Analysis of human papillomavirus type 16 E6, E7 sequence variation in primary cervical cancer from Korean women and its relationship to the expression of immunomodulatory gene

Kim, Jae Won; Park, In Ae; Lee, Hyo Pyo

Abstract

OBJECTIVES
As one of the possible mechanisms of viral evasion in the HPV-infected cervical cancer cells, the role of amino acid sequence change in epitope region has not been reported yet. In this study, sequence variations of HPV 16 E6, E7 gene, especially focused on epitope region, were analysed, the status of immunomodulatory factors were documented, and finally the possible correlation between the sequence variations and the loss of HLA class I expression was examined. METHODS: The entire ORF(open reading frame)s of HPV 16 E6, E7 were sequenced by the fluorescent dideoxy termination method. In addition proteins and transcripts of HLA-ABC, beta2-microglobulin(beta2-m), TAP (transporter associated with antigen processing), and LMP(large multi-functional proteasome) were evaluated by immunohistochemistry and RT-PCR, respectively in 40 clinical specimens of primary cervical cancer and 6 cervical cancer cell-lines. Medical records including pathologic reports were reviewed. RESULTS: Among the 27 cases confirmed as harboring HPV 16 DNA, only one(3.7%) found as a prototype. Among 11 kind of variants identified in total, 4 variants(5 nucleotide sites) which were never reported before has been found, registered firstly to GenBank. The most frequently found one(16 cases, 59.3%) contains D25E, N29S in E6, E7 region, respectively and the most common variation in E6, E7 ORFs found concurrently(p<0.05). Down-regulation of HLA-ABC and beta2-m was identified in 32(86.5%) and 35 cases(89.7%), respectively and transcripts of TAP, LMP were identified in over 85% of cases. However, there was no significant difference in HPV 16 infection, D25E in E6 and so on between HLA-ABC, beta2-m positive and negative groups. The well-known clinicopathologic parameters did not correlate with sequence variations and immunomodulatory factors. Five sequence variations in HPV 16 E6, E7 ORFs that were not previously reported worldwide were found, registered firstly to GenBank. CONCLUSION: It seems that multiple mechanisms are operated in down-regulation of HLA class I molecules and the phenotypic profile of immunomodulatory factors seems to be unrelated in vivo to the naturally occurring HPV 16 E6, E7 variations in epitope region.