The aim of this study was to analyse the correlation between HPV16 E6 variants and the physical status of viral genome (integrated, mixed, episomal) among patients with cervical cancer (n=40) and low-grade squamous intraepithelial lesions - LSIL (n=40). The study was performed on 80 HPV16 positive samples. HPV16 E6 variants were identified using PCR and DNA sequencing. Nucleotide sequences of E6 were compared with the prototype sequence (EUR-350T). The physical state of HPV DNA was determined as the ratio of E2/E6 copy number per cell. Twelve different intratypic variants were identified as belonging to European (in 77 samples) and North-American 1 (in 3 samples) sublineages. The most prevalent non-synonymous variant was EUR-350G, which occurred with similar frequency in cervical cancer and LSIL. The frequencies of additional mutations in variants with EUR-350T or EUR-350G sequences differed significantly. For the first time, missense mutations G122A, C153T and G188A were discovered in EUR-350G variant. The integrated viral genome was predominant in women with cervical cancer. The EUR-350T prototype and EUR-350G without additional mutations variants were prevalent in cervical cancer samples with the HPV16 characterized by integrated DNA. In summary, European variants of HPV16 E6 dominated in both cancer and LSIL group. The presence of EUR-350G favoured the occurrence of additional nucleotide changes. We showed that nucleotide changes occur significantly more often in the mixed form of viral DNA and in LSIL group and that the variants without additional mutations may promote integration of HPV16 genome.
Keywords: HPV type 16 E6 variants; HPV16 physical state; invasive cervical carcinoma; real-time PCR; squamous intraepithelial lesions.