Microsatellite Instability And Loss Of Heterozygosity Of HLA-I Gene In Cervical Carcinoma

Cervical carcinoma (CC) is the second most malignancy among women worldwide. The highest incidence rates are observed in the developing countries. Previous numerous studies have strongly shown that certain high-risk HPV such as HPV16 and HPV18 infection clearly plays an important role in the development of cervical carcinoma; however, several experimental studies have suggested that HPV infection alone is insufficient to initiate malignant transformation of the cervical epithelium because many more women are infected with HPV than develop cervical cancer. Furthermore, the transition from precursor lesion, cervical intraepithelial neoplasia(CIN), to invasive cancer has a long latency period.Thus, like many other malignant tumors, other genetic events are likely to be critical in the pathogenesis of cervical cancer. Genetic events such as the inactivation of multiple tumor suppressor genes (TSGs), activation of oncogenes, have been involved in the carcinogenesis of cervical carcinoma. In recent years, studies of tumor genetics have suggested the genetic instability of cell genome including LOH and MSI is associated with cell transformation, which produced a new hot-spot in the study of tumor carcinogenesis. LOH and MSI were believed to be the main two characteristics of cell genome instability. Analysis of allelic losses of specific chromosomal regions offers an available method to reveal TSGs associated with the development of cancer. A high frequency of allelic deletions on the short arm of chromosome 6(6p21.3) in which HLA-I gene is located has been noted in cervical carcinoma. This observation indicates that some tumor suppressor genes may be located in this region special in the HLA-I region and play an important role in cervical carcinogenesis. Meanwhile, HLA class I antigen plays a central role in the tumor immunology. Down-regulation and loss of HLA class I antigen gene expression is a common phenomenon in most cervical cancers.Therefore, in this study, our aim is to determine the frequency of microsatellite instability(MSI) and loss of heterozygosity(LOH) at HLA-I loci in cervical carcinoma in a high-incidence population group in which HPV infection is endemic by 8 microsatellitemarkers (D6S265, D6S258, D6S276, C1_2C, Cl₂₅, C3_2ll, MIB, Cl₄₄) in or around the HLA-I gene and to study the role of genome instability of HLA-I loci in the carcinogensis and development of cervical cancer. Meanwhile, we also constructed the detailed deletion map of this region and provided important guidance and foundation for discovering new tumor suppressor genes by investigating the expression of expressed sequence tags (ESTs) localized within this smallest common region using difference display RT-PCR and Northern hybridization and cloning the full-length cDNA of candidated EST through cDNA clone sequencing and bioinformatics.Thirty paired blood and tumor samples were collected from County Wufeng, Hubei Province, a high-incidence area of CC and analyzed by PCR based single-stranded length polymorphism (PCR-SSLP) with eight-polymorphism markers in or around the gene. PCR products of tumor tissue and its corresponding peripheral blood sample were separated by electrophoresis through 6-8% denaturing acrylamide gel and visualized by silver staining detection. Loss of heterozygosity was assigned if one band vanished or more than 50% of a band signal reduction of one allele was shown in the tumor samples when compared to the control peripheral blood samples. MSI was accounted if the shifted or additional band occurred in the tumor samples. The results indicate that 23(76.7%) of the 30 cervical carcinoma cases showed LOH at one or more loci. Higher frequencies of LOH were found at four loci: C3_2ll (50%), Cl₄₄ (37%), Cl₂₅ (36.7%), D6S276 (48.3%). MSI was found in 20 out of 30 cases (66.7%). The data suggest that the LOH and MSI of HLA-I gene might participate in the carcinogenesis of cervical carcinoma. The findings also further show that HLA-I gene LOH may be an important mechanism of down-regulation of HLA-I gen...