Assessing HER2 testing quality in breast cancer: variables that influence HER2 positivity rate from a large, multicenter, observational study in Germany.

Despite >10 years of routine human epidermal growth factor receptor 2 (HER2) testing in breast cancer, testing quality is still an issue. Guidelines recommend assessing HER2 positivity rates as a quality indicator; however, the extent to which patient- or tumor-related factors influence HER2 positivity is still unknown. The present study analyzed these influences to identify pathology centers with HER2 positivity rates unexplained by patient- or tumor-related factors. This observational, prospective study monitored routine HER2 testing at 57 institutes of pathology in Germany (January 2013-August 2014). Data collected included HER2 test result, patient- and tumor-related factors, sample source, and method of sample retrieval. Factors influencing HER2 positivity rates were identified by multiple logistic regression. Individual center effects were assessed in an extended multiple logistic regression model by their statistical significance after adjusting for the combined effect of patient- or tumor-related covariates and multiple testing. Analyses included 15 332 invasive breast cancer samples. Histologic grade showed the strongest influence on HER2 positivity, followed by hormone receptor status, histologic subtype, age, and nodal status (all P<0.0001). The overall HER2 positivity rate across centers was 14.4% (range 7.1-27.3%). A statistically significant center effect on the HER2 positivity rate was identified for three centers (P<0.05), with a trend toward a center effect for a further three (P<0.2). This study, the first of its kind, highlights that assessing HER2 testing quality with HER2 positivity rates should include standardized assessment of patient- or tumor-related characteristics to identify centers with HER2 testing quality issues more effectively. As treatment options for HER2-positive breast cancer continue to evolve, identifying the right patients is key.

Genomic characterization of small cell carcinomas of the uterine cervix.

Small cell carcinoma (SCC) of the uterine cervix is a rare and aggressive form of neuroendocrine carcinoma, which resembles small cell lung cancer (SCLC) in its histology and poor survival rate. Here, we sought to define the genetic underpinning of SCCs of the uterine cervix and compare their mutational profiles with those of human papillomavirus (HPV)-positive head and neck squamous cell carcinomas, HPV-positive cervical carcinomas, and SCLCs using publicly available data. Using a combination of whole-exome and targeted massively parallel sequencing, we found that the nine uterine cervix SCCs, which were HPV18-positive (n = 8) or HPV16-positive (n = 1), harbored a low mutation burden, few copy number alterations, and other than TP53 in two cases no recurrently mutated genes. The majority of mutations were likely passenger missense mutations, and only few affected previously described cancer-related genes. Using RNA-sequencing, we identified putative viral integration sites on 18q12.3 and on 8p22 in two SCCs of the uterine cervix. The overall nonsilent mutation rate of uterine cervix SCCs was significantly lower than that of SCLCs, HPV-driven cervical adeno- and squamous cell carcinomas, or HPV-positive head and neck squamous cell carcinomas. Unlike SCLCs, which are reported to harbor almost universal TP53 and RB1 mutations and a dominant tobacco smoke-related signature 4, uterine cervix SCCs rarely harbored mutations affecting these genes (2/9, 22% TP53; 0% RB1) and displayed a dominant aging (67%) or APOBEC mutational signature (17%), akin to HPV-driven cancers, including cervical adeno- and squamous cell carcinomas and head and neck squamous cell carcinomas. Taken together, in contrast to SCLCs, which are characterized by highly recurrent TP53 and RB1 alterations, uterine cervix SCCs were positive for HPV leading to inactivation of the suppressors p53 and RB, suggesting that these SCCs are convergent phenotypes.

Case report: extracranial metastasis from gliosarcoma--the influence of immune system.

Extracranial metastasis of malignant glioma is an extremely rare event. We report on a 67-year-old patient with a primary gliosarcoma that was treated by open resection. The concomitant radio-chemotherapy which followed induced an unusually severe and early leucocytopenia. Ten months after diagnosis, the patient presented with multiple metastases in the lung and the skeletal system. The clinical, radiological and neuropathological findings are described. In addition, we discuss the possible role of a compromised immune system in the development of extracranial glioma metastasis.

Cutaneous lymphatic malformations in disappearing bone (Gorham-Stout) disease: a novel clue to the pathogenesis of a rare syndrome.

BACKGROUND: Gorham-Stout disease is an unusual, progressive syndrome of unknown etiology characterized by mono- or polyostotic osteolysis most often affecting children and young adults. The onset is insidious and the disease progresses to extensive and potentially disabling osteolysis often unresponsive to therapeutic intervention. Although bone and soft tissue lesions are the most frequent manifestations of Gorham-Stout disease, skin lesions can occur and may provide a clue to the pathogenesis of this rare syndrome. OBJECTIVE: Our aim was to describe characteristics of vascular skin lesions of this rare condition using magnetic resonance imaging and histomorphological analysis. METHODS: The case of a 36-year-old man with Gorham-Stout disease of the left leg and foot is reported. RESULTS: This case was remarkable for its prominent lymphatic vascular malformations involving the skin and soft tissues adjacent to the diseased bone-a previously undescribed abnormality, which preceded osteolysis for several years. Magnetic resonance imaging played a key role in defining the extent of disease in skin and soft tissues. LIMITATIONS: It is difficult to assess the true incidence of hemangiomatosis in the earlier reports on Gorham-Stout disease in which hemangiomatous cutaneous lesions were mentioned but not described or illustrated. CONCLUSION: A vascular process with angiomatous histological features is considered to be the pathological hallmark of Gorham-Stout disease, but the specific type of this vascular process is still under debate. Our report highlights a lymphatic malformative nature of Gorham-Stout disease, thereby contributing to a better understanding and characterization of this rare disease entity.

Methylation assay for the diagnosis of lung cancer on bronchial aspirates: a cohort study.

PURPOSE: Recent studies have detected aberrant promoter methylation of adenomatous polyposis coli promoter 1 A (APC), cyclin-dependent kinase inhibitor-2A (p16(INK4a)), retinoic acid receptor beta2, and RAS association domain family protein 1 (RASSF1A) in bronchial aspirates and suggested their use as biomarkers for lung cancer diagnostics. The purpose of this study was to validate these candidate marker genes in a retrospective cohort study. EXPERIMENTAL DESIGN: Bronchial aspirates collected from a cohort comprising 247 patients with suspected lung cancer were investigated retrospectively regarding aberrant promoter methylation using a quantitative methylation-specific real-time PCR (QMSP). RESULTS: Eighty-nine patients were diagnosed with primary lung cancer, 102 had benign lung disease, and 56 showed miscellaneous other conditions. A panel consisting of APC, p16(INK4a), and RASSF1A emerged as useful combination. This panel detected aberrant methylation in bronchial aspirates of 22 of 35 (63%) and 21 of 44 (44%) centrally and peripherally located primary lung cancers, respectively. Bronchial aspirates also showed aberrant methylation in 5 of 7 (71%) patients with a recurrent lung cancer and in 8 of 30 (27%) cases without tumor recurrence. In contrast, only 1 of 102 patients with benign lung disease displayed a (false) positive test result. Rarely, aberrant methylation was found in patients with other malignancies (3 of 16). The QMSP assay correctly confirmed lung cancer in 8 of 12 (67%) cases with an ambiguous cytology. Moreover, it disclosed 9 of 26 (35%) of peripheral tumors lacking simultaneous cytologic or histologic diagnosis of malignancy. CONCLUSIONS: Our findings suggest that the QMSP assay could be applied as a reflex test in cases of suspected lung cancer that defy a definite diagnosis by conventional methods. Thus, the assay could be a useful diagnostic adjunct especially regarding peripheral tumors.

Paclitaxel/Taxol sensitivity in human renal cell carcinoma is not determined by the p53 status.

In this study, we analyzed the role of the p53 status for paclitaxel/Taxol sensitivity in renal cell carcinomas (RCCs) of the clear cell type. Using immunohistochemistry, nuclear p53 accumulation could not be correlated to the paclitaxel/Taxol sensitivity. DNA sequencing detected a p53 gene mutation in two out of eight RCC cell lines, i.e. in exon 8 (cell line clearCa-6), and in exon 9 (cell line clearCa-5). No correlation, however, was found between the p53 status of our RCC cell lines and their paclitaxel/Taxol sensitivity as indicated by the IC50 values. However, paclitaxel-induced growth inhibition in paclitaxel-sensitive RCC cell lines was accompanied by an increase in apoptosis, irrespective of their p53 status. Although CD95 up-regulation was observed in renal cell carcinoma with wild-type p53 upon paclitaxel treatment, paclitaxel-induced apoptosis itself is triggered independently from the CD95 system. In conclusion, the p53 status cannot predict paclitaxel/Taxol sensitivity in RCC cell lines of the clear cell type.

Prognostic implications of CD95 receptor expression in clear cell renal carcinomas.

The CD95 (Apo-1/Fas) receptor-ligand system is a key regulator of apoptosis. Down-regulation of CD95 receptor and up-regulation of CD95 ligand has been reported in a variety of human tumors and is thought to confer a selective survival advantage. To explore the relevance of the CD95 system for tumor progression and prognosis in clear cell renal cell carcinomas (RCCs), we analyzed CD95 receptor and ligand expression in formalin-fixed tissue from 149 clear cell RCCs by immunohistochemistry. CD95 ligand expression could not be detected in nonneoplastic tubule epithelia and in clear cell RCCs. In contrast, CD95 receptor expression was found in the great majority of clear cell RCCs, and no down-regulation of CD95 receptor protein was evident when compared with nonneoplastic tubule epithelia. Although a significant increase (P = 0.004) of CD95 receptor expression was evident from well-differentiated (G1) to poorly differentiated (G3) RCCs, CD95 receptor expression was not correlated with tumor stage or survival of RCC patients. In conclusion, clear cell RCCs differ from other types of human cancer by their failure to down-regulate CD95 receptor expression or up-regulate CD95 ligand expression during tumor progression. These ex vivo observations suggest that down-regulation of CD95 receptor expression may not provide an additional selective growth advantage to RCC cells and thus further confirm our previous in vitro observations on a functional impairment of CD95-mediated apoptosis in RCC.

p27KIP1-expression in human renal cell cancers: implications for clinical outcome.

BACKGROUND: p27Kip1 (p27) protein is an inhibitor of cyclin-dependent kinase complexes and prevents progression of cells from the G1- to the S-phase of the cell cycle. Decreased p27 expression has been shown to be associated with aggressive tumor behavior and decreased patient survival in numerous human malignancies. The aim of this study was to evaluate p27 expression in renal cell cancer and to assess its association with stage and grade as well as its relationship to patient outcome. MATERIALS AND METHODS: One hundred and fifty-four renal cell carcinoma specimens were evaluated for p27 expression by immunohistochemical staining. Immunohistochemical findings were correlated with tumor grade, tumor stage and patient outcome. RESULTS: A progressive loss of nuclear p27 expression was observed with increasing tumor grade. In poorly-differentiated tumors, p27 expression was significantly lower compared to well- and moderately-differentiated tumors (p = 0.025). p27 expression tended to decrease with increasing tumor stage, but the correlation was not statistically significant (p = 0.068). CONCLUSION: The present study suggests that renal cell carcinomas showed increased aggressiveness with loss of p27 expression. A longer follow-up period will demonstrate whether this cell cycle regulator will provide additional prognostic information in patients with renal cell carcinoma.

Equivocal cytology in lung cancer diagnosis: improvement of diagnostic accuracy using adjuvant multicolor FISH, DNA-image cytometry, and quantitative promoter hypermethylation analysis.

BACKGROUND: Sometimes, cytological lung cancer diagnosis is challenging because equivocal diagnoses are common. To enhance diagnostic accuracy, fluorescent in situ hybridization (FISH), DNA-image cytometry, and quantitative promoter hypermethylation analysis have been proposed as adjuncts. METHODS: Bronchial washings and/or brushings or transbronchial fine-needle aspiration biopsies were prospectively collected from patients who were clinically suspected of having lung carcinoma. After routine cytological diagnosis, 70 consecutive specimens, each cytologically diagnosed as negative, equivocal, or positive for cancer cells, were investigated with adjuvant methods. Suspicious areas on the smears were restained with the LAVysion multicolor FISH probe set (Abbott Molecular, Des Plaines, Illinois) or according to the Feulgen Staining Method for DNA-image cytometry analysis. DNA was extracted from residual liquid material, and frequencies of aberrant methylation of APC, p16(INK4A) , and RASSF1A gene promoters were determined with quantitative methylation-specific polymerase chain reaction (QMSP) after bisulfite conversion. Clinical and histological follow-up according to a reference standard, defined in advance, were available for 198 of 210 patients. RESULTS: In the whole cohort, cytology, FISH, DNA-image cytometry, and QMSP achieved sensitivities of 83.7%, 78%, 79%, and 49.6%, respectively (specificities of 69.8%, 98.2%, 98.2%, and 98.4%, respectively). Subsequent to cytologically equivocal diagnoses, FISH, DNA-image cytometry, and QMSP definitely identified malignancy in 79%, 83%, and 49%, respectively. With QMSP, 4 of 22 cancer patients with cytologically negative diagnoses were correctly identified. CONCLUSIONS: Thus, adjuvant FISH or DNA-image cytometry in cytologically equivocal diagnoses improves diagnostic accuracy at comparable rates. Adjuvant QMSP in cytologically negative cases with persistent suspicion of lung cancer would enhance sensitivity.

Penile metastasis secondary to follicular thyroid carcinoma.

Penile metastases are rare and are considered to reflect end-stage malignant disease. The first case of a follicular thyroid carcinoma metastasizing to the penis is described. Local tumor control and probably enhanced survival was achieved by extended surgery of a previous pelvic recurrence and the penile metastasis and this procedure may be justified in selected cases.

Decrease of DNA methyltransferase 1 expression relative to cell proliferation in transitional cell carcinoma.

In many common cancers such as transitional cell carcinoma (TCC), specific genes are hypermethylated, whereas overall DNA methylation is diminished. Genome-wide DNA hypomethylation mostly affects repetitive sequences such as LINE-1 retrotransposons. Methylation of these sequences depends on adequate expression of DNA methyltransferase I (DNMT1) during DNA replication. Therefore, DNMT1 expression relative to proliferation was investigated in TCC cell lines and tissue as well as in renal carcinoma (RCC) cell lines, which also display hypomethylation, as indicated by decreased LINE-1 methylation. Cultured normal uroepithelial cells or normal bladder tissue served as controls. In all tumor cell lines, DNMT1 mRNA as well as protein was decreased relative to the DNA replication factor PCNA, and DNA hypomethylation was present. However, the extents of hypomethylation and DNMT1 downregulation did not correlate. Reporter gene assays showed that the differences in DNMT1 expression between normal and tumor cells were not established at the level of DNMT1 promoter regulation. Diminished DNMT1:PCNA mRNA ratios were also found in 28/45 TCC tissues but did not correlate with the extent of DNA hypomethylation. In addition, expression of the presumed de novo methyltransferases DNMT3A and DNMT3B mRNAs was investigated. DNMT3B overexpression was observed in about half of all high-stage TCC (DNMT3B vs. tumor stage, chi(2): p = 0.03), whereas overexpression of DNMT3A was rarer and less pronounced. Expression of DNMT3A and DNMT3B in most RCC lines was higher than in TCC lines. Our data indicate that DNMT1 expression does not increase adequately with cell proliferation in bladder cancer. This relative downregulation probably contributes to hypomethylation of repetitive DNA but does not determine its extent alone.