Molecular classification of non-muscle-invasive bladder cancer (pTa low-grade, pT1 low-grade, and pT1 high-grade subgroups) using methylation of tumor-suppressor genes.

The role of epigenetics in distinguishing pathological and clinical subgroups in bladder cancer is not fully characterized. We evaluated whether methylation of tumor-suppressor genes (TSGs) would classify non-muscle-invasive (NMI) bladder cancer subgroups and predict outcome. A retrospective design included the following paraffin-embedded primary NMI tumor types (n = 251): pTa low grade (LG) (n = 79), pT1LG (n = 81), and pT1 high grade (HG) (n = 91). Methylation of 25 TSGs was measured using methylation-specific, multiplex, ligation-dependent probe amplification. The TSGs most frequently methylated in the overall series were STK11 (96.8%), MGMT2 (64.5%), RARB (63.0%), and GATA5 (63.0%). TSG methylation correlated to clinicopathological variables in each subgroup and in the overall NMI series. Methylation of RARB, CD44, PAX5A, GSTP1, IGSF4 (CADM1), PYCARD, CDH13, TP53, and GATA5 classified pTa versus pT1 tumors whereas RARB, CD44, GSTP1, IGSF4, CHFR, PYCARD, TP53, STK11, and GATA5 distinguished LG versus HG tumors. Multivariate analyses indicated that PAX5A, WT1, and BRCA1 methylation independently predicted recurrence in pTaLG, PAX6, ATM, CHFR, and RB1 in pT1LG disease; PYCARD, in pT1HG disease; and PAX5A and RB1, in the overall series. Methylation of TSGs provided a molecular classification of NMI disease according to clinicopathological factors. Furthermore, TSG methylation predicted recurrence in NMI subgroups.

Analysis of MYB oncogene in transformed adenoid cystic carcinomas reveals distinct pathways of tumor progression.

Adenoid cystic carcinomas can occasionally undergo dedifferentiation, a phenomenon also referred to as high-grade transformation. However, cases of adenoid cystic carcinomas have been described showing transformation to adenocarcinomas that are not poorly differentiated, indicating that high-grade transformation may not necessarily reflect a more advanced stage of tumor progression, but rather a transformation to another histological form, which may encompass a wide spectrum of carcinomas in terms of aggressiveness. The aim of this study was to gain more insight in the biology of this pathological phenomenon by means of genetic profiling of both histological components. Using microarray comparative genomic hybridization, we compared the genome-wide DNA copy-number changes of the conventional and transformed area of eight adenoid cystic carcinomas with high-grade transformation, comprising four with transformation into moderately differentiated adenocarcinomas and four into poorly differentiated carcinomas. In general, the poorly differentiated carcinoma cases showed a higher total number of copy-number changes than the moderately differentiated adenocarcinoma cases, and this correlated with a worse clinical course. Special attention was given to chromosomal translocation and protein expression of MYB, recently being considered to be an early and major oncogenic event in adenoid cystic carcinomas. Our data showed that the process of high-grade transformation is not always accompanied by an accumulation of genetic alterations; both conventional and transformed components harbored unique genetic alterations, which indicate a parallel progression. Our data further demonstrated that the MYB/NFIB translocation is not necessarily an early event or fundamental for the progression to adenoid cystic carcinoma with high-grade transformation.

Polyamine-modulated factor-1 methylation predicts Bacillus Calmette-Guérin response in patients with high-grade non-muscle-invasive bladder carcinoma.

BACKGROUND: Bacillus Calmette-Guérin (BCG) is a standard treatment to reduce tumor recurrence and delay progression of high-risk non-muscle-invasive (NMI) bladder tumors. However, it is not clear yet which patients are more likely to respond to BCG. OBJECTIVE: The aim was to evaluate the role of polyamine-modulated factor-1 (PMF-1) methylation in predicting clinical outcome of T1 high-grade (T1HG) bladder tumors treated with BCG. DESIGN, SETTING, AND PARTICIPANTS: In a retrospective design, PMF-1 methylation was analyzed on tumor specimens belonging to 108 patients with T1HG NMI bladder cancer undergoing BCG treatment. Median follow-up was 77 mo (range: 5-235 mo). OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: PMF-1 methylation was assessed by methylation-specific polymerase chain reactions. Recurrence, progression into muscle-invasive tumors, and disease-specific survival rates were analyzed using competing risks regression analysis. RESULTS AND LIMITATIONS: Among the 108 patients analyzed, 35 had recurring disease (32.4%), 21 progressed (19.4%), and 16 died of disease (14.8%); 71.3% of tumors had PMF-1 methylation. Univariate analyses using cumulative incidence curves revealed that an unmethylated PMF-1 was significantly associated with increased recurrence (p=0.026), progression (p=0.01), and shorter disease-specific survival (log-rank, p=0.03). Multivariate analyses indicated that among sex, age, focality, tumor size, and concomitant carcinoma in situ, only PMF-1 methylation provided significant hazard ratios (HRs) for recurrence of (HR: 2.032; p=0.042), and progression (HR: 2.910; p=0.020). Limitations of the study include its retrospective design, lymphovascular invasion status not available, short maintenance BCG, and that a second transurethral resection was not performed. CONCLUSIONS: Epigenetic analyses revealed that the methylation status of PMF-1 was associated with the clinical outcome of patients with T1HG tumors undergoing BCG treatment. An unmethylated PMF-1 correlated to recurrence and progression in T1HG disease using univariate and multivariate analyses. Thus, assessing the methylation status of PMF-1 may serve to distinguish patients responding to BCG from those who may require more aggressive therapeutic approaches.

Genetic profile of adenoid cystic carcinomas (ACC) with high-grade transformation versus solid type.

BACKGROUND: ACC can occasionally undergo dedifferentiation also referred to as high-grade transformation (ACC-HGT). However, ACC-HGT can also undergo transformation to adenocarcinomas which are not poorly differentiated. ACC-HGT is generally considered to be an aggressive variant of ACC, even more than solid ACC. This study was aimed to describe the genetic changes of ACC-HGT in relation to clinico-pathological features, and to compare results to solid ACC. METHODS: Genome wide DNA copy number changes were analyzed by microarray CGH in ACC-HGT, four with transformation into moderately differentiated adenocarcinoma (MDA) and two into poorly differentiated carcinoma (PDC), and five solid ACC. In addition, Ki67 index and p53 immunopositivity was assessed. RESULTS: ACC-HGT carried fewer copy number changes compared to solid ACC. Two ACC-HGT cases harboured a breakpoint at 6q23, near the cMYB oncogene. The complexity of the genomic profile concurred with the clinical course of the patient. Among the ACC-HGT, p53 positivity significantly increased from the conventional to the transformed (both MDA and PDC) component. CONCLUSION: ACC-HGT may not necessarily reflect a more advanced stage of tumor progression, but rather a transformation to another histological form in which the poorly differentiated forms (PDC) presents a genetic complexity similar to the solid ACC.

Genetic profile of adenoid cystic carcinomas (ACC) with high-grade transformation versus solid type.

BACKGROUND: ACC can occasionally undergo dedifferentiation also referred to as high-grade transformation (ACC-HGT). However, ACC-HGT can also undergo transformation to adenocarcinomas which are not poorly differentiated. ACC-HGT is generally considered to be an aggressive variant of ACC, even more than solid ACC. This study was aimed to describe the genetic changes of ACC-HGT in relation to clinico-pathological features and to compare results to solid ACC. METHODS: genome-wide DNA copy number changes were analyzed by microarray CGH in ACC-HGT, 4 with transformation into moderately differentiated adenocarcinoma (MDA) and two into poorly differentiated carcinoma (PDC), 5 solid ACC. In addition, Ki-67 index and p53 immunopositivity was assessed. RESULTS: ACC-HGT carried fewer copy number changes compared to solid ACC. Two ACC-HGT cases harboured a breakpoint at 6q23, near the cMYB oncogene. The complexity of the genomic profile concurred with the clinical course of the patient. Among the ACC-HGT, p53 positivity significantly increased from the conventional to the transformed (both MDA and PDC) component. CONCLUSION: ACC-HGT may not necessarily reflect a more advanced stage of tumor progression, but rather a transformation to another histological form in which the poorly differentiated forms (PDC) presents a genetic complexity similar to the solid ACC.

Myopodin methylation is associated with clinical outcome in patients with T1G3 bladder cancer.

PURPOSE: Bacillus Calmette-Guerin is standard treatment to decrease tumor recurrence and delay progression of high risk, nonmuscle invasive bladder tumors. However, it is not yet clear which T1G3 cases are more prone to more aggressive clinical behavior or susceptible to respond to bacillus Calmette-Guerin. We evaluated the role of myopodin methylation as a clinical outcome prognosticator and predictive biomarker for the bacillus Calmette-Guerin response in patients with T1G3 bladder tumors. MATERIALS AND METHODS: We analyzed the methylation status of myopodin in tumor specimens from 170 patients with T1G3 bladder cancer, including a subset of 108 who underwent bacillus Calmette-Guerin treatment. Myopodin methylation was assessed by methylation specific polymerase chain reactions. Recurrence, progression to muscle invasive tumors and disease specific overall survival were analyzed using competing risks regression analysis. RESULTS: Of the 170 cases analyzed 72 recurred (42.4%) and 36 progressed (21.2%). A total of 24 patients (14.1%) died of the disease. Univariate and multivariate survival analysis revealed that myopodin methylation was significantly associated with an increased recurrence rate (p = 0.004), progression (p = 0.002) and shorter disease specific overall survival (p = 0.020). In a subset treated with bacillus Calmette-Guerin myopodin methylation was also related to an increased recurrence rate (p = 0.011), progression (p = 0.030) and shorter disease specific overall survival (p = 0.028). CONCLUSIONS: Epigenetic analysis revealed that myopodin methylation was associated with tumor aggressiveness and clinical outcome in patients with T1G3 disease. Myopodin methylation distinguished patients responding to bacillus Calmette-Guerin from those who may require a more aggressive therapeutic approach.

[Nasosinusal adenocarcinoma: molecular and genetic analysis by MLPA]. / Análisis genético molecular con MLPA en los adenocarcinomas nasosinusales.

INTRODUCTION AND AIM: Intestinal-type sinonasal adenocarcinomas (ITACs) are rare epithelial tumours, primarily originating in the nasal cavities and paranasal sinuses and characterized by glandular structures. The aims of this study are: to determine the genetic alterations in ITACs by MLPA (Multiplex ligation-dependent probe amplification) and to correlate the findings to the clinical behavior and follow-up information of the patients. MATERIAL AND METHOD: We performed a longitudinal prospective study on 20 patients with ITAC, seen in our department between 1998 and 2004. DNA was extracted from primary tumor samples and analyzed by MLPA. RESULTS: The T stage of our series was T2: 4 (20 %), T3: 6 (30 %), T4a: 3 (15 %) and T4b: 7 (35 %). All cases initially were N0 and M0. Seventeen patients (85 %) had professional exposure to wood dust. All patients underwent surgical intervention and 70 % received complementary radiotherapy. Overall 5 and 10 year survival was 42 % and 22 %, respectively. Gains were found most frequently for PTP4A3 and PDCD8 (65 %), TNRFSF7 (50 %), RECQL4 and LMO2 (45 %), and losses for BCL2 (70 %), IL13 (55 %), ABCB1 and RB1 (50 %), PIK3CA and CDH1 (45 %). CONCLUSIONS: Losses of F3, MIF, and BRCA1 significantly correlated with the posterior development of metastases and with worse survival. Also gains of PIK3CA, UTY, and RELA correlated with poor clinical outcome. Losses of BRCA1 and F3 were significant in multivariate analysis.

Análisis genético molecular con MLPA en los adenocarcinomas nasosinusales / Nasosinusal adenocarcinoma: molecular and genetic analysis by MLPA

Introducción y objetivo: Los adenocarcinomas nasosinusales (ACNS) son tumores epiteliales raros, primarios de las fosas nasales y los senos paranasales, que se caracterizan por presentar estructuras glandulares. Los objetivos de este trabajo son: determinar las alteraciones génicas en los ACNS mediante MLPA (multiplex ligation-dependent probe amplification) y establecer la relación entre los cambios génicos del tumor y el comportamiento clínico-evolutivo de los pacientes. Material y método: Realizamos un estudio longitudinal prospectivo a 20 pacientes con ACNS controlados en nuestro servicio entre 1998 y 2004. A las muestras tumorales se les extrajo el ADN para realizar la MLPA. Resultados: La categoría T de los pacientes fue para T2: 4 (20 %), T3: 6 (30 %), T4a: 3 (15 %) y T4b: 7 (35 %). Todos eran inicialmente N0 y M0. El 85 % había estado expuesto, en el medio laboral, al polvo de madera. El tratamiento indicado fue quirúrgico (100 %) con radioterapia complementaria (70 %). La supervivencia global a los 5 y 10 años fue del 42 y el 22 %, respectivamente. Las ganancias génicas más frecuentes fueron: PTP4A3 y PDCD8 (65 %), TNRFSF7 (50 %), RECQL4 y LMO2 (45 %); las pérdidas: BCL2 (70 %), IL13 (55 %), ABCB1 y RB1 (50 %), PIK3CA y CDH1 (45 %). Conclusiones: La aparición de metástasis se correlacionó de forma significativa con pérdidas en F3, MIF y BRCA1. La peor supervivencia con pérdidas en F3, BCRA1 y MIF y ganancias en PIK3CA, UTY y RELA. En el análisis multivariable alcanzaron significación estadística las pérdidas en BRCA1 y F3

Introduction and aim: Intestinal-type sinonasal adenocarcinomas (ITACs) are rare epithelial tumours, primarily originating in the nasal cavities and paranasal sinuses and characterized by glandular structures. The aims of this study are: to determine the genetic alterations in ITACs by MLPA (Multiplex ligation-dependent probe amplification) and to correlate the findings to the clinical behavior and follow-up information of the patients. Material and method: We performed a longitudinal prospective study on 20 patients with ITAC, seen in our department between 1998 and 2004. DNA was extracted from primary tumor samples and analyzed by MLPA. Results: The T stage of our series was T2: 4 (20 %), T3: 6 (30 %), T4a: 3 (15 %) and T4b: 7 (35 %). All cases initially were N0 and M0. Seventeen patients (85 %) had professional exposure to wood dust. All patients underwent surgical intervention and 70 % received complementary radiotherapy. Overall 5 and 10 year survival was 42 % and 22 %, respectively. Gains were found most frequently for PTP4A3 and PDCD8 (65 %), TNRFSF7 (50 %), RECQL4 and LMO2 (45 %), and losses for BCL2 (70 %), IL13 (55 %), ABCB1 and RB1 (50 %), PIK3CA and CDH1 (45 %). Conclusions: Losses of F3, MIF, and BRCA1 significantly correlated with the posterior development of metastases and with worse survival. Also gains of PIK3CA, UTY, and RELA correlated with poor clinical outcome. Losses of BRCA1 and F3 were significant in multivariate analysis

Prognostic value of micrometastases in esophageal and colorectal carcinoma (a clinical experience).

BACKGROUND/AIMS: The term "micrometastases" has been confused in many aspects. While the influence of lymph node metastases in esophageal and colorectal cancer is well known, the presence and importance of micrometastases is under debate. We investigated micro lymph node invasion in two different kinds of digestive tumors with very high mortality, and identified its possible repercussion on patient survival. METHODOLOGY: Lymph nodes of two groups of patients N0 on routine histopathology after radical resection (R0): 21 with esophageal carcinoma (Group I), and 21 with colorectal carcinoma (Group II), were studied by immunohistochemistry using monoclonal antibodies directed against cytokeratins of wide spectrum. The results were classified as positive or negative and compared with patient survival. RESULTS: Five of twenty-one (5/21) patients in group I and eight of twenty-one (8/21) in group II were positive for micrometastases. Median survival time in the positive esophageal group was 9.5 months vs. 68 in the negative one (p=0.16). Median survival time in the positive subset colorectal group was 54.5 months vs. 76.8 in the negative subgroup (p=0.5). Our results did not show statistical differences in survival time between patients positive or negative for micrometastases; however it is evident, especially in the esophageal cancer group, that there is a negative tendency of positive micrometastases on survival time. CONCLUSIONS: The presence of micrometastases in lymph nodes of patients N0 after conventional histopathology is frequent. Our preliminary results did not allow definitive conclusions but we may suppose its negative influence on patient survival.

Schwannoma of the clitoris.

Schwannoma is a tumor of the peripheral nerve sheath, which rarely affects the female genitalia. These tumors are often associated with von Recklinghuasen neurofibromatosis; however, they have also been reported in patients without that disease, and its most common location is the posterior mediastinum, the retroperitoneum, the head and neck, and the extremities. Simple surgical excision and follow up is the most convenient treatment (1).