Usefulness of droplet digital PCR and Sanger sequencing for detection of FGFR3 mutation in bladder cancer

Abstract

Objective: The purpose of our research was to determine the usefulness of different methods for detecting Y373C mutation of gene FGFR3.

Patients and methods total: 138 primary bladder cancer patients (71cases G1 and 67 cases G2-G3) were included in the study. Tumor tissue and urine samples were collected and kept frozen until the isolation of DNA. Sanger sequencing was applied for detecting mutation in cancer and ddPCR was utilized for urine assessment.

Results: ddPCR appears to be more effective and it identified FGFR3 mutation (Y373C) in urinary sediment in 20.3% of cases whereas Sanger sequencing did in 15.5%. Only in 8/39 (20.5%) cases the mutation was observed both in urine and tissue. In 12/39 (30.8%) cases (5G1 and 7 G2-G3) we did not detect any FGFR3 mutation in urine although it was confirmed by sequencing. We only found mutation in urine in 20/39 cases (15 G1, 5 G2-G3) (51.3%). The correlation between the presence of FGFR3 mutations and better survival was confirmed. The Log-Rank test indicates a significant difference in the likelihood of survival for patients with the FGFR3 mutation but without recurrence (Cox's F-test P = 0.17006; Log-Rank Test P = 0.00059).

Conclusion: ddPCR appeared to be more sensitive method for detection FGFR3 gene mutation particularly for detecting low levels of tumor DNA amongst a large excess of nontumor DNA. It is significant as the implementation of such markers into routine practice could be beneficial. The prospective study in larger cohort is needed.

Keywords: Bladder cancer; FGFR3 mutation; Sequencing; ddPCR.