A simple and fast method for the simultaneous detection of nine fibroblast growth factor receptor 3 mutations in bladder cancer and voided urine.
OBJECTIVE
Mutations in the fibroblast growth factor receptor 3 (FGFR3) occur in 50% of primary bladder tumors. An FGFR3 mutation is associated with good prognosis, illustrated by significantly lower percentage of patients with progression and disease-specific mortality. FGFR3 mutations are especially prevalent in low grade/stage tumors, with pTa tumors harboring mutations in 85% of the cases. These tumors recur in 70% of patients. Efficient FGFR3 mutation detection for prognostic purposes and for detection of recurrences in urine is an important clinical issue. In this paper, we describe a simple assay for the simultaneous detection of nine different FGFR3 mutations.


METHODS
The assay consists of one multiplex PCR, followed by extension of primers for each mutation with a labeled dideoxynucleotide. The extended primers are separated by capillary electrophoresis, and the identity of the incorporated nucleotide indicates the presence or absence of a mutation.


RESULTS
The assay was found to be more sensitive than single-strand conformation polymorphism analysis. Mutations could still be detected with an input of only 1 ng of genomic DNA and in a 20-fold excess of wild-type DNA. Moreover, in urine samples from patients with a mutant tumor, the sensitivity of mutation detection was 62%.


CONCLUSIONS
We have developed a fast, easy to use assay for the simultaneous detection of FGFR3 mutations, which can be of assistance in clinical decision-making and as an alternative for the follow-up of patients by invasive cystoscopy for the detection of recurrences in urine.
van Oers
Johanna M M
JM
Department of Pathology, Josephine Nefkens Institute, Erasmus MC, Rotterdam, the Netherlands.
Lurkin
Irene
I
van Exsel
Antonius J A
AJ
Nijsen
Yvette
Y
van Rhijn
Bas W G
BW
van der Aa
Madelon N M
MN
Zwarthoff
Ellen C
EC
D004247:DNA / Q000378:metabolism; D004252:DNA Mutational Analysis; D017931:DNA Primers / Q000235:genetics; D003854:Deoxyribonucleotides / Q000235:genetics; D019075:Electrophoresis, Capillary; D006801:Humans; D009154:Mutation; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D011379:Prognosis; D051498:Receptor, Fibroblast Growth Factor, Type 3 / Q000235:genetics*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000652:urine*
D017931:DNA Primers; D003854:Deoxyribonucleotides; D004247:DNA; D051498:Receptor, Fibroblast Growth Factor, Type 3