RESUMO
Flavopiridol, the first inhibitor of cyclin-dependent kinases to enter clinical trials, has shown promising antineoplastic activity and is currently undergoing Phase II testing. Little is known about mechanisms of resistance to this agent. In the present study, we have characterized an ovarian carcinoma cell line [OV202 high passage (hp)] that spontaneously developed drug resistance upon prolonged passage in tissue culture. Standard cytogenetic analysis and spectral karyotyping revealed that OV202 hp and the parental low passage line OV202 shared several marker chromosomes, confirming the relatedness of these cell lines. Immunoblotting demonstrated that OV202 and OV202 hp contained similar levels of a variety of polypeptides involved in cell cycle regulation, including cyclin-dependent kinases 2 and 4; cyclins A, D1, and E; and proliferating cell nuclear antigen. Despite these similarities, OV202 hp was resistant to flavopiridol and cisplatin, with increases of 5- and 3-fold, respectively, in the mean drug concentrations required to inhibit colony formation by 90%. In contrast, OV202 hp and OV202 displayed indistinguishable sensitivities to oxaliplatin, paclitaxel, topotecan, 1,3-bis(2-chloroethyl)-1-nitrosourea, etoposide, doxorubicin, vincristine, and 5-fluorouracil, suggesting that the spontaneously acquired resistance was not attributable to altered P-glycoprotein levels or a general failure to engage the cell death machinery. After incubation with cisplatin, whole cell platinum and platinum-DNA adducts measured using mass spectrometry were lower in OV202 hp cells than OV202 cells. Similarly, after flavopiridol exposure, whole cell flavopiridol concentrations measured by a newly developed high performance liquid chromatography assay were lower in OV202 hp cells. These data are consistent with the hypothesis that acquisition of spontaneous resistance to flavopiridol and cisplatin in OV202 hp cells is due, at least in part, to reduced accumulation of the respective drugs. These observations not only provide the first characterization of a flavopiridol-resistant cell line but also raise the possibility that alterations in drug accumulation might be important in determining sensitivity to this agent.
Assuntos
Antineoplásicos/toxicidade , Aberrações Cromossômicas , Cisplatino/toxicidade , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Flavonoides/toxicidade , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Piperidinas/toxicidade , Carmustina/toxicidade , Mapeamento Cromossômico , Cisplatino/farmacocinética , Adutos de DNA/análise , Feminino , Humanos , Cariotipagem , Compostos Organoplatínicos/farmacocinética , Compostos Organoplatínicos/toxicidade , Oxaliplatina , Células Tumorais CultivadasRESUMO
Medulloblastomas are highly malignant primitive neuroectodermal tumors of the cerebellum that display a wide variety of histopathological patterns. However, these patterns do not provide an accurate prediction of clinical-biological behavior and no satisfactory morphological grading system has ever been presented. Genetic alterations may provide additional diagnostic information and allow clinically relevant subgrouping of primitive neuroectodermal tumors. We examined 10 medulloblastomas and one medulloblastoma cell line. One amplification site on chromosome 8q24 was detected in the cell line corresponding to the known amplification of the c-myc gene in this cell line. The gain of 2p21-24 in two tumors was shown to represent amplification of the N-myc gene by Southern blot hybridization and fluorescence in situ hybridization. The data show that the isochromosome 17 can be recognized using comparative genomic hybridization (CGH) by the typical combination of loss of 17p combined with gain of 17q. No specific pattern of genetic alterations could be linked to the clinical behavior of the tumors. We have compared our results with previous CGH studies on medulloblastomas.
Assuntos
Neoplasias Cerebelares/genética , Amplificação de Genes/genética , Genes myc/genética , Meduloblastoma/genética , Adolescente , Adulto , Southern Blotting , Neoplasias Cerebelares/patologia , Criança , Pré-Escolar , Cromossomos Humanos Par 17 , Cromossomos Humanos Par 8 , Sondas de DNA , Progressão da Doença , Feminino , Humanos , Hibridização in Situ Fluorescente , Lactente , Masculino , Meduloblastoma/patologia , Células Tumorais CultivadasRESUMO
PURPOSE: Metanephric adenoma has recently been recognized as a unique renal tumor characterized by an unusual degree of cellular differentiation and maturation. We recently studied metanephric adenoma using metaphase analysis and observed concomitant chromosome Y loss and chromosome 7 and 17 gain. To determine if these chromosomal anomalies are consistently present in renal metanephric adenoma, we studied all 11 tumors in the pathology tissue registry at our institution using fluorescence in situ hybridization (FISH). MATERIALS AND METHODS: FISH, using deoxyribonucleic acid probes for chromosomes 1, 7, 8, 17, X and Y, was performed in isolated nuclei from 11 paraffin embedded renal metanephric adenoma specimens. RESULTS: Of the 11 tumors (73%) 8 demonstrated chromosome 7 and 17 gain by FISH, and the remaining 3 were found to have an apparently normal chromosomal content. Of the 8 tumors (75%) from men showed 6 chromosome 7 and 17 gain with Y chromosome loss. Of the 3 tumors (33%) from women 1 had chromosome 7 and 17 gain with X chromosome loss, while 1 had chromosome 7 and 17 gain without sex chromosome aneusomy. Metaphase analysis performed on 2 tumors revealed chromosome 7 and 17 gain and Y chromosome loss in 1, and no apparent, chromosome anomaly in the other, confirming the results of FISH analysis. CONCLUSIONS: FISH analysis of renal metanephric adenoma identified frequent chromosome 7 and 17 gain and sex chromosome loss. These results are consistent with a clonal neoplastic disorder in which chromosomes 7, 17, X and Y are likely to be involved in the pathogenesis of this tumor. These characteristic chromosomal alterations have also been observed in papillary renal cell adenoma and papillary renal cell carcinoma, providing evidence that these tumors may be related.
Assuntos
Adenoma/genética , Carcinoma Papilar/genética , Carcinoma de Células Renais/genética , Aberrações Cromossômicas/genética , Cromossomos Humanos Par 17/genética , Cromossomos Humanos Par 7/genética , Neoplasias Renais/genética , Cromossomos Sexuais/genética , Cromossomos Humanos Par 1/genética , Cromossomos Humanos Par 8/genética , Feminino , Humanos , Hibridização in Situ Fluorescente , MasculinoRESUMO
PURPOSE: Cytogenetic studies of a small number of renal oncocytomas have indicated that loss of chromosomes 1 and Y may be involved in the pathogenesis of this tumor. To evaluate these observations further we selected paraffin embedded renal oncocytoma specimens from 20 male and 10 female patients for fluorescence in situ hybridization analysis. MATERIALS AND METHODS: Isolated nuclei were prepared from paraffin embedded specimens, and fluorescence in situ hybridization was performed with enumeration probes for chromosomes 1, 12, X and Y. RESULTS: Tumors from 10 male (50%) and 4 female (40%) patients demonstrated chromosomal alterations. Loss of chromosome Y was observed in specimens from all 10 male patients, and loss of chromosome 1 or gain of chromosome 12 was noted in 5 and 2 of these specimens, respectively. Of the 4 female patients with chromosomal abnormalities 2 had loss of chromosome 1, 1 had gain of chromosome 1 and 1 had gain of chromosome 12. CONCLUSIONS: Our results confirm that loss of chromosomes Y and 1 is common in renal oncocytoma, and that the alterations are probably involved in the pathogenesis of this tumor.