FGFR testing and urine-based risk straticfication from matched tissue and urine samples within the prospective real-world clinicopathological register trial: BRIDGister.
Authors: Wirtz Ralph M. , Watts Richard , Kellner Ronny , Ortmann Reinhard , Horns Torsten , Enderle Daniel , Meyer Lisa , Noerholm Mikkel , Morken Mario , Veltrup Elke , Hake Roland , Eidt Sebastian , Roggisch Jenny , Koch Stefan , Ecke Thorsten 
Published: February 2022

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Background: The objective of the present study was to assess FGFR mutattions and fusions from matched urine and tissue samples from patients suspicious of bladder cancer and undergoing first TURB within BRIDGister RealWorld Experience trial. Methods: FFPE samples from the first TURB of 39 pts participating in the BRIDGister trial and matched urine samples were prospectively collected and analyzed. RNA from FFPE tissues were extracted by commercial kits and analyzed by Therascreen FGFR IVD kit (Qiagen GmbH, Hilden). In addition extracellular vesicles were centrally isolated for subsequent RNA extraction (exoRNA.Exosome Diagnostics GmbH, Martinsried) and centrall analysis by QIAcuity digital PCR (Qiagen, Hilden). In addition mRNA based profiling of urine was done by dPCR. Concordance, Spearman, Kruskal-Wallis and MannWhitney were analyzed by JMP 9.0.0 (SAS software). Results: The pilot cohort of the BRIDGister trial consisted of 39 patients (median age: 75, male 69% vs female 31%) of diverse clinical stages (Benign lesions/no tumor 23%, pTa 31%, pT1 26%, pT2 21%) and WHO 1973 grade (G1 8%, G2 39%, G3 34%). Based on FFPE tissue testing using Therascreen FGFR IVD kit and exosomal RNA extraction follwoed by dPCR 12 out of 39 patients exhibited FGFR alterations (31%). Comparison with tissue testing as probable gold standard revealed 67% sensitivity, 85% specificity, 67% PPV and 85%NPV. There were 4 patients being FGFR positive for exoRNA from urine with no mutation found in the corresponding TUR biopsy. Determining ERBB2 mRNA by dPCR from urine revealed that high ERBB2 mRNA correlated with higher WHO1973grade, while high FGFR3 mRNA correlated with lower grade tumors (Spearman r=0.4386 p=0.0075 and r=-0.4663 p=0.0042). Similiarly, trends were seen for association of ERBB2 and FGFR3 mRNA with clinical stage tumors in this pilote cohort (Spearman r=0.2359 p=0.0923 and r=-0.2249 p=0.1089). Conclusions: Extraction of exosomal RNA from urine followed by highly sensitive dPCR mutation testing is feasible with good concordance to matched tissue testing. Urine testing might evolve as alternative approach for FGFR3 screening in a non invasive fashion without the need of transurethral biopsy. Interestingly, mRNA assessment of exosomal RNA from urine before TURB correlated with clinical parameters such as WHO Grade 1973 with ERBB2 mRNA being associated with high grade tumors and FGFR3 mRNA being associated with low grade tumors, which is in line with previous tissue tsting results. This indicates the potential to clinically characterize tumor grade and stage before TUR biopsy or surgery which might be helpful for future risk stratification and planning of surgical intervention. Further exploration is warranted and includes the potential of monitoring patients with regard to urine based mutation detection and risk stratification.