RESUMO
Sophoridine (1), a natural anticancer drug, has been used in China for decades. A series of novel N-substituted sophoridinic acid derivatives were synthesized and evaluated for their cytotoxicity with 1 as the lead. The structure-activity relationship indicated that introduction of an aliphatic acyl on the nitrogen atom might significantly enhance the anticancer activity. Among the compounds, 6b bearing bromoacetyl side-chain afforded a potential effect against four human tumor cell lines (liver, colon, breast, and lung). The mechanism of action of 6b is to inhibit the activity of DNA topoisomerase I, followed by the S-phase arrest and then cause apoptotic cell death, similar to that of its parent 1. We consider 6b promising for further anticancer investigation.
Assuntos
Alcaloides/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Neoplasias/tratamento farmacológico , Quinolizinas/farmacologia , Alcaloides/síntese química , Alcaloides/química , Antineoplásicos/química , Morte Celular/efeitos dos fármacos , Técnicas de Química Sintética , DNA Topoisomerases Tipo I/metabolismo , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Estrutura Molecular , Neoplasias/enzimologia , Neoplasias/patologia , Quinolizinas/síntese química , Quinolizinas/química , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
To investigate the antitumor activities of the immunoconjugates composed of anti-type IV collagenase monoclonal antibody Fab' fragment and lidamycin (LDM) prepared with different linkers. The immunoconjugates were prepared by linking Fab' to lysine-69 of LDM apoprotein by SPDP, LCSPDP, SMBS or SSMPB as the intermediate drug linkers. Immunoreactivities of the conjugates were determined by ELISA. The cytotoxicities of the conjugates were examined by clonogenic assay. In vivo antitumor effects of the conjugates were evaluated in nude mice bearing subcutaneously implanted HT-1080 tumor. ELISA assay showed that the conjugates retained part of the immunoreactivity of 3G11 against the antigen. The cytotoxicities of the Fab'-SMBS-LDM and Fab'-SSMPB-LDM to HT-1080 cells were significantly potent, compared with Fab'-SPDP-LDM, Fab'-LCSPDP-LDM and free LDM. In animal models at the same condition, free LDM, Fab'-SPDP-LDM and Fab'-LCSPDP-LDM inhibited the growth of HT-1080 tumor by 70.9%, 74.8% and 72.3%, while Fab'-SMBS-LDM and Fab'-SSMPB-LDM reached 78.0% and 87.7%, respectively. The median survival time of the mice treated with free LDM, Fab'-SPDP-LDM and Fab'-LCSPDP-LDM were prolonged by 71.9%, 82.2% and 107.5%, respectively, compared with that of untreated group. Whereas, the median survival time of Fab'-SMBS-LDM and Fab'-SSMPB-LDM were prolonged by 145.2% and 165.8%, respectively, indicating that Fab'-SSMPB-LDM was more effective than Fab'-SMBS-LDM in tumor suppression and life span prolongation. Fab'-SSMPB-LDM has more marked selective antitumor efficacy and lower toxicity, and might be a novel candidate for cancer therapy.
Assuntos
Aminoglicosídeos/farmacologia , Antibióticos Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Colagenases/imunologia , Enedi-Inos/farmacologia , Fibrossarcoma/patologia , Imunoconjugados/farmacologia , Animais , Anticorpos Monoclonais/imunologia , Linhagem Celular Tumoral/efeitos dos fármacos , Humanos , Fragmentos Fab das Imunoglobulinas/imunologia , Inibidores de Metaloproteinases de Matriz , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transplante de Neoplasias , Carga Tumoral/efeitos dos fármacosRESUMO
Hepatocellular carcinoma (HCC) is a major challenge because of its resistance to conventional cytotoxic chemotherapy and radiotherapy. Multi-targeted therapy might be a new option for HCC treatment. Our previous study showed that N-ras gene was activated in HCC and was inhibited by RNA interference. In the present study, we investigated the alternation of gene expression by microarray in N-Ras-siRNA-treated HepG2 cells. The results revealed that the EREG gene, encoding epiregulin, was dramatically up-regulated in response to silence of N-ras. We speculated that the up-regulation of epiregulin was involved in the compensatory mechanism of N-ras knockdown for cell growth. Therefore, we evaluated whether dual silence of N-ras and epiregulin display a greater suppression of cell growth. The results confirmed that dual knockdown of N-ras and epiregulin synergistically inhibited cell growth. Our results also showed that dual knockdown of N-ras and epiregulin significantly induced cell arrest at G0/G1 phase. Furthermore, Western blot assay showed that dual knockdown of N-ras and epiregulin markedly reduced the phosphorylations of ERK1/2, Akt and Rb, and inhibited the expression of cyclin D1. Our findings imply that multi-targeted silence of oncogenes might be an effective treatment for HCC.
Assuntos
Carcinoma Hepatocelular/patologia , Fator de Crescimento Epidérmico/genética , Neoplasias Hepáticas/patologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/terapia , Linhagem Celular Tumoral , Proliferação de Células , Ciclina D1/antagonistas & inibidores , Ciclina D1/biossíntese , Epirregulina , Fase G1/genética , Técnicas de Silenciamento de Genes , Inativação Gênica , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/terapia , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 1 Ativada por Mitógeno/biossíntese , Proteína Quinase 3 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno/biossíntese , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/biossíntese , RNA Interferente Pequeno/genética , Fase de Repouso do Ciclo Celular/genética , Proteína do Retinoblastoma/antagonistas & inibidores , Proteína do Retinoblastoma/biossínteseRESUMO
Recent advances in cell cycle regulation have led to a suggestion of therapeutically targeting cell cycle checkpoint pathways in cancer cells to increase the toxicity of DNA-damaging agents. In this study, we investigate whether knockdowns of checkpoint kinases Chk1 and Chk2 by RNA interfering potentiate the cytotoxicity and abrogate G(2)/M checkpoint induced by DNA-damaging agent lidamycin (LDM) in HCT116 cells with different p53 status. Our results showed that Chk1 knockdown enhanced the cytotoxicity of LDM through abrogating G(2)/M arrest and increasing apoptosis to a greater extent in HCT116 p53(-/-) cells than in p53(wt) cells. Abrogation of LDM-induced G(2)/M arrest by Chk1 knockdown was associated with reducing the inactivated phosphorylations of Cdc25C and Cdc2. LDM-induced gamma-H2AX was increased in cells with Chk1 knockdown, indicating that DNA double-strand breaks (DSBs) were enhanced. Furthermore, knockdown of Chk1 also increased LDM-mediated apoptotic cell death in p53 knockout cells with activation of caspase-2 and caspase-3. On the contrary, knockdown of Chk2 had no impact on G(2)/M arrest or apoptosis induced by LDM. Moreover, dual knockdown of Chk1 and Chk2 failed to achieve better efficacy than Chk1 alone. Taken together, we suggest that Chk1 is a potential therapeutic target to sensitize human p53 deficient cancer cells to LDM.
Assuntos
Aminoglicosídeos/farmacologia , Antibióticos Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/enzimologia , Enedi-Inos/farmacologia , Proteínas Quinases/deficiência , Proteína Supressora de Tumor p53/metabolismo , Apoptose/genética , Western Blotting , Caspase 2/metabolismo , Caspase 3/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Divisão Celular/efeitos dos fármacos , Divisão Celular/genética , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/genética , Quinase 1 do Ponto de Checagem , Quinase do Ponto de Checagem 2 , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Cisteína Endopeptidases/metabolismo , Fase G2/efeitos dos fármacos , Fase G2/genética , Técnicas de Silenciamento de Genes , Células HCT116 , Humanos , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Proteínas Serina-Treonina Quinases/deficiência , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , RNA Interferente Pequeno/genética , TransfecçãoRESUMO
It has been reported that tetrandrine induces cell cycle arrest and apoptosis in human cancer cells. In the present study, we investigated the role of PI3K/AKT/GSK3beta pathway in tetrandrine- induced G(1) arrest and apoptosis. In HT-29 cells, tetrandrine induced dephosphorylation of AKT, activation and nuclear translocation of GSK3beta as well as upregulation of p27(kip1). Activation of GSK3beta via AKT inhibitoion induced by tetrandrine resulted in enhanced phosphorylation and proteolysis of cyclin D(1), activation of caspase 3 and subsequent cleavage of PARP. Selective GSK3beta inhibitiors and GSK3beta siRNA attenuated tetrandrine-induced G(1) arrest and apoptosis. Similar to tetrandrine, transfection of wild-type GSK3beta led to G(1) arrest and apoptosis via downregulation of cyclin D(1) and cleavage of PARP. These findings suggest that tetrandrine induces G(1) arrest and apoptosis through PI3K/AKT/GSK3beta pathway and identify GSK3beta as an important mediator in the processes.