RESUMO
In the last decade, advances in molecular biology have provided evidence of the genotypic heterogeneity of melanoma. We analysed BRAF, NRAS and c-KIT alterations in tissue samples from 63 stage III/IV melanoma patients and autologous cell-lines, using either allele-specific or quantitative PCR. The expression of BRAF V600E protein was also investigated using an anti-BRAF antibody in the same tissue samples. 81% of FFPE samples and tumor cell-lines harboured a genetic alteration in either BRAF (54%) or NRAS (27%) oncogenes. There was a strong concordance (100%) between tissue samples and tumor cell-lines. The BRAF V600E mutant-specific antibody showed high sensitivity (96%) and specificity (100%) for detecting the presence of a BRAF V600E mutation. The correlation was of 98% between PCR and immunohistochemistry results for BRAF mutation. These results suggest that BRAF and NRAS mutation status of tumor cells is not affected by culture conditions.
Assuntos
Análise Mutacional de DNA , GTP Fosfo-Hidrolases/genética , Melanoma/metabolismo , Proteínas de Membrana/genética , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-kit/genética , Neoplasias Cutâneas/metabolismo , Adulto , Idoso , Alelos , Linhagem Celular Tumoral , Células Cultivadas , Feminino , GTP Fosfo-Hidrolases/metabolismo , Genótipo , Humanos , Imuno-Histoquímica , Masculino , Melanoma/patologia , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Mutação , Oncogenes , Reação em Cadeia da Polimerase , Proteínas Proto-Oncogênicas B-raf/metabolismo , Proteínas Proto-Oncogênicas c-kit/metabolismo , Sensibilidade e Especificidade , Neoplasias Cutâneas/patologia , Adulto JovemRESUMO
We previously defined quantitative trait loci (QTLs) that control susceptibility to 7,12-dimethylbenz(alpha)anthracene-induced mammary carcinoma in SPRD-Cu3 (susceptible) and WKY (resistant) rats. Two of these QTLs, assigned to chromosomes (Chr) 10 and 18, control tumor growth rate and invasiveness. In this study we characterized a congenic strain in which a large segment of WKY Chr 10 was introduced in the SPRD-Cu3 genetic background and demonstrated that this chromosome segment controls this tumor trait. The WKY allele at this QTL (Mcsta1) reduces the growth rate of the fastest growing tumors by 26%. We also previously showed that two SPRD-Cu3-WKY congenic strains containing a WKY chromosome segment derived either from Chr 5 or from Chr 18 exhibit a reduction in tumor multiplicity (QTLs Msctm1 and Mcstm2, respectively) (with no reduction in tumor growth rate in the Chr 18 congenic). In this study we generated a double congenic strain, which contains the two WKY differential segments from Chr 5 and 18, to determine how these two segments interact with one another. Interestingly, two types of epistatic interactions were found: no additive effect was seen with respect to tumor multiplicity, while a reduction in tumor growth rate was observed. It thus appears that WKY alleles located on Chr 5 and Chr 8 interact epistatically in a contrasting manner to modulate tumor multiplicity (in a nonadditive manner) and growth rate (in a synergic manner). Tumor growth rate is thus influenced by two QTLs, on Chr 10 (Mcsta1) and on Chr 18 (Mcsta2), the action of the latter being dependent on the presence of the Chr5 QTL (Mcstm1). The expression level of positional and functional candidate genes was also analyzed. On Chr 5, Pla2g2a is subject to a syntenic control while expression of the Tp53 (Chr 10) and Pmai1/Noxa (Chr 18) genes appears to be controlled by several mammary cancer resistance QTLs.
Assuntos
Epistasia Genética/fisiologia , Neoplasias Mamárias Animais/genética , Locos de Características Quantitativas , Animais , Animais Congênicos , Sequência de Bases , Mapeamento Cromossômico , Feminino , Expressão Gênica , Genes p53/fisiologia , Predisposição Genética para Doença , Fosfolipases A2 do Grupo II/genética , Masculino , Dados de Sequência Molecular , Proteínas Proto-Oncogênicas c-bcl-2/genética , Ratos , Ratos Endogâmicos , Ratos Endogâmicos WKY , Homologia de Sequência do Ácido NucleicoRESUMO
The COP and WKY rat strains are resistant to mammary cancer. It has shown previously that upon chemical carcinogen treatment, COP females exhibit mammary preneoplastic lesions which disappear within a few weeks. We show here that in similar conditions, WKY females do not exhibit any visible preneoplastic lesions. WKY females are characterized by precocious mammary tissue differentiation, including active expression of the beta-casein gene in young virgin females. This trait might be critical in resistance to mammary carcinogenesis of WKY rats. To test this hypothesis, we took advantage of 2 congenic strains that contain a limited chromosome segment of WKY origin, derived either from chromosome 5 or from chromosome 18, introgressed in the susceptible genetic background (SPRD-Cu3). Each of these congenic strains has been shown to be partially resistant to chemically induced mammary carcinogenesis (reduction in tumour multiplicity with respect to the susceptible SPRD-Cu3 rats). We show here that these 2 congenic strains also exhibit precocious mammary differentiation, though to a lower extent than the WKY females. The conclusion of this study is thus 2-fold: (i) eradication of preneoplastic lesions is not a general phenomenon in mammary cancer resistance; (ii) the same segment of rat chromosomes 5 or 18 that controls mammary cancer resistance also contains a quantitative trait locus imposing precocious mammary differentiation. These 2 traits are thus associated, supporting the hypothesis that there might be a cause-effect relationship between precocious mammary differentiation and cancer resistance.
