Loss of heterozygosity and mutations are the major mechanisms of RB1 gene inactivation in Chinese with sporadic retinoblastoma.
We investigated sequence alternation, promoter methylation, and loss of heterozygosity (LOH) of the RB1 gene as possible mechanisms of its inactivation in retinoblastoma. In 42 Chinese patients with sporadic retinoblastoma, the promoter and entire coding region of RB1 were examined for sequence changes. Status of methylation of the CpG-rich island at the 5'end was determined by methylation specific PCR assay. We detected 15 RB1 mutations in 38% (16/42) of the retinoblastoma patients, among them 19% (8/42) were germ-line mutations. A total of nine novel mutations were identified: E54X, S114X, I126S, g73779insG, D718N, IVS2+1G>C, IVS14+1G>C, IVS21+1G>C, and a complex alteration g78177G>T/g78176insTT leading to 543X. Most of them are likely to affect the RB1large pocket domain through the production of truncated gene products. None of the DNA samples showed methylation at the RB1promoter. In 15 cases where both normal and cancerous retinoblastoma tissue specimens were available, allelic loss according to microsatellite markers within or distal to the RB1 locus was analyzed and immunohistological staining for RB1 expression performed. Among them, frequency of LOH at 13q14 was found to be high at 60% (9/15) with no segregation with unilateral tumors. All these nine tumors did not express RB1 protein, showing an association of LOH at the RB1 locus with its loss of expression in retinoblastoma. Our results indicate that the RB1 gene in sporadic retinoblastoma is commonly inactivated because of loss-of-function mutations and loss of heterozygosity but not by the epigenetic phenomenon of promoter hypermethylation.
Choy
Kwong Wai
KW
Department of Ophthalmology & Visual Sciences, the Chinese University of Hong Kong, Hong Kong, China.
Pang
Chi Pui
CP
Yu
Christopher B O
CB
Wong
Hing Lok
HL
Ng
Joan S K
JS
Fan
Dorothy S P
DS
Lo
Kwok Wai
KW
Chai
Joshua T Y
JT
Wang
Jianhua
J
Fu
Weiling
W
Lam
Dennis S C
DS
D002675:Child, Preschool; D002681:China; D002882:Chromosomes, Human, Pair 13; D018899:CpG Islands; D019175:DNA Methylation; D004252:DNA Mutational Analysis; D005260:Female; D020868:Gene Silencing; D016161:Genes, Retinoblastoma; D006801:Humans; D007150:Immunohistochemistry; D007223:Infant; D019656:Loss of Heterozygosity; D008297:Male; D008875:Middle Aged; D009154:Mutation; D011401:Promoter Regions, Genetic; D019572:Retinal Neoplasms / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism; D012175:Retinoblastoma / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D014407:Tumor Cells, Cultured
D016160:Retinoblastoma Protein