RESUMO
Ras activating mutations result in constitutive activation of Ras signalling pathways and occur in 30% of human malignancies. K-ras encodes two splice variants, K-ras 4A and 4B, and K-ras activating mutations which jointly affect both isoforms are prevalent in lung, pancreatic and colorectal cancers. Using RT-PCR we examined their expression in normal adult human tissues and addressed whether K-ras splicing is altered in sporadic colorectal cancer by comparing normal colon with colon carcinoma cell lines, and 'matched' tumour and tumour-free colon tissues from the same patient. K-ras 4B was expressed ubiquitously and was the predominant splice variant. K-ras 4A was expressed differentially, with detection in colorectal tumours and cell lines, and normal colon, pancreas and lung--sites where tumours with K-ras activating mutations arise. Both K-ras splice variants were co-expressed by single colon carcinoma cells. The K-ras 4A/4B ratio was significantly reduced in all 6 cell lines examined, including two that lacked K-ras activating mutations, and in 4/9 primary adenocarcinomas. We conclude that K-ras activating mutations do not affect K-ras splicing per se, both isoforms may play a role in neoplastic progression, and altered splicing of either the K-ras proto-oncogene or oncogene, in favour of K-ras 4B, may modulate tumour development.
Assuntos
Adenocarcinoma/genética , Neoplasias Colorretais/genética , Genes ras/genética , Mutação , Adenocarcinoma/metabolismo , Colo/metabolismo , Neoplasias Colorretais/metabolismo , Análise Mutacional de DNA , Humanos , Isoformas de Proteínas/genética , Proto-Oncogene Mas , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Células Tumorais CultivadasRESUMO
Small cell lung cancer (SCLC) is known to express the HuD protein, the neuronal antigen homologous to Drosophila Elav and Sxl genes involved in neuronal and sex development. HuD is the target of an immune response including high titered antibodies causing paraneoplastic encephalomyelitis and sensory neuropathy. Because the p53 recessive oncogene is mutated and anti-p53 antibodies frequently occur in cancer patients, we wondered if the development of anti-HuD antibodies signaled the presence of HuD mutations in lung cancer. The HuD gene was mapped to chromosome region 1p using a human-mouse hybrid cell panel. We confirmed that 26 of 46 cancer (43 lung cancer and 3 mesothelioma) cell lines expressed HuD mRNA and that this expression, as well as protein expression by Western blot, correlated strongly with the SCLC neuroendocrine phenotype. Southern blot and single-strand conformation polymorphism analyses showed that HuD was not mutated in 78 lung cancers, including patients with the severe paraneoplastic syndrome. Northern blot analysis showed that lung cancer cell lines expressed two major mRNAs (4.3 and 4.0 kilobases) of HuD. We found the three previously described alternative spliced mRNA forms (HuDpro, HuD, and HuDmex). In addition, we also found HuD mRNA had an alternative splicing form in its 5'-coding region. This alternative splice introduced 87 base pairs of sequence and a termination codon resulting in a predicted small, truncated protein (11 amino acids) reminiscent of the male-specific truncated protein in the Sex-lethal (Sxl) gene of Drosophila. However, mRNAs encoding both full-length and truncated proteins were expressed in all SCLCs. These results show that the HuD gene is not mutated in lung cancer, including tumors from patients producing anti-HuD antibodies, but HuD expression is an independent marker or determinant of the neuroendocrine differentiation seen in SCLC.
Assuntos
Antígenos de Neoplasias/genética , Carcinoma de Células Pequenas/imunologia , Cromossomos Humanos Par 1 , Encefalomielite/imunologia , Neoplasias Pulmonares/imunologia , Síndromes Paraneoplásicas/imunologia , Sequência de Bases , Carcinoma de Células Pequenas/genética , Mapeamento Cromossômico , Análise Mutacional de DNA , Encefalomielite/genética , Humanos , Neoplasias Pulmonares/genética , Dados de Sequência Molecular , Síndromes Paraneoplásicas/genética , RNA Mensageiro/genética , Células Tumorais CultivadasRESUMO
We evaluated primary lung cancers, tumor cell lines, and preneoplastic bronchial lesions for molecular genetic abnormalities in the candidate tumor suppressor gene FHIT, which spans the FRA3B fragile site at 3p14.2. 3p14.2 allele loss was very frequent in 32 lung cancer cell lines [100% of small cell lung cancer and 88% of non-small cell lung cancer (NSCLC)] and 108 primary NSCLC cancers (45%), with numerous breakpoints indicating involvement of several distinct regions in the FRA3B site. 3p14 allele loss was least frequent in the adenocarcinoma subtype and occurred at the relatively late carcinoma in situ stage of preneoplastic bronchial lesions found in NSCLC patients. Homozygous deletions within the FHIT/FRA3B region were found in 6 of 135 (4.4%) thoracic cancer cell lines. Northern blot showed low or absent FHIT expression in most thoracic cancer cell lines tested, whereas reverse transcription-PCR showed that 59-62% exhibited aberrant FHIT transcripts but nearly always (93-100%) also expressing the wild-type transcripts. Aberrant transcripts included precise deletions of FHIT exons, insertion of non-FHIT sequences between exons and insertions replacing exons. Complete open reading frame single-strand conformational polymorphism analysis of 102 lung cancer cDNAs revealed only one nonsplicing mutation. Normal cells including bronchial epithelium, lung, and trachea expressed wild-type FHIT transcript and a variant transcript deleted for exon 8 but not the other aberrant transcripts, arguing against exon 8-deleted FHIT transcripts being tumor specific. Our findings support the conclusion that FHIT/FRA3B abnormalities are associated with lung cancer pathogenesis but that FHIT abnormalities differ from the types of mutations and lack of wild-type transcript found in classic tumor suppressor genes, and functional studies are needed to define the role of FHIT in thoracic tumorigenesis.
Assuntos
Hidrolases Anidrido Ácido , Neoplasias Brônquicas/genética , Fragilidade Cromossômica , Neoplasias Pulmonares/genética , Proteínas de Neoplasias , Lesões Pré-Cancerosas/genética , Proteínas/genética , Deleção de Sequência , Alelos , Northern Blotting , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma de Células Pequenas/genética , Sítios Frágeis do Cromossomo , DNA Complementar/metabolismo , Éxons , Humanos , Íntrons , Mutagênese Insercional , Fases de Leitura Aberta , Mutação Puntual , Reação em Cadeia da Polimerase , Polimorfismo Genético , Polimorfismo Conformacional de Fita Simples , Splicing de RNA , Análise de Sequência de DNA , Transcrição Gênica , Células Tumorais CultivadasRESUMO
The beta-catenin gene (CTNNB1) has been shown to be genetically mutated in various human malignancies. To determine whether the beta-catenin gene is responsible for oncogenesis in thoracic malignancies, we searched for the mutation in 166 lung cancers (90 primary tumors and 76 cell lines), one blastoma and 10 malignant mesotheliomas (two primary tumors and eight cell lines). Among the lung cancers, including 43 small cell lung cancers (SCLCs) and 123 non-small cell lung cancers (NSCLCs), we identified four alterations in exon 3, which is the target region of mutation for stabilizing beta-catenin. One primary adenocarcinoma had a somatic mutation from C to G, leading to an amino acid substitution from Ser to Cys at codon 37. Among the cell lines, SCLC NCI-H1092 had a mutation from A to G, leading to an Asp to Gly substitution at codon 6, NSCLC HCC15 had a mutation from C to T, leading to a Ser to Phe substitution at codon 45, and NSCLC NCI-H358 had a mutation from A to G, leading to a Thr to Ala substitution at codon 75. One blastoma also had a somatic mutation from C to G, leading to a Ser to Cys substitution at codon 37. Among the 10 malignant mesotheliomas, we identified a homozygous deletion in the NCI-H28 cell line. Cloning of the rearranged fragment from NCI-H28 indicated that all the exons except exon 1 of the beta-catenin gene are deleted and that the deletion junction is 13 kb downstream from exon 1. Furthermore, Northern blot analysis of 26 lung cancer and eight mesothelioma cell line RNAs detected ubiquitous expression of the beta-catenin messages except NCI-H28, although Western blot analysis showed that relatively less amounts of protein products were expressed in some of lung cancer cell lines. Our findings suggest that the beta-catenin gene is infrequently mutated in lung cancer and that the NCI-H28 homozygous deletion of the beta-catenin gene might indicate the possibility of a new tumor suppressor gene residing in this region at 3p21.3, where various types of human cancers show frequent allelic loss.
Assuntos
Deleção Cromossômica , Cromossomos Humanos Par 3 , Proteínas do Citoesqueleto/genética , Homozigoto , Neoplasias Pulmonares/genética , Mesotelioma/genética , Mutação , Transativadores , Sequência de Bases , DNA de Neoplasias , Éxons , Rearranjo Gênico , Humanos , Dados de Sequência Molecular , beta CateninaRESUMO
The amassing of health information on the Internet and World Wide Web continues unabated. Patients anxious to participate in decisions about their own treatment have turned to the Internet to confirm diagnoses, validate physician-recommended treatment, or seek alternative therapies. While increased information for patients has been linked to improved outcomes, there are inherent dangers associated with the kind of unauthenticated information available on the Web. The authors discuss the nature of these dangers as well as review the advantages for patients of "information therapy" (improved access to health information). They also examine how the Internet has begun to affect the physician-patient relationship, and describe how the Internet and information technology can be effectively used by physicians in patient care. They recommend that the academic health sciences community seize the opportunity to take the lead in ensuring that patients have access to reliable health information, and suggest that "patient informatics" be integrated by academic physicians and educators into the teaching of clinical skills.
Assuntos
Tomada de Decisões , Informática Médica , Participação do Paciente , Relações Médico-Paciente , Centros Médicos Acadêmicos , Competência Clínica , Comunicação , Redes de Comunicação de Computadores , Diagnóstico , Educação Médica , Educação em Saúde , Humanos , Internato e Residência , Liderança , Aplicações da Informática Médica , Assistência ao Paciente , Educação de Pacientes como Assunto , Estudantes de Medicina , Ensino/métodos , TerapêuticaAssuntos
Mobilidade Ocupacional , Instrução por Computador , Educação Médica/métodos , Avaliação de Desempenho Profissional/métodos , Docentes de Medicina , Faculdades de Medicina/organização & administração , Atitude , Coleta de Dados , Eficiência Organizacional , Guias como Assunto , Humanos , Política Organizacional , Editoração , Pesquisa , Estados UnidosRESUMO
MBD4 binds to methylated DNA and acts as a thymine DNA glycosylase in base excision repair. Deficiency of MBD4 in mice enhances mutation at CpG sites and alters apoptosis in response to DNA damage, but does not increase tumorigenesis in mismatch repair-deficient mice. However, in humans, frameshift mutation of MBD4, rather than deletion, is what occurs in up to 43% of microsatellite unstable colon cancers. There is no murine equivalent of this mutation. We now show that recombinant truncated MBD4 (MBD4(tru)) inhibits glycosylase activities of normal MBD4 or Uracil DNA glycosylase in cell-free assays as a dominant negative effect. Furthermore, overexpression of MBD4(tru) in Big Blue (lacI)-transfected, MSI human colorectal carcinoma cells doubled mutation frequency, indicating that the modest dominant negative effect on DNA repair can occur in living cells in short-term experiments. Intriguingly, the whole mutation spectrum was increased, not only at CpG sites, suggesting that truncated MBD4 has a more widespread effect on genomic stability. This demonstration of a dominant negative effect may be of significance in tumour progression and acquisition of drug resistance.
Assuntos
Neoplasias Colorretais/metabolismo , DNA Glicosilases/antagonistas & inibidores , Reparo do DNA/genética , DNA de Neoplasias/metabolismo , Endodesoxirribonucleases/genética , Endodesoxirribonucleases/metabolismo , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , DNA Glicosilases/metabolismo , Metilação de DNA/efeitos dos fármacos , DNA de Neoplasias/efeitos dos fármacos , DNA de Neoplasias/genética , Genes Dominantes/efeitos dos fármacos , Humanos , Mutação , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia , Deleção de Sequência , TransfecçãoRESUMO
The migration of traditional anatomy and pathology wet labs to a digital format requires significant planning and collaboration amongst key faculty, administrative, and technology personnel. In addition to conversion of course materials, the changed format necessitated the refurbishment of a traditional bench lab as a state-of-the art digital classroom, significant IT investment, and ongoing coordination by library, classroom and computer resources personnel. Standardized policies and procedures were developed to guide additional requests for software to support new educational initiatives.
Assuntos
Anatomia/educação , Instrução por Computador , Currículo , Patologia/educação , Instrução por Computador/instrumentação , Instrução por Computador/métodos , Humanos , Internet , Microscopia , Projetos Piloto , Saúde Pública/educação , SoftwareRESUMO
Necrosis in glioblastoma is often associated with high levels of Fas (APO-1), HIF-1alpha and PARP expression. The presence of such molecules suggests a regulative element to cell death within this tissue, which may involve p53. We aimed to establish whether p53 and its downstream targets Bax, MDM2 and p21 play a role in perinecrotic cell death in glioblastoma. Following sequencing of the p53 gene in U87 and U373 glioma cell lines, p53 was found to be reactive in the p53 wild-type line U87 in response to hypoxia but not in the p53 mutant line, U373. Although no increase in perinecrotic p53 expression was detected in spheroid cultures derived from these lines, a 60 kDa MDM2 isoform lacking a C-terminal domain showed perinecrotic localization, irrespective of p53 status. Similar findings were observed surrounding regions of necrosis in 80% of glioblastoma biopsies examined. Increasing levels of wild-type p53 did not affect cell death in U87 spheroid cultures but killed all U373 cells 3 days post transfection. Dominant negative p53 did not affect cell death in U373 and U87 spheroid cultures. Although p53 accumulation appeared not to be important for the onset of cell death both in spheroid and biopsy cases, high levels of perinecrotic 60 kDa MDM2 may have implications for glioma cell death susceptibility in both p53 mutant and wild-type tumour cell populations.
Assuntos
Neoplasias Encefálicas/metabolismo , Glioblastoma/metabolismo , Glioma/metabolismo , Isoenzimas/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Esferoides Celulares/metabolismo , Apoptose/fisiologia , Western Blotting , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Expressão Gênica , Genes p53/fisiologia , Glioblastoma/patologia , Glioma/patologia , Humanos , Imuno-Histoquímica , Necrose/fisiopatologia , Estresse Oxidativo/fisiologia , Reação em Cadeia da Polimerase , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Proteínas Proto-Oncogênicas c-mdm2 , Proteínas Proto-Oncogênicas p21(ras)/biossíntese , Transfecção , Proteína X Associada a bcl-2RESUMO
A generalized perception exists that faculty will not be properly rewarded for efforts in developing computer-based educational materials. Faculty governed by traditional promotion and tenure systems thus may be reluctant to devote energies towards development of these materials. Recent national panels on educational reform have called for a reexamination of academic reward structures to insure that faculty receive appropriate scholarly recognition for materials developed in these new formats. A study of policy documents from accredited medical colleges in the United States was conducted to determine the extent to which academic health science institutions have adopted policies to grant recognition of computer-based materials equivalent to that accorded traditional print publications. Results revealed that while some progress has been made by leading-edge institutions, in three-quarters of the institutions, development of computer-based educational materials is considered evidence in support of teaching, not the more highly rewarded research or scholarly activity.
Assuntos
Instrução por Computador , Avaliação de Desempenho Profissional , Editoração , Faculdades de Medicina/organização & administração , Docentes de Medicina , Humanos , Pesquisa , Ensino , Estados UnidosRESUMO
Popularity of the personal computer has prompted physician interest in direct access to the medical literature from sophisticated electronic data retrieval systems. This paper describes the educational programs developed by the library at the George Washington University Medical Center in response to this interest and growing national trend. As a result of these programs, the role of the librarian has changed permanently. Librarians are now perceived by faculty as educational specialists and information consultants. The evolution in libraries parallels the technological changes taking place in other biomedical communications units. The authors recommend the positive outcomes from adapting traditional roles to meet user's needs in a changing environment.
Assuntos
Informática Médica/educação , Médicos , HumanosRESUMO
RT112 cells, a line derived from a human bladder carcinoma, produce epitectin at very low levels in standard culture media; but production and secretion of this mucin are greatly increased when the cells are exposed to hyperosmotic conditions. It appears that hyperosmolarity, by inducing an increase in intracellular sodium, entrains an increase in intracellular free calcium. Evidence is presented for the view that it is the increase in intracellular free calcium that provides the more direct stimulus for the enhanced production of epitectin.
Assuntos
Antígenos de Neoplasias/metabolismo , Células Tumorais Cultivadas/metabolismo , Amilorida/farmacologia , Antígenos Glicosídicos Associados a Tumores , Cálcio/metabolismo , Linhagem Celular , Humanos , Técnicas Imunoenzimáticas , Líquido Intracelular/metabolismo , Lactatos/farmacologia , Nigericina/farmacologia , Concentração Osmolar , Cloreto de Sódio/farmacologia , Sacarose/farmacologia , Neoplasias da Bexiga Urinária/metabolismoRESUMO
Organizations can no longer rely on a single management solution as they respond to a complex world. Biomedical communications managers need to employ a variety of management tools as they cope with a rapidly changing landscape and an increasing set of business pressures. This article describes two management strategies, strategic planning and total quality management, which can be used to achieve success in the 21st century.
Assuntos
Recursos Audiovisuais , Comunicação , Serviços de Informação/organização & administração , Gestão da Qualidade Total , Humanos , Técnicas de PlanejamentoRESUMO
MBD3: is a member of the methyl-CpG-binding domain family and is located on chromosome 19p13.3, a region of loss of heterozygosity in colon and lung cancers. We therefore screened samples for abnormalities in MBD3. Our results indicate that MBD3 is not a major target of genetic and epigenetic alteration in these cancers.
Assuntos
Neoplasias do Colo/genética , Proteínas de Ligação a DNA/genética , Neoplasias Pulmonares/genética , Northern Blotting , Análise Mutacional de DNA , Proteínas de Ligação a DNA/análise , Epigênese Genética , Humanos , Perda de Heterozigosidade , Reação em Cadeia da Polimerase , Células Tumorais CultivadasRESUMO
A meiosis-activated myelin basic protein (MBP) kinase was purified approximately 8700-fold from soluble post-germinal vesicle breakdown extracts from maturing oocytes of the sea star Pisaster ochraceus. Purification to apparent homogeneity was achieved by sequential chromatography on DEAE-cellulose, hydroxylapatite, phosphocellulose, phenyl-Sepharose, heparin-Sepharose, polylysine-Sepharose, and Mono-Q. The final product exhibited an apparent molecular mass of approximately 42 kDa by both native gradient and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and this precisely correlated with the chromatographic behavior of the recovered MBP kinase activity on a Superose 6/12 column. The kinase utilized the MBP as the major substrate with little or no phosphorylation of histones (H1, H2A, or H2B), casein, phosvitin, protamine, or 40 S ribosomal proteins. The purified enzyme was relatively insensitive to high concentrations of beta-glycerol phosphate, calmodulin, EGTA, NaCl, sodium fluoride, dithiothreitol, spermine, and heparin but was quite sensitive to inhibition by metal ions such as Mn2+, Zn2+, and Ca2+. The true Km values for ATP and myelin basic protein were determined to be 58 and 25 microM, respectively, using double-reciprocal plots. The purified enzyme was unable to utilize GTP in place of ATP. The enzyme was shown to rapidly undergo autophosphorylation. The autophosphorylation was sensitive to alkali treatment implying that phosphate was incorporated on serine/threonine residues. The properties of this MBP kinase are reminiscent of a protein kinase that is also activated in a cyclic fashion at M-phase during the early cell divisions of sea star and sea urchin embryos (Pelech, S. L., Tombe, R., Meijer, L., and Krebs, E. G. (1988) Dev. Biol. 130, 26-36).
Assuntos
Meiose , Oócitos/enzimologia , Proteínas Quinases/isolamento & purificação , Estrelas-do-Mar , Trifosfato de Adenosina/metabolismo , Animais , Cálcio/farmacologia , Cátions Bivalentes , Cromatografia , Ativação Enzimática , Feminino , Quinase 3 da Glicogênio Sintase , Cinética , Manganês/farmacologia , Peso Molecular , Oócitos/fisiologia , Fosforilação , Inibidores de Proteínas Quinases , Proteínas Quinases/metabolismo , Especificidade por Substrato , Zinco/farmacologiaRESUMO
The development of human neuroblastoma is associated with abnormalities of the short arm of chromosome 1 (1p). To determine the importance of sequences on that part of chromosome 1, we transferred translocated chromosomes containing normal portions of chromosome 1p or 1q into the neuroblastoma cell line NGP.1A.TR1 and also normal intact chromosomes 11 and 17 as putative controls. Transfer of chromosome t(X;1q) had no effect on any property we studied of the neuroblastoma cells. It was found that the transfer of chromosome t(X;1p) induced neuronal differentiation, but most of those cells died. The cell lines that grew out from the survivors lacked the t(X;1p) chromosome and were still tumorigenic. Transfer of chromosome 11 induced differentiation but did not affect cell proliferation or the tumorigenic phenotype. We have also demonstrated the existence of a new tumor suppressor gene on chromosome 17 that does not induce differentiation in vitro but completely suppresses the tumor-forming ability of the neuroblastoma cells. Differentiation of neuroblastoma cells in vitro and suppression of tumorigenicity are therefore controlled by multiple genes on different chromosomes and are not necessarily correlated.
Assuntos
Cromossomos Humanos Par 1 , Neuroblastoma/patologia , Animais , Diferenciação Celular , Fusão Celular , Mapeamento Cromossômico , Genes Supressores de Tumor , Marcadores Genéticos , Humanos , Técnicas In Vitro , Camundongos , Camundongos Nus , Transplante de Neoplasias , Neuroblastoma/genética , Polimorfismo de Fragmento de Restrição , Transfecção , Células Tumorais CultivadasRESUMO
CD44 is a widely expressed cell adhesion molecule that binds the extracellular matrix component, hyaluronan, in a tightly regulated manner. Previous studies have shown that the CD44-hyaluronan interaction is affected by changes in the glycosylation state of CD44. In this study, we take advantage of several well-characterized murine L cell mutants defective in heparan sulfate synthesis (gro2C cells), heparan sulfate and chondroitin sulfate synthesis (sog9 cells), and glycosaminoglycan and oligosaccharide processing (sog8 cells) to assess the effects of these defects on the hyaluronan binding ability of CD44. In parental L cells and gro2C cells, CD44 was induced to bind hyaluronan after addition of the activating, anti-CD44 monoclonal antibody, IRAWB 14. By contrast, no inducible binding was observed in sog9 cells. Treatment of L cells with sodium chlorate, an inhibitor of sulfation, also abolished inducible hyaluronan binding. However, inducible and some constitutive hyaluronan binding was observed in sog8 cells. This indicates that sulfation and, in particular, the addition of chondroitin sulfate are required for inducible hyaluronan binding by CD44 in L cells. However, in the absence of fully processed oligosaccharides, chondroitin sulfate is not essential for hyaluronan binding, indicating that the effect of chondroitin sulfate is dependent upon the glycosylation state of the cell. Thus, in addition to glycosylation, chondroitin sulfate biosynthesis is an important post-translational modification that can affect the hyaluronan binding ability of CD44.
Assuntos
Sulfatos de Condroitina/fisiologia , Glicosaminoglicanos/biossíntese , Glicosaminoglicanos/deficiência , Receptores de Hialuronatos/metabolismo , Ácido Hialurônico/metabolismo , Células 3T3 , Animais , Anticorpos Monoclonais/metabolismo , Sítios de Ligação de Anticorpos/genética , Glicosaminoglicanos/genética , Glicosilação , Receptores de Hialuronatos/imunologia , Células L , Camundongos , Mutação , Ligação Proteica/genéticaRESUMO
A survey of automated records management in the United States and Canada was developed to identify existing on-line library systems and technical expertise. Follow-up interviews were conducted with ten libraries. Tables compare the features and availability of four main frame and four minicomputer systems. Results showed: a trend toward vendor-supplied systems; little coordination of efforts among schools; current system developments generally on a universitywide basis; and the importance of having the cooperation of campus computer facilities to the success of automation efforts.
Assuntos
Computadores , Bibliotecas Médicas , Serviços Técnicos de Biblioteca , Automação , Minicomputadores , Estados Unidos , UniversidadesRESUMO
Chromosomal or allelic losses at 3p14 are common in a variety of human tumors, including those of the lung, breast, kidney, and head and neck. This suggests the existence of a tumor suppressor gene in this band. A promising candidate is the recently cloned FHIT gene, which spans the common fragile site, FRA3B, at 3p14.2. We previously identified a region of fragility at 3p14.2 (FRA3B) of > 85 kb by cloning DNA flanking pSV2neo integrations and constructed a partial genomic contig of the region. Using probes from the contig, we tested for deletions within this region in DNA from 105 human tumor cell lines, predominantly derived from lung cancers. We identified one gastric and four lung cancer cell lines with homozygous interstitial deletions involving the FRA3B region. The deletion in one lung cancer cell line lies entirely within our contig and is < 65 kb. We have identified, cloned, and sequenced this breakpoint junction. We have also shown that our probes lie within intron S of the FHIT gene and, furthermore, that exon 5 is located approximately 1 kb from one of our probes and, thus, lies within the region of fragility. Two lines with entirely intronic deletions yield FHIT transcripts of normal size. In one of these, this was the sole transcript identified. In the other line, an FHIT transcript completely normal in sequence was accompanied by two larger abnormal transcripts. These results leave open the possibility that some homozygous deletions within the FHIT gene are without phenotypic effect and result from genetic instability of this region. However, taken together, our results provide evidence that breakage and rearrangement within the FRA3B fragile site sequences result in alterations of FHIT and are likely to be involved in carcinogenesis.