Tumor proteomics by multivariate analysis on individual pathway data for characterization of vulvar cancer phenotypes.

Vulvar squamous cell carcinoma (VSCC) is the fourth most common gynecological cancer. Based on etiology VSCC is divided into two subtypes; one related to high-risk human papilloma virus (HPV) and one HPV negative. The two subtypes are proposed to develop via separate intracellular signaling pathways. We investigated a suggested link between HPV infection and relapse risk in VSCC through in-depth protein profiling of 14 VSCC tumor specimens. The tumor proteomes were analyzed by liquid-chromatography tandem mass spectrometry. Relative protein quantification was performed by 8-plex isobaric tags for relative and absolute quantification. Labeled peptides were fractionated by high-resolution isoelectric focusing prior to liquid-chromatography tandem mass spectrometry to reduce sample complexity. In total, 1579 proteins were regarded as accurately quantified and analyzed further. For classification of clinical groups, data analysis was performed by comparing protein level differences between tumors defined by HPV and/or relapse status. Further, we performed a biological analysis on individual tumor proteomes by matching data to known biological pathways. We here present a novel analysis approach that combines pathway alteration data on individual tumor level with multivariate statistics for HPV and relapse status comparisons. Four proteins (signal transducer and activator of transcription-1, myxovirus resistance protein 1, proteasome subunit alpha type-5 and legumain) identified as main classifiers of relapse status were validated by immunohistochemistry (IHC). Two of the proteins are interferon-regulated and on mRNA level known to be repressed by HPV. By both liquid-chromatography tandem mass spectrometry and immunohistochemistry data we could single out a subgroup of HPV negative/relapse-associated tumors. The pathway level data analysis confirmed three of the proteins, and further identified the ubiquitin-proteasome pathway as altered in the high risk subgroup. We show that pathway fingerprinting with resolution on individual tumor level adds biological information that strengthens a generalized protein analysis.

Presence of human papillomavirus (HPV) in vulvar squamous cell carcinoma (VSCC) and sentinel node.

OBJECTIVE: To investigate the presence of HPV in VSCC and sentinel nodes (SN) in patients in Sweden and the possible influence of HPV on prognosis. PATIENTS AND MATERIALS: Primary tumors from 75 VSCC patients undergoing the SN procedure and SNs from 69 patients were tested for HPV DNA. Analyses were performed by PCR using general (GP5+/6+ and CPI/IIG) type-specific primers, and sequencing in paraffin-embedded VSCC and SN. RESULTS: HPV was detected in 23/75 (31%) of the tumors and in 10/23 (43%) of the SNs in patients with HPV-positive tumors and in one SN of a patient with an HPV-negative tumor. Patients with HPV-positive VSCC were younger at diagnosis (p<0.001) and had better survival (p=0.030), adjusted for age and lesion size, than those with HPV-negative tumors. In patients with HPV-positive tumors, SNs with metastases were more frequently HPV positive (5/5) than those without metastases (5/18) (p=0.007). CONCLUSION: The rate of 31% HPV-positive VSCC in Sweden is similar to other reports. As far as we know, HPV in SN in VSCC never been investigated previously. The differences in age, tumor size, prevalence of HPV in SN and survival of patients with HPV-positive and negative VSCC support the assumption that VSCC develops through two different pathways, with better survival for patients with HPV-positive tumors. Presence of HPV DNA in SN was related to metastatic disease but did not affect survival in this study.

Types of human papillomavirus revealed in cervical adenocarcinomas after DNA sequencing.

In a collection of 173 cervical adenocarcinomas, the prevalence of HPV in relation to the age of women and the distribution of the various oncogenic types of HPV were evaluated. The tumour material was analysed by PCR of the HPV L1 gene followed by direct DNA sequencing and/or the polymerase chain reaction and single-strand conformation polymorphism (PCR-SSCP) technique for the identification and typing of HPV. In 68% (117/173) of the adenocarcinomas, HPV was present. A significant correlation was observed between the prevalence of HPV and age: in women younger than 40 years, HPV was present in 88%, whereas in women 60 years and older, it was found in only 39% (p<0.001). Among the HPV-positive tumours, type 18 predominated (52%) followed by type 16 (35%) and type 45 (7.7%) while other oncogenic types of HPV (31 and 59) were rarely found. HPV 16 was relatively more frequent in older women but this observation was not significant (p=0.06). HPV-typing by PCR and direct DNA sequence analysis is more specific than analysis by PCR-SSCP, especially among the less frequently occurring types of HPV. Our results further show that the prevalence of HPV in women with cervical adenocarcinomas is age-related and that the most frequently occurring types of HPV are 18, followed by 16 and 45. We have concluded that the oncogenic role of HPV in cervical squamous carcinomas and adenocarcinomas is, in some respects, discrepant.

Evaluation of preoperative lymphoscintigraphy and sentinel node procedure in vulvar cancer.

OBJECTIVE: To assess the value of preoperative lymphoscintigraphy, and to evaluate the validity and feasibility of the sentinel node (SN) procedure in vulvar carcinoma. STUDY DESIGN: Retrospective clinical and histopathological review of 77 patients with invasive squamous cell carcinoma in vulva who were treated at Karolinska University Hospital Stockholm, Sweden, from 2000 to 2007. The patients underwent SN mapping preoperatively with radioactive tracer and blue dye (n=60) or only blue dye (n=17). The SN was removed separately followed by complete inguinofemoral lymphadenectomy. RESULTS: The relation between SNs detected on the scintigram and those found during surgery showed good agreement using weighted kappa. The detection rate of SN was 98% for radioisotope plus blue dye, and 94% for blue dye alone. Two cases of false negative SN (false negative rate 2.7%) were found, both with large midline tumors. CONCLUSION: Preoperative scintigram is a valuable help to identify and localize the SNs and gives the best estimate of the accurate number but cannot determine if unilateral or bilateral groins should be explored in cases of midline tumors. Our results are in favor of using radioisotope and blue dye to identify the SNs. This study support previous reports that the method is not recommended for tumors larger than 40 mm to optimize detection of SN and minimize the false negative detection rate.

Erythropoietin and intravenous iron therapy in postpartum anaemia.

BACKGROUND: We assessed whether recombinant human erythropoietin (rhEPO) enhances a rise in haemoglobin concentration in postpartum anaemia compared to intravenous iron alone. DESIGN: Some 60 patients with haemoglobin values

Adenocarcinoma of the uterine cervix: the presence of human papillomavirus and the method of detection.

BACKGROUND: Effective screening programs have contributed to a decrease in the incidence of cervical squamous cell carcinomas but have had a limited sensitivity in the detection of adenocarcinoma precursor lesions. The aim of our study was to analyze cervical adenocarcinoma in greater detail: symptoms preceding the detection, the method of detection and the prevalence of human papillomavirus (HPV) with respect to age at diagnosis. MATERIAL AND METHODS: Clinical data were abstracted from the medical records of 82 women with pure invasive cervical adenocarcinomas. As diagnostic tools we used polymerase chain reaction (PCR)-based single-strand conformation polymorphism (SSCP) and/or direct DNA sequencing for HPV detection. RESULTS: Age at diagnosis predicting factors were HPV status, positive lymph nodes, histology and stage. HPV-negativity, lymph node metastases, advanced stage and poor differentiation were all associated with a high diagnostic age. In the multivariate analysis only HPV status was shown to have an independent impact on age at diagnosis, while stage showed only borderline significance. Twenty-three percent of the cancers were detected by screening and the remaining were due to different symptoms. Among the women considered, 93% had a normal Papanicolaou (Pap) smear 3 years before diagnosis and 60% within 1 year. There was no significant correlation between smoking, oral contraceptives and HPV-positivity. CONCLUSIONS: The absence of HPV was significantly associated with a high age at diagnosis. Pap screening had a limited effect in detecting adenocarcinoma at an early stage.