Oral Human Papillomavirus in Women With High-Grade Cervical Intraepithelial Neoplasia.

OBJECTIVE: This study was designed to investigate the co-prevalence of cervical and oropharyngeal human papillomavirus (HPV) infection in patients with HPV-related high-grade disease of the uterine cervix (high-grade squamous intraepithelial lesion [HSIL]). MATERIALS AND METHODS: In a prospective cohort study, women with abnormal cervical cytology admitted to our colposcopy units received HPV testing of the uterine cervix and the oropharynx via smear. From a subset of patients, oral lavage was collected to compare detection rates of HPV DNA between lavage and swab. Patients with confirmed high-risk HPV (HR-HPV)-positive HSIL of the cervix were further investigated. Sexual behavior and lifestyle factors were documented with a standardized questionnaire. RESULTS: Two hundred thirty-five women were included in the study. Of the 235 women, 135 (57.5%) were cervically HR-HPV positive with histologically confirmed high-grade cervical intraepithelial lesion (median [range] age = 30 [21-45] years). Of these, only 6 (4.4%) also had a positive oral specimen. In 3 (50%) of the 6 cases, the same HPV type was detected in oral and cervical samples (HPV 16, 35, and 45). Oral HPV detection was not higher when combining swab and lavage compared with swab alone. A relation between sexual behavior and oral HPV detection could not be demonstrated. CONCLUSIONS: Oral HPV prevalence in women with cervical HPV infection and HSIL is low. Simultaneous testing of oropharyngeal and cervical HPV infection does not seem promising as future screening strategy.

The Risk of Contralateral Non-sentinel Metastasis in Patients with Primary Vulvar Cancer and Unilaterally Positive Sentinel Node.

BACKGROUND: In patients with primary vulvar cancer and bilateral sentinel lymph node (SLN) biopsy, bilateral complete inguino-femoral lymphadenectomy (LAE) is recommended, even in cases with only unilaterally positive SLN by most guidelines. The risk of contralateral non-SLN metastasis is unclear. METHODS: All patients with primary vulvar cancer receiving an SLN dissection with radioactive tracer ± blue dye at the University Medical Center Hamburg-Eppendorf between 2001 and 2013 were retrospectively evaluated. Median follow-up was 33 months. RESULTS: A total of 140 patients were included; 124 with bilateral and 16 with unilateral SLN dissection. A median number of two SLNs (range 1-7) per groin were dissected. Overall, 53 (53/140, 37.9 %) patients received a complete inguino-femoral LAE, 41 of whom (77.4 %) had previously presented with a positive SLN (33 unilaterally, 8 bilaterally). Of the 33 patients with unilaterally positive SLN, 28 (84.9 %) underwent complete bilateral inguino-femoral LAE despite a contralateral negative SLN. Of these patients, none presented a contralateral non-SLN metastasis (0/28, 0 %) in full dissection; however, one developed groin recurrence in the initially SLN-negative, fully dissected groin after 19 months (1/28, 3.6 %). CONCLUSION: In case of bilateral SLN biopsy for clinically node-negative disease and only unilaterally positive SLN, the risk for contralateral non-SLN metastases appears to be low. These data support the omission of contralateral LAE to reduce surgical morbidity.

DNA-methylation of the homeodomain transcription factor PITX2 reliably predicts risk of distant disease recurrence in tamoxifen-treated, node-negative breast cancer patients--Technical and clinical validation in a multi-centre setting in collaboration with the European Organisation for Research and Treatment of Cancer (EORTC) PathoBiology group.

Our aim was to identify and validate DNA-methylation markers associated with very good outcome in node negative, hormone receptor positive breast cancer patients after adjuvant endocrine therapy which might allow identifying patients who could be spared the burden of adjuvant chemotherapy. Using a methylation microarray, we analysed 117 candidate genes in hormone receptor-positive tumours from 109 breast cancer patients treated by adjuvant tamoxifen. Results were validated in an independent cohort (n=236, 5 centres). Independent methodological validation was achieved by a real-time polymerase chain reaction (PCR)-based technique. DNA methylation of PITX2 showed the strongest correlation with distant recurrence. Its impact on patient outcome was validated in the independent cohort: 86% of patients with low PITX2 methylation were metastasis-free after 10 years, compared to 69% with elevated PITX2 methylation. Moreover, PITX2 methylation added significant independent information to established clinical factors. All clinical and technical findings were confirmed by quantitative DNA-methylation PCR. These results provide strong evidence that DNA-methylation analysis allows clinically relevant risk assessment in tamoxifen-treated primary breast cancer. Based on PITX2 methylation, about half of hormone receptor-positive, node-negative breast cancer patients receiving adjuvant tamoxifen monotherapy can be considered low-risk regarding development of distant recurrences and may thus be spared adjuvant chemotherapy. In addition, these low-risk postmenopausal patients seem to respond sufficiently well to tamoxifen so that they may not require up-front aromatase inhibitor therapy.

Short description of an alternative simplified method for screening recombinant clones within the "AdEasy-System" by Duplex-PCR.

BACKGROUND: Recombinant adenoviral vectors are highly efficient for in vitro and in vivo gene delivery. They can easily be produced in large numbers, transduce a wide variety of cell types and generate high levels of transgene expression. The AdEasy system is a widely used system for generating recombinant adenoviral vectors, which are created with a minimum of enzymatic manipulations and by employing homologous recombination in E. coli. In this paper we describe an alternative simplified method for screening recombinant DNA within the AdEasy system. This Duplex-PCR-method is independent of the transgene or insert and can be used for the complete AdEasy system. It is characterized by a simple standard protocol and the results can be obtained within a few hours. The PCR is run with two different primer sets. The primers KanaFor and KanaRev hybridizise with the Kanamycin resistence gene and AdFor and AdRev detect the adenoviral backbone. In case of recombinant clones, two diagnostic fragments with a size of 384 bp and 768 bp are generated. RESULTS: The practicability of this method was verified with three different transgenes: Cytosin Deaminase (AdCD), p53 (Adp53) and Granulocyte Macrophage Colony Stimulating Factor (AdGM-CSF). Recombinant clones are indicated by two diagnostic fragments and are then suitable for further processing. CONCLUSION: In summary, the presented protocol allows fast detection of recombinants with an easy technique by minimizing the amount of necessary steps for generating a recombinant adenovirus. This method is time sparing and cost-effective.