RESUMO
The alpha1beta1 (VLA-1) integrin is a cell-surface receptor for collagen and laminin and has been implicated in biological pathways involved in several pathological processes. These processes may be inhibited by the monoclonal antibody AQC2, which binds with high affinity to human alpha1beta1 integrin. To understand the structural basis of the inhibition we determined the crystal structure of the complex of a chimeric rat/human I domain of the alpha1beta1 integrin and the Fab fragment of humanized AQC2 antibody. The structure of the complex shows that the antibody blocks the collagen binding site of the I domain. An aspartate residue, from the CDR3 loop of the antibody heavy chain, coordinates the MIDAS metal ion in a manner similar to that of a glutamate residue from collagen. Substitution of the aspartate residue by alanine or arginine results in significant reduction of antibody binding affinity. Interestingly, although the mode of metal ion coordination resembles that of the open conformation, the I domain maintains an overall closed conformation previously observed only for unliganded I domains.
Assuntos
Integrina alfa1beta1/química , Cristalografia por Raios X , Humanos , Fragmentos Fab das Imunoglobulinas , Mutagênese , Conformação Proteica , Proteínas Recombinantes de Fusão/químicaRESUMO
Interferon-beta (IFN-beta) induces various antiproliferative activities. In solid tumor cells, IFN-beta inhibits cell cycle progression, which mainly occurs as S phase accumulation. The IFN-beta-induced cell cycle effect has been implicated in the antitumor effect of combinations of IFN-beta and chemotherapeutic drugs. In this report, we characterized the viability of various human tumor cells in vitro after combination treatment with IFN-beta protein and the chemotherapeutic drugs, cis-platinum (II) diamine dichloride (cisplatin), 5-fluorouracil (5-FU), paclitaxel (Taxol) and gemcitabine. IFN-beta could significantly potentiate the cytotoxicity of these chemotherapeutic drugs. The potentiating effect was observed after pretreatment of tumor cells with IFN-beta but did not require the constant presence of IFN-beta. The potentiating effect correlated with the sensitivity of the tumor cells to the IFN-beta-induced cytotoxicity. Furthermore, chemotherapeutic drugs also potentiated the cytotoxicity of IFN-beta. We conclude that the cell cycle effect per se did not determine the ability of IFN-beta to potentiate the cytotoxicity of chemotherapeutic drugs. We suggest that the combination of local IFN-beta gene therapy with chemotherapy could be an effective cancer treatment.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Desoxicitidina/análogos & derivados , Interferon beta/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Neoplasias da Mama/patologia , Carcinoma/patologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Ciclo Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Cisplatino/administração & dosagem , Cisplatino/farmacologia , Desoxicitidina/administração & dosagem , Desoxicitidina/farmacologia , Sinergismo Farmacológico , Feminino , Fluoruracila/administração & dosagem , Fluoruracila/farmacologia , Humanos , Interferon beta/administração & dosagem , Neoplasias Pulmonares/patologia , Paclitaxel/administração & dosagem , Paclitaxel/farmacologia , Células Tumorais Cultivadas/efeitos dos fármacos , Neoplasias do Colo do Útero/patologia , GencitabinaRESUMO
The lymphotoxin-beta receptor (LT beta R) is a tumor necrosis factor receptor family member critical for the development and maintenance of various lymphoid microenvironments. Herein, we show that agonistic anti-LT beta R monoclonal antibody (mAb) CBE11 inhibited tumor growth in xenograft models and potentiated tumor responses to chemotherapeutic agents. In a syngeneic colon carcinoma tumor model, treatment of the tumor-bearing mice with an agonistic antibody against murine LT beta R caused increased lymphocyte infiltration and necrosis of the tumor. A pattern of differential gene expression predictive of cellular and xenograft response to LT beta R activation was identified in a panel of colon carcinoma cell lines and when applied to a panel of clinical colorectal tumor samples indicated 35% likelihood a tumor response to CBE11. Consistent with this estimate, CBE11 decreased tumor size and/or improved long-term animal survival with two of six independent orthotopic xenografts prepared from surgical colorectal carcinoma samples. Targeting of LT beta R with agonistic mAbs offers a novel approach to the treatment of colorectal and potentially other types of cancers.
Assuntos
Adenocarcinoma/terapia , Anticorpos Monoclonais/uso terapêutico , Neoplasias do Colo/terapia , Receptor beta de Linfotoxina/agonistas , Neoplasias do Colo do Útero/terapia , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/patologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Camptotecina/análogos & derivados , Camptotecina/uso terapêutico , Linhagem Celular Tumoral , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Terapia Combinada , Sinergismo Farmacológico , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Imunoglobulina G/imunologia , Imunoglobulina G/uso terapêutico , Imunoglobulina M/imunologia , Imunoglobulina M/uso terapêutico , Irinotecano , Linfócitos do Interstício Tumoral/imunologia , Receptor beta de Linfotoxina/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Distribuição Aleatória , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/uso terapêutico , Método Simples-Cego , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The integrin alpha1beta1 (very late antigen-1; CD49a/CD29) is a major adhesion receptor for collagen I, IV, and VI, and its induced expression on activated monocytes and lymphocytes plays a central role in their retention and activation at inflammatory sites in autoimmune pathologies. However, the role of alpha1beta1 in allergic settings has not been explored. In this study, we show that a single 45-mg dose of aerosolized monoclonal antibody AQC2 to the alpha1 chain of human and sheep very late antigen-1, given 30 minutes before challenge, blocks both the allergen-induced late response and the associated airway hyperresponsiveness, functional indicators of allergen-induced inflammation, in sheep. AQC2 does not affect the early response. Consistent with these effects, AQC2 tended to reduce the cell response associated with local antigen instillation. An isotype-matched control antibody had no protective effects. Two humanized versions of AQC2, a wild-type IgG1 and an aglycosyl form of the same monoclonal antibody, which has reduced Fc receptor-mediated effector functions, are equally effective in blocking the antigen-induced late response and airway hyperresponsiveness in the sheep model. These data suggest that mononuclear leukocyte adhesion-dependent pathologies contribute to allergic lung disease and provide proof-of-concept that antagonists of alpha1 integrins may be useful in preventing these events.