A therapeutic DNA vaccine and gemcitabine act synergistically to eradicate HPV-associated tumors in a preclinical model.

Although active immunotherapies are effective strategies to induce activation of CD8+ T cells, advanced stage tumors require further improvements for efficient control. Concerning the burden of cancer-related to Human papillomavirus (HPV), particularly the high incidence and mortality of cervical cancer, our group developed an approach based on a DNA vaccine targeting the HPV-16 E7 oncoprotein (pgDE7h). This immunotherapy is capable of inducing an antitumour CD8+ T cell response but show only partial control of tumors in more advanced growth stages. Here, we combined a chemotherapeutic agent (gemcitabine- Gem) with pgDE7h to overcome immunosuppression and improve antitumour responses in a preclinical mouse tumor model. Our results demonstrated that administration of Gem had synergistic antitumor effects when combined with pgDE7h leading to eradication of both early-stages and established tumors. Overall, the antiproliferative effects of Gem observed in vitro and in vivo provided an optimal window for immunotherapy. In addition, the enhanced antitumour responses induced by the combined therapeutic regimen included enhanced frequencies of antigen-presenting cells (APCs), E7-specific IFN-γ-producing CD8+ T cells, and cytotoxic CD8+ T cells and, concomitantly, less pronounced accumulation of immunosuppressive myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs). These findings demonstrated that the combination of Gem and an active immunotherapy strategy show increased effectiveness, leading to a reduced need for multiple drug doses and, therefore, decreased deleterious side effects avoiding resistance and tumor relapses. Altogether, our results provide evidence for a new and feasible chemoimmunotherapeutic strategy that supports future clinical translation.

UVB-induced cell death signaling is associated with G1-S progression and transcription inhibition in primary human fibroblasts.

DNA damage induced by ultraviolet (UV) radiation can be removed by nucleotide excision repair through two sub-pathways, one general (GGR) and the other specific for transcribed DNA (TCR), and the processing of unrepaired lesions trigger signals that may lead to cell death. These signals involve the tumor suppressor p53 protein, a central regulator of cell responses to DNA damage, and the E3 ubiquitin ligase Mdm2, that forms a feedback regulatory loop with p53. The involvement of cell cycle and transcription on the signaling to apoptosis was investigated in UVB-irradiated synchronized, DNA repair proficient, CS-B (TCR-deficient) and XP-C (GGR-deficient) primary human fibroblasts. Cells were irradiated in the G1 phase of the cell cycle, with two doses with equivalent levels of apoptosis (low and high), defined for each cell line. In the three cell lines, the low doses of UVB caused only a transient delay in progression to the S phase, whereas the high doses induced permanent cell cycle arrest. However, while accumulation of Mdm2 correlated well with the recovery from transcription inhibition at the low doses for normal and CS-B fibroblasts, for XP-C cells this protein was shown to be accumulated even at UVB doses that induced high levels of apoptosis. Thus, UVB-induced accumulation of Mdm2 is critical for counteracting p53 activation and apoptosis avoidance, but its effect is limited due to transcription inhibition. However, in the case of XP-C cells, an excess of unrepaired DNA damage would be sufficient to block S phase progression, which would signal to apoptosis, independent of Mdm2 accumulation. The data clearly discriminate DNA damage signals that lead to cell death, depending on the presence of UVB-induced DNA damage in replicating or transcribing regions.