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A Gemma

Publications and source records attributed to A Gemma.

At least 19 recordsLinked to original sources
PubMed2000

Somatic mutation of the hBUB1 mitotic checkpoint gene in primary lung cancer.

Mutations in mitotic checkpoint genes have been detected in several human cancers, and these cancers exhibit chromosomal instability. Aneuploid stem cells seem to result from chromosomal instability and have been reported in many lung cancers. To determine whether alteration of mitotic checkpoint regulators is involved in carcinogenesis and tumor progression in primary lung cancer, we screened the genomic DNA sequence of 30 human lung cancer cell lines and 30 primary lung cancer tumors for a mutation in the hBUB1 mitotic checkpoint gene. First, we designed 26 sets of intron-based primers to amplify each of the 25 exons of the hBUB1 gene to examine the entire coding region of the hBUB1 gene. Using these primers, we performed polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis as well as direct sequencing in the mutation analysis of the hBUB1 gene. Three different nucleotide substitutions were detected in the coding region of the hBUB1 gene in some of the cancer cell lines and primary tumors as follows. The hBUB1 gene of one adenocarcinoma tumor contained a somatic missense mutation, a cytosine-to-guanine substitution in codon 51 of exon 5 that resulted in a histidine-to-aspartic acid amino acid substitution. The hBUB1 gene of three lung cancer cell lines contained a thymine-to-cytosine substitution in codon 430 of exon 12, which did not result in an amino-acid substitution. We were unable to determine whether the nucleotide substitution in exon 12 was a polymorphism or a silent mutation because matched normal tissue was not available. A polymorphism in codon 93 of exon 4, a guanine-to-thymine substitution, in hBUB1 was found in one lung cancer cell line and one primary lung tumor. This is the first report of a somatic missense mutation of a gene involved in a mitotic checkpoint in primary lung cancer. The presence of a point mutation in the hBUB1 gene is consistent with the hypothesis that alteration of mitotic checkpoint genes is involved in the development of primary lung cancers. Because the frequency of hBUB1 gene mutations was low, future studies should focus on other mechanisms of inactivation of the hBUB1 gene as well as mutation analysis of other mitotic checkpoint genes in lung cancers.

Adenocarcinoma↗
PubMed2000

Mutation analysis of the gene encoding the human mannose 6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R) in human cell lines resistant to growth inhibition by transforming growth factor beta(1) (TGF-beta(1)).

The mannose 6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R) is involved in activating the transforming growth factor beta(1) (TGF-beta(1)), an inhibitor of the cell proliferation, and limiting the insulin-like growth factor 2 mediated-growth stimulation. The M6P/IGF2R gene has been reported to be mutated and deleted in various cancers, and is a candidate tumor suppressor gene. We studied the genomic structure of the M6P/IGF2R gene and designed the intron primers to detect mutations in the M6P/IGF2R gene of genomic DNA samples. The M6P/IGF2R gene consists of 48 exons. The previously reported 23 mutations of the M6P/IGF2R gene in human cancers, liver, breast, and gastrointestinal tumors, are located in five exons, exon 27, 28, 31, 40, 48. Using the intron primers designed in this study, polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis, and direct sequencing, we performed an initial analysis of the complete coding sequences of the M6P/IGF2R gene in 21 human cell lines resistant to growth inhibition by TGF-beta(1). An adenine-to-guanine transition, resulting in an asparagine-to-serine amino acid substitution, was found in one lung adenocarcinoma cell line at exon 40 where the mutation has been previously reported in human cancers. This is the first report of a mutation of the M6P/IGF2R gene in lung tumor. These results indicated that the mutation in M6P/IGF2R may be involved in human lung cancinogenesis.

Cell Line↗
PubMed2000

Altered expression and function of beta1 integrins in a highly metastatic human lung adenocarcinoma cell line.

In order to investigate the relationship between beta1 integrins and the metastatic ability of cancer cells, we established a novel and highly metastatic cell line designated PC9-f9 from a poorly metastatic human lung adenocarcinoma cell line (PC9) in nude mice. PC9-f9 cells showed higher invasive activity in the Matrigel invasion assay than PC9 cells. Additionally, in cell adhesion assays, PC9-f9 cells adhered to laminin more strongly than PC9 cells and, unlike PC9 cells, adhered to collagen type IV and fibronectin. FACS analysis showed expression of the integrins alpha2beta1, alpha3beta1 and alpha6beta1 on both of the cell lines but alpha4beta1 and alpha5beta1 were neo-expressed on PC9-f9 cells. In cell adhesion inhibition assays, alpha3beta1 was the major laminin receptor for PC9 cells but not for PC9-f9 cells. Alternatively, PC9-f9 cells adhered to collagen type IV via alpha2beta1 and adhered to fibronectin mainly via alpha5beta1 but also moderately via alpha4beta1. The pretreatment of PC9-f9 cells with anti-beta1 monoclonal antibodies suppressed lung metastases by more than 50%. These data suggest that the altered expression and function of beta1 integrins allow PC9-f9 cells to become more adhesive and invasive, and lead to increased metastatic potential.

Adenocarcinoma↗
PubMed2000

[Mechanism of resistance to growth inhibition by transforming growth factor-beta 1 (TGF-beta 1) in primary lung cancer and new molecular targets in therapy].

Transforming Growth Factor-beta 1 (TGF-beta 1) regulates the proliferation of normal epithelial cells, and resistance to TGF-beta 1 growth inhibition is a common feature of human cancers including lung cancer. In order to understand the mechanism of resistance to growth inhibition by TGF-beta 1 and to reverse the regulation of proliferation in lung cancer, we determined the genomic structure of the genes involved in the signal transduction pathway of TGF-beta 1 and performed an initial mutation survey of the complete coding region of the genes in lung cancer and cell lines with the resistance to growth inhibition by TGF-beta 1. First, a mutation analysis of the TGF-beta type II receptor (TGF-beta RII) was performed. Point mutations of the gene were detected in several colon cancers and an adenocarcinoma of the lung in the poly-A sequence. No mutations of Smad 2, 3, 4, 5 and TGF-beta type I receptor (TGF-beta IR) genes were detected in a series of the tumors we tested, although several mutations of Smad 2 and 4 were previously reported. Frequent alterations of the p15 gene and reduced expression of p21 we already reported from our previous studies. We also determined the genomic structure of the mannose 6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R), which is involved in activating TGF-beta 1, and performed an initial mutation survey of the complete coding sequences of the gene. A point mutation at exon 40 was found in one lung adenocarcinoma cell line. In summary, alterations in the many genes involved in the signal transduction of TGF-beta 1 were found and may mediate the loss of TGF-beta 1 responsiveness in lung cancer. The molecular targets for the regulation of the proliferation of lung cancer are thought to be p15, p21 and the transcriptional regulators.

Cell Division↗
PubMed2000

Increase in the frequency of p16INK4 gene inactivation by hypermethylation in lung cancer during the process of metastasis and its relation to the status of p53.

The p16INK4 gene, which is a tumor suppressor gene, is frequently altered in lung cancers. Hypermethylation of the promoter region of the p16INK4 gene seems to be the major mechanism through which p16INK4 become inactivated. Hypermethylation of the p16INK4 gene was reported to occur at an early stage in lung cancer. To determine whether the change in p16INK4 methylation status occurs at the late stage in the progression of primary lung cancers, we analyzed the primary and metastatic tumor tissues and normal lung samples from 29 cases of advanced lung cancer with distant metastasis. In each tissue sample, we analyzed the p16INK4 and p15INK4b genes for mutations and the methylation status of both genes using PCR-single strand conformation polymorphism, direct sequencing, and methylation-specific PCR analysis. We also analyzed a subset of the samples for p16INK4 protein expression. Genetic mutations in the coding region of the p16INK4 and p15INK4b genes were not found in any of the examined specimens. The promoter region of the p16INK4 gene was hypermethylated in the tumor samples of the primary or metastatic site of 37.0% (10 of 27) of the subjects. The promoter region of the p16INK4 gene was hypermethylated at both the primary and metastatic sites in two of the 10 cases and at only the metastatic site in 8 cases. By immunohistochemical analysis, we confirmed the presence of p16INK4 protein at the primary site of all cases in which the promoter region of the p16INK4 gene was hypermethylated at only the metastatic site. Interestingly, all 8 cases with a hypermethylated p16INK4 promoter region, at only the metastatic site, did not have p53 mutation. The results of this study indicate that tumor cells in which the p16INK4 gene has been inactivated by hypermethylation of the promoter region could have an advantage in progression and metastasis in non-small cell lung cancers, especially in the tumors with normal p53, and that the frequency of p16INK4 gene inactivation by hypermethylation could vary in clinical course.

Adenocarcinoma↗
PubMed1999

Cognitive impairment and (CTG)n expansion in myotonic dystrophy patients.

BACKGROUND: Myotonic dystrophy (DM) is a genetic multisystemic disease with muscular, endocrine, ocular, cardiac and cognitive impairment. The molecular basis of the disease has been identified in an unstable base triplet (CTG)n repeat located in the 3' untranslated region of the miotonin protein-kinase (MT-PK) gene on the long arm of chromosome 19. Cognitive impairment could be a direct expression of this genetic alteration at the central nervous system (CNS) level rather than a consequence of the neuromuscular impairment. To explore this hypothesis, we tested a group of genetically diagnosed, adult onset DM, of their nonaffected relatives (NAR), of patients with spinal muscle atrophy (SMA), and of normal controls using the Wechsler Adult Intelligence Scale (WAIS). METHODS: Seventeen adult-onset DM patients, 9 NAR, 10 SMA patients and 20 unrelated normal controls (NC) were studied. Clinical, neuromuscular and neuropsychiatric evaluation, which included WAIS and the Schedule for Affective Disorders and Schizophrenia (SADS), were performed on the four groups. DM, NAR and NC were also assessed by a neurophysiological (P300) evaluation. A DNA analysis was performed in DM and in NAR to measure presence and magnitude of CTG expansion. RESULTS: We found a statistically significant difference between verbal (p < .0003), nonverbal (p < .0001) and total (p < .0001) IQ of DM patients compared to IQs of NAR, SMA and NC. Seven out of 11 WAIS subtests were significantly and consistently lower in DM patients compared to SMA and/or NC. In DM patients there was a statistically significant negative correlation between nonverbal (r = -.68; p < .002) and total (r = .59; p < .01) IQ and (CTG)n. Patients with DM had a significantly lower P300 amplitude compared to NAR and NC. CONCLUSIONS: Our study indicates that in DM there is a mild but significant cognitive impairment which correlates with the degree of CTG expansion and it is not dependent on the neuromuscular impairment; however further studies with larger groups of patients and controls are suggested to confirm our results, due to the small sample size and to a possible effect of educational level in our patients.

Adolescent↗
PubMed1999

Alteration of the PTEN/MMAC1 gene locus in primary lung cancer with distant metastasis.

The PTEN/MMAC1 gene located at 10q23, has been proposed to be a tumor suppressor gene. To determine the involvement of alteration of the PTEN/MMAC1 gene in carcinogenesis and the progression of primary lung cancers, we analyzed tumor samples of primary and distant metastatic sites and normal lung tissue samples of 30 patients with advanced lung cancer with distant metastasis. The tissues were analyzed for allelic deletion and mutational inactivation of PTEN/MMAC1 by loss of heterozygosity (LOH) analysis, polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP), and direct sequence analysis. LOH of the PTEN/MMAC1 locus was common in each histologic type of primary lung cancer. In this study, the overall allelic deletion rate was 33.3% (7/21). Allelic loss at the primary site and that at the metastatic site of each patient, were identical; in most cases, it seemed that the allelic loss had occurred before metastasis. Sequence analysis of the PTEN/MMAC1 gene revealed a G to C substitution located 8 bp upstream of the coding region of exon 1 and which seems to be a polymorphism, in 4 of the 30 cases. Somatic mutations of the PTEN/MMAC1 gene were not identified in any of the tumors at the primary and metastatic sites. These data indicate that point mutations in the PTEN/MMAC1 gene are probably not an important factor in tumorigenesis and the progression of a major subset of lung cancers. Due to frequent allelic loss at the PTEN/MMAC1 locus occurring at a stage earlier than the metastatic process, alternative mechanisms in which the remaining allele is inactivated such as methylation or homozygous deletion of a small region of the gene that can not be detected by the usual analysis, or alteration of other important tumor suppressor genes lying close to the PTEN/MMAC1 gene on 10q23, may be involved in the tumorigenesis of lung cancers of all histologic subtypes.

Alleles↗
PubMed1998

Allelic deletion analysis of the FHIT gene predicts poor survival in non-small cell lung cancer.

The fragile histidine triad (FHIT) gene at chromosome 3p14.2 is a candidate tumor suppressor gene linked to cancers of the lung, breast, colon, pancreas, and head and neck. Reports of frequent allelic deletion and abnormal transcripts in primary lung tumors plus recent evidence that it is targeted by tobacco smoke carcinogens suggest that it plays an important role in lung carcinogenesis. Non-small cell lung carcinoma still maintains a poor 5-year survival rate with the stage of disease at presentation as a major determinant of prognosis. We examined for allelic deletion at the FHIT locus in a series of 106 non-small cell lung carcinomas for which a full clinical, epidemiological, and 5-year survival profile was available. We found an allelic deletion frequency of 38% at one or two intragenic microsatellites. Allelic deletion of FHIT was related to tumor histology with 4 of 20 adenocarcinomas (20%) displaying loss of heterozygosity (LOH) compared with 12 of 22 (55%) nonadenocarcinomas (P = 0.03). We found that 63% of tumors with LOH of FHIT also had p53 missense mutations whereas only 26% with LOH had wild type p53 negative sequence (P = 0.02). We also found a significant trend toward poorer survival in patients with LOH of at least one locus of the FHIT gene (log rank, P = 0.01). This survival correlation is independent of tumor stage, size, histological subtype, degree of differentiation, and p53 mutation status. Our data support the hypothesis that the loss of the FHIT contributes to the molecular pathogenesis of human lung cancer and is an indicator of poor prognosis.

Acid Anhydride Hydrolases↗
PubMed1998

hSmad5 gene, a human hSmad family member: its full length cDNA, genomic structure, promoter region and mutation analysis in human tumors.

hSmad (mothers against decapentaplegic)-related proteins are important messengers within the Transforming Growth Factor-beta1 (TGF-beta1) superfamily signal transduction pathways. To further characterize a member of this family, we obtained a full length cDNA of the human hSmad5 (hSmad5) gene by rapid amplification of cDNA ends (RACE) and then determined the genomic structure of the gene. There are eight exons and two alternative transcripts; the shorter transcript lacks exon 2. We identified the hSmad5 promoter region from a human genomic YAC clone by obtaining the nucleotide sequence extending 1235 base pairs upstream of the 5' end of the cDNA. We found a CpG island consistent with a promoter region, and we demonstrated promoter activity in a 1232 bp fragment located upstream of the transcription initiation site. To investigate the frequency of somatic hSmad5 mutations in human cancers, we designed intron-based primers to examine coding regions by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis. Neither homozygous deletions or point mutations were found in 40 primary gastric tumors and 51 cell lines derived from diverse types of human cancer including 20 cell lines resistant to the growth inhibitory effects of TGF-beta1. These results suggest that the hSmad5 gene is not commonly mutated and that other genetic alterations mediate the loss of TGF-beta1 responsiveness in human cancers.

Alternative Splicing↗
PubMed1998

Changes in nutritional status during the hospital stay: a predictor of long-term survival.

The objectives of this prospective observational study were to assess whether: 1) midarm circumference (MAC), previously shown to predict in-hospital mortality, maintains its prognostic implication after discharge; 2) in-hospital changes in aspecific indicators of the health status are predictors of long-term survival. The study population consisted of 249 patients from the general community [mean age 80 +/- 7 (70-99) years], consecutively discharged from geriatric and medical wards of an acute care hospital. Changes in health status during hospitalization were recorded (dynamic or delta variables) and health-related variables were collected at discharge (discharge variables). The relationship of both sets of variables to survival over a 3-year period was assessed by Cox's proportional hazards regression analysis. The discriminatory efficacy of predictive models was estimated by the Hanley and McNeil method. Survival curves were drawn with the patients alternatively grouped according to the presence or absence of each of the predictive variables. Serum albumin < 3.5 g/dL (hazard rate = 0.57, 95% confidence limits = 0.33-0.96) and dependency in at least one ADL (h.r. = 0.87, c.l. = 0.79-0.98) were found to be associated with increased mortality, and delta MAC (h.r. = 1.03, c.l. = 1.01-1.05), i.e., there was a positive change or no change in MAC from admission to discharge, with increased survival. A slightly weaker predictive model was obtained using only discharge variables. However, Hanley and McNeil's analysis showed that both models were far from achieving the optimal discrimination of high from low risk subjects. Effects on survival of individual variables varied in magnitude and dependency on time. We concluded that measuring in-hospital changes in nutritional status might improve prediction of long-term survival. Attempts should be made to identify variables having the strongest prognostic implications, and to tailor dynamic assessment to the needs of selected categories of patients.

Activities of Daily Living↗
PubMed1998

Predicting cognitive decline in patients with hypoxaemic COPD.

The objective was to identify predictors of cognitive decline in patients with hypoxaemic COPD on continuous oxygen therapy. Eighty-four consecutive ambulatory hypoxaemic COPD patients in stable clinical conditions were prospectively studied over the course of 2 yr. Baseline multidimensional assessment included respiratory function tests, blood gas analysis, Mini Mental Status (MMS) test, Geriatric Depression Scale (GDS), Activities of Daily Living (ADLs) and Charlson's index of comorbidity. Reassessments were made 1 yr and 2 yr thereafter. Sequential changes in MMS, GDS and ADLs were assessed by Friedman's ANOVA by rank test. Forty patients completed the study (group A), while 44 died or were lost to follow-up (group B). Group B was characterized by more severe respiratory function impairment and worse performances on ADLs and GDS. In group A, MMS deteriorated from baseline to the 1 yr and 2 yr reassessments (27 +/- 2.9 vs. 25.8 +/- 4.1 and 25.4 +/- 4, P < 0.005), whereas GDS and ADLs did not change significantly; the 23 patients experiencing a decline of MMS had baseline lower percentage predicted FVC (52.3 +/- 17.1 vs. 66.9 +/- 13.4, P < 0.03) and FEV1 (27.2 +/- 8.6 vs. 44 +/- 26.8, P < 0.02) values and better affective status (GDS score: 11.9 +/- 7.7 vs. 16.5 +/- 5.6, P < 0.04). Two-year changes in MMS and in GDS scores were inversely correlated (Spearman's p = -0.32, P = 0.04). Cognitive decline is faster in the presence of severe bronchial obstruction and parallels the worsening of the affective status in COPD patients on oxygen therapy. The onset of depression rather than baseline depressive symptoms seems to be a risk factor for cognitive decline.

Activities of Daily Living↗
PubMed1997

Mutation analysis of the transforming growth factor-beta type II receptor in human cell lines resistant to growth inhibition by transforming growth factor-beta.

The transforming growth factor-beta (TGF-beta) binds the type II TGF-beta growth factor receptor (RII) to inhibit the growth of most epithelial tissues. Most human colon and gastric cancers with microsatellite instability (MI) have frameshift mutations in polynucleotide repeats within the RII coding region; these mutations truncate the receptor protein and disable the serine/threonine kinase to produce TGF-beta resistance. To further investigate the type, frequency and tissue distribution of RII mutations, we selected 24 human cancer cell lines from various tissues which were previously reported to be resistant to the inhibitory effects of TGF-beta. We developed protocols for non-isotopic SSCP analysis of PCR products from genomic DNA samples, and we tested them for microsatellite instability. PCR-SSCP analysis followed by DNA sequencing identified deletion mutations in the exon 3 poly-adenine tract in three colon tumor cell lines: LS174T and SW48 had a single base deletion and LS411 had a two base deletion. Among the 24 previously unreported cell lines, only these three demonstrated microsatellite instability. These and other recent data indicate that RII mutations are essentially confined to colon and gastric cancers with microsatellite instability. The narrow spectrum of tissues containing RII mutations illustrates the complexity of genetic checkpoints in human carcinogenesis.

Cell Division↗