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1.
Cell ; 183(7): 1867-1883.e26, 2020 12 23.
Artículo en Inglés | MEDLINE | ID: mdl-33248023

RESUMEN

Biliary atresia (BA) is a severe cholangiopathy that leads to liver failure in infants, but its pathogenesis remains to be fully characterized. By single-cell RNA profiling, we observed macrophage hypo-inflammation, Kupffer cell scavenger function defects, cytotoxic T cell expansion, and deficiency of CX3CR1+effector T and natural killer (NK) cells in infants with BA. More importantly, we discovered that hepatic B cell lymphopoiesis did not cease after birth and that tolerance defects contributed to immunoglobulin G (IgG)-autoantibody accumulation in BA. In a rhesus-rotavirus induced BA model, depleting B cells or blocking antigen presentation ameliorated liver damage. In a pilot clinical study, we demonstrated that rituximab was effective in depleting hepatic B cells and restoring the functions of macrophages, Kupffer cells, and T cells to levels comparable to those of control subjects. In summary, our comprehensive immune profiling in infants with BA had educed that B-cell-modifying therapies may alleviate liver pathology.


Asunto(s)
Atresia Biliar/inmunología , Atresia Biliar/terapia , Hígado/inmunología , Animales , Antígenos CD20/metabolismo , Linfocitos B/inmunología , Atresia Biliar/sangre , Atresia Biliar/tratamiento farmacológico , Biopsia , Receptor 1 de Quimiocinas CX3C/metabolismo , Muerte Celular , Línea Celular , Proliferación Celular , Transdiferenciación Celular , Niño , Preescolar , Estudios de Cohortes , Citotoxicidad Inmunológica , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunoglobulina G/metabolismo , Lactante , Inflamación/patología , Células Asesinas Naturales/inmunología , Macrófagos del Hígado/patología , Hígado/patología , Cirrosis Hepática/sangre , Cirrosis Hepática/complicaciones , Cirrosis Hepática/inmunología , Cirrosis Hepática/patología , Depleción Linfocítica , Linfopoyesis , Masculino , Ratones Endogámicos BALB C , Fagocitosis , ARN/metabolismo , Rituximab/administración & dosificación , Rituximab/farmacología , Rituximab/uso terapéutico , Rotavirus/fisiología , Análisis de la Célula Individual , Células TH1/inmunología , Células Th17/inmunología
2.
J Immunol ; 212(4): 529-533, 2024 Feb 15.
Artículo en Inglés | MEDLINE | ID: mdl-38149924

RESUMEN

One reason for a lack of response to rituximab as well as infusion-related anaphylactic adverse events is the development of antidrug Abs to rituximab. Besides rituximab, a number of other therapeutic Abs targeting CD20 are nowadays available as alternatives. In this study, we investigated the potential cross-reactivity of (human) anti-rituximab Abs to three other anti-CD20 mAbs: ofatumumab, obinutuzumab, and ocrelizumab. In 25 cases of anti-rituximab Abs, cross-reactivity was examined using both direct binding assays and inhibition immunoassays. Although no cross-reactivity was observed to ofatumumab or obinutuzumab, 8 of 25 samples also showed reactivity toward ocrelizumab in at least one of the two assays. Furthermore, in three cases of anti-ocrelizumab Abs, cross-reactivity to rituximab was observed in an inhibition immunoassay, albeit not in a direct binding assay. Our results suggest that obinutuzumab or ofatumumab are safe anti-CD20 alternatives in case of the presence of anti-rituximab Abs. It is advisable to proceed cautiously if switching from rituximab to ocrelizumab (or vice versa) is considered in case these alternatives may not be available.


Asunto(s)
Anticuerpos Monoclonales , Antígenos CD20 , Humanos , Rituximab/uso terapéutico , Antígenos CD20/metabolismo
3.
Nature ; 577(7791): 561-565, 2020 01.
Artículo en Inglés | MEDLINE | ID: mdl-31942071

RESUMEN

Checkpoint blockade therapies that reactivate tumour-associated T cells can induce durable tumour control and result in the long-term survival of patients with advanced cancers1. Current predictive biomarkers for therapy response include high levels of intratumour immunological activity, a high tumour mutational burden and specific characteristics of the gut microbiota2,3. Although the role of T cells in antitumour responses has thoroughly been studied, other immune cells remain insufficiently explored. Here we use clinical samples of metastatic melanomas to investigate the role of B cells in antitumour responses, and find that the co-occurrence of tumour-associated CD8+ T cells and CD20+ B cells is associated with improved survival, independently of other clinical variables. Immunofluorescence staining of CXCR5 and CXCL13 in combination with CD20 reveals the formation of tertiary lymphoid structures in these CD8+CD20+ tumours. We derived a gene signature associated with tertiary lymphoid structures, which predicted clinical outcomes in cohorts of patients treated with immune checkpoint blockade. Furthermore, B-cell-rich tumours were accompanied by increased levels of TCF7+ naive and/or memory T cells. This was corroborated by digital spatial-profiling data, in which T cells in tumours without tertiary lymphoid structures had a dysfunctional molecular phenotype. Our results indicate that tertiary lymphoid structures have a key role in the immune microenvironment in melanoma, by conferring distinct T cell phenotypes. Therapeutic strategies to induce the formation of tertiary lymphoid structures should be explored to improve responses to cancer immunotherapy.


Asunto(s)
Melanoma/inmunología , Melanoma/terapia , Estructuras Linfoides Terciarias/inmunología , Antígenos CD20/metabolismo , Linfocitos B/inmunología , Linfocitos B/metabolismo , Antígeno B7-H1/antagonistas & inhibidores , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Quimiocina CXCL13/metabolismo , Humanos , Memoria Inmunológica/inmunología , Melanoma/genética , Melanoma/patología , Metástasis de la Neoplasia/genética , Metástasis de la Neoplasia/patología , Fenotipo , Pronóstico , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Proteómica , RNA-Seq , Receptores CXCR5/metabolismo , Análisis de la Célula Individual , Tasa de Supervivencia , Factor 1 de Transcripción de Linfocitos T/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Estructuras Linfoides Terciarias/genética , Resultado del Tratamiento , Microambiente Tumoral/inmunología
4.
Mol Ther ; 32(5): 1238-1251, 2024 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-38414244

RESUMEN

Chimeric antigen receptor (CAR) T cell therapies have demonstrated immense clinical success for B cell and plasma cell malignancies. We tested their impact on the viral reservoir in a macaque model of HIV persistence, comparing the functions of CD20 CAR T cells between animals infected with simian/human immunodeficiency virus (SHIV) and uninfected controls. We focused on the potential of this approach to disrupt B cell follicles (BCFs), exposing infected cells for immune clearance. In SHIV-infected animals, CAR T cells were highly functional, with rapid expansion and trafficking to tissue-associated viral sanctuaries, including BCFs and gut-associated lymphoid tissue (GALT). CD20 CAR T cells potently ablated BCFs and depleted lymph-node-associated follicular helper T (TFH) cells, with complete restoration of BCF architecture and TFH cells following CAR T cell contraction. BCF ablation decreased the splenic SHIV reservoir but was insufficient for effective reductions in systemic viral reservoirs. Although associated with moderate hematologic toxicity, CD20 CAR T cells were well tolerated in SHIV-infected and control animals, supporting the feasibility of this therapy in people living with HIV with underlying B cell malignancies. Our findings highlight the unique ability of CD20 CAR T cells to safely and reversibly unmask TFH cells within BCF sanctuaries, informing future combinatorial HIV cure strategies designed to augment antiviral efficacy.


Asunto(s)
Antígenos CD20 , Linfocitos B , Modelos Animales de Enfermedad , Infecciones por VIH , Inmunoterapia Adoptiva , Receptores Quiméricos de Antígenos , Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Animales , Antígenos CD20/metabolismo , Antígenos CD20/inmunología , Linfocitos B/inmunología , Linfocitos B/metabolismo , Virus de la Inmunodeficiencia de los Simios/inmunología , Inmunoterapia Adoptiva/métodos , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/terapia , Infecciones por VIH/terapia , Infecciones por VIH/inmunología , Receptores Quiméricos de Antígenos/inmunología , Receptores Quiméricos de Antígenos/metabolismo , Humanos , Linfocitos T/inmunología , Linfocitos T/metabolismo , VIH-1/inmunología , Carga Viral , Macaca mulatta
5.
Protein Expr Purif ; 215: 106392, 2024 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-37952787

RESUMEN

Cluster of differentiation 20 (CD20) is a nonglycosylated, multispanning transmembrane protein specifically integrated by B lymphocytes. Similar to CD20, another four-pass transmembrane protein, claudin 18.2, has attracted attention as an emerging therapeutic target for cancer. However, their poor solubility and toxic nature often hinder downstream applications, such as antibody drug development. Therefore, developing a cost-effective method for producing drug targets with multiple membrane-spanning domains is crucial. In this study, a high yield of recombinant CD20 was achieved through an E. coli-based in vitro coupled transcription-translation system. Surface plasmon resonance results showed that rituximab (an antileukemia drug) has nanomolar affinity with the CD20 protein, which aligns with published results. Notably, a previously hard-to-express claudin 18.2 recombinant protein was successfully expressed in the same reaction system by replacing its membrane-spanning domains with the transmembrane domains of CD20. The folding of the extracellular domain of the chimeric protein was verified using a commercial anti-claudin 18 antibody. This study provides a novel concept for promoting the expression of four-pass transmembrane proteins and lays the foundation for the large-scale industrial production of membrane-associated drug targets, similar to claudin 18.2.


Asunto(s)
Antígenos CD20 , Escherichia coli , Antígenos CD20/genética , Antígenos CD20/metabolismo , Escherichia coli/metabolismo , Rituximab/genética , Rituximab/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Claudinas/metabolismo
6.
Bioorg Med Chem ; 110: 117831, 2024 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-39004051

RESUMEN

The CD20 antigen is a key target for several diseases including lymphoma and autoimmune diseases. For over 20 years, several monoclonal antibodies were developed to treat CD20-related disorders. As many therapeutic proteins, their clinical use is however limited due to their nature with a costly biotechnological procedure and side effects such as the production of anti-drug neutralizing antibodies. Nucleic acid aptamers have some advantages over mAbs and are currently investigated for clinical use. We herein report the selection of DNA aptamer by using a peptide-based CE-SELEX (Capillary Electrophoresis-Systematic Evolution of Ligands by Exponential Enrichment) method. It was demonstrated that these aptamers bind specifically a CD20-expressing human cell line, with Kd estimated from isothermal titration calorimetry experiments in the micromolar range. This study demonstrates that the CE-SELEX is suitable as alternative method to the conventional Cell-SELEX to discover new cell-targeting compounds.


Asunto(s)
Antígenos CD20 , Aptámeros de Nucleótidos , Electroforesis Capilar , Péptidos , Técnica SELEX de Producción de Aptámeros , Humanos , Antígenos CD20/metabolismo , Antígenos CD20/química , Aptámeros de Nucleótidos/química , Péptidos/química , Péptidos/farmacología , Péptidos/metabolismo , Péptidos/aislamiento & purificación , Línea Celular Tumoral
7.
J Immunol ; 209(2): 379-390, 2022 07 15.
Artículo en Inglés | MEDLINE | ID: mdl-35768150

RESUMEN

NK cells are promising cellular therapeutics against hematological and solid malignancies. Immunogenetic studies have identified that various activating killer cell Ig-like receptors (KIRs) are associated with cancer outcomes. Specifically, KIR2DS2 has been associated with reduced incidence of relapse following transplant in hematological malignancies and improved outcomes in solid tumors, but the mechanism remains obscure. Therefore, we investigated how KIR2DS2 expression impacts NK cell function. Using a novel flow cytometry panel, we show that human NK cells with high KIR2DS2 expression have enhanced spontaneous activation against malignant B cell lines, liver cancer cell lines, and primary chronic lymphocytic leukemia cells. Surface expression of CD16 was increased on KIR2DS2high NK cells, and, accordingly, KIR2DS2high NK cells had increased activation against lymphoma cells coated with the clinically relevant anti-CD20 Abs rituximab and obinutuzumab. Bulk RNA sequencing revealed that KIR2DS2high NK cells have upregulation of NK-mediated cytotoxicity, translation, and FCGR gene pathways. We developed a novel single-cell RNA-sequencing technique to identify KIR2DS2+ NK cells, and this confirmed that KIR2DS2 is associated with enhanced NK cell-mediated cytotoxicity. This study provides evidence that KIR2DS2 marks a population of NK cells primed for anticancer activity and indicates that KIR2DS2 is an attractive target for NK-based therapeutic strategies.


Asunto(s)
Células Asesinas Naturales , Receptores KIR , Antígenos CD20/metabolismo , Línea Celular Tumoral , Citometría de Flujo , Humanos , Células Asesinas Naturales/metabolismo , Receptores KIR/genética , Receptores KIR/metabolismo , Rituximab/metabolismo , Rituximab/farmacología , Rituximab/uso terapéutico
8.
Mol Biol Rep ; 51(1): 751, 2024 Jun 14.
Artículo en Inglés | MEDLINE | ID: mdl-38874667

RESUMEN

BACKGROUND: Recently, new and advanced techniques have been adopted to design and produce nanobodies, which are used in diagnostic and immunotherapy treatments. Traditionally, nanobodies are prepared from camelid immune libraries that require animal treatments. However, such approaches require large library sizes and complicated selection procedures. The current study has employed CDR grafting and site-directed mutagenesis techniques to create genetically engineered nanobodies against the tumor marker CD20 (anti-CD20 nanobodies) used in leukemia treatment. METHODS AND RESULTS: In this study, we utilized the swapping method to graft CDRs from the VH Rituximab antibody to VHH CDRs. We aimed to enhance the binding affinity of the nanobodies by substituting the amino acids (Y101R-Y102R-Y107R) in the VHH-CDR3. To assess the binding capacity of the mutated nanobodies, we conducted an ELISA test. Moreover, through flow cytometry analysis, we compared the fluorescence intensity of the grafted CD20 and mutant nanobodies with that of the commercially available human anti-CD20 in Raji cells. The results showed a significant difference in the fluorescence intensity of the grafted nanobodies and mutant nanobodies when compared to the commercially available human anti-CD20. CONCLUSION: The approach we followed in this study makes it possible to create multiple anti-CD20 nanobodies with varying affinities without the need for extensive selection efforts. Additionally, our research has demonstrated that computational tools are highly reliable in designing functional nanobodies.


Asunto(s)
Afinidad de Anticuerpos , Antígenos CD20 , Regiones Determinantes de Complementariedad , Mutagénesis Sitio-Dirigida , Rituximab , Anticuerpos de Dominio Único , Anticuerpos de Dominio Único/genética , Anticuerpos de Dominio Único/inmunología , Mutagénesis Sitio-Dirigida/métodos , Antígenos CD20/inmunología , Antígenos CD20/genética , Antígenos CD20/metabolismo , Humanos , Rituximab/farmacología , Regiones Determinantes de Complementariedad/genética , Regiones Determinantes de Complementariedad/inmunología , Línea Celular Tumoral , Animales
9.
Cell Mol Biol (Noisy-le-grand) ; 70(5): 155-160, 2024 May 27.
Artículo en Inglés | MEDLINE | ID: mdl-38814221

RESUMEN

In order to explore a new mode for the diagnosis of angioimmunoblastic T-cell lymphoma (AITL), 31 cases of AITL and 28 cases of peripheral T-cell lymphoma, not otherwise specified (PTCL-NOS) were used as the study subjects. Identifying T follicular helper (TFH) cells with CD4, CD10, Bcl-6, and PD-1, identifying proliferative B cells with CD20 and EZH2, identifying proliferative follicular dendritic cells (FDCs) with CD21 and CD23, and analyzing the value of TFH/B/FDC proliferation and immunolocalization in the diagnosis of AITL. (1) Outside the inherent lymphoid follicles, simultaneous proliferation of TFH/B/FDC (a new diagnostic mode) were observed in AITL [83.87%; 26/31], with their immunolocalizations in the same site [83.87%; 26/31], while this phenomenon was not observed in 28 cases of PTCL-NOS (P<0.05). (2) The sensitivity and specificity of using this new mode to diagnose AITL were both high (83.87%, 100%), which was superior to CD2 (100%, 0%), CD3 (100%, 0%), CD4 (100%, 32.14%), CD5 (100%, 25%), CD10 (61.9%, 100%), Bcl-6 (42.86%, 100%), PD-1 (83.87%, 96.43%), and its Youden Index (0.84) was the highest. The areas under the curve (AUC) of CD10, Bcl-6, PD-1, and new mode to diagnosis AITL were 0.81, 0.71, 0.90, and 0.92, respectively, while the new mode had the highest AUC. The simultaneous proliferation of TFH/B/FDC cells outside the inherent lymphoid follicles can be used to assist in the diagnosis of AITL, and the simultaneous spatiotemporal proliferation of TFH/B/FDC cells is a specific immunomorphology of AITL.


Asunto(s)
Proteínas Proto-Oncogénicas c-bcl-6 , Humanos , Femenino , Masculino , Persona de Mediana Edad , Anciano , Proteínas Proto-Oncogénicas c-bcl-6/metabolismo , Neprilisina/metabolismo , Linfadenopatía Inmunoblástica/diagnóstico , Linfadenopatía Inmunoblástica/patología , Células Dendríticas Foliculares/patología , Células Dendríticas Foliculares/metabolismo , Receptor de Muerte Celular Programada 1/metabolismo , Adulto , Linfoma de Células T/diagnóstico , Linfoma de Células T/patología , Linfoma de Células T/metabolismo , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Proliferación Celular , Linfocitos B/inmunología , Linfocitos B/metabolismo , Células T Auxiliares Foliculares/inmunología , Células T Auxiliares Foliculares/metabolismo , Receptores de Complemento 3d/metabolismo , Receptores de Complemento 3d/análisis , Antígenos CD20/metabolismo , Antígenos CD20/análisis , Linfoma de Células T Periférico/diagnóstico , Linfoma de Células T Periférico/patología , Antígenos CD4/metabolismo , Sensibilidad y Especificidad , Anciano de 80 o más Años , Inmunohistoquímica/métodos , Curva ROC
10.
J Cutan Pathol ; 51(8): 614-617, 2024 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-38725374

RESUMEN

Mycosis fungoides (MF) represents the most common type of primary cutaneous T-cell lymphoma. Recognition of MF variants with divergent immunophenotypes is important for accurate diagnosis and appropriate management, as they can be confused with other lymphoma subtypes. We present a case of a 49-year-old male previously diagnosed with a cutaneous lymphoproliferative disorder with an unusual NK/T-cell phenotype. He presented with a 10-year history of pelvic girdle rash involving the right hip and upper thigh. The lesions were characterized as atrophic patches concentrated in sun-protected areas and involving 10% of the body surface area. Shave biopsies revealed an atypical epidermotropic infiltrate composed of hyperchromatic small to medium-sized lymphocytes with perinuclear halos and "tagging" along the dermal-epidermal junction. The immunophenotype was unusual in that the neoplastic lymphocytes showed complete loss of pan T-cell antigens along with expression of CD56, cytotoxic markers, and weak CD20. All other B-cell markers were negative. The combination of clinical findings, in addition to the histopathologic and immunophenotypic profile, were diagnostic of null T-cell phenotype MF with aberrant expression of CD56 and CD20. Null T-cell phenotype MF is very uncommon, can be diagnostically challenging, and can mislead the diagnosis of aggressive lymphoma subtypes.


Asunto(s)
Antígenos CD20 , Antígeno CD56 , Micosis Fungoide , Neoplasias Cutáneas , Humanos , Micosis Fungoide/patología , Micosis Fungoide/diagnóstico , Micosis Fungoide/metabolismo , Masculino , Persona de Mediana Edad , Antígeno CD56/metabolismo , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/diagnóstico , Neoplasias Cutáneas/metabolismo , Antígenos CD20/metabolismo , Inmunofenotipificación/métodos , Fenotipo , Linfocitos T/patología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Diagnóstico Diferencial , Biomarcadores de Tumor/metabolismo
11.
J Endocrinol Invest ; 47(11): 2865-2871, 2024 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-38642306

RESUMEN

BACKGROUND: CD20+ T cells represent up to 5% of circulating T lymphocytes. These cells have been shown to produce higher levels of IL-17A and IFN-γ than those of CD20- T lymphocytes. Some reports described the role of CD20+ T cells in autoimmune disorders such as multiple sclerosis and rheumatoid arthritis possibly due to their ability to produce these inflammatory cytokines. This study is aimed at describing the behavior of CD20+ T lymphocytes in the most frequent autoimmune disorder, i.e., Hashimoto's thyroiditis (HT), presenting isolated or associated to further autoaggressive disorders in a frame of poly-autoimmunity. METHODS: The study group encompasses 65 HT patients: 23 presenting in isolated form (IT) and 42 with an associated non-endocrine autoimmune disorder [16 with chronic atrophic gastritis (CAG), 15 with nonsegmental vitiligo (VIT), and 11 with celiac disease (CD)]. Twenty healthy donors act as control group (HD). Chronic use of interfering drugs, severe or chronic disorders, and pregnancy and lactation were used as exclusion criteria. Whole blood samples (100 µl) were stained with fluorescent-labeled antibodies (anti-CD45, anti-CD3, anti-CD19, anti-CD16, anti-CD56, anti-CD4, anti-CD8, anti-CD20). Red blood cells were then lysed by adding 1 ml of hypotonic buffer, and samples were acquired on a Flow Cytometer. RESULTS: CD3+CD8+CD20+ T lymphocytes' percentages, were significantly higher in the whole group of autoimmune patients compared to healthy donors (p = 0.0145). Dividing HT patients based on the type of presentation of autoimmune thyroiditis, CAG group showed the highest percentage of these cells as compared to HD and CD (p = 0.0058). IT patients showed higher percentages of CD3+ CD8+CD20+ cells than those of HD patients although not reaching statistical significance. However, dividing IT group based on thyroid function, hypothyroid patients showed higher CD8+CD20+ cell percentages than those of HD and euthyroid patients (p = 0.0111). Moreover, in IT patients, these cells were negatively correlated with FT4 levels (p = 0.0171; r = -0.4921). CONCLUSIONS: These preliminary findings indicate that CD8+CD20+ T cells are activated in patients with autoimmune thyroiditis and may behave differently according to the presence of poly-autoimmunity and hypothyroidism.


Asunto(s)
Antígenos CD20 , Enfermedad de Hashimoto , Poliendocrinopatías Autoinmunes , Humanos , Enfermedad de Hashimoto/inmunología , Enfermedad de Hashimoto/sangre , Femenino , Proyectos Piloto , Masculino , Adulto , Poliendocrinopatías Autoinmunes/inmunología , Poliendocrinopatías Autoinmunes/sangre , Poliendocrinopatías Autoinmunes/diagnóstico , Persona de Mediana Edad , Antígenos CD20/inmunología , Antígenos CD20/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Estudios de Casos y Controles , Anciano , Vitíligo/inmunología , Vitíligo/patología , Vitíligo/sangre , Gastritis Atrófica/inmunología , Gastritis Atrófica/patología , Gastritis Atrófica/sangre , Enfermedad Celíaca/inmunología , Enfermedad Celíaca/sangre , Enfermedad Celíaca/diagnóstico , Enfermedad Celíaca/patología
12.
Am J Dermatopathol ; 46(10): 672-674, 2024 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-39293664

RESUMEN

ABSTRACT: Although CD20 expression is typically scarce in mycosis fungoides (MF), it is more commonly associated with T-cell lymphomas. Nevertheless, isolated instances of CD20-positive MF have been documented infrequently. Here, we present a unique case of CD20-positive MF in a 30-year-old man who manifested with a hypopigmented patch on the anterior chest. Histopathological examination revealed epidermotropic infiltrates of small- to medium-sized lymphocytes with hyperchromatic and cerebriform nuclei aligned along the basal and low-mid layers of the epidermis. Immunophenotypic analysis demonstrated neoplastic T cells expressing CD4+, CD8+, and CD3+ with the loss of CD7. Intriguingly, a notable subset of the neoplastic T cells exhibited CD20 expression. This case contributes to the sparse literature on CD20-positive MF and underscores its diagnostic and clinical ramifications. The role of B cells has been more thoroughly characterized in T-cell lymphomas other than MF. However, its significance in MF remains unclear due to the scarcity of reported cases. Some hypotheses propose that the B cells' expression might indicate immune dysregulation or complex interactions within the tumor microenvironment. Another perspective suggests it could signify a progression of the disease towards a more aggressive lymphoma phenotype. Further investigation and documentation of similar cases is imperative to elucidate the clinical features, prognosis, and optimal therapeutic strategies. The long-term prognosis and outcomes in patients with hypopigmented MF and CD20 positivity remain ambiguous, underscoring the necessity for continued research and scrutiny of analogous cases.


Asunto(s)
Antígenos CD20 , Micosis Fungoide , Neoplasias Cutáneas , Humanos , Micosis Fungoide/patología , Micosis Fungoide/inmunología , Micosis Fungoide/metabolismo , Masculino , Adulto , Antígenos CD20/metabolismo , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/inmunología , Neoplasias Cutáneas/metabolismo , Biomarcadores de Tumor/análisis , Biomarcadores de Tumor/metabolismo , Inmunofenotipificación , Relevancia Clínica
13.
Proc Natl Acad Sci U S A ; 118(7)2021 02 16.
Artículo en Inglés | MEDLINE | ID: mdl-33563755

RESUMEN

CD20 is a B cell-specific membrane protein and represents an attractive target for therapeutic antibodies. Despite widespread usage of anti-CD20 antibodies for B cell depletion therapies, the biological function of their target remains unclear. Here, we demonstrate that CD20 controls the nanoscale organization of receptors on the surface of resting B lymphocytes. CRISPR/Cas9-mediated ablation of CD20 in resting B cells resulted in relocalization and interaction of the IgM-class B cell antigen receptor with the coreceptor CD19. This receptor rearrangement led to a transient activation of B cells, accompanied by the internalization of many B cell surface marker proteins. Reexpression of CD20 restored the expression of the B cell surface proteins and the resting state of Ramos B cells. Similarly, treatment of Ramos or naive human B cells with the anti-CD20 antibody rituximab induced nanoscale receptor rearrangements and transient B cell activation in vitro and in vivo. A departure from the resting B cell state followed by the loss of B cell identity of CD20-deficient Ramos B cells was accompanied by a PAX5 to BLIMP-1 transcriptional switch, metabolic reprogramming toward oxidative phosphorylation, and a shift toward plasma cell development. Thus, anti-CD20 engagement or the loss of CD20 disrupts membrane organization, profoundly altering the fate of human B cells.


Asunto(s)
Antígenos CD20/metabolismo , Linfocitos B/inmunología , Antígenos CD19/metabolismo , Línea Celular Tumoral , Membrana Celular/metabolismo , Células Cultivadas , Humanos , Activación de Linfocitos , Receptores de Antígenos de Linfocitos B/metabolismo
14.
Int J Mol Sci ; 25(16)2024 Aug 18.
Artículo en Inglés | MEDLINE | ID: mdl-39201672

RESUMEN

Multiple sclerosis (MS) is a disease of the central nervous system (CNS) characterized by inflammation and autoimmune responses. This review explores the participation of T cells, particularly certain CD3+CD20+ T cells, in the clinical manifestations of MS and highlights their presence in diagnosed patients. These T cells show aberrant expression of CD20, normally considered a B-cell marker. In this review, relevant journal articles available in PubMed and CINAHL were identified by employing diverse search terms, such as MS, CD3+CD20+ T cells, the incidence and significance of CD3+CD20+ T cells in MS patients, and the impact of rituximab treatment. The search was limited to articles published in the ten-year period from 2014 to 2024. The results of this review suggest that most scholars agree on the presence of CD3+CD20+ T cells in cerebrospinal fluid. Emerging concepts relate to the fundamental role of CD20-expressing T cells in determining the target and efficacy of MS therapeutics and the presence of T cells in the cerebrospinal fluid of MS patients. The results clearly show that CD20+ T cells indicate disease chronicity and high disease activity.


Asunto(s)
Antígenos CD20 , Complejo CD3 , Esclerosis Múltiple , Rituximab , Linfocitos T , Humanos , Esclerosis Múltiple/inmunología , Esclerosis Múltiple/metabolismo , Esclerosis Múltiple/patología , Antígenos CD20/metabolismo , Antígenos CD20/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Complejo CD3/metabolismo , Complejo CD3/inmunología , Rituximab/uso terapéutico
15.
Prep Biochem Biotechnol ; 53(3): 239-246, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-35579623

RESUMEN

Several monoclonal antibodies targeting the CD20 have been produced and Ofatumumab is a case in point. Although whole antibodies target cancer cells effectively, their applications are restricted in some ways. Single-chain fragment variable antibodies, rather than employing the entire structure of antibodies, have proven a practical approach for creating completely functional antigen-binding fragments. In current research, the DNA coding sequence of VL and VH of the wild and mutant forms of ofatumumab were joined with a flexible linker (GGGGS)3 separately. Using the E. coli BL21 (DE3) expression system, the VL-linker-VH genes were cloned into the pET-28 a (+), and the associated recombinant proteins were produced. Purified and refolded scFvs (scFv-C and scFv-V3) represented a concentration of around 0.7 mg/ml from 1 L of initial E. coli culture with a molecular weight of about 27 kDa. Affinity measurement disclosed anti-CD20 scFv-V3 possesses a higher affinity constant compared to anti-CD20 scFv-C. The recombinant scFvs exclusively attach to Raji cells but not to Jurkat cells, according to a cell-ELISA analysis. The MTT test signified anti-CD20 scFvs could affect cell viability in Raji cells but had no impact on Jurkat cells and also, Raji cells viability was affected more significantly by anti-CD20 scFv-V3.


Asunto(s)
Antígenos CD20 , Anticuerpos de Cadena Única , Humanos , Antígenos CD20/genética , Antígenos CD20/metabolismo , Clonación Molecular , Escherichia coli/genética , Escherichia coli/metabolismo , Anticuerpos Monoclonales Humanizados/genética , Anticuerpos Monoclonales Humanizados/farmacología , Anticuerpos Monoclonales Humanizados/metabolismo , Anticuerpos de Cadena Única/genética
16.
Inflamm Res ; 71(10-11): 1181-1189, 2022 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-35951029

RESUMEN

INTRODUCTION: Although CD20 is classically a B cell marker, in the last three decades, dim expression has been noted on a subset of T cells as well that has been independently verified by a number of groups. Our understanding of these cells and their function is not well established. METHODS: A thorough review of original articles on CD20+ T cells was undertaken of Pubmed by using combination of phrases including "CD20+", "CD20-positive" and "T cells". Articles in English were considered, and there was no time restriction. RESULTS: CD20+ T cells express the standard T cell markers and, in comparison to CD20¯ T cells, appear to express greater inflammatory cytokines and markers of effector function. Although the ontogeny of these cells is still being established, the current theory is that CD20 may be acquired by trogocytosis from B cells. CD20+ T cells may be found in healthy controls and in a wide range of pathologies including autoimmune diseases, haematological and non-haematological malignancies and human immunodeficiency virus (HIV) infections. One of the best studied diseases where these cells are found is multiple sclerosis (MS) where a number of therapeutic interventions, including anti-CD20 depletion, have been shown to effectively deplete these cells. CONCLUSION: This review summarises the latest understanding of CD20+ T cells, their presence in various diseases, their putative function and how they may be an ongoing target of CD20-depleting agents. Unfortunately, our understanding of these cells is still at its infancy and ongoing study in a wider range of pathologies is required.


Asunto(s)
Antígenos CD20 , Esclerosis Múltiple , Humanos , Antígenos CD20/metabolismo , Antígenos CD20/uso terapéutico , Subgrupos de Linfocitos T , Linfocitos B , Recuento de Linfocitos
17.
J Immunol ; 205(7): 1743-1751, 2020 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-32839239

RESUMEN

Myasthenia gravis (MG) is an autoimmune disease in which Abs target neuromuscular junction proteins, in particular the acetylcholine receptor. We previously identified the antiapoptotic protein survivin in the autoreactive B cells and plasma cells of MG patients. To further define the role of survivin in MG, we have assessed PBMCs from 29 patients with MG and 15 controls. We confirmed the increased expression of survivin in CD20+ lymphocytes from MG patients compared with controls. Furthermore, the CD20+ population of cells from MG patients contained a higher percentage of extracellular survivin compared with controls. The analysis of CD4+ cells showed an increased percentage of intracellular survivin in MG patients compared with controls, whereas the extracellular survivin CD4+ percentage was unaffected. In an experimental mouse model of MG, we assessed the therapeutic potential of an Ab raised to a modified survivin peptide but cross-reactive to survivin. Ab treatment reduced disease severity, lowered acetylcholine receptor-specific Abs, and decreased CD19+ survivin+ splenocytes. The ability to target survivin through Ab recognition of autoreactive cells offers the potential for a highly specific therapeutic agent for MG.


Asunto(s)
Linfocitos B/metabolismo , Miastenia Gravis/inmunología , Survivin/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Animales , Anticuerpos/metabolismo , Antígenos CD20/metabolismo , Autoantígenos/inmunología , Linfocitos B/inmunología , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunidad Humoral , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Péptidos/inmunología , Adulto Joven
18.
Proc Natl Acad Sci U S A ; 116(51): 25800-25807, 2019 12 17.
Artículo en Inglés | MEDLINE | ID: mdl-31748274

RESUMEN

CD8+ T cells are believed to play an important role in multiple sclerosis (MS), yet their role in MS pathogenesis remains poorly defined. Although myelin proteins are considered potential autoantigenic targets, prior studies of myelin-reactive CD8+ T cells in MS have relied on in vitro stimulation, thereby limiting accurate measurement of their ex vivo precursor frequencies and phenotypes. Peptide:MHC I tetramers were used to identify and validate 5 myelin CD8+ T cell epitopes, including 2 newly described determinants in humans. The validated tetramers were used to measure the ex vivo precursor frequencies and phenotypes of myelin-specific CD8+ T cells in the peripheral blood of untreated MS patients and HLA allele-matched healthy controls. In parallel, CD8+ T cell responses against immunodominant influenza epitopes were also measured. There were no differences in ex vivo frequencies of tetramer-positive myelin-specific CD8+ T cells between MS patients and control subjects. An increased proportion of myelin-specific CD8+ T cells in MS patients exhibited a memory phenotype and expressed CD20 compared to control subjects, while there were no phenotypic differences observed among influenza-specific CD8+ T cells. Longitudinal assessments were also measured in a subset of MS patients subsequently treated with anti-CD20 monoclonal antibody therapy. The proportion of memory and CD20+ CD8+ T cells specific for certain myelin but not influenza epitopes was significantly reduced following anti-CD20 treatment. This study, representing a characterization of unmanipulated myelin-reactive CD8+ T cells in MS, indicates these cells may be attractive targets in MS therapy.


Asunto(s)
Anticuerpos Monoclonales/metabolismo , Antígenos CD20 , Linfocitos T CD8-positivos , Esclerosis Múltiple , Proteínas de la Mielina/metabolismo , Adolescente , Adulto , Antígenos CD20/inmunología , Antígenos CD20/metabolismo , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/metabolismo , Células Cultivadas , Femenino , Humanos , Masculino , Persona de Mediana Edad , Esclerosis Múltiple/inmunología , Esclerosis Múltiple/metabolismo , Adulto Joven
19.
Cancer Sci ; 112(9): 3585-3597, 2021 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-34252986

RESUMEN

Diffuse large B cell lymphoma (DLBCL) heterogeneity promotes recurrence and anti-CD20-based therapeutic resistance. Previous studies have shown that downregulation of MS4A1/CD20 expression after chemoimmunotherapy with rituximab leads to rituximab resistance. However, the mechanisms of CD20 loss remain unknown. We identified that pyruvate dehydrogenase kinase 4 (PDK4) is markedly elevated in DLBCL cells derived from both patients and cell lines with R-CHOP (rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone) resistance. We found that overexpression of PDK4 in DLBCL cells resulted in cell proliferation and resistance to rituximab in vitro and in vivo. Furthermore, loss of PDK4 expression or treatment with the PDK4 inhibitor dichloroacetate was able to significantly increase rituximab-induced cell apoptosis in DLBCL cells. Further studies suggested PDK4 mediates a metabolic shift, in that the main energy source was changed from oxidative phosphorylation to glycolysis, and the metabolic changes could play an important role in rituximab resistance. Importantly, by knocking down or overexpressing PDK4 in DLBCL cells, we showed that PDK4 has a negative regulation effect on MS4A1/CD20 expression. Collectively, this is the first study showing that targeting PDK4 has the potential to overcome rituximab resistance in DLBCL.


Asunto(s)
Antígenos CD20/metabolismo , Antineoplásicos Inmunológicos/administración & dosificación , Reprogramación Celular/genética , Resistencia a Antineoplásicos/genética , Glicoproteínas/metabolismo , Linfoma de Células B Grandes Difuso/tratamiento farmacológico , Linfoma de Células B Grandes Difuso/metabolismo , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/metabolismo , Rituximab/administración & dosificación , Transducción de Señal/genética , Adolescente , Adulto , Anciano , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Resistencia a Antineoplásicos/efectos de los fármacos , Femenino , Humanos , Linfoma de Células B Grandes Difuso/genética , Linfoma de Células B Grandes Difuso/patología , Masculino , Ratones , Persona de Mediana Edad , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/genética , Transfección , Resultado del Tratamiento , Carga Tumoral/efectos de los fármacos , Carga Tumoral/genética , Ensayos Antitumor por Modelo de Xenoinjerto , Adulto Joven
20.
Br J Cancer ; 125(12): 1687-1698, 2021 12.
Artículo en Inglés | MEDLINE | ID: mdl-34743199

RESUMEN

BACKGROUND: Small extracellular vesicles (sEVs) including exosomes, carrying the CD20, could be involved in immunotherapy resistance in diffuse large B cell lymphoma (DLBCL). We have reported endogenous brain-derived neurotrophic factor/TrkB (tropomyosin-related kinase B) survival axis in DLBCL. Here, we performed a comparative study of sEV production by germinal centre B cell (GCB) and activated B cell (ABC)-DLBCL cell lines, and analysed TrkB activation on this process. METHODS: GCB (SUDHL4 and SUDHL6) and ABC (OCI-LY3, OCI-LY10 and U2932) cell lines were used. sEVs were characterised using nanoparticle tracking analysis technology and western blot. CD20 content was also analysed by enzyme-linked immunoassay, and complement-dependent cytotoxicity of rituximab was investigated. 7,8-Dihydroxyflavone (7,8-DHF) was used as a TrkB agonist. In vivo role of sEVs was evaluated in a xenograft model. RESULTS: sEVs production varied significantly between DLBCL cells, independently of subtype. CD20 level was consistent with that of parental cells. Higher CD20 expression was found in sEVs after TrkB activation, with a trend in increasing their concentration. sEVs determined in vitro and in vivo protection from rituximab, which seemed CD20 level-dependent; the protection was enhanced when sEVs were produced by 7,8-DHF-treated cells. CONCLUSIONS: DLBCL-derived sEVs have the differential capacity to interfere with immunotherapy, which could be enhanced by growth factors like neurotrophins. Evaluating the sEV CD20 level could be useful for disease monitoring.


Asunto(s)
Antígenos CD20/metabolismo , Vesículas Extracelulares/metabolismo , Linfoma de Células B Grandes Difuso/genética , Glicoproteínas de Membrana/metabolismo , Receptor trkB/metabolismo , Animales , Línea Celular Tumoral , Xenoinjertos , Humanos , Ratones , Ratones SCID
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