Xavier Puig

Clinicopathological and molecular characterization of colorectal micropapillary carcinoma.

Invasive micropapillary carcinoma is associated with frequent lymph node metastasis and adverse clinical outcome. Initially described as a variant of breast and ovarian carcinoma, it has subsequently been found in other organs, most recently the colon. Reports of colorectal micropapillary carcinoma to date are limited in number, and their molecular profile has not been established. The aims of the present study were to analyze their clinicopathological features and molecular profile, and compare them with those of conventional adenocarcinoma. Clinicopathological features of a cohort of 379 patients with primary colorectal cancer were retrospectively reviewed for the presence of the pattern characteristic of micropapillary carcinoma. We also assessed the expression of KRT7, KRT20, CEACAM5, MUC1 (EMA, clone E29), MUC1 (clone MA695), MLH1, MSH2, MSH6 and TP53 by immunohistochemistry. Genetic assessments of microsatellite instability, chromosomes 17p and 18q, and mutations in TP53, BRAF and KRAS were performed using DNA extracted from formalin-fixed, paraffin-embedded sections. In all, 60 of the reviewed cases (16%) had a micropapillary component that ranged from 5 to 95% of the tumor, characterized by a higher frequency of an infiltrative pattern, lymphovascular and perineural invasion, a higher depth of invasion and more positive lymph nodes than conventional adenocarcinoma. Immunohistochemistry for MUC1 (clone MA695) and MUC1 (EMA, clone E29) enhanced the characteristic inside–out staining pattern of the micropapillary carcinoma component, whereas the rest of the tumor showed luminal staining patterns. KRT7 expression was slightly increased in micropapillary carcinoma, but did not reach significance (17–3%, P=0.1967). The molecular parameters showed a higher frequency of TP53 alterations and a low incidence of microsatellite instability and RER phenotype (loss of mismatch repair protein) in micropapillary carcinoma. With regard to the histological parameters, micropapillary carcinoma appears to be more aggressive than conventional colorectal adenocarcinoma. The molecular profile supports the hypothesis that micropapillary carcinoma carcinogenesis develops through the classical chromosomal instability pathway.

Genetic and immunophenotype analyses of TP53 in bladder cancer: TP53 alterations are associated with tumor progression.

Altered p53 status is a frequent event in bladder cancer and reported to have prognostic significance. We studied the TP53 gene and its product in 76 patients affected with urinary bladder carcinomas by immunohistochemistry (mAb DO-7), polymerase chain reaction single-strand conformational polymorphism (exons 4-8) followed by direct sequencing of shifted bands, and loss of heterozygosity in 17p (p53CA). H-RAS mutations were also studied. The receiver operating characteristic curve and the logistic-regression analysis were used to evaluate the validity of immunohistochemistry in predicting TP53 mutations. A p53-positive nuclear phenotype was defined by a cutoff of 20% tumor cells being immunoreactive and was found in 23 cases, while TP53 mutations were detected in 22 cases, four of them with a negative p53 phenotype. TP53 deletions were identified in 23 cases. No H-RAS gene mutations were observed. There was a significant association between phenotype and genotype results. Moreover, a significant association was observed between p53 status and tumor stage and grade, being alterations more common in high-stage and high-grade tumors (both χ2 test; P < .01). Deletion of 17p significantly correlated with tumor stage (P < .01) and grade (P = .01), allelic losses being more common in advanced disease. Data from these studies suggest that genetic assays are necessary for the optimal determination of TP53 alterations, mainly in tumors with a p53 negative phenotype, and especially in early stage tumors for which p53 status may assist in determining its progression to invasive disease. Since p53 alterations are significantly associated to clinicopathological features of poor prognosis, the inclusion of both p53 phenotype and TP53 mutation status into a predictive panel of tumor markers for bladder cancer is recommended.

Lack of p53 nuclear immunostaining is not indicative of absence of TP53 gene mutations in colorectal adenocarcinomas.

Multiple studies using primary tumors have reported that alterations in p53 expression and detection of TP53 mutations are associated with clinical aggressiveness and poor response to specific therapies. However, there is no general agreement regarding the optimal technical approach to the analysis of p53. We have studied a series of 100 primary colorectal adenocarcinomas by immunohistochemistry with the monoclonal antibody PAb1801, and single-stranded conformation polymorphism (PCR-SSCP, exons 4–8) followed by direct sequencing of shifted bands. p53 Nuclear staining was undetectable (score 0) in 29 of 100 cases. However, gene mutations were detected in 15 of these cases, with all of these mutations leading to abnormal proteins. p53 Nuclear staining was detectable and scored as less than 10% tumor cells positive in 15 of 100 cases but was still considered to be displaying a p53-negative phenotype because the cut-off value for positivity was 10% positive tumor cells. Nevertheless, TP53 gene mutations were detected in 2 of these cases. p53 Nuclear immunoreactivities were detectable and scored as more than 10% tumor cells positive in 56 cases, considered the p53-positive phenotype. TP53 gene mutations were identified in 51 of these 56 cases. These results reveal that immunohistochemical assessment does not predict TP53 mutation status in colorectal adenocarcinoma, mainly in cases displaying absence of nuclear staining. It is thus concluded that molecular profiling should be conducted in parallel with immunophenotyping when analyzing colorectal tumors for p53 status.

Microsatellite instability of the colorectal carcinoma can be predicted in the conventional pathologic examination. A prospective multicentric study and the statistical analysis of 615 cases consolidate our previously proposed logistic regression model

High microsatellite instability (MSI-H) allows the identification of a subset of colorectal carcinomas associated with good prognosis and a higher incidence of Lynch syndrome. The aim of this work was to assess the interobserver variability and optimize our MSI-H prediction model previously published based on phenotypic features. The validation series collected from five different hospitals included 265 primary colorectal carcinomas from the same number of patients. The eight clinicopathological parameters that integrate our original model were evaluated in the corresponding centers. Homogeneity assessment revealed significant differences between hospitals in the estimation of the growth pattern, presence of Crohn-like reaction, percentage of cribriform structures, and Ki-67 positivity. Despite this observation, our model was globally able to predict MSI-H with a negative predictive value of 97.0%. The optimization studies were carried out with 615 cases and resulted in a new prediction model RERtest8, which includes the presence of tumor infiltrating lymphocytes at the expense of the percentage of cribriform structures. This refined model achieves a negative predictive value of 97.9% that is maintained even when the immunohistochemical parameters are left out, RERtest6. The high negative predictive value achieved by our models allows the reduction of the cases to be tested for MSI to less than 10%. Furthermore, the easy evaluation of the parameters included in the model renders it a useful tool for the routine practice and can reinforce other published models and the current clinical protocols to detect the subset of colorectal cancer patients bearing hereditary nonpolyposis colorectal cancers risk and/or MSI-H phenotype.

A novel logistic model based on clinicopathological features predicts microsatellite instability in colorectal carcinomas.

High-frequency microsatellite instability has been reported to be associated with good prognosis in colorectal adenocarcinoma. However, methods to assess microsatellite instability (MIN) are based on genetic assays and are not ideally suited to most histopathology laboratories. The aim of the present study was to develop a model for prediction of MIN status in colorectal cancer based on phenotypic characteristics. Clinicopathological features of a cohort of 204 patients with primary colon cancer were retrospectively reviewed following predetermined criteria. Genetic assessment of MIN status was performed on DNA extracted from sections of formalin-fixed, paraffin-embedded specimens by testing a panel of 11 microsatellite markers. Logistic regression analysis generated a mathematical tool capable of identifying colorectal tumors displaying MIN status with a sensitivity of 77.8% and a specificity of 96.8%. Features associated with instability included the proximal location of the lesions, occurrence of solid and/or mucinous differentiation, absence of cribriform structures, presence of peritumoral Crohn-like reaction, expansive growth pattern, high Ki67 proliferative index, and p53-negative phenotype. This approach predicts microsatellite instability in colorectal carcinoma with an overall assigned accuracy of 95.1% and a negative predictive value of 97.8%. Implementation of this tool to routine histopathological studies could improve the management of patients with colorectal cancer, especially those presenting with stage II and III of the disease. It will also assist in identifying a subset of patients likely to benefit from adjuvant chemotherapy.

Distribución geográfica y evolución temporal de la mortalidad evitable en Cataluña (1986-2001)

Objetivos: Analizar la evolucion temporal y la distribucion geografica por sectores sanitarios de la mortalidad evitable en Cataluna. Material y metodos: Se analizo la mortalidad evitable segun la clasificacion utilizada en el Departamento de Salud de Cataluna por sectores sanitarios durante el periodo 1986-2001 y se agruparon las causas en tratables o prevenibles. Se calcularon las tasas de mortalidad estandarizadas por el metodo directo e indirecto y la razon de mortalidad comparativa para el grupo de tratables y prevenibles y para los 46 sectores sanitarios. Tambien se calculo el promedio de cambio anual ajustado por edad mediante la regresion de Poisson de la mortalidad evitable y general. Resultados: El total de defunciones evitables fue 61.261 (el 7,3% de la mortalidad general). 10.623 (17,34%) se clasificaron como tratables y 50.638 (82,65%) como prevenibles. El promedio de cambio anual para las causas evitables fue del -2,43% (intervalo de confianza [IC] del 95%, -2,60 a -2,26), superior al -1,57% (IC del 95%, -1,61 a -1,52) de la mortalidad general. Las tasas fueron mas elevadas para las causas prevenibles que para las tratables, aunque en ambos grupos se aprecio un descenso de la mortalidad. El sector sanitario del Segria destaca por presentar sobremortalidad en los 2 periodos y en los 2 grupos de causas. Cuatro sectores sanitarios presentan un aumento significativo de la mortalidad por causas prevenibles, pero ninguno por causas tratables. Conclusiones: En Cataluna, durante el periodo 1986-2001 se produce un descenso de la mortalidad evitable, mayor que el de la mortalidad general. La distribucion geografica muestra una gran dispersion, aunque se identifican claramente zonas en las que es necesaria la intervencion preventiva.

Mortalidad por cáncer de estómago en Cataluña: distribución geográfica y evolución temporal entre 1986 y 2000

Fundamento y objetivo Este estudio analiza la evolucion temporal y los cambios en la distribucion espacial, por comarcas, de la mortalidad por cancer de estomago en Cataluna, para cada sexo, durante el periodo 1986-2000. Material y metodo Las defunciones por cancer de estomago de los anos 1986-2000 proceden del Registro de Mortalidad de Cataluna del Departament de Salut y la poblacion del Institut d’Estadistica de Cataluna. Para el analisis de la evolucion temporal se ajusto un modelo de regresion de Poisson para cada sexo. Para el analisis espacial en cada comarca se ha utilizado un modelo jerarquico bayesiano. Resultados En los anos 1986-2000 se produjeron 8.627 defunciones por cancer de estomago en varones y 5.831 en mujeres. Durante este periodo se ha estimado un descenso de la mortalidad del 3,13% en los varones y del 3,91% en las mujeres. El analisis espacial muestra que las zonas con menor riesgo de mortalidad son las del litoral y este riesgo aumenta de forma gradual hacia las zonas del interior. Este patron geografico es muy similar para los 2 sexos y se ha ido difuminando con el paso del tiempo. Conclusiones La evolucion temporal y la distribucion geografica de la mortalidad por cancer de estomago en Cataluna son similares en ambos sexos y coherentes con las tendencias observadas en otros paises desarrollados. Este patron indica una mejora en los habitos alimentarios y una mejor accesibilidad a los servicios sanitarios de las zonas con mayor riesgo.

Cross-reactivity of EGFR mutation-specific immunohistochemistry assay in HER2-positive tumors.

The coexpression of HER2 and EGFR L858R in a solitary nodule removed from the lung, whose mutation was not confirmed by molecular techniques, made us think about the possible existence of a cross-reaction between HER2 and the EGFR L858R-specific antibody. Our study was designed to further analyze the existence of this cross-reaction and stress the need to exclude a metastatic breast cancer when dealing with EGFR L858R-positive cases. The series consists of 42 primary breast carcinomas, 22 HER2 positive for overexpression and amplification, and 20 negative for both. EGFR mutations were studied by immunohistochemistry and confirmed using real-time PCR when positive. Immunohistochemistry assay with EGFR L858R was positive in 19 (86%) of the HER2-positive breast carcinomas and negative in all HER2-negative carcinomas. The EGFR L858R antibody gives false-positive results in most of the breast carcinomas with HER2 overexpression/amplification. As a consequence, it is essential to confirm any EGFR L858R-positive cases by molecular methods or at least discard the presence of HER2 overexpression/amplification before rendering a diagnosis. It is also important to consider that HER2 has been described in other carcinomas such as urothelial, gastric or ovarian, as well as lung, although infrequently.

Geographical distribution and time trends in avoidable mortality in Catalonia, Spain (1986-2001)

OBJECTIVES To analyze time trends and geographical variation in avoidable mortality by health areas in Catalonia. MATERIAL AND METHODS Avoidable mortality was analyzed according to the classification used by the Health Department of the Regional Government of Catalonia from 1986-2001 for health areas and causes were grouped as treatable and preventable. Standardized mortality rates were calculated by the direct and indirect method and the comparative mortality figures were calculated for the treatable and preventable groups and for the 46 health areas. The mean annual change adjusted for age was also calculated using a Poisson regression of avoidable and general mortality. RESULTS The total number of avoidable deaths was 61,261 (7.3% of overall deaths). 10,623 cases (17.34%) were classified as treatable and 50,638 (82.65%) as preventable. The mean annual change for avoidable causes was -2.43% (95% CI, -2.60 to -2.26), higher than the -1.57% (95% CI, -1.61 to -1.52) change for general mortality. The rates were higher for preventable causes than for treatable causes, although mortality decreased in both groups. The health area of Segrià was notable for its significantly higher mortality from both treatable and preventable causes in both periods. Four health areas showed a significant increase in mortality from preventable causes but none showed an increase in mortality from treatable causes. CONCLUSIONS In Catalonia, the decrease in avoidable mortality was greater than that in general mortality from 1986 to 2001. The geographical distribution shows wide dispersion but allows areas requiring preventive interventions to be identified.

Correlation of mRNA-PCA3 urine levels with the new grading system in prostate cancer

PURPOSE To evaluate the PCA3 (Prostate Cancer 3 gene) as a tool to improve prostate cancer (PCa) screening and its capability to predict PCa aggressiveness. PATIENTS AND METHODS A retrospective study with data from consecutive patients with suspected PCa seen in the urology department between November 2009 and April 2016 and who were candidates for prostate biopsy. A total of 1038 urine samples were tested in our laboratory with a kit that generated a PCA3 score (s-PCA3). A prostate biopsy was recommended only in those patients with s-PCA3≥35. Associations between variables were analyzed using the R software. RESULTS In patients with a positive s-PCA3 (44.5%), a subsequent biopsy was recommended. Of a total of 151 biopsies studied, 56.3% yielded a diagnosis of PCa. The probability of a positive biopsy increased as the s-PCA3 increased (p=0.041). The percentage of affected cylinders increased as the s-PCA3 increased (p=0.015). A statistically significant relationship was observed between s-PCA3 and both the Gleason score and the Grade Group (p=0.001 and 0.008, respectively). The best log-linear models and a logistic model confirmed the relationships shown previously with Fishers exact tests. CONCLUSIONS S-PCA3 may serve as an additional marker to reduce the indication for biopsies and avoid overdiagnosis and overtreatment of patients with suspected PCa. The prognostic significance of s-PCA3 was confirmed, as it was associated with tumor volume and Gleason score. Importantly, to our knowledge this is the first time that an association has been demonstrated between s-PCA3 and the new Grade Group.