Somatic Mutations of PI3K in Early and Advanced Gallbladder Cancer: Additional Options for an Orphan Cancer.

Gallbladder cancer (GBC) is the second-leading cause of death from malignant tumors in Chilean women. The phosphatidylinositol 3-kinase (PI3K) pathway is involved in proliferation, cell survival, and growth. We investigated mutations in exons 9 and 20 of the PI3K gene in GBC. Mutations in exons 9 (E542K, E545G, E545K) and 20 (H1047L and H1047R) of PI3K were determined by direct sequencing in 130 cases of GBC. The patient group consisted of 110 women and 20 men, and mutations were found in 22 cases (16.9%). Of these, 14 cases had mutations in exon 9 (63.6%) (E542K, 64%; E545K, 29%; and E545G, 7%) and 8 in exon 20 (37.4%; H1047L, 50%; H1047R, 50%). No differences were noted in the frequency and type of mutations analyzed by sex, age, or histologic features. We observed mutations in 22% of the early-stage GBC and 14.6% of the advanced cases. In this series of GBC, 17% of cases were noted as having mutations in either exons 9 or 20 of PI3K. These results suggest that therapeutic testing of inhibitors of the PI3K/AKT pathway may be of benefit in advanced GBC patients.

Inactivation of tumor suppressor gene pten in early and advanced gallbladder cancer.

BACKGROUND: PTEN is a tumor suppressor gene that regulates the PTEN/PI3k/AKT/mTOR pathway, which is frequently altered in human cancers including gallbladder cancer (GBC). To determine the frequency of PTEN expression in GBC and to establish its relation to clinical and morphological parameters and survival in GBC. METHODS: The immunohistochemical expression of PTEN was studied in 108 GBC. All the cases included areas of non-tumor mucosa adjacent to the tumor. RESULTS: The group was comprised of 108 patients, 91 women (84.3%) and 17 men (15.7%) with an average age of 65.2 years (SD ± 12.3 years). Thirty-five cases (33%) were early carcinomas (EC) and the remaining 73 (67%) were advanced cases (AC). All the internal controls were positive (moderate or intense in 96.3%). Only in three AC (4.1%) was there a complete absence of PTEN immunohistochemical expression. There were no significant differences in relation between PTEN expression and tumor infiltration or degree of differentiation. The three patients with PTEN inactivation died before 10 months; however, the other patients with AC had a survival of 53% at 10 months. DISCUSSION: Loss of PTEN expression was observed in 4.1% of the advanced GBC. All the patients with this alteration died before 10 months. PTEN inactivation could be a rare event, but with a poor prognosis in advanced GBC.

[BRAF gene mutation in wild-type KRAS patients with colorectal cancers]. / Mutación del gen BRAF en pacientes con cánceres de colon y recto con KRAS no mutado.

BACKGROUND: In colorectal cancer, BRAF and KRAS mutation are mutually exclusive, but both are independent prognostic factors for the disease. AIM: To determine the frequency of BRAF V600E mutation in colorectal cancer. MATERIAL AND METHODS: A KRAS mutation study was carried out in 100 tissue samples of primary and metastatic adenocarcinomas of colon and rectum from patients aged 61.1 ± 62 years (56 women). Negative KRAS mutation cases underwent study of BRAF V600E mutation by restriction fragment length polymorphism (RFLP) and direct sequencing. RESULTS: Primary tumors were located in the colon and rectum in 88 and six cases respectively. Five were liver metastases and in one case, the sample location was undetermined. Forty two samples were KRAS positive (mutated). In 12 of the 58 KRAS negative (wild type) samples, the V600E mutation in codon 15 of the BRAF gene was demonstrated. No differences in the frequency and distribution of mutations, stratified by gender, age, primary tumor versus metastasis, or tumor location were observed. CONCLUSIONS: Twelve percent of KRAS negative colorectal cancer samples showed BRAF gene mutation. Considering that 42% of samples have a KRAS mutation, 54% of patients should not respond to therapies with monoclonal antibodies directed against epidermic growth factor (EGFR) pathway.

Mutación del gen BRAF en pacientes con cánceres de colon y recto con KRAS no mutado / BRAF gene mutation in wild-type KRAS patients with colorectal cancers

Background: In colorectal cancer, BRAF and KRAS mutation are mutually exclusive, but both are independent prognostic factors for the disease. Aim: To determine the frequency of BRAF V600E mutation in colorectal cancer. Material and Methods: A KRAS mutation study was carried out in 100 tissue samples of primary and metastatic adenocarcinomas of colon and rectum from patients aged 61.1 ± 62 years (56 women). Negative KRAS mutation cases underwent study of BRAF V600E mutation by restriction fragment length polymorphism (RFLP) and direct sequencing. Results: Primary tumors were located in the colon and rectum in 88 and six cases respectively. Five were liver metastases and in one case, the sample location was undetermined. Forty two samples were KRAS positive (mutated). In 12 of the 58 KRAS negative (wild type) samples, the V600E mutation in codon 15 of the BRAF gene was demonstrated. No differences in the frequency and distribution of mutations, stratified by gender, age, primary tumor versus metastasis, or tumor location were observed. Conclusions: Twelve percent of KRAS negative colorectal cancer samples showed BRAF gene mutation. Considering that 42% of samples have a KRAS mutation, 54% of patients should not respond to therapies with monoclonal antibodies directed against epidermic growth factor (EGFR) pathway.

¿Es posible utilizar las muestras de colecistectomías con cáncer en investigación?: Calidad del ADN de muestras obtenidas del sistema público y privado de salud / Quality of DNA from archival pathological samples of gallbladder cancer

Background: The quality of the archival samples stored at pathology services could be a limiting factor for molecular biology studies. Aim: To determine the quality of DNA extracted from gallbladder cancer samples at different institutions. Material and Methods: One hundred ninety four samples coming from fve medical centers in Chile, were analyzed. DNA extraction was quantifed determining genomic DNA concentration. The integrity of DNA was determined by polymerase chain reaction amplification of different length fragments of a constitutive gene (β-globin products of 110, 268 and 501 base pairs). Results: The mean DNA concentration obtained in 194 gallbladder cancer samples was 48 ± 43.1 ng/µl. In 22% of samples, no amplification was achieved despite obtaining a mean DNA concentration of 58.3 ng/ul. In 81, 67 and 22% of samples, a DNA amplification of at least 110, 268 or 501 base pairs was obtained, respectively. No differences in DNA concentration according to the source of the samples were demonstrated. However, there were marked differences in DNA integrity among participating centers. Samples from public hospitals were of lower quality than those from private clinics. Conclusions: Despite some limitations, in 80% of cases, the integrity of DNA in archival samples from pathology services in our country would allow the use of molecular biology techniques.

Expresión y amplificación del gen HER2 en el cáncer gástrico avanzado / HER2 gene amplification and overexpression in advanced gastric cancer

Background:Overexpression/amplification of the HER2 gene in advanced gastric cancer is a predictor of response to adjuvant therapy with monoclonal antibodies.Aim: To determine the frequency of HER2 gene overexpression and amplificationin advanced gastric cancer. Material and Methods: One hundred nine advancedgastric cancer biopsy specimens, from 76 men and 33 women aged 67 ± 14 and 62± 12 years respectively, were selected. Three histological patterns (diffuse, intestinaland mixed) were recognized. Automated immunohistochemistry was performedwith monoclonal c-erbB-2 (NCL-356) Novocastra. Fluorescent in situ hybridization (FISH) for HER2 was performed in positive cases. Results: In 39% of cases,immunohistochemical staining was negative. It was 1+, 2+ and 3+ positive in 15,36 and 11% of cases, respectively. It was positive in 16% and 3% of intestinal typeand mixed carcinomas, respectively. It was negative in all diffuse carcinomas. FISHwas performed in 39 (2 +) cases and in 11 (3 +) cases. The gene amplification waspositive in two (2 +) and 11 (3 +) cases (11.9%). The overall concordance betweenimmunohistochemical staining and in situ hybridization was 85%. Conclusions: Inadvanced gastric cancer, HER2 gene overexpression or amplification was observed in11% and 12% of cases, respectively.

[HER2 gene amplification and overexpression in advanced gastric cancer]. / Expresión y amplificación del gen HER2 en el cáncer gástrico avanzado.

BACKGROUND: Overexpression/amplification of the HER2 gene in advanced gastric cancer is a predictor of response to adjuvant therapy with monoclonal antibodies. AIM: To determine the frequency of HER2 gene overexpression and amplification in advanced gastric cancer. MATERIAL AND METHODS: One hundred nine advanced gastric cancer biopsy specimens, from 76 men and 33 women aged 67 ± 14 and 62 ± 12 years respectively, were selected. Three histological patterns (diffuse, intestinal and mixed) were recognized. Automated immunohistochemistry was performed with monoclonal c-erbB-2 (NCL-356) Novocastra. Fluorescent in situ hybridization (FISH) for HER2 was performed in positive cases. RESULTS: In 39% of cases, immunohistochemical staining was negative. It was 1+, 2+ and 3+ positive in 15, 36 and 11% of cases, respectively. It was positive in 16% and 3% of intestinal type and mixed carcinomas, respectively. It was negative in all diffuse carcinomas. FISH was performed in 39 (2 +) cases and in 11 (3 +) cases. The gene amplification was positive in two (2 +) and 11 (3 +) cases (11.9%). The overall concordance between immunohistochemical staining and in situ hybridization was 85%. CONCLUSIONS: In advanced gastric cancer, HER2 gene overexpression or amplification was observed in 11% and 12% of cases, respectively.

[Quality of DNA from archival pathological samples of gallbladder cancer]. / ¿Es posible utilizar las muestras de colecistectomías con cáncer en investigación?: Calidad del ADN de muestras obtenidas del sistema público y privado de salud.

BACKGROUND: The quality of the archival samples stored at pathology services could be a limiting factor for molecular biology studies. AIM: To determine the quality of DNA extracted from gallbladder cancer samples at different institutions. MATERIAL AND METHODS: One hundred ninety four samples coming from five medical centers in Chile, were analyzed. DNA extraction was quantified determining genomic DNA concentration. The integrity of DNA was determined by polymerase chain reaction amplification of different length fragments of a constitutive gene (ß-globin products of 110, 268 and 501 base pairs). RESULTS: The mean DNA concentration obtained in 194 gallbladder cancer samples was 48 ± 43.1 ng/µl. In 22% of samples, no amplification was achieved despite obtaining a mean DNA concentration of 58.3 ng/ul. In 81, 67 and 22% of samples, a DNA amplification of at least 110, 268 or 501 base pairs was obtained, respectively. No differences in DNA concentration according to the source of the samples were demonstrated. However, there were marked differences in DNA integrity among participating centers. Samples from public hospitals were of lower quality than those from private clinics. CONCLUSIONS: Despite some limitations, in 80% of cases, the integrity of DNA in archival samples from pathology services in our country would allow the use of molecular biology techniques.