p=0.049) and miR-30c-5p (p=0.03). Interestingly, miRPathDABv2.0 analysis predicted those microRNAs be related to cancer, including bladder cancer. They showed targeting important pathways as PI3K.Akt, Wnt/B catenin and P53 signaling related with disease progression and treatment response.
Conclusion: Our study supports the value of urinary exosomal microRNAs as non-invasive biomarkers to predict BCG treatment response in nonmuscle-invasive bladder cancer.
Citation Format: Vladimir Valera, Beatriz Walter, Maria J. Merino. Urinary exosome analysis as a marker of treatment response in bladder cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5831.