The late region of human neurotropic JC virus encodes a small 71-amino-acid agnoprotein that is also found in the polyomaviruses simian virus 40 and BK virus. Several functions of agnoprotein have been identified, including roles in regulating viral transcription and virion maturation. Earlier studies showed that agnoprotein expressed alone induced p21/WAF-1 expression and caused cells to accumulate in the G₂/M stage of the cell cycle. Here we report that agnoprotein expression sensitized cells to the cytotoxic effects of the DNA-damaging agent cisplatin. Agnoprotein reduced the viability of cisplatin-treated cells and increased chromosome fragmentation and micronucleus formation. Whereas cisplatin-treated control cells accumulated in S phase, cells expressing agnoprotein did not, instead becoming aneuploid. Agnoprotein expression correlated with impaired double-strand-break repair activity in cellular extracts and reduced expression of the Ku70 and Ku80 DNA repair proteins. After agnoprotein expression, much of the Ku70 protein was located in the perinuclear space, where agnoprotein was also found. Results from binding studies showed an interaction of agnoprotein with Ku70 which was mediated by the N terminus. The ability of agnoprotein to inhibit double-strand break repair activity when it was added to cellular extracts was also mediated by the N terminus. We conclude that agnoprotein inhibits DNA repair after DNA damage and interferes with DNA damage-induced cell cycle regulation. Since Ku70 is a subunit of the DNA-dependent protein kinase that is responsible both for double-strand break repair and for signaling damage-induced cell cycle arrest, the modulation of Ku70 and/or Ku80 by agnoprotein may represent an important event in the polyomavirus life cycle and in cell transformation.

The aim of the present study was to examine, using the micronucleus (MN) assay, the low-dose radiation response of normal skin cells from cancer patients and to determine whether the hyper-radiosensitivity (HRS)-like phenomenon occurs in cells of these patients. Primary skin fibroblasts and keratinocytes derived from 40 patients with cervix cancer were studied. After in vitro gamma irradiation with single doses ranging from 0.05 to 4 Gy, MN induction was assessed. For each patient, the linear-quadratic (LQ) model and the induced repair (IR) model were fitted over the whole data set. In fits of the IR model, an HRS-like response after low doses (seen as the deviation over the LQ curve) was demonstrated for the fibroblasts of two patients and for the keratinocytes of four other patients. The alpha(s)/alpha(r) ratio for the six patients ranged from 2.7 to 15.4, whereas the values of the parameter d(c) ranged from 0.13 to 0.36 Gy. No relationship was observed between chromosomal radiosensitivity of fibroblasts and keratinocytes derived from the same donor in the low-dose (0.1-0.25 Gy) region. In conclusion, the fact that low-dose chromosomal hypersensitivity was observed for cells of only six of the patients studied suggests that it is not a common finding in human normal cells and can represent an individual characteristic.

Purpose: HPV is involved in the development of some head and neck squamous-cell carcinomas (HNSCC). It was suggested that only transcriptionally active virus can induce carcinogenesis, therefore, the aim of our study was to analyze the frequency of active HPV infection, virus type, and its prognostic role in HNSCC patients.Methods: Status of active HPV infection was assessed for 155 HNSCC patients based on p16 expression and HPV DNA presence. Univariate and multivariate analyses with Cox proportional regression model were performed to select independent prognostic factors.Results: Active HPV infection was detected in 20.65% of patients. We identified 16.0, 40.9 and 1.7% of HPV positive oral cavity, oropharyngeal, and laryngeal cancer cases, respectively. HPV16 was dominant (81.25%) followed by HPV35 (9.38%) and double infections with HPV16 and 35 (6.25%) or HPV35 and 18 (3.12%). Patients with active HPV infection demonstrated significantly higher survival than HPV negative ones (OS 80.89% vs. 37.08%,p = 0.000; DFS 93.0% vs. 53.35%,p = 0.000, respectively). Longer OS and DFS were maintained for infected patients when oropharyngeal and non-oropharyngeal cases were analyzed separately. Interestingly, all patients infected with other than HPV16 types survived 5 years without cancer progression. In the analyzed group of 155 patients the strongest independent favourable prognostic factor for both OS and DFS was HPV presence.Conclusions: High prevalence of HPV-driven HNSCC (mostly within oropharynx) was detected, with HPV16 type the most frequent, followed by HPV35 and HPV18. The presence of active HPV infection improved survival of both oropharyngeal and non-oropharyngeal cancer patients and should be taken into account in treatment planning.

To investigate whether the in vitro radiosensitivity of normal lymphocytes and fibroblasts evaluated by the micronucleus (MN) assay predicts acute and late reactions after radio-therapy in cancer patients. Studies were performed on blood samples from 31 cervical and head and neck cancer patients and on skin fibroblasts from eight of the cancer patients. The radiosensitivity of lymphocytes and of fibroblasts was also assessed in 24 and five healthy donors, respectively. Radiosensitivity was measured after in vitro irradiation with doses ranging from 2 to 5 Gy using micronucleus frequency (the number of micronuclei per single binucleated (BN) cell) and the percentage of BN cells with micronuclei. The in vitro results were compared with the maximum grade of acute and late reactions. There was no significant difference in cellular radiosensitivity between cancer patients and healthy donors. Although cancer patients differed considerably in normal-cell radiosensitivity, no correlation was found between radiosensitivity, either of lymphocytes or fibroblasts, and acute and late clinically observed side effects. In addition, no relationship was observed between the radiosensitivity of lymphocytes and fibroblasts derived from the same donors. The data do not support the usefulness of the MN assay in predicting normal-tissue response to radiotherapy in cancer patients.