The Isothermal Amplification AmpFire Assay for Human Papillomavirus (HPV) Detection and Genotyping in Formalin-Fixed, Paraffin-Embedded Oropharyngeal Cancer Samples.

Human papillomavirus (HPV)-related oropharyngeal squamous cell carcinomas (OPSCCs) represent a distinct clinical entity compared with HPV-negative tumors with particular regard to treatment response and survival outcome. The aim of this study was to assess the AmpFire Multiplex HR-HPV tests, for the detection and genotyping of 15 high-risk (HR) HPV types in formalin-fixed, paraffin-embedded (FFPE) samples and identify HPV-driven OPSCC. DNA from 160 OPSCC FFPE specimens plus 23 samples from other head and neck primary sites was tested. Results were compared with those obtained using Linear Array HPV-DNA Genotyping Test. Linear Array and AmpFire Multiplex HR-HPV tests showed, for all samples and specifically for OPSCCs, an overall concordance agreement of 98.9% and 99.4% and a Cohen κ coefficient of 0.972 and 0.984, respectively. A concordance of 100% for HPV16 and HPV18 was observed. The overall agreement between p16INK4a overexpression and HPV detection by the AmpFire Multiplex HR-HPV assay in 145 OPSCC samples was 93.8%, with a Cohen κ coefficient of 0.848. The AmpFire HPV Tests are simple assays for detection and genotyping of HPV-DNA in OPSCC FFPE samples and can be easily implemented in the clinical practice setting for HPV-DNA detection.

VAV2 signaling promotes regenerative proliferation in both cutaneous and head and neck squamous cell carcinoma.

Regenerative proliferation capacity and poor differentiation are histological features usually linked to poor prognosis in head and neck squamous cell carcinoma (hnSCC). However, the pathways that regulate them remain ill-characterized. Here, we show that those traits can be triggered by the RHO GTPase activator VAV2 in keratinocytes present in the skin and oral mucosa. VAV2 is also required to maintain those traits in hnSCC patient-derived cells. This function, which is both catalysis- and RHO GTPase-dependent, is mediated by c-Myc- and YAP/TAZ-dependent transcriptomal programs associated with regenerative proliferation and cell undifferentiation, respectively. High levels of VAV2 transcripts and VAV2-regulated gene signatures are both associated with poor hnSCC patient prognosis. These results unveil a druggable pathway linked to the malignancy of specific SCC subtypes.

Human papillomavirus DNA detected in fingertip, oral and bathroom samples from unvaccinated adolescent girls in Tanzania.

OBJECTIVE: Human papillomavirus (HPV) DNA has been detected in vaginal samples from adolescent girls who report no previous sex and, in high-income settings, from fingertips, raising the possibility of non-sexual transmission. No such studies originate from East Africa which bears among the highest cervical cancer incidence and HPV prevalence worldwide. HPV-related oral cancer incidence is increasing, but oral HPV prevalence data from East Africa are limited. We aimed to describe the HPV DNA prevalence in genital and non-genital sites and in the bathroom of unvaccinated adolescent girls, and examine genotype concordance between sites. METHODS: We nested a cross-sectional study of HPV in genital and extragenital sites within a cohort study of vaginal HPV acquisition. Unvaccinated girls age 16-18 years in Tanzania, who reported ever having had sex, were consented, enrolled and tested for the presence of HPV DNA in vaginal samples collected using self-administered swabs, oral samples collected using an oral rinse, and on fingertips and bathroom surfaces collected using a cytobrush. RESULTS: Overall, 65 girls were enrolled and 23 (35%, 95% CI 23% to 47%) had detectable vaginal HPV. Adequate (ß-globin positive) samples were collected from 36 girls' fingertips and HPV was detected in 7 (19%, 95% CI 6% to 33%). 63 girls provided adequate oral samples, 4 (6%, 95% CI 0% to 13%) of which had HPV DNA detected. In bathroom samples from 58 girls, 4 (7%, 95% CI 0% to 14%) had detectable HPV DNA. Of the 11 girls with extragenital HPV, six had the same genotype in >1 site. CONCLUSION: We found a high prevalence of HPV in non-genital sites in adolescent girls and in their bathrooms, in this region with a high cervical cancer incidence. Concordance of genotypes between sites supports the possibility of autoinoculation.

Subcutaneous preconditioning increases invasion and metastatic dissemination in mouse colorectal cancer models.

Mouse colorectal cancer (CRC) models generated by orthotopic microinjection of human CRC cell lines reproduce the pattern of lymphatic, haematological and transcoelomic spread but generate low metastatic efficiency. Our aim was to develop a new strategy that could increase the metastatic efficiency of these models. We used subcutaneous implantation of the human CRC cell lines HCT116 or SW48 prior to their orthotopic microinjection in the cecum of nude mice (SC+ORT). This subcutaneous preconditioning significantly enhanced metastatic dissemination. In the HCT116 model it increased the number and size of metastatic foci in lymph nodes, lung, liver and peritoneum, whereas, in the SW48 model, it induced a shift from non-metastatic to metastatic. In both models the number of apoptotic bodies in the primary tumour in the SC+ORT group was significantly reduced compared with that in the direct orthotopic injection (ORT) group. Moreover, in HCT116 tumours the number of keratin-positive tumour buddings and single epithelial cells increased at the invasion front in SC+ORT mice. In the SW48 tumour model, we observed a trend towards a higher number of tumour buds and single cells in the SC+ORT group but this did not reach statistical significance. At a molecular level, the enhanced metastatic efficiency observed in the HCT116 SC+ORT model was associated with an increase in AKT activation, VEGF-A overexpression and downregulation of ß1 integrin in primary tumour tissue, whereas, in SW48 SC+ORT mice, the level of expression of these proteins remained unchanged. In summary, subcutaneous preconditioning increased the metastatic dissemination of both orthotopic CRC models by increasing tumour cell survival and invasion at the tumour invasion front. This approach could be useful to simultaneously study the mechanisms of metastases and to evaluate anti-metastatic drugs against CRC.

High RAB25 expression is associated with good clinical outcome in patients with locally advanced head and neck squamous cell carcinoma.

Currently there are no molecular markers able to predict clinical outcome in locally advanced head and neck squamous cell carcinoma (HNSCC). In a previous microarray study, RAB25 was identified as a potential prognostic marker. The aim of this study was to analyze the association between RAB25 expression and clinical outcome in patients with locally advanced HNSCC treated with standard therapy. In a retrospective immunohistochemical study (n = 97), we observed that RAB25-negative tumors had lower survival (log-rank, P = 0.01) than patients bearing positive tumors. In an independent prospective mRNA study (n = 117), low RAB25 mRNA expression was associated with poor prognosis. Using classification and regression tree analysis (CART) we established two groups of patients according to their RAB25 mRNA level and their risk of death. Low mRNA level was associated with poor local recurrence-free (log-rank, P = 0.005), progression-free (log-rank, P = 0.002) and cancer-specific (log-rank, P < 0.001) survival. Multivariate Cox model analysis showed that low expression of RAB25 was an independent poor prognostic factor for survival (hazard ratio: 3.84, 95% confidence interval: 1.93-7.62, P < 0.001). Patients whose tumors showed high RAB25 expression had a low probability of death after treatment. We also found lower RAB25 expression in tumors than in normal tissue (Mann-Whitney U, P < 0.001). Moreover, overexpression of RAB25 in the UM-SCC-74B HNSCC cell line increased cisplatin sensitivity, and reduced cell migration and invasion. Our findings support a tumor suppressor role for RAB25 in HNSCC and its potential use to identify locally advanced patients with a high probability of survival after genotoxic treatment.