Biotecnología aplicada para la artritis reumatoide en el Centro de Reumatología

Caracterización de Moléculas HLA tipo II y evaluación de citocinas en pacientes cubanos con Artritis Reumatoide

Revista Cubana de reumatología
Vol. 8, No. 9-10 (2006)

María del Carmen Domínguez Horta, N. Lorenzo, A. Barbera, María Victoria Hernandez, Ana María Torres Lima, M. Nazabal, H. Camacho, I. Hernández, Neisy Ortiz, V. Morera, G. Padrón

Resumen

La artritis reumatoide es una enfermedad de etiología multifactorial, que involucra la presencia de factores genéticos, ambientales, inmunológicos y hormonales. En los últimos años se ha establecido una asociación entre la predisposición a padecer esta enfermedad y la existencia de determinados haplotipos del HLA clase II. El objetivo fundamental de este trabajo es la caracterización del polimorfismo de las moléculas HLA-DQB1 y HLA-DRB de un grupo de pacientes cubanos con Artritis Reumatoide, además analizar una posible correlación entre los niveles de varias citocinas proinflamatorias en un grupo de estos pacientes, con los alelos HLA tipo II genotipados, el sexo y el tiempo de diagnosticada la enfermedad. El estudio se llevó a cabo en 50 pacientes cubanos con el diagnóstico de artritis reumatoide y un grupo control, compuesto de 211 donantes sanos. Los haplotipos HLA-DQ y HLA-DRB1 fueron determinados a través de la reacción en cadena de la polimerasa. La cuantificación de las citocinas se realizó empleando inmuno-ensayos comerciales. Los resultados obtenidos en este análisis indican que los alelos con un radio de la razón de probabilidades superior a 2 fueron para el caso de la molécula HLA-DQB1: HLADQB1 *03 y *06, y para la molécula HLA-DRB1 los alelos: *01, *04, *09 y *10. Encontramos además que los niveles de la citocina interferon ganma están significativamente aumentados en los pacientes con menos tiempo de diagnosticada la enfermedad. Este trabajo constituye el primer reporte de caracterización de las moléculas HLA tipo II, a través de técnicas moleculares, en pacientes cubanos con artritis reumatoide

Palabras claves: Artritis Reumatoide; HLA-DQ; HLA-DR; IFN.

Un ligando peptídico alterado como novedoso fármaco para el tratamiento de la artritis idiopática juvenil

Revista Cubana de Reumatología
Vol. 11, No. 13-14 (2009)

Lorenzo Pérez Noraylis, Barberá Betancourt Ariana, Cantera Oceguera Dolores, Alonso Amarys, Chall Elsy, Franco Lourdes, Padrón Palomares Ramón Gabriel, Domínguez Horta María del Carmen

Resumen

La inducción de tolerancia periférica usando autoantígenos constituye una terapia muy atractiva para las enfermedades autoinmunes. El objetivo de este trabajo es demostrar el potencial terapéutico de un ligando peptídico alterado derivado de la proteína de estrés térmico de 60 humana en el modelo animal de artritis inducida por adyuvante y en ensayos in vitro con células mononucleares en pacientes con artritis idiopática juvenil. Se comprobó a través del examen clínico e histopatológico de las ratas, que el péptido inhibe eficientemente el desarrollo de la enfermedad; además, la terapia de los animales con el péptido induce una reducción significativa de la expresión del TNF alfa. Por otra parte, la estimulación con ligandos peptídicos alterados de las células mononucleares de pacientes con artritis idiopática juvenil incrementó significativamente la frecuencia de células T reguladoras CD4+CD25+Foxp3+. Estos resultados avalan el potencial terapéutico de esta molécula para el tratamiento de la artritis idiopática juvenil y otras enfermedades autoinmunes.

Palabras clave: ligando peptídico, artritis idiopática juvenil, células T reguladoras

Observaciones clínicas en pacientes con artritis reumatoide tratados con anticuerpo monoclonal T1h (anti CD6)

Revista Cubana de reumatología
Vol. 11, No. 13-14 (2009)

Dinorah Marisabel Prada Hernández, Claudino Molinero Rodríguez, Roberto Vicente Torres Moya, Ana Marta López Mantecón, Jorge Alexis Gómez Morejón, Isabel María Hernández Cuellar, José Pedro Martínez Larrarte, Yusimí Reyes Pineda, Joel Rodríguez Milera

A clinical exploratory study with itolizumab, an anti-CD6 monoclonal antibody, in patients with rheumatoid arthritis

Results Immunol 2012 Nov 21;2:204-11.
doi: 10.1016/j.rinim.2012.11.001

Pedro C Rodriguez, Roberto Torres-Moya, Gil Reyes, Claudino Molinero, Dinorah Prada, Ana M Lopez, Isabel M Hernandez, Maria V Hernandez, Jose P Martinez, Xochel Hernandez, Angel Casaco, Mayra Ramos, Yisel Avila, Yinet Barrese, Enrique Montero, Patricia Hernandez

Abstract

T cells are involved in the pathogenesis of rheumatoid arthritis (RA). CD6 is a co-stimulatory molecule, predominantly expressed on lymphocytes, that has been linked to autoreactive responses. The purpose of this study was to evaluate the safety, immunogenicity and preliminary efficacy of itolizumab, a humanized anti-CD6 monoclonal antibody, in patients with active rheumatoid arthritis. Fifteen patients were enrolled in a phase I, open-label, dose-finding study. Five cohorts of patients received a weekly antibody monotherapy with a dose-range from 0.1 to 0.8 mg/kg. Itolizumab showed a good safety profile, with no severe or serious adverse events reported so far. No signs or symptoms associated with immunosuppression were observed in the study. Objective clinical responses were achieved in more than 80% of patients after treatment completion, and these responses tend to be sustained afterwards. This clinical study constitutes the first evidence of the safety and positive clinical effect of a monotherapy using an anti-CD6 antibody in patients with rheumatoid arthritis.

Keywords: ACR, American College of Rheumatology; AE, adverse events; CD6; CRP, C reactive protein; DMARD, disease-modifying antirheumatic drug; ESR, eritrosedimentation rate; Exploratory study; NSAIDs, nonsteroidal antiinflammatory drugs; RA, rheumatoid arthritis; RF, rheumatoid factor; Rheumatoid arthritis; SAE, serious adverse event.; T lymphocyte; iv, intravenous; mAbs, monoclonal antibodies.

Therapeutic effect of two altered peptide ligands derived from the human heat shock protein 60 in experimental models of rheumatoid arthritis

Biotecnología Aplicada
2013;30:153-156

María del C Domínguez, Norailys Lorenzo, Ariana Barberá, Gabriel Padrón, Ana María Torres, María V Hernández, Isabel Hernández, Rafael Gil, Aniel Sánchez, Vladimir Besada, Luis J González, Hilda Garay, Osvaldo Reyes, Ever Pérez, Matilde López, Yuliet Mazola, Karelia Cosme, Julio Ancizar

Abstract

Induction of immune tolerance as therapeutic approach for autoimmune diseases constitutes a current research focal point. In this sense, two Altered Peptide Ligands (APLs) were evaluated for the induction of peripheral tolerance in patients with Rheumatoid Arthritis (RA). Two novel T cell epitopes from human heat-shock protein 60 (hHsp60), an autoantigen involved in the pathogenesis of RA, were identified by bioinformatics tools and two APLs were designed from these epitopes (APL-1 and APL-2). APL-1 increases the proportions of the CD4+CD25highFoxP3+ regulatory T cells in ex vivo assays using PBMCs isolated from RA patients. While, APL-2 increased the IL-10 level and suppressed IL-17 secretion, and induces the activation of T cells through his ability to modify cell cycle phase’s distribution of CD4+ T cells from RA patients. Additionally, the therapeutic effect of these APLs in two animal models was evaluated: adjuvant induced arthritis (AA) in Lewis rat and collagen induced arthritis (CIA) in DBA/1 mice. Our approach was compared to metotrexate (MTX), the treatment of reference for RA, in CIA model. Clinical score, TNF-α levels and histopathology were monitored. Both APLs efficiently inhibited the course of AA and CIA, with significant reduction of the clinical and histopathology scores. The therapeutic effect induced by APLs is mediated by different molecular mechanisms, associ- ated with immunologic tolerance. These results indicate a therapeutic potentiality of these APLs and support further investigation for treatment of RA. This study won the Annual Award of the Academy of Sciences of Cuba in 2012.

Keywords: rheumatoid arthritis, altered peptide ligand, hHsp60, immune tolerance, regulatory T cells

Tratamiento actual de la artritis reumatoide. Perspectivas para el desarrollo de las terapias antígeno-específicas

Biotecnología Aplicada
2012;29:137-145

Ariana Barberá, Noraylis Lorenzo, María del Carmen Domínguez

Resumen

La artritis reumatoide es una enfermedad degenerativa caracterizada por la inflamación crónica de las articulaciones periféricas. La primera línea de tratamiento de esta enfermedad implica el uso de potentes antinflamatorios y drogas que provocan una supresión global del sistema inmune. Sin embargo, estos fármacos no inducen una remisión sostenida, y su uso puede causar una inmunosupresión importante que puede conducir a complicaciones. Por ello es necesario el desarrollo de nuevas modalidades terapéuticas para esta enfermedad. Las terapias antígeno-específicas suprimen las células patogénicas, sin afectar la propiedad del sistema inmune de responder ante las infecciones. Las proteínas de estrés térmico son candidatas promisorias en esta modalidad de tratamiento. Aunque se ha avanzado en el desarrollo de terapias antígeno-específicas eficientes en modelos animales con excelentes resultados, ha sido difícil trasladarlas a los seres humanos. El uso combinado de las terapias antígeno-específicas con los fármacos actuales puede ser una estrategia muy atractiva en el futuro cercano para lograr la remisión completa de la enfermedad. Algunas de estas combinaciones de tratamiento ya han comenzado a evaluarse en modelos animales y en pacientes con artritis reumatoide.

Palabras clave: artritis reumatoide, células T reguladoras, terapia biológica, citocina proinflamatoria,
terapia combinada

Therapeutic effect of two altered peptide ligands derived from the human heat shock protein 60 in experimental models of rheumatoid arthritis

Biotecnología Aplicada 2013;30:153-156

María del C Domínguez, Norailys Lorenzo, Ariana Barberá, Gabriel Padrón, Ana María Torres, María V Hernández, Isabel Hernández, Rafael Gil, Aniel Sánchez, Vladimir Besada, Luis J González, Hilda Garay, Osvaldo Reyes,  Ever Pérez, Matilde López, Yuliet Mazola, Karelia Cosme, Julio Ancizar

Induction of immune tolerance as therapeutic approach for autoimmune diseases constitutes a current research focal point. In this sense, two Altered Peptide Ligands (APLs) were evaluated for the induction of peripheral tolerance in patients with Rheumatoid Arthritis (RA). Two novel T cell epitopes from human heat-shock protein 60 (hHsp60), an autoantigen involved in the pathogenesis of RA, were identifi ed by bioinformatics tools and two APLs were designed from these epitopes (APL-1 and APL-2). APL-1 increases the proportions of the CD4+CD25highFoxP3+ regulatory T cells in ex vivo assays using PBMCs isolated from RA patients. While, APL-2 increased the IL-10 level and suppressed IL-17 secretion, and induces the activation of T cells through his ability to modify cell cycle phase’s distribution of CD4+ T
cells from RA patients. Additionally, the therapeutic effect of these APLs in two animal models was evaluated: adjuvant induced arthritis (AA) in Lewis rat and collagen induced arthritis (CIA) in DBA/1 mice. Our approach was compared to metotrexate (MTX), the treatment of reference for RA, in CIA model. Clinical score, TNF-α levels and histopathology were monitored. Both APLs effi ciently inhibited the course of AA and CIA, with signifi cant reduction of the clinical and histopathology scores. The therapeutic effect induced by APLs is mediated by different molecular mechanisms, associated with immunologic tolerance. These results indicate a therapeutic potentiality of these APLs and support further investigation for treatment of RA. This study won the Annual Award of the Academy of Sciences of Cuba in 2012.

Keywords: rheumatoid arthritis, altered peptide ligand, hHsp60, immune tolerance, regulatory T cells,
collagen induced arthritis, adjuvant induced arthritis

APL-2, an altered peptide ligand derived from heat-shock protein 60, induces interleukin-10 in peripheral blood mononuclear cell derived from juvenile idiopathic arthritis patients and downregulates the inflammatory response in collagen-induced arthritis model

Clin Exp Med
DOI 10.1007/s10238-014-0273-x

Norailys Lorenzo, Dolores Cantera, Ariana Barbera, Amaris Alonso, Elsy Chall, Lourdes Franco, Julio Ancizar, Yanetsy Nun˜ez, Fiorella Altruda, Lorenzo Silengo, Gabriel Padro´n, Maria del Carmen Dominguez

Abstract

Juvenile idiopathic arthritis (JIA) is a heterogeneous group of diseases characterized by autoimmune arthritis of unknown cause with onset before age of 16 years. Methotrexate provides clinical benefits in JIA. For children who do not respond to methotrexate, treatment with anti-tumor necrosis factor (TNF)-a is an option. However, some patients do not respond or are intolerant to anti-TNF therapy. Induction of peripheral tolerance has long been considered a promising approach to the treatment of chronic autoimmune diseases. We aimed to evaluate the potentialities of two altered peptide ligands (APLs) derived from human heat-shock protein 60, an autoantigen involved in the pathogenesis of autoimmune arthritis, in JIA patients. Interferon (IFN)-c, TNF-a and interleukin (IL)-10 levels were determined in ex vivo assays using peripheral blood mononuclear cells (PBMC) from these patients. Wild-type peptide and one of these APLs increased IFN-c and TNF-a levels. Unlike, the other APLs (called APL2) increased the IL-10 level without affecting IFN-c and TNF-a levels. On the other hand, APL2 induces a marked activation of T cells since it transforms cell cycle phase’s distribution of CD4? T cells from these patients. In addition, we evaluated the therapeutic effect of APL2 in collagen-induced arthritis model. Therapy with APL2 reduced arthritis scores and histological lesions in mice. This effect was associated to a decrease in TNF-a and IL-17 levels. These results indicate a therapeutic potentiality of APL2 for JIA.

Keyword: Altered peptide ligand Juvenile idiopathic arthritis Collagen-induced arthritis Tolerance Interleukin-10

An altered peptide ligand corresponding to a novel epitope from heat-shock protein 60 induces regulatory T cells and suppresses pathogenic response in an animal model of adjuvant-induced arthritis

Autoimmunity, 2011; Early Online: 1–12
DOI: 10.3109/08916934.2010.550590

Maria Del Carmen Domínguez, Noraylis Lorenzo, Ariana Barbera, Guillaume Darrasse-Jeze, Maria Victoria Hernádez, Ana Torres, Isabel Herna´Ndez, Rafael Gil, David Klatzmann,  Gabriel Padrón

Abstract

Induction of immune tolerance as therapeutic approach for autoimmune diseases constitutes a current research focal point. In this sense, we aimed to evaluate an altered peptide ligand (APL) for induction of peripheral tolerance in patients with rheumatoid arthritis (RA). A novel T-cell epitope from human heat-shock protein 60 (Hsp60), an autoantigen involved in the pathogenesis of RA, was identified by bioinformatics tools and an APL was design starting from this epitope. We investigated the ability of this APL for inducing regulatory T cells (Treg cells) in mice and evaluated the therapeutic effect of this peptide in an adjuvant-induced arthritis (AA) rat model. Clinical score, TNFa levels and histopathology were monitored, as well as the capacity of this APL for inducing Treg cells. Finally, the potentialities of the APL for inducing Treg cells were evaluated in ex vivo assays using mononuclear cells isolated from peripheral blood (PBMC). The APL induced an increase of the proportions of Treg cells in the draining lymph nodes of the injected site in mice. The APL efficiently inhibited the course of AA, with significant reduction of the clinical and histopathology score. This effect was associated with an increase of the proportions of Treg cells and a decrease of TNFa levels in spleen. Finally, stimulation of PBMCs from RA patients by the APL increases the proportions of the CD4þCD25highFoxP3þ Treg cells. These results indicate a therapeutic potentiality of APL and support further investigation of this candidate drug for treatment of RA.

Keywords: Rheumatoid arthritis, APL, HSP60, tolerance, regulatory T cells

Therapeutic effect of an altered peptide ligand derived from heat-shock protein 60 by suppressing of inflammatory cytokines secretion in two animal models of rheumatoid arthritis

Autoimmunity, 2012; Early Online: 1–11
DOI: 10.3109/08916934.2012.697592

N. Lorenzo, A. Barbera, MC. Domínguez, AM. Torres, MV. Hernández, I. Hernández, R. Gil, H. Garay, O. Reyes, F. Altruda, L. Silengo, G. Padrón

Abstract

Rheumatoid arthritis is a systemic autoinmmune disease mediated by T cells. Productive engagement of T cell receptors by major histocompatibility complex-peptide leads to proliferation, differentiation and the definition of effector functions. Altered peptide ligands (APL) generated by amino acid substitutions in the antigenic peptide have diverse effects on T cell response.We predicted a novel T cell epitope from human heat-shock protein 60, an autoantigen involved in the pathogenesis of rheumatoid arthritis. Three APLs were designed from this epitope and it was demostrated that these peptides induce the activation of T cells through their ability to modify cell cycle phase’s distribution of CD4 þ T cells from RA patients. Also, IL-17, TNF-a and IL-10 levels were determined in PBMC from these patients. Unlike the wild-type peptide and the other two APLs, APL2 increased the IL-10 level and suppressed IL-17 secretion in these assays. Therapeutic effect of this APL inadjuvant arthritis (AA) and collagen-induced arthritis (CIA) models was also evaluated. Clinical score, histopathology, inflammatory and regulatory cytokine concentration were monitored in the animals. APL2 efficiently inhibited the progression of AA and CIA with a significant reduction of the clinical and histopathogic score. Therapeutic effect of APL2 on CIA was similar to that obtained with MTX; the standard treatment for RA. This effect was associated with a decrease of TNF-a and IL-17 levels. These results suggest that the therapeutic effect of APL2 is mediated in part by down-regulation of inflammatory cytokines and support the potential use of APL2 as a therapeutic drug in RA patients.

Keywords: Rheumatoid Arthritis, AA, HSP60, CIA, APL, inflammatory cytokines

Phase I Clinical Trial with a Novel Altered Peptide Ligand Derived from Human Heat-Shock Protein 60 for Treatment of Rheumatoid Arthritis: Safety, Pharmacokinetics and Preliminary Therapeutic Effects

J Clin Trials 2018, 8:1
DOI: 10.4172/2167-0870.1000339

Dinorah Prada, Jorge Gómez, Norailys Lorenzo, Oreste Corrales, Ana López, Evelio González, Ania Cabrales, Yusimy Reyes, Yuliet Bermudez, Yisel Avila, Lina Pérez, Claudio Molinero, Osmel Martinez, Leonardo Oramas, Yaysel Miñoso, Yassel Ramos, Hilda Garay, Ever Pérez, Matilde López, Osvaldo Reyes, Yolanda Cruz, Alfredo Hernández, Cabal Carlos, Vladimir Besada, Luis Javier González, Gabriel Padrón, and Maria del Carmen Domínguez Horta

Abstract

Background: CIGB 814 is an Altered Peptide Ligand (APL) from a CD4+ T-cell epitope of human heat shock protein 60 (HSP60), an auto-antigen involved in the pathogenesis of rheumatoid arthritis (RA). It induced mechanisms associated with restoration of peripheral tolerance in preclinical studies. This clinical trial was
conducted to assess safety and pharmacokinetics (PK) of CIGB-814 in patients with RA.
Method: 20 patients with moderated active RA were included in an open label trial. Sequential dose-escalation of 1, 2.5 and 5 mg of CIGB-814 was studied. Consecutive groups of six, five and nine patients received a subcutaneous dose weekly of the peptide during the first month and one dose monthly during the next five months.
Clinical response in patients was evaluated according to the American College of Rheumatology (ACR) and Disease Activity Score in 28 joints (DAS 28) criteria. Function and health-related quality of life, quantification of proinflammatory cytokines and radiographic changes in patients by magnetic resonance imaging (MRI) were also
assessed.
Result: The treatment was well tolerated at all doses. Only mild events were observed. PK study showed that CIGB-814 reached the maximum concentration in plasma in 30 min and was cleared mostly after 4 h. CIGB-814 reduced disease activity and MRI score in patients. This effect was less marked with the dose of 5 mg. Five and
eleven out of 18 patients achieved ACR 50 and ACR 70 respectively at the end of the treatment. In addition, patients showed decreases of DAS28 scores, during treatment and at the end of the follow-up. This therapy improved function and health-related quality of life of patients. CIGB-814 significantly decreased interleukin (IL)-17 in patients treated with 2.5 mg. Therapy with 1 mg and 2.5 mg of CIGB-814 led to significant reduction of interferon gamma (IFN-γ).
Conclusion: Phase I concluded showing safety of CIGB-814. The PK profile revealed that peptide is cleared from plasma very rapidly. Results indicated preliminary evidences of clinical efficacy and support further clinical investigation of this peptide for treatment of RA.

Keywords: APL; HSP60; Rheumatoid arthritis; Clinical trial; Safety

CIGB-814, an altered peptide ligand derived from human heat-shock protein 60, decreases anti-cyclic citrullinated peptides antibodies in patients with rheumatoid arthritis

Clin Rheumatol 2019 Mar;38(3):955-960.
doi: 10.1007/s10067-018-4360-3

Oreste Corrales, Laura Hernández, Dinorah Prada, Jorge Gómez, Yusimy Reyes, Ana Marta López, Luis Javier González, Maria Del Carmen Domínguez Horta  

Abstract

Rheumatoid arthritis (RA) is a chronic T cell-mediated autoimmune disease. Serum autoantibodies against cyclic citrullinated peptides (anti-CCP) are significant markers for diagnosis and prognosis of this disease. Induction of immune tolerance as therapeutic approach for RA constitutes a current research focal point. In this sense, we carried out a phase I clinical trial in RA patients with a new therapeutic candidate (called CIGB-814); which induced mechanisms associated with restoration of peripheral tolerance in preclinical studies. CIGB 814 is an altered peptide ligand (APL), derived from a CD4+ T cell epitope of human heat-shock protein 60 (HSP60), an autoantigen involved in the pathogenesis of RA. Twenty patients with moderate disease activity were included in this open label trial. Sequential dose-escalation of 1, 2.5 and 5 mg of CIGB-814 was studied. Consecutive groups of six, five, and nine patients received a subcutaneous dose weekly of the peptide during the first month and one dose monthly during the next 5 months. The peptide was well tolerated and reduced disease activity. Here, we reported the quantification of anti-CCP antibodies during the treatment with this APL and in the follow-up stage. Anti-CCP antibodies were quantified in the plasma from patients by a commercial enzyme immunoassay at baseline (T0) and at weeks 28 and 48. Results showed that CIGB-814 induced a significant reduction of anti-CCP antibodies. In addition, this decrease correlated with clinical improvement in patients assessed by Disease Activity Score in 28 joints (DAS28) criteria. These findings reinforce the therapeutic potential of CIGB-814.

Keywords: APL; Anti-CCP; CIGB-814; HSP60; Rheumatoid arthritis.