RESUMEN
BACKGROUND: Alemtuzumab efficacy and safety was demonstrated in CARE-MS I and extension studies (CAMMS03409; TOPAZ). OBJECTIVE: Evaluate serum neurofilament light chain (sNfL) in CARE-MS I patients and highly active disease (HAD) subgroup, over 7 and 2 years for alemtuzumab and subcutaneous interferon beta-1a (SC IFNB-1a), respectively. METHODS: Patients received SC IFNB-1a 44 µg 3×/week or alemtuzumab 12 mg/day at baseline and month 12, with further as-needed 3-day courses. sNfL was measured using single-molecule array (Simoa™). HAD definition was ⩾2 relapses in year before randomization and ⩾1 baseline gadolinium-enhancing lesion. RESULTS: Baseline median sNfL levels were similar in alemtuzumab (n = 354) and SC IFNB-1a-treated (n = 159) patients (31.7 vs 31.4 pg/mL), but decreased with alemtuzumab versus SC IFNB-1a until year 2 (Y2; 13.2 vs 18.7 pg/mL; p < 0.0001); 12.7 pg/mL for alemtuzumab at Y7. Alemtuzumab-treated patients had sNfL at/below healthy control median at Y2 (72% vs 47%; p < 0.0001); 73% for alemtuzumab at Y7. HAD patients (n = 102) had higher baseline sNfL (49.4 pg/mL) versus overall population; alemtuzumab HAD patients attained similar levels (Y2, 12.8 pg/mL; Y7, 12.7 pg/mL; 75% were at/below control median at Y7). CONCLUSION: Alemtuzumab was superior to SC IFNB-1a in reducing sNfL, with levels in alemtuzumab patients remaining stable through Y7. CLINICALTRIALS.GOV IDENTIFIER: NCT00530348, NCT00930553, NCT02255656.
Asunto(s)
Filamentos Intermedios , Esclerosis Múltiple Recurrente-Remitente , Alemtuzumab/efectos adversos , Humanos , Interferón beta-1a/uso terapéutico , Esclerosis Múltiple Recurrente-Remitente/tratamiento farmacológico , Proteínas de NeurofilamentosRESUMEN
Alemtuzumab is a humanized monoclonal antibody specific for the CD52 protein present at high levels on the surface of B and T lymphocytes. In clinical trials, alemtuzumab has shown a clinical benefit superior to that of interferon-ß in relapsing-remitting multiple sclerosis patients. Treatment with alemtuzumab leads to the depletion of circulating lymphocytes followed by a repopulation process characterized by alterations in the number, proportions and properties of lymphocyte subsets. Of particular interest, an increase in the percentage of T cells with a regulatory phenotype (Treg cells) has been observed in multiple sclerosis patients after alemtuzumab. Since Treg cells play an important role in the control of autoimmune responses, the effect of alemtuzumab on Treg cells was further studied in vitro. Alemtuzumab effectively mediated complement-dependent cytolysis of human T lymphocytes and the remaining population was enriched in T cells with a regulatory phenotype. The alemtuzumab-exposed T cells displayed functional regulatory characteristics including anergy to stimulation with allogeneic dendritic cells and ability to suppress the allogeneic response of autologous T cells. Consistent with the observed increase in Treg cell frequency, the CD25(hi) T-cell population was necessary for the suppressive activity of alemtuzumab-exposed T cells. The mechanism of this suppression was found to be dependent on both cell-cell contact and interleukin-2 consumption. These findings suggest that an alemtuzumab-mediated increase in the proportion of Treg cells may play a role in promoting the long-term efficacy of alemtuzumab in patients with multiple sclerosis.
Asunto(s)
Anticuerpos Monoclonales Humanizados/farmacología , Antineoplásicos/farmacología , Esclerosis Múltiple/inmunología , Linfocitos T Reguladores/inmunología , Alemtuzumab , Antígenos CD/inmunología , Antígenos de Neoplasias/inmunología , Antígeno CD52 , Comunicación Celular/efectos de los fármacos , Comunicación Celular/inmunología , Supervivencia Celular , Células Dendríticas/inmunología , Células Dendríticas/patología , Femenino , Glicoproteínas/inmunología , Humanos , Interleucina-2/inmunología , Subunidad alfa del Receptor de Interleucina-2/inmunología , Masculino , Esclerosis Múltiple/tratamiento farmacológico , Esclerosis Múltiple/patología , Linfocitos T Reguladores/patologíaRESUMEN
Alemtuzumab, a monoclonal antibody directed against human CD52, is used in the treatment of MS. To characterize the impact of anti-CD52 administration, a monoclonal antibody to mouse CD52 (anti-muCD52) was generated and evaluated in EAE mouse models of MS. A single course of anti-muCD52 provided a therapeutic benefit accompanied by a reduction in the frequency of autoreactive T lymphocytes and production of pro-inflammatory cytokines. Examination of the CNS revealed a decrease in infiltrating lymphocytes, demyelination and axonal loss. Electrophysiological assessment showed preservation of axonal conductance in the spinal cord. These findings suggest that anti-CD52 therapy may help preserve CNS integrity.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Antígenos CD/inmunología , Antígenos de Neoplasias/inmunología , Enfermedades Desmielinizantes/tratamiento farmacológico , Enfermedades Desmielinizantes/inmunología , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Encefalomielitis Autoinmune Experimental/inmunología , Glicoproteínas/inmunología , Secuencia de Aminoácidos , Animales , Anticuerpos Monoclonales/genética , Anticuerpos Monoclonales/farmacología , Axones/efectos de los fármacos , Axones/inmunología , Axones/patología , Antígeno CD52 , Enfermedades Desmielinizantes/patología , Encefalomielitis Autoinmune Experimental/patología , Glicoproteínas/antagonistas & inhibidores , Humanos , Inflamación/tratamiento farmacológico , Inflamación/inmunología , Inflamación/patología , Mediadores de Inflamación/antagonistas & inhibidores , Mediadores de Inflamación/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Datos de Secuencia MolecularRESUMEN
Alemtuzumab is a monoclonal antibody against the CD52 antigen present at high levels on the surface of lymphocytes. While treatment of multiple sclerosis patients with alemtuzumab results in marked depletion of lymphocytes from the circulation, it has not been associated with a high incidence of serious infections. In a human CD52 transgenic mouse, alemtuzumab treatment showed minimal impact on the number and function of innate immune cells. A transient decrease in primary adaptive immune responses was observed post-alemtuzumab but there was little effect on memory responses. These results potentially help explain the level of immunocompetence observed in alemtuzumab-treated MS patients.
Asunto(s)
Inmunidad Adaptativa/genética , Anticuerpos Monoclonales Humanizados/fisiología , Antígenos CD/inmunología , Antígenos de Neoplasias/inmunología , Glicoproteínas/inmunología , Alemtuzumab , Animales , Antígenos CD/genética , Antígenos de Neoplasias/genética , Linfocitos B/inmunología , Antígeno CD52 , Células Cultivadas , Glicoproteínas/genética , Humanos , Inmunidad Innata/genética , Ratones , Ratones Transgénicos , Linfocitos T/inmunología , Resultado del TratamientoRESUMEN
The molecular changes induced by alemtuzumab following binding of CD52 on B tumor cells were investigated. Alemtuzumab alone had no detectable impact on cell signaling but cross-linking of alemtuzumab on the surface of B tumor lines with anti-human Fc antibodies induced a transient Ca(2+) flux followed by phosphorylation of several kinases involved in stress and survival pathways, and expression of associated proteins including TNF-α. Cross-linking of alemtuzumab also induced capping and caspase-dependent apoptosis of the tumor lines. When using primary cells from B-CLL patients, alemtuzumab alone was capable of inducing protein phosphorylation and apoptosis through the cross-linking of alemtuzumab by FcγRIIb receptors on B-CLL cells. Apoptosis was prevented by blocking of FcγRIIb receptors with anti-CD32 antibody. Overall, our results indicate that cross-linking of alemtuzumab on B tumor cells can occur naturally through Fc receptor interaction and leads to the activation of specific cellular pathways and induction of apoptosis.
Asunto(s)
Anticuerpos Monoclonales Humanizados/farmacología , Apoptosis/efectos de los fármacos , Linfocitos B/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Alemtuzumab , Anticuerpos/inmunología , Anticuerpos/farmacología , Antineoplásicos/farmacología , Linfocitos B/metabolismo , Linfocitos B/patología , Calcio/metabolismo , Línea Celular Tumoral , Perfilación de la Expresión Génica , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Humanos , Immunoblotting , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/metabolismo , Leucemia Linfocítica Crónica de Células B/patología , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptores de IgG/inmunología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales Cultivadas , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Overexpression of TMPRSS4, a cell surface-associated transmembrane serine protease, has been reported in pancreatic, colorectal and thyroid cancers, and has been implicated in tumor cell migration and metastasis. Few reports have investigated both TMPRSS4 gene expression levels and the protein products. In this study, quantitative RT-PCR and protein staining were used to assess TMPRSS4 expression in primary non-small cell lung carcinoma (NSCLC) tissues and in lung tumor cell lines. At the transcriptional level, TMPRSS4 message was significantly elevated in the majority of human squamous cell and adenocarcinomas compared with normal lung tissues. Staining of over 100 NSCLC primary tumor and normal specimens with rabbit polyclonal anti-TMPRSS4 antibodies confirmed expression at the protein level in both squamous cell and adenocarcinomas with little or no staining in normal lung tissues. Human lung tumor cell lines expressed varying levels of TMPRSS4 mRNA in vitro. Interestingly, tumor cell lines with high levels of TMPRSS4 mRNA failed to show detectable TMPRSS4 protein by either immunoblotting or flow cytometry. However, protein levels were increased under hypoxic culture conditions suggesting that hypoxia within the tumor microenvironment may upregulate TMPRSS4 protein expression in vivo. This was supported by the observation of TMPRSS4 protein in xenograft tumors derived from the cell lines. In addition, staining of human squamous cell carcinoma samples for carbonic anhydrase IX (CAIX), a hypoxia marker, showed TMPRSS4 positive cells adjacent to CAIX positive cells. Overall, these results indicate that the cancer-associated TMPRSS4 protein is overexpressed in NSCLC and may represent a potential therapeutic target.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Hipoxia de la Célula , Neoplasias Pulmonares/metabolismo , Proteínas de la Membrana/biosíntesis , Serina Endopeptidasas/biosíntesis , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Animales , Antígenos de Neoplasias/análisis , Biomarcadores de Tumor/biosíntesis , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Anhidrasa Carbónica IX , Anhidrasas Carbónicas/análisis , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Línea Celular Tumoral , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Pulmón/metabolismo , Neoplasias Pulmonares/genética , Proteínas de la Membrana/genética , Ratones , Ratones Desnudos , ARN Mensajero/biosíntesis , Serina Endopeptidasas/genética , Transcripción Genética , Microambiente Tumoral , Regulación hacia ArribaRESUMEN
Autoimmunity has long been linked to myocarditis and its sequela, dilated cardiomyopathy, the leading causes of heart failure in young patients. However, the underlying mechanisms are poorly defined, with most clinical investigations focused on humoral autoimmunity as the target for intervention. Here, we show that the α-isoform of myosin heavy chain (α-MyHC, which is encoded by the gene Myh6) is the pathogenic autoantigen for CD4+ T cells in a spontaneous mouse model of myocarditis. Further, we found that Myh6 transcripts were absent in mouse medullary thymic epithelial cells (mTECs) and peripheral lymphoid stromal cells, which have been implicated in mediating central and peripheral T cell tolerance, respectively. Transgenic expression of α-MyHC in thymic epithelium conferred tolerance to cardiac myosin and prevented myocarditis, demonstrating that α-MyHC is a primary autoantigen in this disease process. Remarkably, we found that humans also lacked α-MyHC in mTECs and had high frequencies of α-MyHC-specific T cells in peripheral blood, with markedly augmented T cell responses to α-MyHC in patients with myocarditis. Since α-MyHC constitutes a small fraction of MyHC in human heart, these findings challenge the longstanding notion that autoimmune targeting of MyHC is due to its cardiac abundance and instead suggest that it is targeted as a result of impaired T cell tolerance mechanisms. These results thus support a role for T cell-specific therapies for myocarditis.
Asunto(s)
Autoinmunidad , Linfocitos T CD4-Positivos/inmunología , Miocarditis/inmunología , Miocardio/inmunología , Miosinas Ventriculares/inmunología , Animales , Autoantígenos/genética , Secuencia de Bases , Miosinas Cardíacas/genética , Miosinas Cardíacas/inmunología , Cartilla de ADN/genética , Modelos Animales de Enfermedad , Femenino , Antígenos HLA-DQ/genética , Humanos , Tolerancia Inmunológica , Lactante , Recién Nacido , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones Noqueados , Ratones Transgénicos , Miocarditis/etiología , Miocarditis/prevención & control , Cadenas Pesadas de Miosina/genética , Cadenas Pesadas de Miosina/inmunología , Timo/citología , Timo/inmunología , Miosinas Ventriculares/genéticaRESUMEN
Genome-wide analyses have shown that the MHC class II region is the principal locus that confers susceptibility to a number of human autoimmune diseases. Due to the high degree of linkage disequilibrium across the MHC, it has been difficult to dissect the contribution of individual genes to disease susceptibility. As a result, intensive efforts have been made to generate mice transgenic for human class II molecules as models of autoimmune disease. However, in every case, additional manipulations-such as immunization with Ag in adjuvant, expression of immunostimulants on target tissues, or coexpression of TCR transgenes-have been required to induce disease. In this study, we show that expression of the human HLA-DQ8 (DQA1*0301/DQB1*0302) molecule alone in three lines of transgenic nonobese diabetic murine class II-deficient (mII(-/-)) mice results in the spontaneous development of autoimmune myocarditis. The disease shares key features of human myocarditis and was characterized by lymphocytic infiltrates in the myocardium and cardiac myocyte destruction, circulating IgG autoantibodies against cardiac myosin heavy chain, and premature death due to heart failure. We demonstrate that myocarditis could be transferred into healthy HLA-DQ8(+)RAG-1(-/-)mII(-/-) nonobese diabetic recipients with lymphocytes, but not sera. It has been widely thought that autoimmune myocarditis is of infectious etiology, with the immune responses arising secondary to cardiac damage from pathogens. These studies provide direct experimental evidence that spontaneous autoimmune myocarditis can occur in the absence of infection and that expression of HLA-DQ8 confers susceptibility to this organ-specific autoimmune disease.
Asunto(s)
Enfermedades Autoinmunes/inmunología , Modelos Animales de Enfermedad , Antígenos HLA-DQ/fisiología , Miocarditis/inmunología , Traslado Adoptivo , Animales , Autoanticuerpos/biosíntesis , Autoanticuerpos/sangre , Enfermedades Autoinmunes/genética , Enfermedades Autoinmunes/patología , Linfocitos T CD4-Positivos/inmunología , Cruzamientos Genéticos , Predisposición Genética a la Enfermedad , Antígenos HLA-DQ/biosíntesis , Antígenos HLA-DQ/genética , Humanos , Inmunoglobulina G/biosíntesis , Inmunoglobulina G/sangre , Transfusión de Linfocitos , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones Noqueados , Ratones Transgénicos , Miocarditis/genética , Miocarditis/patología , Bazo/citología , Bazo/trasplanteRESUMEN
Although HLA-DQ8 has been implicated as a key determinant of genetic susceptibility to human type 1 diabetes, spontaneous diabetes has been observed in HLA-DQ8 transgenic mice that lack expression of murine MHC class II molecules (mII(-/-)) only when the potent costimulatory molecule, B7.1, is transgenically expressed on pancreatic beta cells. To study the contribution of HLA-DQ8 to the development of diabetes in this model, we crossed RIP-B7.1mII(-/-) mice with a set of transgenic mouse lines that differed in their HLA-DQ8 expression patterns on APC subpopulations, in particular dendritic cells and cortical thymic epithelial cells. Surprisingly, we found that even in the absence of HLA-DQ8 and CD4 T cells, a substantial fraction of the RIP-B7.1mII(-/-) mice developed diabetes. This disease process was remarkable for not only showing insulitis, but also inflammatory destruction of the exocrine pancreas with diffusely up-regulated expression of MHC class I and ICAM-1 molecules. Expression of HLA-DQ8 markedly increased the kinetics and frequency of diabetes, with the most severe disease in the lines with the highest levels of HLA-DQ8 on cortical thymic epithelial cells and the largest numbers of CD4 T cells. However, the adoptive transfer of diabetes was not HLA-DQ8-dependent and disease could be rapidly induced with purified CD8 T cells alone. Expression of B7.1 in the target tissue can thus dramatically alter the cellular and molecular requirements for the development of autoimmunity.