The last years, cowpox infections are being increasingly reported through Eurasia. Cowpox viruses (CPXVs) have been reported to have different genotypes and may be subdivided in at least five genetically distinct monophyletic clusters. However, little is known about their in vitro and in vivo features. In this report, five genetically diverse CPXVs, including one reference strain (CPXV strain Brighton) and four clinical isolates from human and animal cases, were compared with regard to growth in cells, pathogenicity in mice and inhibition by antivirals. While all CPXVs replicated similarly in vitro and showed comparable antiviral susceptibility, marked discrepancies were seen in vivo, including differences in virulence with recorded mortality rates of 0%, 20% and 100%. The four CPXV clinical isolates appeared less pathogenic than two reference strains, CPXV Brighton and vaccinia virus Western-Reserve. Disease severity seemed to correlate with high viral DNA loads in several organs, virus titers in lung tissues and levels of IL-6 cytokine in the sera. Our study highlighted that the species CPXV consists of viruses that not only differ considerably in their genotypes but also in their in vivo phenotypes, indicating that CPXVs should not be longer classified as a single species. Lung virus titers and IL-6 cytokine level in mice may be used as biomarkers for predicting disease severity. We further demonstrated the potential benefit of cidofovir, CMX001 and ST-246 use as antiviral therapy.

Human papillomavirus (HPV) causes cervical cancer and a large fraction of head and neck squamous cell carcinomas (HNSCC). Cidofovir (CDV) proved efficacious in the treatment of several HPV-induced benign and malignant hyper proliferations. To provide a better insight into how CDV selectively eradicates transformed cells, HPV+ and HPV− cervical carcinoma and HNSCC cell lines were compared to normal cells for antiproliferative effects, CDV metabolism, drug incorporation into cellular DNA, and DNA damage. Incorporation of CDV into cellular DNA was higher in tumor cells than in normal cells and correlated with CDV antiproliferative effects, which were independent of HPV status. Increase in phospho-ATM levels was detected following CDV exposure and higher levels of γ-H2AX (a quantitative marker of double-strand breaks) were measured in tumor cells compared to normal cells. A correlation between DNA damage and CDV incorporation into DNA was found but not between DNA damage and CDV antiproliferative effects. These data indicate that CDV antiproliferative effects result from incorporation of the drug into DNA causing DNA damage. However, the anti-tumor effects of CDV cannot be exclusively ascribed to DNA damage. Furthermore, CDV can be considered a promising broad spectrum anti-cancer agent, not restricted to HPV+ lesions.

Tumor-tumor distant interactions within one organism are of major clinical relevance determining clinical outcome. To investigate this poorly understood phenomenon, a double human cervical xenograft model in nude mice was developed. A first tumor was induced subcutaneously by injection of human papillomavirus positive cervical carcinoma cells into the mouse lower right flank and 3 weeks later, animals were challenged with the same tumor cell line injected subcutaneously into the upper left flank. These tumors had no direct physical contact and we found no systemic changes induced by the primary tumor affecting the growth of a secondary tumor. However, ablation of the primary tumor by local treatment with cidofovir, a nucleotide analogue with known antiviral and antiproliferative properties, resulted not only in a local antitumor effect but also in a temporary far-reaching effect leading to retarded growth of the challenged tumor. Cidofovir far-reaching effects were linked to a reduced tumor-driven inflammation, to increased anti-tumor immune responses, and could not be enhanced by co-administration with immune stimulating adjuvants. Our findings point to the potential use of cidofovir in novel therapeutic strategies aiming to kill tumor cells as well as to influence the immune system to fight cancer.