Delphi project in bronchial asthma. Two stages.

From the paediatric point of view, we have undertaken two Delphi studies into bronchial asthma. The first is related to the consensus known as the consensus document of the five associations. The second is more recent and has been undertaken with GEMA (the Spanish Guidelines on the Management of Asthma). The aim of this paper is to carry out a descriptive study comparing the 2 Delphi processes and to objectively assess if in some way behaviour over the past two years has changed as far as expert opinion is concerned. In the consensus document those points giving rise to most controversy were the treatment of children under three years of age and treatment with immunotherapy in allergic asthma. It is also necessary to highlight how important it was at that particular point in time to define the phenotypes of wheezing and the predictive index of asthma in children of less than 3 years of age. Of the 52 questions in the questionnaire, in 13.6% the panel of experts reached no consensus in their positions. Following GEMA the Delphi methodology, 56 questions were asked in the first round of the questionnaire, and consensus was reached in 87.5%. As regards the paediatric part relating to diagnosis and treatment in children, agreement was reached on all the questions in the first round. Agreement was reached in 8.92% questions in the second round. Clinical guidelines and consensus documents can modify behaviour towards an illness, both in the diagnosis and treatment.

Loss of heterozygosity on the short arm of chromosome 8 in male breast carcinomas.

Identification of loss of heterozygosity (LOH) at specific genetic loci in neoplastic cells suggests the presence of a tumor suppressor gene within the deleted region. LOH on chromosome 8p has been identified in colorectal, bladder, hepatocellular, and prostatic carcinomas. Little is currently known about the molecular events occurring during the development of male breast cancer. We studied LOH on chromosome 8p in 23 male breast carcinomas. Five polymorphic DNA markers were used: D8S136 and D8S137 on 8p12-21.3; and D8S254, D8S258, and D8S349 on 8p22. DNA was extracted from microdissected normal and tumor cells obtained from formalin-fixed, paraffin-embedded tissue sections and amplified by the PCR. LOH was identified in 19 of 23 cases (83%) with at least one marker. Seven cases showed LOH only at 8p22, six cases showed LOH only at 8p12-21.3, and six cases showed LOH at both 8p22 and 8p12-21.3. In five of these last six cases, at least one locus was retained between the two deleted regions; thus, the whole short arm of chromosome 8 was not lost in these tumors. Our results show that there are two discrete areas of deletion on chromosome 8p in male breast cancer, suggesting the presence of one or more tumor suppressor genes that may play a role in the development or progression of the disease.

A 50-gene signature is a novel scoring system for tumor-infiltrating immune cells with strong correlation with clinical outcome of stage I/II non-small cell lung cancer.

PURPOSE: To identify a novel system for scoring intratumoral immune response that can improve prognosis and therapy decisions in early stage non-small cell lung cancer (NSCLC). METHODS/PATIENTS: Eighty-four completely resected stage I/II NSCLC without adjuvant therapy were classified by expression profiling using whole genome microarrays. An external cohort of 162 tumors was used to validate the results. Immune cells present in tumor microenvironment were evaluated semiquantitatively by CD20, CD79, CD3, CD8, CD4 and CD57 immunostaining. Univariate and multivariate analyses of variables associated with recurrence-free survival were performed. RESULTS: Initial molecular classification identified three clusters, one with significantly better RFS. A reduced two-subgroup classification and a 50-gene predictor were built and validated in an external dataset: high and low risk of recurrence patients (HR = 3.44; p = 0.001). Analysis of the predictor´s genes showed that the vast majority were related to a B/plasma cell immune response overexpressed in the low-risk subgroup. The predictor includes genes coding for unique B lineage-specific genes, functional elements or other genes that, although non-restricted to this lineage, have strong influence on B-cell homeostasis. Immunostains confirmed increased B-cells in the low-risk subgroup. Gene signature (p < 0.0001) and CD20 (p < 0.05) were predictors for RFS, while CD79 and K-RAS mutations showed a tendency. CONCLUSIONS: Favorable prognosis in completely resected NSCLC is determined by a B-cell-mediated immune response. It can be differently scored by a 50-gene expression profile or by CD20 immunostaining. That prognosis information not reflected by traditional classifications may become a new tool for determining individualized adjuvant therapies.

Vascular enumeration as a prognosticator for colorectal carcinoma.

Vascular enumeration is thought to be an independent prognosticator for several human tumours, including breast, bladder and colorectal carcinomas. There have been 12 reports on the prognostic influence of vascular enumeration in colorectal carcinoma with different results. To test the prognostic influence of this factor in our patients, we have selected 126 patients with colorectal carcinoma Dukes' stages A to C treated only with curative surgery with no further adjuvant therapy. The minimal follow-up time was 5 years (60 months). After immunostaining with CD34, we performed a manual count of the vessels following Gasparini's criteria. In our series vascular enumeration showed significant association with the histological grade (P = 0.03) with a cut-off point at 77 vessels/200x, but not with tumour staging and vascular and neural invasion (P > 0.05). Vascular enumeration was a prognosticator for RFS (relapse-free survival) (P = 0.009) and OS (overall survival) (P = 0.01) in all Dukes' stages in the univariate analysis, but this prognostic influence was lost in the multivariate analysis, in which only stage, histological differentiation, location and vascular and neural invasion behaved as significant independent prognosticators.

Triton tumor of the parotid area. Case report.

A 27-year-old woman with a Malignant Triton Tumor (MTT), or malignant schwannoma with rhabdomyoblastic differentiation, located in the parotid cell and infiltrating the nasal sinuses and the left orbit is described. The initially resected tumor showed three recurrences within a 2 years follow-up period. During successive recurrences an increase in cellular density, number of mitoses and necrosis was noticed. Immunohistochemical analysis showed that the tumor was composed of a mixed population of cells. Some of them showed positivity for actin, desmin and myoglobin, while others were positive for S-100 protein, glial fibrillary acid protein, and IV-collagen. Cytokeratin stainings were negative. Up to now, 8 benign triton tumors and another 45 cases of MTT have been described. None of them was primarily located in the parotid gland, and infiltration to the orbital cavity has not been previously described.

Case report: esophageal collision tumor (oat cell carcinoma and adenocarcinoma) in Barrett's esophagus: immunohistochemical, electron microscopy and LOH analysis.

We report a case of an esophageal collision tumor composed of adenocarcinoma and oat cell carcinoma. Both tumors appeared to arise from dysplastic Barrett's mucosae in a 75-year-old man. Immunohistochemical stains and electron microscopy demonstrated a separate identity for each of the tumors in collision. Molecular analysis of microsatellite regions was performed in different microdissected areas. Identical loss of heterozygosity (LOH) at 9p21 and 17p13 was determined in the three different microdissected areas of the adenocarcinoma component. LOH was not determined in any area of the oat cell carcinoma. This is the first study that analyzes the allele status of an esophageal collision tumor. Our findings suggest a biclonal origin for both components of the collision tumor.

3p21, 5q21, 9p21 and 17p13 allelic deletions accumulate in the dysplastic spectrum of laryngeal carcinogenesis and precede malignant transformation.

A tissue field of somatic genetic alterations precede the histopathological phenotypic changes of carcinoma. Loss of Heterozygosity (LOH) at the sites of known or putative tumor suppressor genes is a common genetic abnormality detected in precancerous conditions. These genomic changes could be of potential use in the diagnosis and prognosis of pre-malignant laryngeal lesions. Recently the concept of laryngeal intraepithelial neoplasia (LIN) was introduced. To evaluate patients with an increased risk of developing invasive laryngeal carcinoma via a dysplasia-carcinoma progression we investigated 102 microdissected cell populations. Cell populations were procured from 15 laryngectomy specimens with different peritumoral histological changes adjacent to the squamous cell carcinoma cells and 15 laryngeal endoscopic biopsies with no evidence of malignant transformation in a 6-10-year follow-up period. Histological diagnoses were subdivided into keratosis without dysplasia (KWD), with mild dysplasia (LIN 1), with moderate dysplasia (LIN 2), and with severe dysplasia or carcinoma in situ (LIN 3). Microsatellite analysis was performed with the aim of studying LOH of 5q21 (APC), 9p21 (p16), 3p21 and 17p13 (p53) chromosomal regions. Frequent allelic losses were found in carcinoma cells at p53 (54%), p16 (66%), 3p21(87%) and 5q21(58%). Identical LOH patterns were determined in 100% of the LIN3 peritumoral cells, 60% of LIN2, 50% of LIN 1 and 25% of KWD. In contrast, histologically normal mucosae, KWD and LIN1 lesions without malignant progression showed no allelic loss. These results show that dysplasia correlates with LOH at 3p21, 5q21, 9p21 and 17p13 in early laryngeal carcinogenesis. These genomic changes in pre-malignant laryngeal lesions could be of potential use as markers for cancer risk assessment.

Comparative study of tumor angiogenesis and immunohistochemistry for p53, c-ErbB2, c-myc and EGFr as prognostic factors in gastric cancer.

Gastric cancer (GC) continues to be a highly aggressive malignancy with poor prognosis and low survival rates. The survival of patients with GC depends mainly on the stage of the disease, with early GC having a 5 year survival of 90-100% and advanced tumors a 5 year survival of 15-25%. The role of other prognostic factors in these tumors is still under investigation. 28 gastric dysplasia, 45 Early GC and 98 Advanced Gastric Cancers were evaluated for expression of the oncogenes p53, c-ErbB2, c-myc and the EGFr in paraffin-embedded material utilizing Avidin-Biotin immunohistochemistry techniques. In 34 cases of GC microvessel density (MVD) was determined in CD34 stained sections. Statistical correlations with stage, histologic type, differentiation degree, location, size, ploidy patterns and overall survival were done. The Mantel-Cox test was performed to evaluate which factors had an independent prognostic value. Both, tumor angiogenesis and p53 protein expression were statistically associated (95% confidence intervals) with overall survival in patients with GC. p53 protein expression was also correlated with cardial location, nodal involvement and tumor stage. c-ErbB2 may recognize a group of highly aggressive well differentiated adenocarcinomas with worse prognosis. c-myc was also significantly enhanced in well differentiated tumors. EGFr showed no significant associations. Mantel-Cox was performed to compare the prognostic value of tumor stage, p53 protein expression and tumor angiogenesis. Tumor angiogenesis was the most important prognostic indicator to predict overall survival in our series. p53 expression was not independent and did not provide additional prognostic information to tumor stage. Our study suggests that angiogenesis as demonstrated by microvessel counts in CD34 stained sections is a significantly important prognostic factor for predicting survival in gastric cancer.

Hepatocellular carcinomas diagnosed following metastasis to the oral cavity. Report of 2 cases.

Oral metastases from hepatocellular carcinomas are rare. Case 1 was a 66-year-old male without previous history of liver disease who presented with metastasis to the gingival jaw mucosae on the lingual side. Case 2 was a 71-year-old male, with a previous history of diabetes, hepatitis, and cirrhosis who presented with metastasis to the right palatine tonsil. Oral metastases were the first manifestation of the hepatocellular carcinoma in both cases. A review of the literature disclosed 20 cases of hepatocellular carcinoma metastasizing to the oral cavity, 7 affecting the gingival mucosae and none of them affecting the palatine tonsil.

Prognostic significance of microvascular counts in rectal carcinoma.

Numerous studies of many tumor types have demonstrated that microvessel quantitation as a measure of angiogenesis is a powerful prognostic tool. Vascular enumeration has been claimed to be an independent prognosticator for several human tumors, including breast carcinoma, melanoma or bladder carcinoma; however, the studies of colorectal cancer have rendered variable results. To test the prognostic influence of this factor in our patients, we selected 39 patients with rectal carcinoma Dukes' stages A to C treated only with curative surgery, with no further adjuvant therapy. The minimal follow-up time was 5 years (60 months). After immunostaining with CD34, we performed a manual count of the vessels following Gasparini's criteria. In our series, vascular enumeration has been a prognosticator for OS (overall survival) but not for RFS (relapse-free-survival) at all Dukes' stages in the univariate analysis. This prognostic influence was lost in the multivariate analysis, in which only stage as well as vascular and neural invasion behaved as significant independent prognosticators. The presence of hypervascularization did not show any significant association with histologic grade, tumor staging, and vascular or neural invasion.

LOH at the APC/MCC gene (5Q21) in gastric cancer and preneoplastic lesions. Prognostic implications.

The APC/MCC gene (Familial Adenomatous Polyposis) at 5q21 plays a role in colon cancer carcinogenesis. LOH at this locus has also been described in gastric cancer and preneoplastic lesions. The APC locus has been recently related to a cell surface adhesion molecule and its alteration may favour metastatic dissemination. LOH at 5q21 has been associated with poor prognosis in other tumors such as lung cancer. Thirty-six gastric cancers were evaluated for LOH at 5q21 with 2 polymorphic markers from microdissected paraffin-embedded material. All tumors were classified by stage, histologic type, degree of differentiation and survival rates. In 4 cases, intestinal metaplasia cells in the adjacent mucosae were also microdissected. Six cases of moderate-severe gastric dysplasia were also added to the study. LOH was determined in 84% of the informative cases of GC, affecting both early and advanced stages of disease. Genomic instability was assessed in 5 cases, 3 of them associated with LOH. The only case of gastric cancer that did not show LOH or instability at 5q21 was a stage II, poorly differentiated intestinal carcinoma without evidence of recurrence after a 36 month follow-up period (the mean survival rate in our series was 28.3% at 36 months). We also found LOH in 2/6 dysplastic lesions and 1/4 intestinal metaplasias. Our data show that LOH at 5q21 is frequent in gastric cancer and is also present in intestinal metaplasia and dysplastic lesions. LOH at this locus is not a prognostic factor in GC in our study, due to the high incidence of LOH that we found.

LOH at the APC/MCC gene (5Q21) is frequent in early stages of non-small cell lung cancer.

Lung cancer is the leading cause of death in both women and men in the United States and many European countries. Molecular cytogenetic and LOH analyses of non-small cell lung cancer have shown somatic genetic alterations in a variety of chromosomes, such as 1p, 3p, 5q, 8p, 9p, 11p, 11q and 17p. Allelic deletions of the known tumor suppressor gene APC at 5q21 are frequently observed in advanced stages of lung cancer and have been correlated with poor prognosis in previous reports. We investigated 33 cases of NSCL for LOH at 5q21: 22 squamous cell and 11 adenocarcinomas. Normal and tumor cells were microdissected from paraffin embedded tissues and PCR amplification was performed utilising the specific markers D5S299 and D5S346 at 5q21 and PYGM at 11q13, respectively. Clinicopathological data, survival and recurrence rates were obtained in all cases. We detected LOH at 5q21 in 4/9 (44%) informative adenocarcinomas and in 13/16 (81%) informative SCC. LOH was frequent in early stages (12/15 stage I cases) and did not correlate with recurrence or poor survival. Our results show that LOH at 5q21 is more frequent in squamous cell carcinomas than in adenocarcinomas, is frequent in early stages of the disease, and does not have prognostic significance.

17p13 (p53 locus), 5q21 (APC locus) and 9p21 (p16 locus) allelic deletions are frequently found in oral exfoliative cytology cells from smoker patients with non-small-cell lung cancer.

Molecular cytogenetic and LOH analyses of non-small cell lung cancer (NSCLC) have shown frequent allelic deletions in a variety of chromosomes where tumour suppressor genes are located. Allelic loss at 9p21 (p16 locus), 17p13 (p53) and 5q21(APC) has been frequently described in NSCLC and has also been described in premalignant epithelial lesions of the bronchus and normal bronchial cells. These findings suggest that a tissue field of somatic genetic alterations precedes the histopathological phenotypic changes of carcinoma. Similar changes have been described in oral and laryngeal epithelial tumours associated with smoke exposure. We previously reported frequent LOH at 5q21, 9p21 and TP53 in tumor cells and peritumoral normal bronchial cells from surgically resected NSCLC. We now analyze 96 cases of normal oral exfoliative cytology in which normal epithelial cells were obtained: 43 cases from smoker patients with NSCLC diagnosis, 33 smoker patients with no evidence of malignancy and 20 non-smoker patients with no evidence of tumour. All groups had a similar age and sex distribution. PCR amplification was performed utilising the specific markers D5S346, D9S157 and TP53. In normal oral mucosae cells from patients with NSCLC, we found that 21% of the informative cases showed LOH at any of the three analyzed loci distributed as follows: 14.3% of the informative cases showed LOH at 5q21, 7.7% at 9p21 and 22.2% at TP53. Within the smoker risk group only one case (4% of the informative cases) showed LOH at TP53, while no LOH was found at 5q21 or 9p21. No LOH was found in non-smokers. In conclusion, our results show that a significant number of patients with NSCLC have LOH at TP53, 5q21 and 9p21 in normal oral mucosae, while LOH at these loci is unusual in similar cells obtained from patients with no evidence of malignancy. Our study demonstrates that LOH studies can detect smoker patients with a mutated genotype in normal epithelial cells. Further prospective studies may confirm whether LOH studies can detect patients with a higher risk of NSCLC.

A 50-gene signature is a novel scoring system for tumor-infiltrating immune cells with strong correlation with clinical outcome of stage I/II non-small cell lung cancer

Purpose: To identify a novel system for scoring intratumoral immune response that can improve prognosis and therapy decisions in early stage non-small cell lung cancer (NSCLC). Methods/patients: Eighty-four completely resected stage I/II NSCLC without adjuvant therapy were classified by expression profiling using whole genome microarrays. An external cohort of 162 tumors was used to validate the results. Immune cells present in tumor microenvironment were evaluated semiquantitatively by CD20, CD79, CD3, CD8, CD4 and CD57 immunostaining. Univariate and multivariate analyses of variables associated with recurrence-free survival were performed. Results: Initial molecular classification identified three clusters, one with significantly better RFS. A reduced two-subgroup classification and a 50-gene predictor were built and validated in an external dataset: high and low risk of recurrence patients (HR = 3.44; p = 0.001). Analysis of the predictor´s genes showed that the vast majority were related to a B/plasma cell immune response overexpressed in the low-risk subgroup. The predictor includes genes coding for unique B lineage-specific genes, functional elements or other genes that, although non-restricted to this lineage, have strong influence on B-cell homeostasis. Immunostains confirmed increased B-cells in the low-risk subgroup. Gene signature (p < 0.0001) and CD20 (p < 0.05) were predictors for RFS, while CD79 and K-RAS mutations showed a tendency. Conclusions: Favorable prognosis in completely resected NSCLC is determined by a B-cell-mediated immune response. It can be differently scored by a 50-gene expression profile or by CD20 immunostaining. That prognosis information not reflected by traditional classifications may become a new tool for determining individualized adjuvant therapies (AU)

No disponible