RESUMO
Interactions between germline-encoded natural killer (NK) cell receptors and their respective ligands on tumorigenic or virus-infected cells determine NK cell cytotoxic activity and/or cytokine secretion. NK cell cytokine responses can be augmented in and can potentially contribute to multiple sclerosis (MS), an inflammatory disease of the central nervous system focused upon the oligodendrocytes (OLs). To investigate mechanisms by which NK cells may contribute to MS pathogenesis, we developed an in vitro human model of OL-NK cell interaction. We found that activated, but not resting human NK cells form conjugates with, and mediate cytotoxicity against, human oligodendrocytes. NK cells, when in conjugate with OLs, rapidly synthesize and polarize IFN-γ toward the OLs. IFN-γ is capable of reducing myelin oligodendrocyte and myelin associated glycoproteins (MOG and MAG) content. This activity is independent of MHC class-I mediated inhibition via KIR2DL1, but dependent upon the interaction between NK cell-expressed KIR2DL4 and its oligodendrocyte-expressed ligand, HLA-G. NK cells from patients with MS express higher levels of IFN-γ following conjugation to OLs, more actively promote in vitro reduction of MOG and MAG and have higher frequencies of the KIR2DL4 positive population. These data collectively suggest a mechanism by which NK cells can promote pathogenic effects upon OLs.
Assuntos
Interferon gama/imunologia , Células Matadoras Naturais/imunologia , Oligodendroglia/imunologia , Receptores KIR2DL4/imunologia , Linhagem Celular , Citotoxicidade Imunológica/imunologia , Antígenos HLA-G/imunologia , Humanos , Esclerose Múltipla/imunologia , Glicoproteína Associada a Mielina/imunologia , Receptores de Células Matadoras Naturais/imunologiaRESUMO
The mountain peacock pheasant (Polyplectron inopinatum), the Malayan peacock pheasant (Polyplectron malacense), and the Congo peafowl (Afropavo congensis) are all listed as vulnerable to extinction under the International Union for Conservation of Nature Red List of Threatened Species. Here the authors report fatal infection with a novel herpesvirus in all 3 species of birds. DNA was extracted from the livers of birds with hepatocellular necrosis and intranuclear eosinophilic inclusions consistent with herpesvirus infection. Based on degenerate herpesvirus primers and polymerase chain reaction, 220- and 519-base pair products of the herpes DNA polymerase and DNA terminase genes, respectively, were amplified. Sequence analysis revealed that all birds were likely infected with the same virus. At the nucleotide level, the pheasant herpesvirus had 92% identity with gallid herpesvirus 3 and 77.7% identity with gallid herpesvirus 2. At the amino acid level, the herpes virus had 93.8% identity with gallid herpesvirus 3 and 89.4% identity with gallid herpesvirus 2. These findings indicate that the closest relative to this novel herpesvirus is gallid herpesvirus 3, a nonpathogenic virus used widely in a vaccine against Marek's disease. In situ hybridization using probes specific to the peacock pheasant herpesvirus DNA polymerase revealed strong intranuclear staining in the necrotic liver lesions of an infected Malayan peacock pheasant but no staining in normal liver from an uninfected bird. The phasianid herpesvirus reported here is a novel member of the genus Mardivirus of the subfamily Alphaherpesvirinae and is distinct from other galliform herpesviruses.
Assuntos
Animais de Zoológico/virologia , Doenças das Aves/virologia , Espécies em Perigo de Extinção , Galliformes , Infecções por Herpesviridae/veterinária , Fígado/virologia , Mardivirus/genética , Animais , Sequência de Bases , Doenças das Aves/mortalidade , Primers do DNA/genética , DNA Polimerase Dirigida por DNA/genética , Endodesoxirribonucleases/genética , Infecções por Herpesviridae/mortalidade , Hibridização In Situ/veterinária , Fígado/patologia , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/veterinária , Análise de Sequência de DNA/veterinária , Homologia de Sequência , Especificidade da EspécieRESUMO
Changes in the proteome of colon mucosal cells accompany the transition from normal mucosa via adenoma and invasive cancer to metastatic disease. Samples from 15 patients with sporadic sigmoid cancers were analyzed. Proteins were separated by two-dimensional gel electrophoresis. Relative differences in expression levels between normal tissue, adenoma, carcinoma and metastasis were evaluated in both intra- and inter-patient comparisons. Up- and down-regulated proteins (> twofold) during development to cancer or metastasis were excised and submitted to peptide mass fingerprinting and MS/MS sequence analysis, facilitated by the use of a compact disc workstation. In total, 112 protein spots were found to be differentially regulated, of which 72 were determined as to protein identity, 46 being up-regulated toward the progression of cancer, and 26 down-regulated. Several of the identifications correlate with proteins of the cell cycle, cytoskeleton or metabolic pathways. The pattern changes now identified have the potential for design of marker panels for assistance in diagnostics and therapeutic strategies in colorectal cancer.
Assuntos
Neoplasias do Colo/metabolismo , Proteoma , Adenoma/metabolismo , Idoso , Idoso de 80 Anos ou mais , Carcinoma/metabolismo , Núcleo Celular/metabolismo , Neoplasias do Colo/patologia , Biologia Computacional , Progressão da Doença , Regulação para Baixo , Eletroforese em Gel Bidimensional , Feminino , Humanos , Masculino , Espectrometria de Massas , Pessoa de Meia-Idade , Mucosa/patologia , Metástase Neoplásica , Peptídeos/química , Ploidias , Regulação para CimaRESUMO
T cell-mediated destruction of the myelin sheath causes inflammatory damage of the CNS in multiple sclerosis (MS). The major T and B cell responses in MS patients who are HLA-DR2 (about two-thirds of MS patients) react to a region between residues 84 and 103 of myelin basic protein (1 ). The crystal structure of HLA-DR2 complexed with myelin basic protein(84-102) confirmed that Lys(91) is the major TCR contact site, whereas Phe(90) is a major anchor to MHC and binds the hydrophobic P4 pocket (2 ). We have tested peptides containing repetitive 4-aa sequences designed to bind critical MHC pockets and to interfere with T cell activation. One such sequence, EYYKEYYKEYYK, ameliorates experimental autoimmune encephalomyelitis in Lewis rats, an animal model of MS.
Assuntos
Encefalomielite Autoimune Experimental/imunologia , Antígenos de Histocompatibilidade/metabolismo , Peptídeos/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Adjuvantes Imunológicos/farmacologia , Sequência de Aminoácidos , Animais , Sítios de Ligação/genética , Encefalomielite Autoimune Experimental/tratamento farmacológico , Feminino , Antígeno HLA-DR2/metabolismo , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Esclerose Múltipla/tratamento farmacológico , Esclerose Múltipla/imunologia , Proteína Básica da Mielina/genética , Proteína Básica da Mielina/imunologia , Proteína Básica da Mielina/farmacologia , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Fragmentos de Peptídeos/farmacologia , Peptídeos/genética , Peptídeos/farmacologia , Ratos , Ratos Endogâmicos Lew , Homologia de Sequência de Aminoácidos , Linfócitos T/imunologiaRESUMO
Progress in developing a small animal model of human immunodeficiency virus type 1 (HIV-1) disease would greatly facilitate studies of transmission, pathogenesis, host immune responses, and antiviral strategies. In this study, we have explored the potential of rats as a susceptible host. In a single replication cycle, rat cell lines Rat2 and Nb2 produced infectious virus at levels 10- to 60-fold lower than those produced by human cells. Rat-derived cells supported substantial levels of early HIV-1 gene expression, which was further enhanced by overexpression of human cyclin T1. Rat cells displayed quantitative, qualitative, and cell-type-specific limitations in the late phase of the HIV-1 replication cycle including relative expression levels of HIV-1 Gag proteins, intracellular Gag processing, and viral egress. Nb2 cells were rendered permissive to HIV-1 R5 viruses by coexpression of human CD4 and CCR5, indicating that the major restriction on HIV-1 replication was at the level of cellular entry. We also found that primary rat lymphocytes, macrophages, and microglia expressed considerable levels of early HIV-1 gene products following infection with pseudotyped HIV-1. Importantly, primary rat macrophages and microglia, but not lymphocytes, also expressed substantial levels of HIV-1 p24 CA and produced infectious virions. Collectively, these results identify the rat as a promising candidate for a transgenic small animal model of HIV-1 infection and highlight pertinent cell-type-specific restrictions that are features of this species.
Assuntos
Infecções por HIV/virologia , HIV-1/fisiologia , Macrófagos/virologia , Glicoproteínas de Membrana , Microglia/virologia , Linfócitos T/virologia , Replicação Viral , Animais , Antígenos CD4/metabolismo , Linhagem Celular , Células Cultivadas , Ciclina T , Ciclinas/metabolismo , Modelos Animais de Doenças , Repetição Terminal Longa de HIV/fisiologia , HIV-1/genética , Humanos , Camundongos , Ratos , Receptores CCR5/metabolismo , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/metabolismo , Proteínas Virais/genética , Proteínas Virais/metabolismoRESUMO
Multigenic programs controlling susceptibility to apoptosis in response to ionizing radiation have not yet been defined. Here, using DNA microarrays, we show gene expression patterns in an apoptosis-sensitive and apoptosis-resistant murine B cell lymphoma model system both before and after irradiation. From the 11,000 genes interrogated by the arrays, two major patterns emerged. First, before radiation exposure the radioresistant LYar cells expressed significantly greater levels of message for several genes involved in regulating intracellular redox potential. Compared with LYas cells, LYar cells express 20- to 50-fold more mRNA for the tetraspanin CD53 and for fructose-1,6-bisphosphatase. Expression of both of these genes can lead to the increase of total cellular glutathione, which is the principle intracellular antioxidant and has been shown to inhibit many forms of apoptosis. A second pattern emerged after radiation, when the apoptosis-sensitive LYas cells induced rapid expression of a unique cluster of genes characterized by their involvement in mitochondrial electron transport. Some of these genes have been previously recognized as proapoptotic; however others, such as uncoupling protein 2, were not previously known to be apoptotic regulatory proteins. From these observations we propose that a multigenic program for sensitivity to apoptosis involves induction of transcripts for genes participating in mitochondrial uncoupling and loss of membrane potential. This program triggers mitochondrial release of apoptogenic factors and induces the "caspase cascade." Conversely, cells resistant to apoptosis down-regulate these biochemical pathways, while activating pathways for establishment and maintenance of high intracellular redox potential by means of elevated glutathione.
Assuntos
Apoptose/genética , Proteínas de Membrana Transportadoras , Mitocôndrias/genética , Proteínas Mitocondriais , Proteínas de Neoplasias , Proteínas do Tecido Nervoso , Análise de Sequência com Séries de Oligonucleotídeos , Oxirredução , Animais , Anexinas/metabolismo , Antígenos CD/genética , Antígenos de Diferenciação de Linfócitos T/genética , Proteínas de Transporte/genética , Análise por Conglomerados , Proteína 7 de Ligação a Ácidos Graxos , Proteínas de Ligação a Ácido Graxo , Frutose-Bifosfatase/genética , Regulação Neoplásica da Expressão Gênica , Canais Iônicos , Cinética , Camundongos , Modelos Biológicos , Proteína P2 de Mielina/genética , Porinas/genética , Proteínas/genética , Espectrometria de Fluorescência , Tetraspanina 25 , Fatores de Tempo , Células Tumorais Cultivadas , Proteína Desacopladora 2 , Regulação para Cima , Canais de Ânion Dependentes de VoltagemRESUMO
Molecular mimicry refers to structural homologies between a self-protein and a microbial protein. A major epitope of myelin basic protein (MBP), p87-99 (VHFFKNIVTPRTP), induces experimental autoimmune encephalomyelitis (EAE). VHFFK contains the major residues for binding of this self-molecule to T cell receptor (TCR) and to the major histocompatibility complex. Peptides from papilloma virus strains containing the motif VHFFK induce EAE. A peptide from human papilloma virus type 40 (HPV 40) containing VHFFR, and one from HPV 32 containing VHFFH, prevented EAE. A sequence from Bacillus subtilis (RKVVTDFFKNIPQRI) also prevented EAE. T cell lines, producing IL-4 and specific for these microbial peptides, suppressed EAE. Thus, microbial peptides, differing from the core motif of the self-antigen, MBPp87-99, function as altered peptide ligands, and behave as TCR antagonists, in the modulation of autoimmune disease.
Assuntos
Proteínas de Bactérias/imunologia , Encefalomielite Autoimune Experimental/imunologia , Epitopos/imunologia , Ligantes , Fragmentos de Peptídeos/imunologia , Linfócitos T/imunologia , Sequência de Aminoácidos , Animais , Divisão Celular/imunologia , Encefalomielite Autoimune Experimental/prevenção & controle , Feminino , Interferon gama/metabolismo , Interleucina-4/metabolismo , Camundongos , Camundongos Endogâmicos , Dados de Sequência Molecular , Proteína Básica da Mielina/imunologia , Receptores de Antígenos de Linfócitos T/antagonistas & inibidoresRESUMO
The p53 molecule might serve as a common tumor-associated antigen, as the tumor suppressor gene p53 is mutated and the p53 protein is often over-expressed in tumor cells. We report that effective immunity to p53 can be induced through an idiotypic network by immunization of mice with a monoclonal antibody (PAb-240) specific for mutated p53, or with a peptide derived from the complementarity determining region (CDR) 3 of the variable domain of the light chain (VL) of this antibody. The immunized mice produced IgG antibodies to p53 and mounted a cytotoxic reaction to a tumor line bearing mutated p53. The idiotypically immunized mice were resistant to challenge with the tumor cells. Thus antibodies to p53 might serve as immunogens for activating resistance to some tumors. At the basic level, these findings indicate that a network of p53 immunity may be organized naturally within the immune system.
Assuntos
Anticorpos Anti-Idiotípicos/imunologia , Anticorpos Monoclonais/imunologia , Neoplasias Experimentais/imunologia , Proteína Supressora de Tumor p53/imunologia , Animais , Anticorpos Anti-Idiotípicos/administração & dosagem , Anticorpos Monoclonais/administração & dosagem , Formação de Anticorpos/efeitos dos fármacos , Epitopos/administração & dosagem , Epitopos/química , Epitopos/imunologia , Feminino , Rejeição de Enxerto/genética , Rejeição de Enxerto/imunologia , Imunidade/efeitos dos fármacos , Cadeias Leves de Imunoglobulina/genética , Cadeias Leves de Imunoglobulina/imunologia , Região Variável de Imunoglobulina/genética , Região Variável de Imunoglobulina/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Mutação/genética , Transplante de Neoplasias/imunologia , Neoplasias Experimentais/fisiopatologia , Oligopeptídeos/administração & dosagem , Oligopeptídeos/síntese química , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/imunologia , Proteína Supressora de Tumor p53/genética , VacinaçãoRESUMO
Macrophages have long been known to play a key role in the healing processes of tissues that regenerate after injury; however, the nature of their involvement in healing of the injured central nervous system (CNS) is still a subject of controversy. Here we show that the absence of regrowth in transected rat optic nerve (which, like all other CNS nerves in mammals, cannot regenerate after injury) can be overcome by local transplantation of macrophages preincubated ex vivo with segments of a nerve (e.g., sciatic nerve) that can regenerate after injury. The observed effect of the transplanted macrophages was found to be an outcome of their stimulated activity, as indicated by phagocytosis. Thus, macrophage phagocytic activity was stimulated by their preincubation with sciatic nerve segments but inhibited by their preincubation with optic nerve segments. We conclude that the inability of nerves of the mammalian CNS to regenerate is related to the failure of their macrophages recruited after injury to acquire growth-supportive activity. We attribute this failure to the presence of a CNS resident macrophage inhibitory activity, which may be the biochemical basis underlying the immune privilege of the CNS. The transplantation of suitably activated macrophages into injured nerves may overcome multiple malfunctioning aspects of the CNS response to trauma, and thus may be developed into a novel, practical, and multipotent therapy for CNS injuries.
Assuntos
Macrófagos/fisiologia , Regeneração Nervosa , Nervo Óptico/fisiologia , Animais , Ativação de Macrófagos , Macrófagos/transplante , Masculino , Traumatismos do Nervo Óptico , Ratos , Ratos Sprague-DawleyRESUMO
Compared to the peripheral nervous system (PNS), the central nervous system (CNS) of mammals has a poor prospect for regeneration. Accumulating evidence suggests that this is due, in part, to differences in how the immune and nervous systems communicate in response to injury. The macrophage is one of the central cells in this communication with the capacity to respond in a variety of ways depending on the conditions of stimulation. After injury, macrophages enter the CNS much later and in fewer numbers than they do the PNS. It is possible that this late and reduced response is not sufficient to modify the CNS environment to one that is conducive to successful regeneration. In the present study we investigated whether the limited macrophage invasion of injured CNS is due to the presence of an endogenous inhibitory factor that is persistent after injury. Using an in vitro migration assay, we show that rat optic nerve (CNS) is deficient in its ability to attract monocytes as compared to rat sciatic nerve (PNS). We further demonstrate that this deficiency is due to the presence of a soluble inhibitory factor in the CNS. This factor may also cause a subsequent effective difference in those macrophages that are recruited, as is shown by morphological data. The brain-resident factor that inhibits macrophage migration may be the physiological basis of an immune-brain barrier underlying the known phenomenon of immune privilege.
Assuntos
Macrófagos/imunologia , Nervo Óptico/imunologia , Animais , Quimiotaxia de Leucócito , Técnicas In Vitro , Ativação de Macrófagos , Macrófagos/citologia , Neuroimunomodulação , Ratos , Ratos Sprague-Dawley , Nervo Isquiático/imunologiaAssuntos
Sistema Nervoso Central/fisiologia , Sistema Imunitário/fisiologia , Macrófagos/fisiologia , Regeneração Nervosa/fisiologia , Animais , Sistema Nervoso Central/citologia , Sistema Nervoso Central/imunologia , Humanos , Sistema Imunitário/citologia , Sistema Imunitário/metabolismo , Macrófagos/metabolismo , Regeneração Nervosa/imunologiaAssuntos
Sistema Nervoso Central/lesões , Citocinas/fisiologia , Macrófagos/fisiologia , Neuroglia/fisiologia , Animais , Axônios/fisiologia , Sistema Nervoso Central/fisiopatologia , Interleucina-2/fisiologia , Fator Estimulador de Colônias de Macrófagos/fisiologia , Compressão Nervosa , Degeneração Neural , Regeneração Nervosa , Neuroimunomodulação , Nervo Óptico/fisiologia , Traumatismos do Nervo Óptico , Células PC12 , Coelhos , Ratos , Nervo Isquiático/fisiologia , Fator de Necrose Tumoral alfa/fisiologia , Vertebrados/fisiologiaRESUMO
Measurements in cancer patients showed that the pH of tumors averages 0.8 unit lower than that of the surrounding normal tissues, confirming published work. Based on this, the anti-carcinoma monoclonal antibody (mAb) L6 was used to prepare immunoconjugates with daunomycin (DM), the drug being released at the acidic pH of the tumor. A direct linking of the aconitic derivative of DM (AcoDM) to mAb L6 led to conjugates that either had a low drug/antibody ratio (less than 5:1) or precipitated in vitro. In order to increase the drug load and avoid precipitation, several biopolymers were tested as spacers between the drug and the L6. To attach the polymer derivative to the mAb, the former was maleimidized and the mAb was thiolated. The AcoM/mAb ratio obtained was 20, and the mAb retained its highly specific binding to tumor cells. At pH 6 the AcoDM-L6 conjugate was toxic to cultured C-3347 carcinoma cells with an inhibitory concentration (IC50) of 5 micrograms/ml. The conjugate was less effective than the free DM with an IC50 of 0.2 micrograms/ml. The L6 alone was not toxic. At a tumor pH of 6.5, 15% of the AcoDM was released. The amount of released drug reached a maximum 24-48 h after exposure to the acidic medium. In vivo localization studies demonstrated a similar tumor uptake of the conjugate and mAb L6 with 18% of the injected dose/g tumor and a maximum uptake in tumor 48 h after injection. Our data indicate that it is possible to construct conjugates based on a pH-sensitive linker that can be targeted successfully to a tumor with release of a portion of the drug at the tumor site, but testing is needed to establish whether such release has anti-tumor activity in vivo and offers an advantage over treatment with unconjugated drug.
Assuntos
Daunorrubicina/farmacocinética , Imunotoxinas/farmacocinética , Neoplasias/metabolismo , Idoso , Animais , Anticorpos Monoclonais/administração & dosagem , Daunorrubicina/administração & dosagem , Feminino , Humanos , Concentração de Íons de Hidrogênio , Imunotoxinas/química , Masculino , Camundongos , Pessoa de Meia-Idade , Distribuição Tecidual , Células Tumorais CultivadasRESUMO
Mammalian central nervous system (CNS) axons are virtually incapable of regenerating after injury. However, CNS neurons of lower vertebrates, such as fish and amphibians, are endowed with a high regenerative capacity. Lately, the glial cells have been credited with the regenerative ability of any specific CNS. We have previously demonstrated that many oligodendrocytes are recovered in cultures of injured rat optic nerve, while only a few oligodendrocytes are recovered from injured fish optic nerve in culture. We further demonstrated that medium conditioned by regenerating fish optic nerves (CM), which has been shown to cause axonal elongation in injured rabbit optic nerves, causes a decrease in the number of oligodendrocytes in rat glial cultures. In the present study, we demonstrate that soluble factors in the CM are capable of reducing the number of fish oligodendrocytes in fish optic nerve cultures. In addition, an inverse relationship was found between the number of macrophages and the number of oligodendrocytes. These results thus suggest that macrophages and/or activated resident microglial cells are directly or indirectly responsible for the presence of these soluble factor(s) that regulate the postinjury number of oligodendrocytes in the fish optic nerves.
Assuntos
Carpa Dourada/fisiologia , Macrófagos/fisiologia , Oligodendroglia/citologia , Traumatismos do Nervo Óptico , Animais , Contagem de Células , Regeneração Nervosa , Nervo Óptico/metabolismo , Nervo Óptico/fisiologia , Técnicas de Cultura de ÓrgãosRESUMO
The two monoclonal antibodies (mAb), L6 (anti-carcinoma), and 1F5 [anti-(B-cell-lymphoma)], were chemically linked to the enzyme penicillin-V amidase (PVA), which hydrolyzes phenoxyacetamides, to explore the potential of using mAb-enzyme conjugates for the localization of chemotherapeutic drugs at tumor cells. The phenoxyacetamide derivatives of doxorubicin and melphalan were prepared, yielding the less toxic amides, doxorubicin-N-p-hydroxyphenoxyacetamide (DPO) and melphalan-N-p-hydroxyphenoxyacetamide (MelPO). These were hydrolyzed by PVA to doxorubicin and melphalan respectively. In vitro studies with the L6-positive lung carcinoma cell line, H2981, and the 1F5-positive B-cell lymphoma line, Daudi, showed that DPO was 80-fold less toxic to H2981 cells and 20-fold less toxic to Daudi cells than doxorubicin, and its toxicity was substantially increased when the H2981 cells were pretreated with L6-PVA or the Daudi cells were pretreated with 1F5-PVA. The cytotoxic effect was antigen-specific, since only the binding mAb-enzyme conjugate increased the cytotoxicity of the prodrug. MelPO was more than 1000-fold less toxic than melphalan to H2981 cells and more than 100-fold less toxic than melphalan to Daudi cells. Pretreatment with the mAb-PVA conjugates did not enhance the toxicity of MelPO in either cell line, because PVA hydrolyzes the phenoxyacetamide bond of MelPO too slowly to generate a toxic level of melphalan.
Assuntos
Amidoidrolases/farmacologia , Imunotoxinas/farmacologia , Penicilina Amidase/farmacologia , Pró-Fármacos/farmacologia , Células Tumorais Cultivadas/efeitos dos fármacos , Anticorpos Monoclonais , Antígenos de Neoplasias/imunologia , Carcinoma/tratamento farmacológico , Carcinoma/imunologia , Carcinoma/patologia , Doxorrubicina/análogos & derivados , Doxorrubicina/metabolismo , Doxorrubicina/farmacologia , Doxorrubicina/uso terapêutico , Humanos , Imunotoxinas/síntese química , Linfoma/tratamento farmacológico , Linfoma/imunologia , Linfoma/patologia , Melfalan/análogos & derivados , Melfalan/metabolismo , Melfalan/farmacologia , Penicilina V/farmacologia , Pró-Fármacos/metabolismoRESUMO
Alkaline phosphatase (AP) was covalently linked to the two antitumor monoclonal antibodies, L6 (anticarcinoma) and 1F5 (anti-B lymphoma), forming conjugates that could bind to antigen-positive tumor cells. The conjugates were able to convert the prodrugs, mitomycin phosphate (MOP) and etoposide phosphate (EP), into an active mitomycin C derivative, mitomycin alcohol, and etoposide, respectively. MOP and EP were less toxic to cultured cells from the H2981 lung adenocarcinoma than their respective hydrolysis products, mitomycin alcohol and etoposide, by a factor greater than 100, and they were also less toxic in mice. Pretreatment of H2981 cells with L6-AP greatly enhanced the cytotoxic effects of MOP and EP, while 1F5-AP caused no such enhancement. A strong antitumor response was observed in H2981-bearing mice that were treated with L6-AP followed 24 h later by either MOP or a combination of MOP and EP. This response was superior to that of MOP or combinations of MOP and EP given alone.
Assuntos
Adenocarcinoma/tratamento farmacológico , Fosfatase Alcalina/uso terapêutico , Etoposídeo/análogos & derivados , Neoplasias Pulmonares/tratamento farmacológico , Mitomicinas/uso terapêutico , Compostos Organofosforados , Células Tumorais Cultivadas/citologia , Adenocarcinoma/patologia , Fosfatase Alcalina/metabolismo , Animais , Anticorpos Monoclonais/uso terapêutico , Biotransformação , Etoposídeo/metabolismo , Etoposídeo/farmacologia , Etoposídeo/uso terapêutico , Feminino , Humanos , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Nus , Mitomicinas/metabolismo , Mitomicinas/farmacologia , Transplante de Neoplasias , Transplante Heterólogo , Células Tumorais Cultivadas/efeitos dos fármacosRESUMO
Two anti-tumor monoclonal antibodies, L6 (anticarcinoma) and 1F5 (anti-B lymphoma), were covalently linked to alkaline phosphatase (AP), forming conjugates that could bind to the surface of antigen-positive tumor cells. The conjugates were capable of converting a relatively noncytotoxic prodrug, etoposide phosphate (EP), into etoposide--a drug with significant antitumor activity. In vitro studies with a human colon carcinoma cell line, H3347, demonstrated that while EP was less toxic than etoposide by a factor of greater than 100, it was equally toxic when the cells were pretreated with L6-AP, a conjugate that bound to the surface of H3347 cells. The L6-AP conjugate localized in H3347 tumor xenografts in nude mice and histological evaluation indicated that the targeted enzyme (AP) was distributed throughout the tumor mass. A strong antitumor response was observed in H3347-bearing mice that were treated with L6-AP followed 18-24 hr later by EP. This response, which included the rejection of established tumors, was superior to that of EP (P less than 0.005) or etoposide (P less than 0.001) given alone. The IF5-AP conjugate did not bind to H3347 cells and did not enhance the toxicity of EP on these cells in vitro. In addition, IF5-AP did not localize to H3347 tumors in nude mice and did not demonstrate enhanced antitumor activity in combination with the prodrug.