RNA based liquid biopsy testing of urine to predict pathological complete response to radiomolecular instillation NAC therapy before surgery: The Bladder BRIDGister experience.

Abstract

643 Background: Patients with MIBC achieving pathological complete response (pCR) upon neoadjuvant chemotherapy (NACT) have improved prognosis. While gene expression profiles from tumor tissues are predictive of response, so far no expression profiling of tumor cells in urine is established to detect tumor cells or monitor response towards ongoing treatment. However, tumor specific gene expression in urine samples may enable non-invasive testing and may distinguish responding from non-responding tumors for early treatment adoption / intensification at each neaoadjuvant treatment cycle. Methods: Tumor cells were isolated from 10 ml urine collected before TUR biopsy and before each cycle of treatment as well as before deep TURB or cystectomy. Similiarly whole urine was drawn before and at each cycle. FFPE tissues as well as matched urine samples from 12 patients receiving systemic chemotherapy +/- intravesical Radioligandtherapy were prospectively collected, RNA extracted, and relative gene expression subtyping markers and targets were analysed by RT-qPCR.Dynamics of urine expression profiles were compared to treatment time point and pathological outcome. Results: RNA expression profiling of matched TUR biopsy tissue and by filtering tumor cells from urine samples allowed robust detection of molecular marker profiles identical to matched tissue profiles. The high dynamic range of KRT5 and KRT20 mRNA allowed detection of 100 tumor cells per 10 ml urine and enabled molecular subtyping into luminal and non luminal subtypes. Interestingly, while non luminal tumors with high KRT5 mRNA expression predicted to not respond to chemotherapy exhibited no KRT5 mRNA decline at sequential time points before each neoadjuvant chemotherapy, the addition of Lu 177 based radioligand therapy broke chemotherapy resistance and resulted in stepwise drop of KRT5 mRNA at each therapy cycle. Fast decline of KRT mRNA expression indicated complete response towards combination therapy already within the first 6 weeks. Conclusions: The results contribute to our findings of treatment efficacy of radioligand instillation therapies in otherwise chemotherapy resistant tumors. Moreover, by comparing gene expression profiles from urine samples before and before each cycle of therapy the response could be recapitulated and detected already after one week of monotherapy radioligand treatment. This suggests that expression profiling from urine samples is feasible andallows non invasive prediction of treatment outcome, which will be further analyzed in upcoming phase I/II intravesical radioligand therapy trials of the Bladder BRIDGister clinical trial group.