RESUMO
The gene encoding PTPROt (truncated isoform of protein tyrosine phosphatase receptor-type O) is methylated and suppressed in chronic lymphocytic leukemia (CLL). PTPROt exhibits in vitro tumor-suppressor characteristics through the regulation of B-cell receptor (BCR) signaling. Here we generated transgenic (Tg) mice with B-cell-specific expression of PTPROt. Although lymphocyte development is normal in these mice, crossing them with TCL1 Tg mouse model of CLL results in a survival advantage compared with the TCL1 Tg mice. Gene expression profiling of splenic B-lymphocytes before detectable signs of CLL followed by Ingenuity Pathway Analysis revealed that the most prominently regulated functions in TCL1 Tg vs non-transgenic (NTg) and TCL1 Tg vs PTPROt/TCL1 double Tg are the same and also biologically relevant to this study. Further, enhanced expression of the chemokine Ccl3, the oncogenic transcription factor Foxm1 and its targets in TCL1 Tg mice were significantly suppressed in the double Tg mice, suggesting a protective function of PTPROt against leukemogenesis. This study also showed that PTPROt-mediated regulation of Foxm1 involves activation of p53, a transcriptional repressor of Foxm1, which is facilitated through suppression of BCR signaling. These results establish the in vivo tumor-suppressive function of PTPROt and identify p53/Foxm1 axis as a key downstream effect of PTPROt-mediated suppression of BCR signaling.
Assuntos
Biomarcadores Tumorais/metabolismo , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/metabolismo , Inflamação/prevenção & controle , Leucemia Linfocítica Crônica de Células B/prevenção & controle , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/fisiologia , Proteína Supressora de Tumor p53/metabolismo , Animais , Linfócitos B/metabolismo , Linfócitos B/patologia , Biomarcadores Tumorais/genética , Western Blotting , Células Cultivadas , Feminino , Citometria de Fluxo , Proteína Forkhead Box M1 , Fatores de Transcrição Forkhead/genética , Perfilação da Expressão Gênica , Regulação Leucêmica da Expressão Gênica , Humanos , Inflamação/genética , Inflamação/patologia , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Proteínas Proto-Oncogênicas/fisiologia , Proteínas Proto-Oncogênicas c-bcr/genética , Proteínas Proto-Oncogênicas c-bcr/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Proteína Supressora de Tumor p53/genéticaRESUMO
T-cell acute lymphoblastic leukemia (T-ALL) is a neoplasia of thymocytes characterized by the rapid accumulation of the precursors of T lymphocytes. HMGA2 (high-mobility group AT-hook 2) gene expression is extremely low in normal adult tissues, but it is overexpressed in many tumors. To identify the biological function of HMGA2, we generated transgenic mice carrying the human HMGA2 gene under control of the VH promoter/Eµ enhancer. Approximately 90% of Eµ-HMGA2 transgenic mice became visibly sick between 4 and 8 months due to the onset and progression of a T-ALL-like disease. Characteristic features included severe alopecia (30% of mice); enlarged lymph nodes and spleen; and profound immunological abnormalities (altered cytokine levels, hypoimmunoglobulinemia) leading to reduced immune responsiveness. Immunophenotyping showed accumulation of CD5+CD4+, CD5+CD8+ or CD5+CD8+CD4+ T-cell populations in the spleens and bone marrow of sick animals. These findings show that HMGA2-driven leukemia in mice closely resembles spontaneous human T-ALL, indicating that HMGA2 transgenic mice should serve as an important model for investigating basic mechanisms and potential new therapies of relevance to human T-ALL.
Assuntos
Proteína HMGA2/biossíntese , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Animais , Proliferação de Células/fisiologia , Feminino , Proteína HMGA2/genética , Proteína HMGA2/metabolismo , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Fenótipo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologiaRESUMO
Breast cancer is the leading cause of cancer-related death in women worldwide. Expression of the WWOX tumor suppressor is absent or reduced in a large proportion of breast tumors suggesting that loss of WWOX may contribute to breast tumorigenesis. Wwox-deficient mice die by 3-4 weeks of age precluding adult tumor analysis. To evaluate the effect of WWOX-altered expression on mammary tumor formation, the Wwox-heterozygous allele was back crossed onto the C3H mammary tumor-susceptible genetic background (Wwox(C3H)+/-) and incidence of mammary tumor formation was evaluated. Although 50% of the female Wwox(C3H)+/- mice developed mammary carcinomas, only 7% of Wwox(C3H)+/+ mice did. Intriguingly, mammary tumors in Wwox(C3H)+/- mice frequently lost WWOX protein expression suggesting a genetic predisposition toward mammary tumorigenesis. Immunohistochemical staining of hormone receptors revealed loss of estrogen receptor-α (ER) and progesterone receptor in the majority of these tumors. In vitro, depletion of WWOX in MCF7 ER-positive cells led to reduced ER expression and reduced sensitivity to tamoxifen and estrogen treatment and was associated with enhanced survival and anchorage-independent growth. Finally, cDNA array analyses of murine normal mammary epithelial cells and mammary tumors identified 163 significantly downreguated and 129 upregulated genes in the tumors. The majority of differentially expressed genes were part of pathways involved in cellular movement, cell-to-cell signaling and interaction, cellular development, cellular growth and proliferation and cell death. These changes in gene expression of mouse mammary tumors in Wwox(C3H)+/- mice resemble, at least in part, human breast cancer development. Our findings demonstrate the critical role that the WWOX tumor suppressor gene has in preventing tumorigenesis in breast cancer.
Assuntos
Neoplasias Mamárias Animais/metabolismo , Oxirredutases/metabolismo , Alelos , Animais , Biomarcadores Tumorais/metabolismo , Morte Celular , Linhagem Celular Tumoral , Cruzamentos Genéticos , Feminino , Humanos , Imuno-Histoquímica/métodos , Camundongos , Camundongos Endogâmicos C3H , Camundongos Transgênicos , Receptores de Estrogênio/metabolismo , Transdução de Sinais , Oxidorredutase com Domínios WWRESUMO
B-cell chronic lymphocytic leukemia (B-CLL) is the most common adult leukemia. Deregulation of the T-cell leukemia/lymphoma 1 (TCL1) oncogene in mouse B cells causes a CD5-positive leukemia similar to aggressive human B-CLLs. We recently reported that levels of TCL1 expression in B-CLL are regulated by miR-29 and miR-181 that target 3' UTR of TCL1. To determine whether treatment with microRNAs targeting TCL1 can inhibit B-CLL in mice, we generated TCL1 transgenic mice using a construct containing the 3' and 5' UTRs of TCL1 under B-cell-specific Emicro promoter (Emicro-TCL1FL). At the age of 16-20 months, these mice showed B-CLL-like disease. Immunophenotyping revealed accumulation of CD5+CD23+B220+ population in spleens and lymph nodes. Our results show that CD5+CD23+ B-cell populations from Emicro-TCL1FL mice actively proliferate and show significantly increased levels of phospho-Akt. Emicro-TCL1FL mice showed immunological abnormalities similar to human B-CLL, including hypoimmunoglobulinemia, abnormal levels of cytokines and impaired immune response. These findings revealed biochemical and immunological similarities between Tcl1-driven B-CLL in mice and human B-CLL.
Assuntos
Antígenos CD5/metabolismo , Proliferação de Células , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Linfocítica Crônica de Células B/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas/fisiologia , Receptores de IgE/metabolismo , Regiões 3' não Traduzidas/genética , Regiões 5' não Traduzidas/genética , Animais , Western Blotting , Citocinas/metabolismo , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Humanos , Imunização , Leucemia Linfocítica Crônica de Células B/imunologia , Linfonodos/metabolismo , Linfonodos/patologia , Camundongos , Camundongos Transgênicos , MicroRNAs/fisiologia , Fosforilação , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Baço/metabolismo , Baço/patologiaRESUMO
FHIT, at a constitutively active chromosome fragile site, is often a target of chromosomal aberrations and deletion in a large fraction of human tumors. Inactivation of murine Fhit allelessignificantly increases susceptibility of mice to spontaneous and carcinogen-induced tumorigenesis. In this study, transgenic mice, carrying a human FHIT cDNA under control of the endogenous promoter, were produced to determine the effect of Fhit expression, from a nonfragile cDNA transgene outside the fragile region, on carcinogen-induced tumor susceptibility of wildtype and Fhit heterozygous mice. Mice received sufficient oral doses of N-nitrosomethybenzylamine (NMBA) to cause forestomach tumors in >80% of nontransgenic control mice. Although the level of expression of the FHIT transgene in the recombinant mouse strains was much lower than the level of endogenous Fhit expression, the tumor burden in NMBA-treated male transgenic mice was significantly reduced, while female transgenic mice were not protected. To determine if the difference in protection could be due to differences in epigenetic changes at the transgene loci in male versus female mice, we examined expression, hypermethylation and induced re-expression of FHIT transgenes in male and female mice or cells derived from them. The transgene was methylated in male and female mice and in cell lines established from male and female transgenic kidneys, the FHIT locus was both hypermethylated and deacetylated. It is likely that the FHIT transgene is more tightly silenced in female transgenic mice, leading to a lack of protection from tumor induction.
Assuntos
Hidrolases Anidrido Ácido/genética , Predisposição Genética para Doença , Proteínas de Neoplasias/genética , Neoplasias Experimentais/genética , Transgenes , Animais , Sequência de Bases , Western Blotting , Carcinógenos/toxicidade , Metilação de DNA , Metilases de Modificação do DNA/antagonistas & inibidores , Primers do DNA , Feminino , Inibidores de Histona Desacetilases , Hibridização in Situ Fluorescente , Masculino , Camundongos , Camundongos Transgênicos , Neoplasias Experimentais/induzido quimicamente , Regiões Promotoras Genéticas , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Five years ago the fragile histidine triad (FHIT) gene including the most common fragile site locus of the human genome, FRA3B, was identified. The gene is altered in many types of cancer and several data support the idea that FHIT has to be considered a tumor suppressor. FHIT abnormalities were investigated in some skin tumors. Fifty-seven per cent of Merkel cell carcinomas displayed abnormal FHIT products but the involvement of FHIT in human non-melanoma skin cancer is still unclear. Because the murine Fhit locus is similar to its human homologue and is altered in cancer cell lines, we have established a strain of Fhit-deficient mice. After N-nitrosomethylbenzylamine treatment, the spectrum of tumors developed by the Fhit-deficient mice was similar to those observed in a familial skin cancer condition, the Muir-Torre syndrome, although there is no clear evidence yet for a relationship of FHIT and the human syndrome. Because cancer cells lacking in FHIT are defective in apoptosis, we propose the Fhit-deficient mouse as a model to understand a possible proapoptotic mechanism deficiency in the human syndrome.
Assuntos
Hidrolases Anidrido Ácido , Proteínas de Neoplasias , Proteínas/genética , Neoplasias Cutâneas/genética , Animais , Apoptose/genética , Modelos Animais de Doenças , Genes Supressores de Tumor/genética , Humanos , Neoplasias Cutâneas/patologiaRESUMO
Mice carrying one inactivated Fhit allele (Fhit +/- mice) are highly susceptible to tumor induction by N-nitrosomethylbenzylamine, with 100% of Fhit +/- mice exhibiting tumors of the forestomach/squamocolumnar junction vs. 25% of Fhit +/+ controls. In the current study a single N-nitrosomethylbenzylamine dose was administered to Fhit +/+, +/-, and -/- mice to compare carcinogen susceptibility in +/- and -/- Fhit-deficient mice. At 29 weeks after treatment, 7.7% of wild-type mice had tumors. Of the Fhit -/- mice 89.5% exhibited tumors (average 3.3 tumors/mouse) of the forestomach and squamocolumnar junction; half of the -/- mice had medium (2 mm diameter) to large (>2 mm) tumors. Of the Fhit +/- mice 78% exhibited tumors (average 2.4 tumors/mouse) and 22% showed medium to large tumors. Untreated Fhit-deficient mice have been observed for up to 2 years for spontaneous tumors. Fhit +/- mice (average age 21 mo) exhibit an average of 0.94 tumors of different types; Fhit -/- mice (average age 16 mo) also showed an array of tumors (average 0.76 tumor/mouse). The similar spontaneous and induced tumor spectra observed in mice with one or both Fhit alleles inactivated suggests that Fhit may be a one-hit tumor suppressor gene in some tissues.
Assuntos
Hidrolases Anidrido Ácido , Proteínas de Neoplasias , Neoplasias Experimentais/genética , Proteínas/genética , Animais , Carcinógenos/toxicidade , Dimetilnitrosamina/análogos & derivados , Dimetilnitrosamina/toxicidade , Feminino , Genótipo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neoplasias Experimentais/induzido quimicamente , Neoplasias Experimentais/patologia , Fenótipo , Neoplasias Gástricas/induzido quimicamente , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologiaRESUMO
PURPOSE: The tumor suppressor gene, FHIT, has been cloned and mapped at chromosome region 3p14.2, one of the regions most frequently deleted in cervical carcinoma. In this report, we show that the expression of the Fhit protein in relation to human papillomavirus (HPV) subtype, the type of the intraepithelial lesion, HIV-induced immunodeficiency, and the DNA content (ploidy) correlates with the biological behavior of the lesions. EXPERIMENTAL DESIGN: To investigate involvement of the FHIT gene in squamous intraepithelial lesions of low and high grade (LGSILs and HGSILs, respectively) of the uterine cervix, we examined the Fhit protein expression by immunocytochemistry in 131 cervical smears of 96 HIV-seropositive patients (42 with LGSILs and 10 with HGSILs) and 35 HIV-seronegative (5 with LGSILs) persons. RESULTS: Fhit protein was detected in normal cells, whereas dysplastic cells (independently of HPV infection and HPV subtypes) showed reduced expression of Fhit (P = 0.00001). Lesions from 52 HIV-seropositive patients, 42 LGSILs and 10 HGSILs, showed diploid DNA content in 63.5%, aneuploid in 32.7%, and polyploid in 3.8%, but 90% of the HGSILs showed an aneuploid DNA content, and all were infected by HPV 16/18 subtypes. 23.8% of LGSIL cases were associated with HPV 16. CONCLUSIONS: These data clearly suggest that loss of Fhit expression occurs in the early stages of cervical carcinogenesis. Pap test represents one of the most convenient and rapid procedures available in identification of cellular changes; hence, Fhit staining might be used as an useful tool in larger population screening to detect early alteration in cellular behaviors.
Assuntos
Hidrolases Anidrido Ácido , Genes Supressores de Tumor/fisiologia , Proteínas de Neoplasias , Papillomaviridae/fisiologia , Ploidias , Proteínas/genética , Displasia do Colo do Útero/genética , Adulto , Feminino , Inativação Gênica , Soronegatividade para HIV , Soropositividade para HIV , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , Papillomaviridae/isolamento & purificação , Proteínas/metabolismo , Displasia do Colo do Útero/virologiaRESUMO
Car-R and Car-S outbred mouse lines, phenotypically selected for resistance and susceptibility to skin carcinogenesis respectively, show significant linkage disequilibrium (LD) at genetic markers mapping on chromosomal regions where skin cancer modifier loci (Skts3, Skts1, and Psl1 on Chrs 5, 7, and 9 respectively) have been mapped in standard crosses. Analysis of these regions for genetic linkage with skin cancer phenotypes in 245 (Car-R x Car-S)F2 intercross mice, by using single nucleotide polymorphisms (SNPs), revealed significant linkage at a possible allelic form of the Skts1 locus, whose mapping region was shortened to a <5.5-cM interval near the Tyr locus. The Car-derived Skts1 locus was linked with papilloma multiplicity and latency by a recessive inheritance of the susceptibility allele. Putative loci on Chr 5 (Skts3) and 9 (Psl1) showed no significant linkage. These results point to the important role of the Stks1 locus in mouse skin tumorigenesis in independent crosses. The shortened Skts1 mapping region should facilitate the identification of candidate genes.
Assuntos
Desequilíbrio de Ligação , Polimorfismo de Nucleotídeo Único/genética , Neoplasias Cutâneas/genética , Animais , Animais não Endogâmicos , Cruzamentos Genéticos , Modelos Animais de Doenças , Marcadores Genéticos , Predisposição Genética para Doença , Haplótipos/genética , Heterozigoto , Camundongos , Camundongos Endogâmicos , Repetições de MicrossatélitesRESUMO
The tumor suppressor gene FHIT spans a common fragile site and is highly susceptible to environmental carcinogens. FHIT inactivation and loss of expression is found in a large fraction of premaligant and malignant lesions. In this study, we were able to inhibit tumor development by oral gene transfer, using adenoviral or adenoassociated viral vectors expressing the human FHIT gene, in heterozygous Fhit(+/-) knockout mice, that are prone to tumor development after carcinogen exposure. We therefore suggest that FHIT gene therapy could be a novel clinical approach not only in treatment of early stages of cancer, but also in prevention of human cancer.
Assuntos
Hidrolases Anidrido Ácido , Dimetilnitrosamina/análogos & derivados , Genes Supressores de Tumor/fisiologia , Terapia Genética/métodos , Proteínas de Neoplasias/genética , Neoplasias Experimentais/prevenção & controle , Adenoviridae/genética , Animais , Carcinógenos/efeitos adversos , Dependovirus/genética , Dimetilnitrosamina/efeitos adversos , Vetores Genéticos/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas de Neoplasias/metabolismo , Proteínas de Neoplasias/fisiologia , Neoplasias Experimentais/induzido quimicamente , Neoplasias Experimentais/patologiaRESUMO
Transgenic mice carrying the RET oncogene under the control of the metallothionein promoter exhibit severe pigmentation of the whole skin and melanocytic tumors. The genetic background influences melanoma development in RET mice; founder mice crossed with BALB/c mice show decreased incidence and increased latency of melanocytic tumors, whereas progeny of C57BL/6 mice show the opposite effect. Using partially congenic RET mice on a C57BL/6 genetic background (N3/RET mice), we studied genetic linkage in (N3/RETxBALB/c)xN3/RET backcross mice. We mapped three melanoma modifier loci, on chromosome 1 (Melm1 and Melm2) and chromosome 11 (Melm3), that are linked with early melanoma incidence and latency. Mapping of Melm loci and of five additional regions on chromosomes 6, 8, 9, 12, and 13 indicated allelic imbalance in N3/RET mice, with a significant excess of BALB/c alleles, suggesting the presence of additional putative melanoma modifier loci on these chromosomes.
Assuntos
Proteínas de Drosophila , Ligação Genética , Melanoma Experimental/genética , Proteínas Proto-Oncogênicas/genética , Receptores Proteína Tirosina Quinases/genética , Desequilíbrio Alélico , Animais , Mapeamento Cromossômico , Cruzamentos Genéticos , Feminino , Endogamia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fenótipo , Proteínas Proto-Oncogênicas c-retRESUMO
Car-R (carcinogenesis-resistant) and Car-S (carcinogenesis-susceptible) outbred mice, obtained by phenotypic selection from an initial intercross of eight inbred strains, show a >100-fold difference in their susceptibility to two-stage skin tumorigenesis. We found that the lines carry a high degree of genetic polymorphism. with an average heterozygosity of 0.39. This polymorphism allowed the use of linkage disequilibrium (LD) and haplotype analysis for the mapping of a skin cancer modifier locus on Chr 7, in a short region of 6 cM, around the Tyr gene. Car-S mice inherited the susceptibility allele at this locus from the A/J, BALB/c, SJL/J, and SWR/J strains. Our results demonstrate the feasibility and usefulness of mapping disease genes by LD in phenotypically selected, genetically heterogeneous animals.
Assuntos
Animais não Endogâmicos/genética , Haplótipos/genética , Desequilíbrio de Ligação , Neoplasias Cutâneas/genética , Animais , Cromossomos , Cruzamentos Genéticos , Predisposição Genética para Doença , Heterozigoto , Camundongos , Camundongos Endogâmicos , Repetições de Microssatélites , Neoplasias Experimentais/genéticaRESUMO
To investigate the role of the Fhit gene in carcinogen induction of neoplasia, we have inactivated one Fhit allele in mouse embryonic stem cells and produced (129/SvJ x C57BL/6J) F(1) mice with a Fhit allele inactivated (+/-). Fhit +/+ and +/- mice were treated intragastrically with nitrosomethylbenzylamine and observed for 10 wk posttreatment. A total of 25% of the +/+ mice developed adenoma or papilloma of the forestomach, whereas 100% of the +/- mice developed multiple tumors that were a mixture of adenomas, squamous papillomas, invasive carcinomas of the forestomach, as well as tumors of sebaceous glands. The visceral and sebaceous tumors, which lacked Fhit protein, were similar to those characteristic of Muir-Torre familial cancer syndrome.
Assuntos
Hidrolases Anidrido Ácido , Neoplasias Primárias Múltiplas/genética , Proteínas/genética , Adenoma/genética , Animais , Carcinógenos , Dimetilnitrosamina/análogos & derivados , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Camundongos Knockout , Proteínas de Neoplasias/deficiência , Proteínas de Neoplasias/genética , Neoplasias Primárias Múltiplas/induzido quimicamente , Neoplasias Primárias Múltiplas/patologia , Papiloma/genética , Proteínas/metabolismo , Mapeamento por Restrição , Neoplasias das Glândulas Sebáceas/genética , Neoplasias Gástricas/induzido quimicamente , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , SíndromeRESUMO
We measured the frequency of mutant (MF) lymphocytes at the hprt locus in a population of 43 coke-oven workers exposed to PAH and in a group of 26 non-exposed workers. A non-significant increase in MF in the exposed group (19.0 +/- 16.3) compared to the non-exposed group (15.8 +/- 14.6) was observed. Moreover, when we considered smoking habits for the overall population, the MF values were higher, although not significantly, in smokers than in non-smokers. For some T-cell mutant clone structural alterations, splicing and coding errors were detected by PCR-based methods. We analysed 161 HPRT- clones, derived from exposed and non-exposed workers by multiplex-PCR and 56 HPRT- clones by reverse transcriptase-PCR. Overall, the percentages of the different types of gene alterations were similar in exposed and non-exposed subjects. Only the frequency of splice mutations in mutant clones derived from coke-oven workers was higher (22%) than in non-exposed donors (11%).
Assuntos
Hipoxantina Fosforribosiltransferase/genética , Mutagênicos/efeitos adversos , Mutação , Exposição Ocupacional/efeitos adversos , Hidrocarbonetos Policíclicos Aromáticos/efeitos adversos , Linfócitos T/efeitos dos fármacos , Adulto , Fatores Etários , Coque , Análise Mutacional de DNA , Humanos , Indústrias , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Linfócitos T/enzimologia , Linfócitos T/ultraestruturaRESUMO
Genetic linkage experiments using crosses between mouse inbred strains with an inherited predisposition and resistance to lung cancer make it possible to investigate the genetics of the complex inheritance of susceptibility and resistance to lung cancer. We have previously mapped a major locus (pulmonary adenoma susceptibility 1, Pas1) affecting inherited predisposition to lung cancer in mice onto chromosome 6, near Kras2. Appropriate crosses that include Pas1/+ mice provide a model system for identifying loci that can modify the lung cancer predisposition phenotype caused by Pas1. Using this approach, we mapped the pulmonary adenoma resistance 1 (Par1) locus on to mouse chromosome 11; this locus selectively inhibits lung tumor development in Pas1/+ animals and therefore behaves like a modulator gene of Pas1. More recently, we have mapped lung tumor modifier loci specifically affecting the initiation and progression of lung cancer. Thus experimental models provide an essential tool for the mapping of lung cancer susceptibility/resistance genes and for the subsequent cloning of candidate genes.
Assuntos
Adenoma/genética , Mapeamento Cromossômico , Predisposição Genética para Doença , Neoplasias Pulmonares/genética , Animais , Ligação Genética , Humanos , CamundongosRESUMO
Mouse inbred strains with inherited predisposition and resistance to lung cancer provide a tool for the dissection of the complex genetics of this disease. In the present report, we have crossed the BALB/c with the SWR/J strain and performed whole-genome scanning for loci affecting lung tumor development in their F2 progeny. Both parental strains carry the pulmonary adenoma susceptibility 1 (Pas1) locus, a major locus affecting predisposition to lung cancer in mice. On distal chromosome 18 and on centromere of chromosome 6, we have mapped two pulmonary adenoma resistance loci (Par2 and Par4, respectively), which reduce lung tumor multiplicity strongly, up to 15-fold. Par2 and Par4, however, do not affect lung tumor size, which is instead controlled by an additional locus that we have mapped on the central region of chromosome 4. We designated this locus as "pulmonary adenoma progression 1" (Papg1), because it specifically modifies lung tumor size but not multiplicity. The present results, therefore, provide evidence for the existence of cancer modifier loci acting on specific stages of lung tumorigenesis.
Assuntos
Neoplasias Pulmonares/genética , Adenoma/genética , Adenoma/patologia , Animais , Mapeamento Cromossômico , Progressão da Doença , Suscetibilidade a Doenças , Feminino , Escore Lod , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos , Neoplasias Primárias Múltiplas/genética , Neoplasias Primárias Múltiplas/patologiaRESUMO
To study the structure and mechanism of deletion mutation in human somatic cells in vivo, we have identified and sequenced the breakpoints of 16 independent deletions at the hypoxanthine-guanine phosphoribosyl transferase (HPRT) locus in human T-lymphocytes. Seven deletions were found in exon 2, three in each of exon 3 and 6, and one in each of intron 3, exon 8 and exon 9. Most of the deletions seemed to result from non-homologous recombination, possibly by a slippage-misalignment mechanism between short repeat sequences. Putative secondary DNA structures, possibly acting as intermediates in the deletion formation, were identified in several mutants. Six of the seven exon 2 deletions had a breakpoint within a 12 bp region (in the 5' end of exon 2) which contains a 9 nucleotide palindrome (AACCAGGTT) and is preceded by a TGA direct repeat tract. One of the mutants had two deletions in tandem, separated by the palindrome. Another mutant, in which 23 bp containing the palindromic sequence was deleted, had an additional base (C) inserted between the breakpoints forming a direct repeat (gACGAC) in the deletion junction. Taken together with previously reported deletion mutations at the HPRT locus, these results suggest that the deletion cluster in the 5' part of HPRT exon 2 in T-cells in vivo is promoted by the 9 nucleotide palindrome sequence and the TGA repeat tract. The former may act as a stabilizer in a putative intermediate structure, and the latter may induce slippage and misalignment during replication.
Assuntos
Hipoxantina Fosforribosiltransferase/genética , Linfócitos/fisiologia , Deleção de Sequência , Sequência de Bases , Éxons , Humanos , Dados de Sequência Molecular , Família Multigênica , Mutação , Reação em Cadeia da Polimerase/métodos , Análise de Sequência de DNARESUMO
Cytotoxicity and mutagenicity were measured in human lymphocytes after treatment in vitro with anti- or syn-benzo[a]pyrene diolepoxide, two diastereoisomer metabolites of benzo[a]pyrene. These compounds were incubated with resting and cycling lymphocytes to determine the inhibition of cell proliferation induced by phytohemoagglutinin and interleukin2 at different times after treatment. Anti-benzo[a]pyrene diolepoxide was more cytotoxic than the syn-adduct under all conditions, and its effect on cell growth was more marked in cycling lymphocytes. In contrast, neither of the compounds induced alteration of the ATP intracellular pool. Cytotoxic effects of anti- and syn-benzo[a]pyrene diolepoxide were also assessed by determining the cloning efficiency. Both compounds affected the cloning efficiency in human lymphocytes and the effect of anti-benzo[a]pyrene was particularly marked. Mutagenic potency of anti- and syn-benzo[a]pyrene diolepoxide at the hgprt locus was measured both in the V79 cell line and in human lymphocytes by selection of mutant cells in medium containing 6-thioguanine. Both compounds increased the mutant frequency in comparison with the control and anti-benzo[a]pyrene diolepoxide was more active than the syn-metabolite.
RESUMO
Human PBL were treated in vitro with the ultimate reactive metabolites of BaP anti- and syn-BaPDE and DNA damage and repair were measured. The incorporation of radioactivity into DNA due to UDS was higher after treatment with anti-BaPDE. Radioactive DNA adduct dosimetry applied to PBL treated with tritiated syn- and anti-BaPDE demonstrated that anti-BaPDE gave more DNA adducts, which were more efficiently removed than syn adducts in the 24 h following the treatment. HPLC analysis of deoxynucleosides obtained from the enzymatic digestion of DNA showed that in treated PBL the major DNA adduct involved deoxyguanosine. DNA strand breaks, detected by FADU, were induced at comparable levels by anti- and syn-BaPDE (0.1-0.4 micrograms/ml), and persisted after 20 h of post-treatment incubation. Only in the case of syn-BaPDE did the percentage of double-stranded DNA tend to increase with time after the treatment.