Sensitive detection of FGFR3 mutations in bladder cancer and urine sediments by peptide nucleic acid-mediated real-time PCR clamping.
Somatic mutations of the fibroblast growth factor receptor 3 (FGFR3) gene were detected by peptide nucleic acid (PNA)-mediated real-time PCR clamping. Mutation was detected in negative control containing only wild-type DNA due to a misincorporation of dNTPs to PNA binding sites when the amount of template DNA was decreased to 1 ng. Thus, the amount of template DNA was critical determinant of the assay sensitivity in PNA-mediated PCR clamping. Assay conditions were optimized to detect FGFR3 mutations in exons 7, 10, and 15, at a concentration of more than 1% mutated DNA using 50 ng of genomic DNA as the template. Mutations were detected in 12 of 13 (92.3%) tumor tissues and 11 of 13 (84.6%) urine samples from patients with superficial bladder cancer, while no mutations were detected in tissues and/or urine samples from patients with muscle-invasive bladder cancer or chronic cystitis.
Miyake
Makito
M
Oncogene Research Unit/Cancer Prevention Unit, Tochigi Cancer Center Research Institute, 4-9-13 Yonan, Utsunomiya, Tochigi 320-0834, Japan.
Sugano
Kokichi
K
Kawashima
Kiyotaka
K
Ichikawa
Hiroki
H
Hirabayashi
Kaoru
K
Kodama
Tetsuro
T
Fujimoto
Hiroyuki
H
Kakizoe
Tadao
T
Kanai
Yae
Y
Fujimoto
Kiyohide
K
Hirao
Yoshihiko
Y
D004252:DNA Mutational Analysis / Q000379:methods*; D015342:DNA Probes / Q000235:genetics; D006801:Humans; D020135:Peptide Nucleic Acids / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D051498:Receptor, Fibroblast Growth Factor, Type 3 / Q000235:genetics* / Q000652:urine*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D016482:Urinalysis / Q000379:methods*; D001749:Urinary Bladder Neoplasms / Q000235:genetics*