RESUMO
Graves' disease (GD), an autoimmune thyroid disease, is one of the main autoimmune diseases in the general population. It is known that the pathophysiology of this disease may be related to immunological mechanisms dysregulation. These mechanisms can be influenced by GD therapies, such as iodide or antithyroid drugs (ATD). OBJECTIVE: Verify relation between clinical, biochemical and treatment modalities used prior to surgery and histopathological characteristics observed in total thyroidectomy products from patients previously diagnosed with Graves' disease. Furthermore, these data were related to composition of lymphocytic infiltrate in terms of proportions of lymphocytes CD4+, CD8+, CD25+ and CD20+. We aim to contribute to the understanding of the evolution pattern of GD, whose pathophysiology is not yet completely understood. METHODS: Cross-sectional study assessing thyroidectomy products for the presence of lymphocytic infiltrate, as well as the proportion and intensity of CD4+, CD8+, CD25+ and CD20+ markers. We selected 50 patients who underwent total or partial thyroidectomy in a tertiary service between 1996 and 2013 due to GD with histopathological confirmation. The control group (non-autoimmune disease group) consisted of 12 patients with histopathological data compatible with normal perilesional thyroid parenchyma. The intensity of lymphocytic infiltrate and immunohistochemical expression of the markers CD4+ (helper T lymphocytes), CD8+ (cytotoxic T lymphocytes), CD25+ (regulatory T lymphocytes) and CD20+ (B lymphocytes) were retrospectively evaluated and relationship with ultrasound, laboratory and clinical data was assessed. RESULTS: No differences were found in intensity, presence of lymphoid follicles, and expression of CD4+/CD8+/CD25+ in patients with GD who did or did not use ATD or iodide. In the group that did not use ATD, a higher proportion of CD20+ expression was found. The GD group was associated with hyperplastic epithelium and the control group was associated with simple epithelium. There was no difference in ultrasound thyroid volume between the groups. In GD patients with mild lymphocytic infiltrate, higher free thyroxin (FT4) levels were observed than those in patients with no infiltrate or moderate infiltrate. CONCLUSION: We found a lower proportion of intrathyroidal CD20+ B lymphocytes in patients under use of methimazole. However, no difference was observed in intrathyroidal lymphocyte subpopulations related to the short-term use of iodide. The understanding of thyroid autoimmunity, as well as identifying points of pharmacological modulation, are very important for advancement and improvement in treatments for these diseases.
Assuntos
Antígenos CD20 , Doença de Graves , Metimazol , Glândula Tireoide , Humanos , Doença de Graves/tratamento farmacológico , Doença de Graves/patologia , Doença de Graves/imunologia , Metimazol/uso terapêutico , Metimazol/farmacologia , Feminino , Masculino , Glândula Tireoide/patologia , Glândula Tireoide/metabolismo , Glândula Tireoide/efeitos dos fármacos , Adulto , Pessoa de Meia-Idade , Antígenos CD20/metabolismo , Iodetos/metabolismo , Estudos Transversais , Linfócitos B/imunologia , Linfócitos B/metabolismo , Linfócitos B/efeitos dos fármacos , Antitireóideos/farmacologia , Antitireóideos/uso terapêutico , Subpopulações de Linfócitos/efeitos dos fármacos , Subpopulações de Linfócitos/metabolismo , Subpopulações de Linfócitos/imunologia , Tireoidectomia , IdosoAssuntos
Antígeno B7-H1/metabolismo , Melanoma/metabolismo , Neoplasias Cutâneas/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos CD20/metabolismo , Biomarcadores Tumorais/metabolismo , Estudos de Coortes , Feminino , Humanos , Imuno-Histoquímica , Linfonodos/metabolismo , Masculino , Melanoma/patologia , Pessoa de Meia-Idade , Metástase Neoplásica , Estudos Retrospectivos , Neoplasias Cutâneas/patologiaRESUMO
BACKGROUND: RTXM83 is a rituximab biosimilar with proven clinical safety and efficacy. It is the first rituximab biosimilar developed and approved in South America and is currently marketed in several Latin American, Middle Eastern and African countries. OBJECTIVE: The aim of this study was to present the physicochemical and biological characterization studies utilized to demonstrate the similarity between RTXM83 and its reference product. METHODS: Primary and higher order protein structures were analysed using peptide mapping with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS), fluorescence spectroscopy and circular dichroism, and micro-differential scanning calorimetry, among other techniques. Charge variants were determined by cation-exchange chromatography (CEX) and capillary isoelectric focusing (cIEF). Glycosylation and glycoforms distribution were analysed using MS, normal phase high-performance liquid chromatography (NP-HPLC) and high-performance anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD). Size variants were evaluated by size-exclusion chromatography (SEC), sedimentation velocity analytical ultracentrifugation (SV-AUC), dynamic light scattering (DLS), and capillary electrophoresis-sodium dodecyl sulfate (CE-SDS). Biological characterization included binding assays for complement C1q, CD20, and several Fc receptors (FcRs), as well as potency determination for in vitro apoptosis induction, complement-dependent cytotoxicity (CDC), and antibody-dependent cell-mediated cytotoxicity (ADCC). RESULTS: RTXM83 and the reference product showed identical primary sequences and disulfide bridge patterns, and similarity at higher order protein structures, post-translational modification profiles (amino acid modifications, charge variants, and glycosylation) and levels of purity and process-related impurities. Functional studies demonstrated that RTXM83 is similar to the reference product regarding the three known mechanisms of action of rituximab: CDC, ADCC, and apoptosis induction. Binding affinities to CD20, complement component C1q, and different FcRs were also equivalent. CONCLUSION: RTXM83 is similar to its reference product in all critical quality attributes.
Assuntos
Medicamentos Biossimilares/química , Medicamentos Biossimilares/uso terapêutico , Rituximab/química , Rituximab/uso terapêutico , Citotoxicidade Celular Dependente de Anticorpos/fisiologia , Antígenos CD20/metabolismo , Cromatografia Líquida de Alta Pressão/métodos , Dicroísmo Circular/métodos , Complemento C1q/metabolismo , Difusão Dinâmica da Luz/métodos , Eletroforese Capilar/métodos , Glicosilação , Humanos , Mapeamento de Peptídeos/métodos , Receptores Fc/metabolismo , Espectrometria de Massas em Tandem/métodos , Ultracentrifugação/métodosRESUMO
BACKGROUND: Among the complex of HTLV-associated diseases, Sjögren's syndrome (SS) is one of the most controversial. This work aims to detect morphological and inflammatory alterations, including clues of the presence of HTLV-1, in minor salivary glands of patients with dryness symptoms. METHODS: We have assessed HTLV-1-seropositive patients (HTLV-1 group) and patients with SS (SS group). We used formalin-fixed, paraffin-embedded minor salivary gland tissue to evaluate the morphological aspects and, by means of immunohistochemistry, the presence of Tax protein, CD4, CD8 and CD20 cells. Additionally, viral particles and proviral load were analysed by PCR. RESULTS: The HTLV-1 group had the highest prevalence of non-specific chronic sialadenitis (85.71%; P = 0.017) and greater amount of T CD8+ cells. In the SS group, focal lymphocytic sialadenitis (80%; P = 0.017) prevailed, with a greater amount of B CD20+ . Both immunohistochemistry and PCR identified the Tax protein and its gene in the salivary glands of both groups and in similar proportions. CONCLUSION: The results indicate that HTLV-1-seropositive patients have different patterns of morphological/inflammatory alterations, suggesting a likely difference in the process of immune activation.
Assuntos
Infecções por HTLV-I/patologia , Glândulas Salivares Menores/patologia , Síndrome de Sjogren/patologia , Antígenos CD20/metabolismo , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD8-Positivos/patologia , Feminino , Produtos do Gene tax/metabolismo , Infecções por HTLV-I/imunologia , Infecções por HTLV-I/metabolismo , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Reação em Cadeia da Polimerase , Glândulas Salivares Menores/imunologia , Glândulas Salivares Menores/metabolismo , Sialadenite , Síndrome de Sjogren/imunologiaRESUMO
BACKGROUND: Survival of adults with B-Acute Lymphoblastic Leukemia requires accurate risk stratification of patients in order to provide the appropriate therapy. Contemporary techniques, using clinical and cytogenetic variables are incomplete for prognosis prediction. METHODS: To improve the classification of adult patients diagnosed with B-ALL into prognosis groups, two strategies were examined and combined: the expression of the ID1/ID3/IGJ gene signature by RT-PCR and the immunophenotypic profile of 19 markers proposed in the EuroFlow protocol by Flow Cytometry in bone marrow samples. RESULTS: Both techniques were correlated to stratify patients into prognostic groups. An inverse relationship between survival and expression of the three-genes signature was observed and an immunophenotypic profile associated with clinical outcome was identified. Markers CD10 and CD20 were correlated with simultaneous overexpression of ID1, ID3 and IGJ. Patients with simultaneous expression of the poor prognosis gene signature and overexpression of CD10 or CD20, had worse Event Free Survival and Overall Survival than patients who had either the poor prognosis gene expression signature or only CD20 or CD10 overexpressed. CONCLUSION: By utilizing the combined evaluation of these two immunophenotypic markers along with the poor prognosis gene expression signature, the risk stratification can be significantly strengthened. Further studies including a large number of patients are needed to confirm these findings.
Assuntos
Antígenos CD20/metabolismo , Cadeias J de Imunoglobulina/genética , Proteína 1 Inibidora de Diferenciação/genética , Proteínas Inibidoras de Diferenciação/genética , Proteínas de Neoplasias/genética , Neprilisina/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras B/classificação , Adolescente , Adulto , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/imunologia , Prognóstico , Análise de Sobrevida , Adulto JovemRESUMO
INTRODUCTION: Rituximab was the first monoclonal antibody approved for the treatment of B-cell non-Hodgkin lymphoma (NHL) expressing CD20 antigen. This antibody has also the potential to be used as a specific fluorescent and radiolabel agent for targeting NHL. OBJECTIVE: To radiolabel rituximab with technetium-99m (99mTc) or Cy7 and evaluate both probes as potential imaging agents for NHL. METHODS: Rituximab was derivatized with the trifluoroacetyl hydrazino protected form of succinimidyl ester of HYNIC and radiolabeled with 99mTc. Radiochemical stability and in vitro cell assays were evaluated. Biodistribution and single-photon emission computed tomography/computed tomography (SPECT/CT) were performed. Raji cells were transfected with luciferase for bioluminescent NHL imaging up to 21 days. Rituximab was labeled with Cy7 for in vivo noninvasive fluorescence imaging up to 96 h. RESULTS: Radiolabeling was carried out in a fast, reproducible, easy, and stable way with high radiochemical purity and did not interfere with epitope recognition. Biodistribution and SPECT/CT studies showed high liver and discrete tumor uptake. Bioluminescence and fluorescence studies helped us evaluate rituximab-Cy7 in Raji subcutaneous engraftment in BALB/c nude mice. CONCLUSIONS: Our results support the potential use of rituximab labeled either with 99mTc or Cy7 as a molecular imaging tool for staging, restaging, and guiding surgical excision of tumors, which merits further evaluation.
Assuntos
Carbocianinas , Linfoma não Hodgkin/diagnóstico por imagem , Imagem Molecular/métodos , Rituximab , Tecnécio , Animais , Antígenos CD20/metabolismo , Carbocianinas/farmacocinética , Linhagem Celular Tumoral , Usos Diagnósticos de Compostos Químicos , Feminino , Humanos , Camundongos Endogâmicos BALB C , Compostos Radiofarmacêuticos/metabolismo , Compostos Radiofarmacêuticos/farmacocinética , Rituximab/química , Rituximab/metabolismo , Rituximab/farmacocinética , Tomografia Computadorizada com Tomografia Computadorizada de Emissão de Fóton Único/métodos , Tecnécio/farmacocinética , Distribuição TecidualRESUMO
BACKGROUND: Biosimilars are described as biological products that resemble the structure of original biologic therapeutic products, with no clinically meaningful differences in terms of safety and effectiveness from the original. A wide range of biosimilars are under development or are already licensed in many countries. Biosimilars are earning acceptance and becoming a reality for immunotherapy treatments mainly based on the alternatives for the commercial anti-CD20 monoclonal antibody rituximab. The most important mechanism of action reported for this antibody is the induction of antibody-dependent cell cytotoxicity (ADCC), which is associated with the polymorphisms present at the 158 position in the IgG receptor FcγRIIIa. OBJECTIVE: The aim of the study was to validate the functional comparability between the proposed rituximab biosimilar RTXM83 and the original product. To achieve this we assessed the binding capacity and ADCC induction of this biosimilar, taking into account the different FcγRIIIa-158 polymorphisms. METHODS: Binding capacity was evaluated by flow cytometry using CD20 positive cells and a wide range of antibody concentrations. The FcγRIIIa-158 polymorphisms were analyzed by polymerase chain reaction (PCR) followed by allele-specific restriction enzyme digestion. ADCC was measured by a colorimetric lactate dehydrogenase-release assay, using effector cells from donors with different FcγRIIIa-158 polymorphisms. RESULTS: Binding capacity assay showed no differences between both products. Regarding ADCC, a similar relative potency was obtained between both antibodies, showing a higher response for the FcγRIIIa-158 valine/valine (V/V) polymorphism compared to the phenylalanine/phenylalanine (F/F), for both rituximab and RTXM83. CONCLUSION: Our data strongly suggest the biocomparability between the proposed biosimilar and the originator rituximab, in antibody recognition and ADCC activity. Additionally, our results suggest that donors with the FcγRIIIa-158V/V polymorphism induce a higher ADCC response, as has been reported.
Assuntos
Medicamentos Biossimilares/farmacologia , Rituximab/farmacologia , Citotoxicidade Celular Dependente de Anticorpos , Antígenos CD20/metabolismo , Medicamentos Biossimilares/metabolismo , Linhagem Celular , Relação Dose-Resposta a Droga , Humanos , L-Lactato Desidrogenase/metabolismo , Polimorfismo Genético , Receptores de IgG/genética , Receptores de IgG/metabolismo , Rituximab/metabolismoRESUMO
BACKGROUND: The Caribbean island of Tobago, which is 97% African ancestry, has one of the highest rates of prostate cancer in the world. We have previously reported that human herpesvirus 8 (HHV-8) infection is significantly associated with prostate cancer in Tobago. In this study, we extend those results testing the hypothesis that HHV-8 seropositive Tobagonian men have a chronic HHV-8 infection in their prostates that is associated with increased inflammation. METHODS: Prostate sections were screened by immunohistochemistry for the expression of HHV-8 proteins K8.1 and LANA-1 and for presence of B cells (CD20) and macrophages (CD68). RESULTS: HHV-8 antigen expression representing lytic and latent infections was seen in 73.9% of prostates from HHV-8 seropositive subjects. Latent infections were seen predominantly in glandular epithelia whereas lytic gene expression was seen mainly in macrophages in prostate stroma. Macrophage infiltrates were significantly increased in sections expressing HHV-8 proteins. CONCLUSION: HHV-8 establishes a chronic latent infection in the prostate, which is associated with an increased macrophage infiltrate.
Assuntos
Infecções por Herpesviridae/patologia , Macrófagos/patologia , Próstata/patologia , Neoplasias da Próstata/virologia , Antígenos CD/metabolismo , Antígenos CD20/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Antígenos Virais/metabolismo , Linfócitos B/metabolismo , Linfócitos B/patologia , Biomarcadores/metabolismo , Glicoproteínas/metabolismo , Infecções por Herpesviridae/virologia , Herpesvirus Humano 8 , Humanos , Macrófagos/metabolismo , Masculino , Proteínas Nucleares/metabolismo , Próstata/metabolismo , Próstata/virologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Trinidad e Tobago , Proteínas Virais/metabolismoRESUMO
We described a 21-year-old woman with a diagnosis of Sjögren syndrome that came for consultation with a localized mass over her left arm of fast growth. Lab results were normal; a Doppler ultrasound showed the presence of partial thrombosis in the left axillary vein; a magnetic resonance imaging showed edema on the biceps muscle and the biopsy of the mass disclosed the presence of severe lymphocyte infiltrate within the connective tissue and scarce muscle fibers. Immunostaining showed positive results for antígeno comun leucocitario in spanish (leukocyte common antigen) (ACL) and CD3. Those results are consistent with the diagnosis of focal myositis. The patient was treated with low doses of prednisone and methotrexate, with good response.
Assuntos
Miosite/diagnóstico , Trombose/diagnóstico , Antígenos CD20/metabolismo , Braço/patologia , Veia Axilar/patologia , Biópsia , Complexo CD3/metabolismo , Antígenos CD8/metabolismo , Diagnóstico Diferencial , Edema/diagnóstico , Feminino , Humanos , Linfócitos/citologia , Imageamento por Ressonância Magnética , Metotrexato/uso terapêutico , Músculo Esquelético/patologia , Miosite/diagnóstico por imagem , Prednisona/uso terapêutico , Trombose/diagnóstico por imagem , Ultrassonografia Doppler , Adulto JovemRESUMO
Cervical intraepithelial neoplasias (CIN) are closely associated with oncogenic subtypes of the human papillomavirus (HPV). In the presence of this virus, it is known that the activation or suppression of immune system is the key to the development, progression and/or regression of cervical lesions. Therefore, the objective of this study is to compare the local immune response among HIV-seropositive and seronegative patients with cervical intraepithelial neoplasia regarding the expression of T lymphocytes (CD3+, CD4+ and CD8+), B lymphocytes (CD20+) and natural killers cells (CD56+) in the cervical stroma. A cross-sectional study of paraffin blocks containing cervical tissue after conization by the Loop Electrosurgical Excision Procedure (LEEP) from 47 HIV-seropositive and 38 seronegative patients with CIN. Cervical stroma immunohistochemistry was performed in the CIN area. The Fisher's exact test was used for the statistical analysis. When HIV-seropositive and seronegative women were compared, the seropositive women had a higher count of CD8+ T lymphocytes (52.1% versus 28.9%, P<0.04). Considering CIN degree (CIN 1 and CIN 2/3), the HIV-seronegative patients with CIN 1 had a low count of CD20+B-lymphocytes (7.1%) in comparison with CIN 1 HIV seropositive and with CIN 2/3 HIV-seronegative patients, respectively 50% (P<0.018) and 54.5% (P<0.0048). The HIV infection and degree of CIN influenced the cytotoxic lymphocytes inducing an increase in the number of cells high count of CD20+ lymphocytes with CIN 1.