Local radiotherapy is the major therapeutic approach to control inoperable cervical cancer. In this study, we analyzed the local immune microenvironment of cervical cancer before and after clinical radiation therapy to investigate whether tumor response due to immunomodulation. A total of 59 patients with pathologically diagnosed cervical cancers classified according to the International Federation of Obstetrics and Gynecology (FIGO) criteria were recruited. The patients were treated according to their disease status with standard radiation regimens. For each patient, tumor biopsies were conducted before, during and after radiation treatment with the doses of 0, 10, 20 and 30 Gy, respectively. All of the tumor samples were then grouped according to the doses delivered and tumor infiltrating lymphocytes with the biomarkers of CD8, CD4, FOXP3 and OX40 were measured by in situ immunohistochemistry. We found that before radiation treatment both CD8(+) T cell and CD4(+) T cell infiltrates were more present in the tumor stroma than in the tumor nests, while OX40(+) and FOXP3(+) T cell infiltrates were present at similar levels in both the tumor nests and stroma. After radiation treatment, the levels of CD8(+) T cells and CD4(+) T cells in the tumor nests and stroma were decreased compared with the levels before irradiation. However, OX40(+) T cells and FOXP3(+) T cells did not show any difference before and after irradiation, which indicates that the FOXP3(+) T cells were more resistant to ionizing radiation than were the cytotoxic effector T cells and demonstrates a dynamic rebalance of infiltrating lymphocytes in the tumor milieu occurs after radiotherapy. This suggest that local antitumor immunity could be compromised due to decreased cytotoxic effector cells and the relatively stable status of FOXP3(+) T cells after irradiation. Therefore, regulation of these FOXP3(+) T cells may be a potentially effective approach to enhance the efficacy of cancer radiation therapy.

Integration of high-risk human papillomavirus (HPV) into the host genome is a key event for cervical carcinogenesis. Different methods have been used to explore the physical states of the HPV genome to reveal the mechanisms for malignant transformation of the infected cells. Consensus has been reached that, although variable portions of the HPV genome are deleted in the integrated HPV sequences, common disruption of the viral E2 gene has been demonstrated in different studies. The head-to-tail concatemers of the full-length HPV16 genome is another typical integration pattern of HPV16, typically found in Caski cell lines, but its prevalence in cervical cancer has never been tested. Here, by introducing a modified PCR, we identified this head-to-tail concatemers of full-length HPV genomes in advanced cervical cancer with HPV16 single positive. Our results show that more than half of the cases contain this integrated head-to-tail concatemers of full-length HPV16 genomes. Further studies in two cervical cell lines, Caski cells and Siha cells, revealed a correlation between the prevalence of the spliced variants of integrated HPV16 sequences and the full-length transcription of the integrated head-to-tail concatemers of the full-length HPV16 genome. Based on these results, we propose that HPV16 integrated into host cells by two mechanisms: one mechanism is shared by other DNA virus and cause integration of the head-to-tail concatemers of the viral genome; another is related to the reverse transcription process, which the integrated HPV sequence is generated by the reverse transcription of the viral mRNA.