RESUMO
Background: Activation of the phosphoinisitide-3 kinase (PI3K) pathway through mutation and constitutive upregulation has been described in renal cell carcinoma (RCC), making it an attractive target for therapeutic intervention. We performed a randomized phase II study in vascular endothelial growth factor (VEGF) therapy refractory patients to determine whether MK-2206, an allosteric inhibitor of AKT, was more efficacious than the mammalian target of rapamycin inhibitor everolimus. Patients and methods: A total of 43 patients were randomized in a 2:1 distribution, with 29 patients assigned to the MK-2206 arm and 14 to the everolimus arm. Progression-free survival (PFS) was the primary endpoint. Results: The trial was closed at the first futility analysis with an observed PFS of 3.68 months in the MK-2206 arm and 5.98 months in the everolimus arm. Dichotomous response rate profiles were seen in the MK-2206 arm with one complete response and three partial responses in the MK-2206 arm versus none in the everolimus arm. On the other hand, progressive disease was best response in 44.8% of MK2206 versus 14.3% of everolimus-treated patients. MK-2206 induced significantly more rash and pruritis than everolimus, and dose reduction occurred in 37.9% of MK-2206 versus 21.4% of everolimus-treated patients. Genomic analysis revealed that 57.1% of the patients in the PD group had either deleterious TP53 mutations or ATM mutations or deletions. In contrast, none of the patients in the non-PD group had TP53 or ATM defects. No predictive marker for response was observed in this small dataset. Conclusions: Dichotomous outcomes are observed when VEGF therapy refractory patients are treated with MK-2206, and MK-2206 does not demonstrate superiority to everolimus. Additionally, mutations in DNA repair genes are associated with early disease progression, indicating that dysregulation of DNA repair is associated with a more aggressive tumor phenotype in RCC.
Assuntos
Antineoplásicos/uso terapêutico , Carcinoma de Células Renais/tratamento farmacológico , Everolimo/uso terapêutico , Compostos Heterocíclicos com 3 Anéis/uso terapêutico , Neoplasias Renais/tratamento farmacológico , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Renais/mortalidade , Intervalo Livre de Doença , Feminino , Humanos , Neoplasias Renais/mortalidade , Masculino , Pessoa de Meia-Idade , Resultado do TratamentoRESUMO
BACKGROUND: KRAS mutations in NSCLC are associated with a lack of response to epidermal growth factor receptor inhibitors. Selumetinib (AZD6244; ARRY-142886) is an oral selective MEK kinase inhibitor of the Ras/Raf/MEK/ERK pathway. PATIENTS AND METHODS: Advanced nonsmall-cell lung cancer (NSCLC) patients failing one to two prior regimens underwent KRAS profiling. KRAS wild-type patients were randomized to erlotinib (150 mg daily) or a combination of selumetinib (150 mg daily) with erlotinib (100 mg daily). KRAS mutant patients were randomized to selumetinib (75 mg b.i.d.) or the combination. The primary end points were progression-free survival (PFS) for the KRAS wild-type cohort and objective response rate (ORR) for the KRAS mutant cohort. Biomarker studies of ERK phosphorylation and immune subsets were carried out. RESULTS: From March 2010 to May 2013, 89 patients were screened; 41 KRAS mutant and 38 KRAS wild-type patients were enrolled. Median PFS in the KRAS wild-type arm was 2.4 months [95% confidence interval (CI) 1.3-3.7] for erlotinib alone and 2.1 months (95% CI 1.8-5.1) for the combination. The ORR in the KRAS mutant group was 0% (95% CI 0.0% to 33.6%) for selumetinib alone and 10% (95% CI 2.1% to 26.3%) for the combination. Combination therapy resulted in increased toxicities, requiring dose reductions (56%) and discontinuation (8%). Programmed cell death-1 expression on regulatory T cells (Tregs), Tim-3 on CD8+ T cells and Th17 levels were associated with PFS and overall survival in patients receiving selumetinib. CONCLUSIONS: This study failed to show improvement in ORR or PFS with combination therapy of selumetinib and erlotinib over monotherapy in KRAS mutant and KRAS wild-type advanced NSCLC. The association of immune subsets and immune checkpoint receptor expression with selumetinib may warrant further studies.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Cloridrato de Erlotinib/administração & dosagem , Proteínas Proto-Oncogênicas p21(ras)/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Benzimidazóis/administração & dosagem , Carcinoma Pulmonar de Células não Pequenas/patologia , Intervalo Livre de Doença , Feminino , Humanos , MAP Quinase Quinase Quinase 1/genética , Masculino , Pessoa de Meia-Idade , Mutação , Inibidores de Proteínas Quinases/administração & dosagemRESUMO
BACKGROUND: We completed a phase I clinical trial to test the safety and toxicity of combined treatment with cixutumumab (anti-IGF-1R antibody) and selumetinib (MEK 1/2 inhibitor). METHODS: Patients with advanced solid tumours, refractory to standard therapy received selumetinib hydrogen sulphate capsules orally twice daily, and cixutumumab intravenously on days 1 and 15 of each 28-day cycle. The study used a 3+3 design, with a dose-finding cohort followed by an expansion cohort at the maximally tolerated dose that included pharmacokinetic and pharmacodynamic correlative studies. RESULTS: Thirty patients were enrolled, with 16 in the dose-finding cohort and 14 in the expansion cohort. Grade 3 or greater toxicities included nausea and vomiting, anaemia, CVA, hypertension, hyperglycaemia, and ophthalmic symptoms. The maximally tolerated combination dose was 50 mg twice daily of selumetinib and 12 mg kg(-1) every 2 weeks of cixutumumab. Two patients achieved a partial response (one unconfirmed), including a patient with BRAF wild-type thyroid carcinoma, and a patient with squamous cell carcinoma of the tongue, and six patients achieved time to progression of >6 months, including patients with thyroid carcinoma, colorectal carcinoma, and basal cell carcinoma. Comparison of pre- and on-treatment biopsies showed significant suppression of pERK and pS6 activity with treatment. CONCLUSIONS: Our study of anti-IGF-1R antibody cixutumumab and MEK 1/2 inhibitor selumetinib showed that the combination is safe and well-tolerated at these doses, with preliminary evidence of clinical benefit and pharmacodynamic evidence of target inhibition.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias/tratamento farmacológico , Idoso , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/efeitos adversos , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais Humanizados , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/farmacocinética , Benzimidazóis/administração & dosagem , Benzimidazóis/efeitos adversos , Benzimidazóis/farmacocinética , Estudos de Coortes , Intervalo Livre de Doença , Feminino , Humanos , MAP Quinase Quinase Quinases/antagonistas & inibidores , Masculino , Pessoa de Meia-Idade , Neoplasias/metabolismo , Receptor IGF Tipo 1/antagonistas & inibidores , Resultado do TratamentoRESUMO
BACKGROUND: Eribulin mesylate is a synthetic macrocyclic ketone analogue of Halichondrin B that has demonstrated high antitumor activity in preclinical and clinical settings. This phase I study aimed to determine the maximum tolerated dose (MTD), dose-limiting toxicities (DLTs), and pharmacokinetics in combination with cisplatin (CP) in patients with advanced solid tumours. METHODS: Thirty-six patients with advanced solid tumours received eribulin mesylate 0.7-1.4 mg m(-2) and CP 60-75 mg m(-2). Eribulin mesylate was administered on days 1, 8, and 15 in combination with CP day 1 every 28-day cycle. The protocol was amended after dose level 4 (eribulin mesylate 1.4 mg m(-2), CP 60 mg m(-2)) when it was not feasible to administer eribulin mesylate on day 15 because of neutropenia; the treatment schedule was changed to eribulin mesylate on days 1 and 8 and CP on day 1 every 21 days. RESULTS: On the 28-day schedule, three patients had DLT during the first cycle: grade (G) 4 febrile neutropenia (1.0 mg m(-2), 60 mg m(-2)); G 3 anorexia/fatigue/hypokalemia (1.2 mg m(-2), 60 mg m(-2)); and G 3 stomatitis/nausea/vomiting/fatigue (1.4 mg m(-2), 60 mg m(-2)). On the 21-day schedule, three patients had DLT during the first cycle: G 3 hypokalemia/hyponatremia (1.4 mg m(-2), 60 mg m(-2)); G 4 mucositis (1.4 mg m(-2), 60 mg m(-2)); and G 3 hypokalemia (1.2 mg m(-2), 75 mg m(-2)). The MTD and recommended phase II dose was determined as eribulin mesylate 1.2 mg m(-2) (days 1, 8) and CP 75 mg m(-2) (day 1), on a 21-day cycle. Two patients had unconfirmed partial responses (PR) (pancreatic and breast cancers) and two had PR (oesophageal and bladder cancers). CONCLUSIONS: On the 21-day cycle, eribulin mesylate 1.2 mg m(-2), administered on days 1 and 8, in combination with CP 75 mg m(-2), administered on day 1 is well tolerated and showed preliminary anticancer activity.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Éteres Cíclicos/uso terapêutico , Macrolídeos/uso terapêutico , Neoplasias/tratamento farmacológico , Adulto , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Cisplatino/administração & dosagem , Cisplatino/efeitos adversos , Éteres Cíclicos/administração & dosagem , Éteres Cíclicos/efeitos adversos , Furanos/administração & dosagem , Furanos/efeitos adversos , Humanos , Cetonas/administração & dosagem , Cetonas/efeitos adversos , Macrolídeos/administração & dosagem , Macrolídeos/efeitos adversos , Pessoa de Meia-Idade , Adulto JovemAssuntos
Colectomia , Doenças do Colo/patologia , Doença de Crohn/cirurgia , Infecções por Vírus Epstein-Barr/patologia , Doenças do Íleo/patologia , Transtornos Linfoproliferativos/patologia , Células de Reed-Sternberg/patologia , Ceco/patologia , Ceco/cirurgia , Colo/patologia , Colo/cirurgia , Doenças do Colo/diagnóstico , Doença de Crohn/complicações , Procedimentos Cirúrgicos do Sistema Digestório , Infecções por Vírus Epstein-Barr/diagnóstico , Herpesvirus Humano 4 , Humanos , Doenças do Íleo/diagnóstico , Íleo/patologia , Íleo/cirurgia , Transtornos Linfoproliferativos/diagnóstico , Masculino , Adulto JovemRESUMO
BACKGROUND: Adrenocortical carcinoma (ACC) is a rare and aggressive endocrine malignancy without an available effective systemic chemotherapy. Insulin growth factor 2 (IGF-2) overexpression leading to the activation of the IGF-1 receptor (IGF-1R)/mammalian target of rapamycin (mTOR) pathway is well described in ACC. Cixutumumab, a fully human IgG1 monoclonal antibody directed at IGF-1R was combined with temsirolimus on the basis of preclinical data. METHODS: Patients received cixutumumab, 3-6 mg kg(-1) intravenously (IV) weekly, and temsirolimus, 25-37.5 mg IV weekly (4-week cycles), with restaging after 8 weeks. RESULTS: Twenty-six patients were enrolled (13 (50%) men); median age, 47 years; median number of prior therapies, 4. Five patients previously received an IGF-1R inhibitor and one, temsirolimus. The most frequent toxicities, at least possibly drug related, were grade 1-2 thrombocytopenia (38%), mucositis (58%), hypercholesterolaemia (31%), hypertriglyceridemia (35%), and hyperglycaemia (31%). In all, 11 of 26 patients (42%) achieved stable disease (SD) >6 months (duration range=6-21 months) with 3 of the 11 having received a prior IGF-1R inhibitor. CONCLUSION: Cixutumumab combined with temsirolimus was well tolerated and >40% of patients achieved prolonged SD.
Assuntos
Anticorpos Monoclonais/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Sirolimo/análogos & derivados , Carcinoma Adrenocortical/tratamento farmacológico , Adulto , Idoso , Anticorpos Monoclonais Humanizados , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Intervalo Livre de Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Sirolimo/administração & dosagem , Adulto JovemRESUMO
BACKGROUND: Cancer cachexia is characterised by skeletal muscle wasting; however, potential for muscle anabolism in patients with advanced cancer is unproven. METHODS: Quantitative analysis of computed tomography images for loss/gain of muscle in cholangiocarcinoma patients receiving selumetinib (AZD6244; ARRY-142886) in a Phase II study, compared with a separate standard therapy group. Selumetinib is an inhibitor of mitogen-activated protein/extracellular signal-regulated kinase and of interleukin-6 secretion, a putative mediator of muscle wasting. RESULTS: Overall, 84.2% of patients gained muscle after initiating selumetinib; mean overall gain of total lumbar muscle cross-sectional area was 13.6 cm(2)/100 days (â¼2.3 kg on a whole-body basis). Cholangiocarcinoma patients who began standard treatment were markedly catabolic, with overall muscle loss of -7.3 cm(2)/100 days (â¼1.2 kg) and by contrast only 16.7% of these patients gained muscle. CONCLUSION: Our findings suggest that selumetinib promotes muscle gain in patients with cholangiocarcinoma. Specific mechanisms and relevance for cachexia therapy remain to be investigated.
Assuntos
Benzimidazóis/efeitos adversos , Neoplasias dos Ductos Biliares/tratamento farmacológico , Ductos Biliares Intra-Hepáticos , Colangiocarcinoma/tratamento farmacológico , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Músculo Esquelético/efeitos dos fármacos , Inibidores de Proteínas Quinases/efeitos adversos , Adulto , Idoso , Neoplasias dos Ductos Biliares/mortalidade , Caquexia/tratamento farmacológico , Colangiocarcinoma/mortalidade , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/metabolismoRESUMO
BACKGROUND: Nasopharyngeal colonization by Streptococcus pneumoniae precedes pneumococcal disease. Elucidation of procedures to prevent or eradicate nasopharyngeal carriage in a model akin to the human would help to diminish the incidence of both pneumonia and invasive pneumococcal disease. METHODS: We conducted a survey of the nasopharynx of infant rhesus macaques from our breeding colony, in search of natural carriers of S. pneumoniae. We also attempted experimental induction of colonization, by nasopharyngeal instillation of a human S. pneumoniae strain (19F). RESULTS: None of 158 colony animals surveyed carried S. pneumoniae in the nasopharynx. Colonization was induced in eight of eight infant rhesus by nasopharyngeal instillation and lasted 2weeks in 100% of the animals and 7weeks in more than 60%. CONCLUSION: Rhesus macaques are probably not natural carriers of S. pneumoniae. The high rate and duration of colonization obtained in our experiments indicates that the rhesus macaque will serve as a human-like carriage model.
Assuntos
Portador Sadio/veterinária , Macaca mulatta/microbiologia , Infecções Pneumocócicas/veterinária , Streptococcus pneumoniae/crescimento & desenvolvimento , Animais , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/microbiologia , Portador Sadio/microbiologia , Contagem de Colônia Microbiana , Masculino , Testes de Sensibilidade Microbiana , Nasofaringe/diagnóstico por imagem , Nasofaringe/microbiologia , Infecções Pneumocócicas/diagnóstico por imagem , Infecções Pneumocócicas/microbiologia , RadiografiaRESUMO
Small-cell lung cancer (SCLC) lines are distinguished from non-small-cell lung cancer (NSCLC) lines by their growth in floating aggregates, in contrast to the adherent monolayers formed by NSCLC cells in culture. Of 50 well-characterized SCLC lines recently described by the National Cancer Institute (NCI)-Navy Medical Oncology Branch, only four variant cell lines (SCLC-v) grew as adherent monolayers. One line, NCI-H446, was unique in growing long-term with coexisting floating and surface adherent subpopulations. We have physically segregated these two populations over many passages in vitro to enrich for relatively pure cultures of floating and adherent cells. No differences in c-myc expression, keratin pattern, or cytogenetic appearance were found between the adherent and floating sublines. However, expression of the neuroendocrine marker neuron-specific enolase in the floating cells was three times that found in the adherent cells. The floating subline also had much greater surface expression of neuroendocrine tumor antigens detected by monoclonal antibodies UJ13A and HNK-1, which have been recently shown to detect the neural cell adhesion molecule (NCAM) on SCLC cells. Two other adherent SCLC-v lines were also found to be unreactive with UJ13A and HNK-1, generalizing the association between NCAM expression and the growth of most SCLC cultures as floating aggregates. In conclusion, we have an interesting model to study expression of NCAM as related to the adhesive properties of SCLC cells.
Assuntos
Carcinoma de Células Pequenas/metabolismo , Moléculas de Adesão Celular Neuronais/metabolismo , Adesão Celular , Neoplasias Pulmonares/metabolismo , Anticorpos Monoclonais , Antígenos de Neoplasias/imunologia , Northern Blotting , Western Blotting , Carcinoma de Células Pequenas/imunologia , Carcinoma de Células Pequenas/patologia , Moléculas de Adesão Celular Neuronais/imunologia , Divisão Celular , Citogenética , Regulação para Baixo , Expressão Gênica , Humanos , Técnicas In Vitro , Queratinas/metabolismo , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Fosfopiruvato Hidratase/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Células Tumorais CultivadasRESUMO
BACKGROUND: Human cancer cell lines grown in the presence of the cytotoxic agent mitoxantrone frequently develop resistance associated with a reduction in intracellular drug accumulation without increased expression of the known drug resistance transporters P-glycoprotein and multidrug resistance protein (also known as multidrug resistance-associated protein). Breast cancer resistance protein (BCRP) is a recently described adenosine triphosphate-binding cassette transporter associated with resistance to mitoxantrone and anthracyclines. This study was undertaken to test the prevalence of BCRP overexpression in cell lines selected for growth in the presence of mitoxantrone. METHODS: Total cellular RNA or poly A+ RNA and genomic DNA were isolated from parental and drug-selected cell lines. Expression of BCRP messenger RNA (mRNA) and amplification of the BCRP gene were analyzed by northern and Southern blot hybridization, respectively. RESULTS: A variety of drug-resistant human cancer cell lines derived by selection with mitoxantrone markedly overexpressed BCRP mRNA; these cell lines included sublines of human breast carcinoma (MCF-7), colon carcinoma (S1 and HT29), gastric carcinoma (EPG85-257), fibrosarcoma (EPF86-079), and myeloma (8226) origins. Analysis of genomic DNA from BCRP-overexpressing MCF-7/MX cells demonstrated that the BCRP gene was also amplified in these cells. CONCLUSIONS: Overexpression of BCRP mRNA is frequently observed in multidrug-resistant cell lines selected with mitoxantrone, suggesting that BCRP is likely to be a major cellular defense mechanism elicited in response to exposure to this drug. It is likely that BCRP is the putative "mitoxantrone transporter" hypothesized to be present in these cell lines.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica , Mitoxantrona/farmacologia , Proteínas de Neoplasias/biossíntese , Northern Blotting , Southern Blotting , Neoplasias da Mama/genética , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/metabolismo , Resistência a Múltiplos Medicamentos/genética , Resistencia a Medicamentos Antineoplásicos/genética , Fibrossarcoma/tratamento farmacológico , Fibrossarcoma/metabolismo , Humanos , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/metabolismo , Proteínas de Neoplasias/genética , RNA Mensageiro/análise , RNA Neoplásico/análise , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/metabolismo , Células Tumorais Cultivadas , Regulação para CimaRESUMO
Monoclonal antibodies (MoAbs) to antigens of human small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) were produced from BALB/c mice immunized with either intact cultured cells or membrane preparations from cultured tumor cells. Of 172 MoAbs produced from two NSCLC immunized mice and reactive to NSCLC cells, 137 bound staphylococcal Protein A directly, and only 11 of these 172 MoAbs were significantly reactive with cultured SCLC cells by a solid-phase radioimmunoassay. In contrast, only 16 of 99 MoAbs produced from two SCLC immunized mice and reactive to SCLC cells directly bound Protein A, and most were of an IgM isotype, but 98 of 99 of these antibodies also reacted with cultured NSCLC target plates. Twenty hybridomas producing antibodies reactive with lung cancer cells but not with a B-lymphoblastoid line were cloned. Eleven of these cloned hybridomas were from NSCLC fusions, and nine were from SCLC fusions. When representative hybridoma supernatants were tested against a broad panel of SCLC and NSCLC target plates a similar pattern was seen, with the supernatants from NSCLC-derived hybridomas only reacting strongly to the NSCLC target plates, but the supernatants from SCLC-derived hybridomas always reacting to both SCLC and NSCLC plates. We conclude that the humoral response to immunization with NSCLC cells or membrane preparations is predominantly IgG and that to SCLC is predominantly IgM. Furthermore, many IgG MoAbs reactive with NSCLC lines are poorly reactive with SCLC cells.
Assuntos
Carcinoma de Células Pequenas/imunologia , Neoplasias Pulmonares/imunologia , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/imunologia , Linhagem Celular , Imunofluorescência , Humanos , Imunização , Imunoglobulina G/análise , Isotipos de Imunoglobulinas/análise , Imunoglobulina M/análise , Camundongos , Camundongos Endogâmicos BALB C , CoelhosRESUMO
Variant small cell lung cancer (SCLC) is distinguished from the classic histology by changes in growth characteristics and morphology, c-myc amplification, a loss of some biochemical markers, and relative chemo- and radioresistance. Three variant SCLC lines were incubated in 1 microM all-trans retinoic acid. After 8-10 days, a marked change in morphology was noted in all three lines, with tight cell aggregation and central necrosis of large floating spheroids similar to classic SCLC. The retinoid-treated cells demonstrated decreases in growth rate and cloning efficiency to levels comparable with classic SCLC cell lines. Retinoic acid incubation caused a reproducible decrease in c-myc expression in variant SCLC cells, but was not noted to increase L-dopa decarboxylase, an enzyme which biochemically distinguishes classic from variant SCLC cell lines. Retinoid exposure led to an increase in HLA and Leu-7 antigens, but a reduction of antibody binding to 3-fucosyllactosamine, a dominant SCLC glycolipid antigen. Clonogenic assays revealed that the variant cells, after incubation in retinoic acid, became more sensitive to etoposide, but more resistant to Adriamycin. We conclude that exposure of variant SCLC cells to retinoic acid can lead to a morphologic phenotype similar to classic SCLC, but with substantial differences in cell biology. ?
Assuntos
Carcinoma de Células Pequenas/patologia , Tretinoína/farmacologia , Antígenos de Neoplasias/análise , Antígenos de Superfície/análise , Northern Blotting , Diferenciação Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Dopa Descarboxilase/metabolismo , Doxorrubicina/farmacologia , Etoposídeo/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Técnicas In Vitro , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-myc , Células Tumorais CultivadasRESUMO
The response of class I major histocompatibility complex antigen expression to in vitro administration of interferon and tumor necrosis factor alpha (TNF-alpha) was measured using class I major histocompatibility complex-deficient small cell lung cancer cell lines. Significant induction also was observed using gamma interferon (IFN-gamma) alone, whereas TNF-alpha alone yielded only modest induction. Classic small cell lung cancer cell lines NCI-H146 and NCI-H209 best demonstrated synergistic HLA and beta 2-microglobulin antigen induction with IFN-gamma and TNF-alpha with the following dose schedule: 3-6 days of TNF-alpha (200 units/ml) followed by 48 h of IFN-gamma (100 IU/ml). Induction was quantitated using an 125I-Protein A radioimmunoassay. Synergistic induction of the HLA and beta 2-microglobulin surface antigens on NCI-H146 was also possible with alpha interferon and TNF-alpha but required a higher concentration of the interferon, i.e., 3-6 days of TNF-alpha (200 units/ml) followed by 48 h of alpha interferon (1000 units/ml). Small cell lung cancer cell line NCI-H146 was further studied for expression of major histocompatibility complex messenger RNA using the optimal doses and sequence of addition of IFN-gamma and TNF-alpha as indicated above. A significant induction with IFN-gamma alone and synergistic induction with both IFN-gamma and TNF-alpha was quantitated for both HLA-A2 and beta 2-microglobulin transcripts using Northern blot analysis. Incubation with relatively low subcytotoxic doses of IFN-gamma and TNF-alpha also resulted in a marked synergistic decrease in c-myc message.
Assuntos
Regulação Neoplásica da Expressão Gênica , Genes MHC Classe I/efeitos dos fármacos , Antígenos de Histocompatibilidade Classe I/genética , Interferon Tipo I/farmacologia , Interferon gama/farmacologia , Células Tumorais Cultivadas/imunologia , Fator de Necrose Tumoral alfa/farmacologia , Animais , Anticorpos Monoclonais , Northern Blotting , Carcinoma de Células Pequenas/imunologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Cães , Sinergismo Farmacológico , Antígenos de Histocompatibilidade Classe I/análise , Humanos , Neoplasias Pulmonares/imunologia , Radioimunoensaio , Proteínas Recombinantes/farmacologia , Células Tumorais Cultivadas/citologia , Células Tumorais Cultivadas/efeitos dos fármacosRESUMO
Deletions of the 3p chromosome region and molecular alterations of the tumor suppressor genes RB1 and TP53, located, respectively, at 13q14 and 17p13, are well-documented in small cell lung cancer (SCLC). Because of technical difficulties, karyotypes of primary SCLC specimens are rarely reported. In this study, detailed cytogenetic analysis was performed on 13 early passage SCLC cell lines and fresh specimens, including 4 lung primaries. Numerous chromosome alterations were found, even in newly diagnosed primary tumors. Consistent with previous molecular studies, chromosomal losses of 3p (13 cases) and 17p13 (12 cases) were frequently observed. Numerical losses of chromosome 13 and structural rearrangements affecting 13q14 were identified in 10 specimens. In addition, losses of chromosome 5 and structural alterations of 5q occurred in 12 tumors; among these, 9 displayed losses of region 5q13-q21. Double minutes were found in 4 cases (3 of 5 specimens from patients who received prior cytotoxic therapy but only 1 of 8 from untreated patients). DNA analysis revealed amplification of either MYC1 or MYCN in cells from each of these 4 tumors. Overall, the cytogenetic findings underscore that progression of SCLC involves multiple genetic changes and suggest further that a tumor suppressor gene(s) on 5q may contribute to SCLC tumorigenesis.
Assuntos
Carcinoma de Células Pequenas/genética , Aberrações Cromossômicas/genética , Bandeamento Cromossômico , Cromossomos Humanos Par 3 , Cromossomos Humanos Par 5 , Neoplasias Pulmonares/genética , Transtornos Cromossômicos , Cromossomos Humanos Par 13 , Feminino , Humanos , Cariotipagem , Masculino , Polimorfismo de Fragmento de Restrição , Células Tumorais CultivadasRESUMO
Fumitremorgin C (FTC) is a potent and specific chemosensitizing agent in cell lines selected for resistance to mitoxantrone that do not overexpress P-glycoprotein or multidrug resistance protein. The gene encoding a novel transporter, the breast cancer resistance protein (BCRP), was recently found to be overexpressed in a mitoxantrone-selected human colon cell line, S1-M1-3.2, which was used to identify FTC. Because the drug-selected cell line may contain multiple alterations contributing to the multidrug resistance phenotype, we examined the effect of FTC on MCF-7 cells transfected with the BCRP gene. We report that FTC almost completely reverses resistance mediated by BCRP in vitro and is a pharmacological probe for the expression and molecular action of this transporter.
Assuntos
Transportadores de Cassetes de Ligação de ATP/efeitos dos fármacos , Resistência a Múltiplos Medicamentos , Indóis/farmacologia , Proteínas de Neoplasias/efeitos dos fármacos , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Antineoplásicos/farmacologia , Neoplasias da Mama/metabolismo , Resistencia a Medicamentos Antineoplásicos , Feminino , Humanos , Proteínas de Neoplasias/genética , Transfecção , Células Tumorais Cultivadas/efeitos dos fármacosRESUMO
Human breast carcinoma MCF-7/AdrVp cells display a novel multidrug resistance phenotype that is characterized by the overexpression of a 95-kDa membrane glycoprotein (p95) and by marked reduction in intracellular anthracycline accumulation, without overexpression of P-glycoprotein or the multidrug resistance protein MRP. p95 is also highly expressed in multidrug-resistant NCI-H1688 cells derived from a human small cell lung carcinoma. Deglycoslyated p95 from NCI-H1688 cells was isolated by two-dimensional gel electrophoresis and then digested with trypsin. The tryptic peptides were analyzed by mass spectrometry and microsequencing. These analyses identified p95 to be identical to NCA-90, the nonspecific cross-reacting antigen related to the carcinoembryonic antigen (CEA). Further confirmation that p95 is indeed NCA-90 was obtained by Northern and Western blot studies using probes or antibodies specific for p95, NCA-90, or CEA family members. Western blot studies also revealed that CEA itself is overexpressed in MCF-7/AdrVp cells compared to parental MCF-7/W cells. The enforced expression of NCA-90 protein in HeLa cells stably transfected with NCA-90 cDNA did not result in increased resistance of the transfected cells to daunorubicin or a decrease in daunorubicin accumulation in the transfected cells compared to cells transfected only with the expression vector. However, a recent report by H. Kawaharata et al. (Int. J. Cancer, 72: 377-382, 1997) of diminished accumulation, retention, and cytotoxicity of doxorubicin in EJNIH3T3 cells in which enforced expression of CEA was accomplished leaves open the possibility that the overexpression of CEA, possibly in combination with that of NCA-90, could account at least in part for the drug resistant phenotype displayed by MCF-7/AdrVp cells.
Assuntos
Antígenos de Neoplasias , Neoplasias da Mama/química , Neoplasias da Mama/metabolismo , Antígeno Carcinoembrionário/imunologia , Moléculas de Adesão Celular , Resistência a Múltiplos Medicamentos , Glicoproteínas de Membrana/biossíntese , Glicoproteínas de Membrana/química , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/química , Células 3T3/metabolismo , Sequência de Aminoácidos , Animais , Antibióticos Antineoplásicos/farmacologia , Western Blotting , Neoplasias da Mama/tratamento farmacológico , Reações Cruzadas , Doxorrubicina/farmacologia , Resistencia a Medicamentos Antineoplásicos , Eletroforese em Gel Bidimensional , Epitopos , Células HeLa , Humanos , Focalização Isoelétrica , Glicoproteínas de Membrana/isolamento & purificação , Camundongos , Dados de Sequência Molecular , Proteínas de Neoplasias/isolamento & purificação , Fenótipo , Dodecilsulfato de Sódio , Células Tumorais CultivadasRESUMO
The multidrug resistance phenotype of human breast carcinoma MCF-7/AdrVp cells is characterized by overexpression of a 95-kilodalton membrane glycoprotein (p95), accompanied by a marked reduction in intracellular anthracycline accumulation, without overexpression of P-glycoprotein or the multidrug resistance protein. We discovered that the mRNA of the H19 gene is overexpressed in MCF-7/AdrVp cells relative to parental MCF-7 cells or drug-sensitive MCF-7/AdrVp revertant cells. H19 is an imprinted gene with an important role in fetal differentiation, as well as a postulated function as a tumor suppressor gene. Another p95-overexpressing multidrug-resistant cell line, human lung carcinoma NCI-H1688, also displays high levels of H19 mRNA. In contrast, several multidrug-resistant cell lines that overexpress P-glycoprotein or the multidrug resistance protein do not have higher levels of H19 mRNA than their drug-sensitive counterparts. This is the first report of H19 gene overexpression accompanying any form of drug resistance. The association of H19 and p95 gene expression in drug resistance warrants further study.
Assuntos
Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Resistência a Múltiplos Medicamentos/genética , Glicoproteínas de Membrana/biossíntese , Proteínas Musculares/biossíntese , Proteínas Musculares/genética , Proteínas de Neoplasias/biossíntese , RNA não Traduzido , Sequência de Bases , Neoplasias da Mama/metabolismo , Clonagem Molecular , Sondas de DNA , DNA de Neoplasias/genética , Expressão Gênica , Humanos , Dados de Sequência Molecular , RNA Longo não Codificante , Células Tumorais CultivadasRESUMO
Cellular resistance to the antifolate methotrexate (MTX) is often caused by target amplification, uptake defects, or alterations in polyglutamylation. Here we have examined MTX cross-resistance in a human breast carcinoma cell line (MCF7/MX) selected in the presence of mitoxantrone, an anticancer agent associated with the multidrug resistance (MDR) phenotype. Examination of protein expression and enzyme activities showed that MCF7/MX cells displayed none of the classical mechanisms of MTX resistance. They did, however, exhibit an ATP-sensitive accumulation defect accompanied by reduced polyglutamylation. Although the kinetics of drug uptake was similar between parental and resistant cells, the resistant cells exhibited increased energy-dependent drug efflux. This suggested the involvement of an ATP-binding cassette (ABC) transporter. However, cells transfected with the breast cancer resistance protein (BCRP)-the ABC transporter known to be highly overexpressed in MCF7/MX cells and to confer mitoxantrone resistance (D. D. Ross et al., J. Natl. Cancer Inst. 91: 429-433, 1999)-were not MTX resistant, which suggested that this transporter is not involved in MTX cross-resistance. Moreover, members of the MRP protein family of transport proteins, which had previously been implicated in MTX resistance, were not found to be overexpressed in the MCF7/MX cells. Thus, our data suggest that a novel MTX-specific efflux pump may be involved in this unusual cross-resistance phenotype.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Antineoplásicos/toxicidade , Resistência a Múltiplos Medicamentos , Metotrexato/farmacocinética , Metotrexato/toxicidade , Mitoxantrona/toxicidade , Proteínas de Neoplasias , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Transporte Biológico , Neoplasias da Mama , Sobrevivência Celular/efeitos dos fármacos , Feminino , Humanos , Mitoxantrona/farmacocinética , Peptídeo Sintases/metabolismo , Proteínas Recombinantes/metabolismo , Transfecção , Células Tumorais CultivadasRESUMO
The breast cancer resistance protein (BCRP) gene, formally known as ATP-binding cassette transporter G2 (ABCG2) gene, encodes an ABC half transporter that causes resistance to certain cancer chemotherapeutic drugs when transfected and expressed in drug sensitive cancer cells. Here we report the organization of the BCRP gene, and the initial characterization of the BCRP promoter. We identified the genomic sequence of BCRP and its promoter by screening a human genomic lambda phage library, as well as a BAC library, and by searching the human genome database. The BCRP gene spans over 66 kb and consists of 16 exons and 15 introns. The exons range in size from 60 to 532 bp. The translational start site is found in the second exon. The first exon contains the majority of the 5' UTR. Promoter activity was characterized by a luciferase reporter assay using transient transfection of the human breast cancer cell line MCF7, and the human choriocarcinoma cell lines JAR, BeWo and JEG-3, which we find to have high endogenous expression of BCRP. The BCRP gene is transcribed by a TATA-less promoter with several putative Sp1 sites, which are downstream from a putative CpG island. The sequence 312 bp directly upstream from the BCRP transcriptional start site conferred basal promoter activity. The 5' region upstream of the basal promoter is characterized by both positive and negative regulatory domains.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Neoplasias da Mama/genética , Proteínas de Neoplasias/genética , Regiões Promotoras Genéticas , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/química , Sequência de Bases , DNA Complementar/química , Resistência a Múltiplos Medicamentos/genética , Biblioteca Genômica , Humanos , Dados de Sequência Molecular , Proteínas de Neoplasias/químicaRESUMO
PURPOSE: The sequence in which chemotherapeutic agents are administered can alter their pharmacokinetics, therapeutic effect, and toxicity. We evaluated the pharmacokinetics and pharmacodynamics of docetaxel and topotecan when coadministered on two different sequences of administration. PATIENTS AND METHODS: On cycle 1, docetaxel was administered as a 1-hour infusion at 60 mg/m(2) without filgrastim and at 60, 70, and 80 mg/m(2) with filgrastim on day 1, and topotecan was administered at 0.75 mg/m(2) as a 0.5-hour infusion on days 1 to 4. On cycle 2, topotecan was administered on days 1 to 4, and docetaxel was administered on day 4. Cycles were repeated every 21 days. Blood samples for high-performance liquid chromatography measurement of docetaxel (CL(DOC)) and topotecan (CL(TPT)) total clearance were obtained on day 1 of cycle 1 and day 4 of cycle 2. CL(DOC) and CL(TPT) were calculated using compartmental methods. RESULTS: Mean +/- SD CL(DOC) in cycles 1 and 2 were 75.9 +/- 79.6 L/h/m(2) and 29.2 +/- 17.3 L/h/m(2), respectively (P: <.046). Mean +/- SD CL(TPT) in cycles 1 and 2 were 8.5 +/- 4.4 L/h/m(2) and 9.3 +/- 3.4 L/h/m(2), respectively (P: >. 05). Mean +/- SD neutrophil nadir in cycles 1 and 2 were 4,857 +/- 6, 738/microL and 2,808 +/- 4,518/microL, respectively (P: =.02). CONCLUSION: Administration of topotecan on days 1 to 4 and docetaxel on day 4 resulted in an approximately 50% decrease in docetaxel clearance and was associated with increased neutropenia.