DHPLC is superior to SSCP in screening p53 mutations in esophageal cancer tissues.
Mutations of the p53 tumor-suppressor gene universally occur on exons 5-8 in human cancer. We analyzed these mutations in esophageal cancer tissue from 207 patients in China using 2 methods, single-strand conformation polymorphism (SSCP), one of the most frequently used methods, and the recently developed denaturing high-performance liquid chromatography (DHPLC), and compared their sensitivity and efficiency. Exons 5-8 of p53 were amplified from esophageal cancer tissue genomes, screened for fragments of mutations and polymorphisms by SSCP and DHPLC in a blind study and confirmed by direct sequencing to detect the mutations and polymorphisms. The numbers detected by DHPLC were greater than those detected by SSCP, though the rate of mutations and polymorphisms was lower in SSCP than in DHPLC, which appeared to detect smaller mutations (substitutions and 1 bp insertions/deletions). Of the mutations with substitutions detected by DHPLC but not by SSCP, 50% substituted adenosine for other nucleotides, suggesting that these mutations are often missed when SSCP is used. According to these data, the sensitivity of SSCP and DHPLC was 81% and 97%, respectively, and the specificity was 97% and 85%, respectively. Our results suggest that DHPLC may be recommended over SSCP when screening gene mutations. Thus, rates of p53 mutations and polymorphisms in esophageal cancer tissue in Chinese patients were 49% and 41% by DHPLC and SSCP, respectively.
Yamanoshita
Osamu
O
Department of Occupational and Environmental Health, Nagoya University Graduate School of Medicine, Nagoya, Japan. yaman@med.nagoya-u.ac.jp
Kubota
Takeo
T
Hou
Jun
J
Ping
Yu-Min
YM
Zhang
Xiu-Lan
XL
Li
Xiang-Ping
XP
Li
Shao-Sen
SS
Li
Xiao-Xian
XX
Zhu
De-Chen
DC
Fukushima
Yoshimitsu
Y
Nakajima
Tamie
T
D000230:Adenocarcinoma / Q000235:genetics; D000328:Adult; D000368:Aged; D044466:Asian People / Q000235:genetics; D002294:Carcinoma, Squamous Cell / Q000235:genetics; D002681:China; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004938:Esophageal Neoplasms / Q000235:genetics*; D005069:Evaluation Studies as Topic; D005260:Female; D005820:Genetic Testing / Q000379:methods*; D006801:Humans; D008297:Male; D008875:Middle Aged; D009154:Mutation; D018807:Polymorphism, Single-Stranded Conformational; D011489:Protein Denaturation; D012680:Sensitivity and Specificity; D016159:Tumor Suppressor Protein p53 / Q000235:genetics*
D016159:Tumor Suppressor Protein p53