RESUMO
The aim of this study was to enhance the effectiveness of photo thermal therapy (PTT) in the targeting of superficial bladder cancers using a green light laser in conjunction with gold nanoparticles (GNPs) conjugated to antibody fragments (anti-EGFR). GNPs conjugated with anti-EGFR-antibody fragments were used as probes in the targeting of tumor cells and then exposed to a green laser (532nm), resulting in the production of sufficient thermal energy to kill urothelial carcinomas both in vitro and in vivo. Nanoparticles conjugated with antibody fragments are capable of damaging cancer cells even at relatively very low energy levels, while non-conjugated nanoparticles would require an energy level of 3 times under the same conditions. The lower energy required by the nanoparticles allows this method to destroy cancerous cells while preserving normal cells when applied in vivo. Nanoparticles conjugated with antibody fragments (anti-EGFR) require less than half the energy of non-conjugated nanoparticles to kill cancer cells. In an orthotopic bladder cancer model, the group treated using PTT presented significant differences in tumor development.
Assuntos
Receptores ErbB/antagonistas & inibidores , Receptores ErbB/imunologia , Terapia com Luz de Baixa Intensidade/métodos , Nanoconjugados/uso terapêutico , Neoplasias da Bexiga Urinária/terapia , Animais , Anticorpos/administração & dosagem , Linhagem Celular Tumoral , Ouro , Humanos , Fragmentos de Imunoglobulinas/administração & dosagem , Nanopartículas Metálicas/uso terapêutico , Nanopartículas Metálicas/ultraestrutura , Camundongos , Camundongos Endogâmicos C3H , Nanoconjugados/ultraestrutura , Nanotecnologia , Neoplasias da Bexiga Urinária/ultraestruturaRESUMO
Chronic or excessive (+)-methamphetamine (METH) use often leads to addiction and toxicity to critical organs like the brain. With medical treatment as a goal, a novel single-chain variable fragment (scFv) against METH was engineered from anti-METH monoclonal antibody mAb6H4 (IgG, kappa light chain, K(d) = 11 nM) and found to have similar ligand affinity (K(d) = 10 nM) and specificity as mAb6H4. The anti-METH scFv (scFv6H4) was cloned, expressed in yeast, purified, and formulated as a naturally occurring mixture of monomer ( approximately 75%) and dimer ( approximately 25%). To test the in vivo efficacy of the scFv6H4, male Sprague-Dawley rats (n = 5) were implanted with 3-day s.c. osmotic pumps delivering 3.2 mg/kg/day METH. After reaching steady-state METH concentrations, an i.v. dose of scFv6H4 (36.5 mg/kg, equimolar to the METH body burden) was administered along with a [(3)H]scFv6H4 tracer. Serum pharmacokinetic analysis of METH and [(3)H]scFv6H4 showed that the scFv6H4 caused an immediate 65-fold increase in the METH concentrations and a 12-fold increase in the serum METH area under the concentration-time curve from 0 to 480 min after scFv6H4 administration. The scFv6H4 monomer was quickly cleared or converted to multivalent forms with an apparent t(1/2lambdaz) of 5.8 min. In contrast, the larger scFv6H4 multivalent forms (dimers, trimers, etc.) showed a much longer t(1/2lambdaz) (228 min), and the significantly increased METH serum molar concentrations correlated directly with scFv6H4 serum molar concentrations. Considered together, these data suggested that the scFv6H4 multimers (and not the monomer) were responsible for the prolonged redistribution of METH into the serum.
Assuntos
Anticorpos Monoclonais/farmacocinética , Fragmentos de Imunoglobulinas/uso terapêutico , Metanfetamina/sangue , Metanfetamina/imunologia , Animais , Anticorpos Monoclonais/uso terapêutico , Especificidade de Anticorpos , Dimerização , Avaliação Pré-Clínica de Medicamentos , Fragmentos de Imunoglobulinas/administração & dosagem , Fragmentos de Imunoglobulinas/farmacologia , Região Variável de Imunoglobulina , Masculino , Ratos , Ratos Sprague-DawleyAssuntos
Células Endoteliais/efeitos dos fármacos , Fragmentos de Imunoglobulinas/administração & dosagem , Molécula 1 de Adesão de Célula Vascular/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/farmacologia , Antineoplásicos , Biomarcadores Tumorais/imunologia , Biomarcadores Tumorais/metabolismo , Avaliação Pré-Clínica de Medicamentos , Células Endoteliais/imunologia , Células Endoteliais/metabolismo , Fragmentos de Imunoglobulinas/farmacologia , Lipossomos , Camundongos , Receptores Fc/metabolismo , Células Tumorais Cultivadas , Molécula 1 de Adesão de Célula Vascular/metabolismoRESUMO
Recombinant immunotoxins are fusion proteins which contain a ligand derived from the immune system fused to a toxin. The protein toxin is truncated to delete its binding domain, allowing selective ligand-directed binding. Growth factor fusion toxins are often considered immunotoxins. One of these molecules, containing the truncated diphtheria toxin and human IL-2 (Ontak), Ligand Pharmaceuticals), has been approved for the treatment of cutaneous T-cell lymphoma. Recombinant immunotoxins have also been produced containing the variable domains (Fv fragment) of monoclonal antibodies fused to toxins. These agents are relatively versatile with respect to the range of antigens possible. Several of these recombinant immunotoxins have showed clinical effectiveness in Phase I testing against haematological malignancies. One of these molecules, BL22, targets CD22 on hairy-cell leukaemia and has enabled patients to achieve complete remissions despite previous treatment and resistance to chemotherapy.
Assuntos
Neoplasias Hematológicas/tratamento farmacológico , Imunotoxinas/uso terapêutico , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral/efeitos dos fármacos , Ensaios Clínicos Fase I como Assunto , Ensaios Clínicos Fase II como Assunto , Terapia Combinada , Desenho de Fármacos , Avaliação Pré-Clínica de Medicamentos , Previsões , Neoplasias Hematológicas/imunologia , Humanos , Fragmentos de Imunoglobulinas/administração & dosagem , Fragmentos de Imunoglobulinas/uso terapêutico , Imunotoxinas/administração & dosagem , Imunotoxinas/química , Macaca fascicularis , Camundongos , Estrutura Molecular , Proteínas de Neoplasias/imunologia , Proteínas Recombinantes de Fusão/uso terapêutico , Indução de Remissão , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The antibody levels in 18 batches of the preparations of human immunoglobulin, Immunovenin and Immunovenin-Intact, for intravenous injection were determined in the enzyme immunoassay with the use of the mixture of P. aeruginosa lipopolysaccharide antigens of seven immunotypes. The average antibody titers in these preparations were identical. The preparations were found to have protective action against P. aeruginosa experimental infection in mice.
Assuntos
Fragmentos de Imunoglobulinas/administração & dosagem , Imunoglobulina G/administração & dosagem , Infecções por Pseudomonas/prevenção & controle , Animais , Anticorpos Antibacterianos/análise , Avaliação Pré-Clínica de Medicamentos , Humanos , Imunização Passiva , Técnicas Imunoenzimáticas , Fragmentos de Imunoglobulinas/análise , Imunoglobulina G/análise , Injeções Intravenosas , Lipopolissacarídeos/imunologia , Camundongos , Infecções por Pseudomonas/imunologia , Pseudomonas aeruginosa/imunologiaRESUMO
(N-(2-Hydroxypropyl)methacrylamide (HPMA)) copolymers have seen extensive development as sophisticated lysosomotropic drug carriers. They can be used for site-specific drug delivery by incorporation of appropriate targeting groups and here we report their conjugation to antitumour monoclonal antibodies (the murine IgG, antibody B72.3 and its Fab' and Fab'2 fragments) and assessment as vehicles for tumour-specific drug delivery. Conjugates were synthesised containing an average 5 copolymer units (Mw 20kD) per antibody molecule. Kinetics of elimination and body distribution of radiolabelled conjugates in mice were substantially modified compared with native antibody and fragments, showing prolonged circulation in the bloodstream. Notably, the half-time for bloodclearance of the Fab' fragment (35min) was extended ten-fold following conjugation (6h). The conjugates provoked only a low immune response in A/J mice, following three injections in adjuvant (IgG titre-1 less than 100), and were resistant (up to 50%) to proteolytic degradation by preparations of rat liver lysosomal enzymes. The parent antibody targeted efficiently to human colorectal carcinoma (LS174T) xenografts in nude mice (up to 25%/g); the conjugates, however, showed no tumour-targeting, probably due to masking by polymer chains (which are attached by non-specific aminolysis). Conjugates designed to maintain immunoreactivity following linkage through oxidised carbohydrates are currently being synthesised. Nevertheless, the conjugates display increased rates of extravasation, compared with proteins of the same hydrodynamic size, and the decreased charge is anticipated to accelerate diffusion through tumour interstitium.
Assuntos
Acrilamidas/farmacocinética , Imunotoxinas/administração & dosagem , Acrilamidas/farmacologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/metabolismo , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Portadores de Fármacos , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Fragmentos de Imunoglobulinas/administração & dosagem , Fragmentos de Imunoglobulinas/metabolismo , Imunoglobulina G/análise , Imunoglobulina M/análise , Imunotoxinas/metabolismo , Radioisótopos do Iodo , Cinética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Microssomos Hepáticos/enzimologia , Transplante de Neoplasias , Peptídeo Hidrolases/metabolismo , Distribuição Tecidual , Transplante HeterólogoRESUMO
Rabbit antisera were prepared against the idiotypic determinants of the mouse IgA myeloma protein-315, its purified heavy and light chains, and the Fv fragment comprising the variable region of both heavy and light chains. Agar diffusion demonstrated a line of identity between protein-315 and its Fv fragment against either homologous antiserum. Protein-315 and Fv fragment were labeled with (125)I and reacted with their anti-idiotypic antisera. Inhibition studies confirmed that the Fv fragment contained all the idiotypic specificities present in the intact protein. Fv was as effective as protein-315 on a weight basis in inhibiting the reaction between anti-idiotypic antiserum and protein-315. Protein-315 has high affinity for the 2,4-dinitrophenyl group, and such ligands can inhibit the reaction between protein-315 and its anti-idiotypic antibodies. Hapten inhibition was also obtained with the Fv fragment and its anti-idiotypic antiserum, implying that Fv contains an intact combining site. In this system with protein-315 the antigenic determinants expressed as idiotypic specificities are entirely within the variable region and are not influenced in their expression by the constant region. We therefore suggest that, in general, idiotypic determinants are antigenic determinants of the Fv protions of immunoglobulins.