Inmunoproteómica aplicada a la presentación de péptidos virales por moléculas de MHC de clase I

Tesis doctoral inédita. Universidad Autónoma de Madrid, Facultad de Ciencias. Fecha de lectura: 21-02-201

H-2Ld class I molecule protects an HIV N-extended epitope from in vitro trimming by endoplasmic reticulum aminopeptidase associated with antigen processing.

In the classical MHC class I Ag presentation pathway, antigenic peptides derived from viral proteins by multiple proteolytic cleavages are transported to the endoplasmic reticulum lumen and are then exposed to ami-nopeptidase activity. In the current study, a long MHC class I natural ligand recognized by cytotoxic T lymphocytes was used to study the kinetics of degradation by aminopeptidase. The in vitro data indicate that this N-extended peptide is efficiently trimmed to a 9-mer, unless its binding to the MHC molecules protects the full-length peptide.We thank Dr. A. K. Stout of the NIH Tetramer Facility for providing the peptide/MHC complex reagent. This work was suppor ted by grants provi ded by Programa Ramón y Cajal, and Fundación FIPSE to D. L.; by grants provided by Comunidad de Madrid and Ministerio de Educación y Ciencia to M. D. V.; and by a joint grant provided by Instituto de Salud Carlos III to D. L., and M. D. V.S

Natural HLA-B*2705 protein ligands with glutamine as anchor motif: implications for HLA-B27 association with spondyloarthropathy

The presentation of short viral peptide antigens by human leukocyte antigen (HLA) class I molecules on cell surfaces is a key step in the activation of cytotoxic T lymphocytes, which mediate the killing of pathogen-infected cells or initiate autoimmune tissue damage. HLA-B27 is a well known class I molecule that is used to study both facets of the cellular immune response. Using mass spectrometry analysis of complex HLA-bound peptide pools isolated from large amounts of HLA-B*2705(+) cells, we identified 200 naturally processed HLA-B*2705 ligands. Our analyses revealed that a change in the position (P) 2 anchor motif was detected in the 3% of HLA-B*2705 ligands identified. B*2705 class I molecules were able to bind these six GlnP2 peptides, which showed significant homology to pathogenic bacterial sequences, with a broad range of affinities. One of these ligands was able to bind with distinct conformations to HLA-B27 subtypes differentially associated with ankylosing spondylitis. These conformational differences could be sufficient to initiate autoimmune damage in patients with ankylosing spondylitis-associated subtypes. Therefore, these kinds of peptides (short, with GlnP2, and similar low affinity to all HLA-B27 subtypes tested but with unlike conformations in differentially ankylosing spondylitis-associated subtypes) must not be excluded from future researches involving potential arthritogenic peptides.This work was supported by grants from the Programa Ramón y Cajal and the Ministerio de Ciencia e Innovación (to D. L.) and from the Israel Science Foundation (ISF 916/05) (to A. A.).N

The viral transcription group determines the HLA class I cellular immune response against human respiratory syncytial virus

The cytotoxic T-lymphocyte-mediated killing of virus-infected cells requires previous recognition of short viral antigenic peptides bound to human leukocyte antigen class I molecules that are exposed on the surface of infected cells. The cytotoxic T-lymphocyte response is critical for the clearance of human respiratory syncytial virus infection. In this study, naturally processed viral human leukocyte antigen class I ligands were identified with mass spectrometry analysis of complex human leukocyte antigen-bound peptide pools isolated from large amounts of human respiratory syncytial virus-infected cells. Acute antiviral T-cell response characterization showed that viral transcription determines both the immunoprevalence and immunodominance of the human leukocyte antigen class I response to human respiratory syncytial virus. These findings have clear implications for antiviral vaccine design.This work was supported by the Spanish Ministry of Economy grants BIO2011-25636 to D.L. and to A. A. from the ISF 916/05. The funding agencies had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. We have no conflicting financial interestsS

Generation of Molecular Diversity from Amino Acids. A Source for the Discovery of New TRP Channel Modulators

Trabajo presentado en el IV RECI: New Horizons in Ion Channel Research, celebrado en Cuenca (España) del 12 al 13 de febrero de 2013.Ion channels are central and challenging targets in medicinal chemistry but, because of the scarce structural knowledge, rational approaches to ion channel modulators are still rare. Moreover, the multimodal activation of some channels, like TRPs, complicates still more the scenario for rational discovery programs. Due to these facts, most strategies directed to identify ion channel modulators rely on the screening of peptide and small-molecule libraries. In this context, we have been involved in the development of synthetic pathways for the generation of diverse, chiral, highly functionalized linear and heterocyclic scaffolds from amino acids, and in the production of discrete libraries from them. The screening of these libraries on different TRP channels has allowed the discovery of some innovative hits that have progressed to hit-to-lead optimization programs. This communication will deal with the synthesis, structural characterization, and biological evaluation of a collection of β,γ–diaminoester derivatives that display significant activity at TRPV1, TRPM8 and TRPA1 channels. Compound RGM04-7, a selective.Supported by MICINN grants: Consolider-Ingenio 2010 (CSD2008-00005 and CSD2006-00015), SAF2009-09323 and BFU2009-08346, and the Generalitat Valenciana (PROMETEO/2010/046)

Discovery of New Antagonists for TRPM8 Channel by High Throughput Assay

Trabajo presentado en el IV RECI: New Horizons in Ion Channel Research, celebrado en Cuenca (España) del 12 al 13 de febrero de 2013.TRP ion channels family is represented by 85 members that can be organized by their sequence homology into seven subfamilies. Some members of these subfamilies play an important role in detecting temperature changes. Within TRPV (vanilloid) subfamily, TRPV1 is the most studied member, and has been related with chronic pain, furthermore its pharmacological blockade and genetic deletion experiments have validated TRPV1 as a therapeutic target. Another member of the TRP family is TRPA1, which is activated by noxious cold and chemical compounds including allyl isothiocyanate (AITC), the pungent principle of wasabi and other mustard oils. TRPA1 appears to have a central role in the pain response but also it has been demonstrated that is essential for asthma [1]. TRP melastatin 8 (TRPM8) is activated by chemical cooling agents (such as menthol) or by temperatures between 28-15 ºC, mediating the detection of innocuous cold thermal stimuli. TRPM8 expression up-regulates has been suggested to play an important role in carcinogenesis and related with prostate cancer [2]. In this study was evaluated the biological activity of a new chemical library, through high throughput screening. We report here the identification of compounds presented a high blockade activity on TRPM8 and share common structure. These hits with notorious antagonistic effect were selected and observed in patch-clamp experiments performed in stable cell lines that expressed TRPV1, TRPM8 and TRPA1 to characterize more accurately their properties. These new pharmacophoric scaffolds can be used as a hit to develop new compounds with better modulator properties interesting to the clinical field or as a research tool.Supported by Consolider-Ingenio 2010 (CSD2008-00005), MICINN (BFU2009- 08346), MICCIN (BES-2010-037112), La Fundacion La Marato de TV3, PrometeoGVA and SAF2009-09323Peer reviewe

Pandemia de Covid-19 en el siglo XXI

The cause of the COVID-19 pandemic of the 21st century is the new coronavirus 2 of severe acute respiratory syndrome (SARS-Cov-2). This coronavirus spreads from person to person through respiratory droplets and produces symptoms such as fever, cough, dyspnea, fatigue and death in severe patients. SARS-Cov-2 can infect the respiratory, digestive, urinary, and blood systems since the presence of the virus has been observed in urine, faeces, blood, and respiratory tract samples. The elderly and people with underlying diseases are more susceptible to infection, and may develop acute respiratory distress syndrome (ARDS), cytokine storm, and even death. The multiple scientific advances on the new coronavirus are progressing at an unprecedented pace to learn about the nature, treatment and prevention of SARS-Cov-2.El causante de la pandemia de COVID-19 del siglo XXI, es el nuevo coronavirus 2 del síndrome respiratorio agudo severo (SARS-CoV-2). Este coronavirus se propaga por contagio de persona a persona a través de patículas respiratorias y produce síntomas como fiebre, tos, disnea, fatiga y la muerte en pacientes graves. El SARS-CoV-2 puede infectar el sistema respiratorio, digestivo, urinario y el sistema hematológico, ya que se ha observado presencia del virus en orina, heces, sangre y en muestras del tracto respiratorio. Los ancianos y personas con enfermedades subyacentes son más susceptibles a la infección, pudiendo desarrollar el síndrome de dificultad respiratoria aguda (SDRA), tormenta de citoquinas e incluso la muerte. Los múltiples avances científicos sobre el nuevo coronavirus, progresan a un ritmo sin precedentes para conocer la naturaleza, tratamiento y prevención del SARS-CoV-2

Una de las mejores terapias contra el cáncer: la Inmunoterapia

La Inmuno-oncología es una modalidad de tratamiento que incluye inmunoterapias que aprovechan el sistema inmune del paciente para luchar contra el cáncer. En estos momentos hay un mayor conocimiento de las interacciones complejas entre el Sistema Inmune y las células tumorales, donde se han identificado moléculas clave que gobiernan estas interacciones. Los tratamientos tradicionales del cáncer, como la radioterapia, la quimioterapia y cirujía, están diseñados para actuar directamente contra los tumores mediante la inhibición de su crecimiento y en última instancia conducen a su destrucción. Las inmunoterapias conllevan unos mecanismos de acción complementarios que pueden ser combinadas con estos tratamientos tradiccionales. La inmunoterapia es un tratamiento en evolución basado en el papel basado en el papel que tiene el sistema inmune en la erradicación del cáncer. Este artículo presenta una revisión sobre los tratamientos en inmunoterapia contra diversas dianas con gran potencial terapeútico para tratar procesos oncológicos. Las inmunoterapias tienen el potencial de ser un arma poderosa contra el cáncer.Immuno-oncology is a modality of treatment that includes immunotherapies that take advantage of the patient's immune system to fight against cancer. At present there is a greater awareness of the complex interactions between the Immune System and tumor cells, where key molecules have been identified that control these interactions. Traditional cancer treatments, such as radiation therapy, chemotherapy and surgery, are designed to actions against directly tumors by inhibiting their growth and ultimately leading to their destruction. Immunotherapies entail complementary mechanisms of action that can be combined with these traditional treatments. Immunotherapy is an evolving treatment based on the role of the immune system in eradication of the cancer. This article presents a review on the treatments in immunotherapy against diverse targets with great therapeutic potential to treat oncological processes. Immunotherapies have the potential to be a powerful weapon against cancer

TLR4-independent upregulation of activation markers in mouse B lymphocytes infected by HRSV.

Human respiratory syncytial virus (HRSV) is the most common cause of severe respiratory infections in infants and young children, often leading to hospitalization. In addition, HRSV poses a serious health risk in immunocompromised individuals and the elderly. It has been reported that this virus can infect mouse antigen-presenting cells, including B lymphocytes. In these B cells, HRSV infection upregulates the expression of activation markers, including MHC class II and CD86, but not MHC class I molecules. Here, we report that HRSV infection of spleen B lymphocytes downregulated TLR4. Either blocking with anti-TLR4 antibody or genetic deletion, but not functional deficiency of TLR4, moderately reduced the infectivity of HRSV in B lymphocytes. HRSV-infected B lymphocytes with deleted TLR4 upregulated MHC class II and CD86 molecules to the same levels as TLR4(+) wild type B cells. Since the activation of monocytes and macrophages by HRSV was previously reported to depend on TLR4, the current study indicates that these cells and B lymphocytes respond to HRSV infection with different activation pathways.Dr. Mark E. Peeples (Department of Immunology/Microbiology, Rush-Presbyterian-St. Luke's Medical Center, Chicago, Illinois, USA) kindly provided the rgHRSV. Technical assistance of Carmen Mir is gratefully acknowledged. This work was supported by grants from Programa Ramón y Cajal, and Fondo de Investigaciones Sanitarias de la Seguridad Social to D. L.; by grant SAF2006-07805 from Ministerio de Educación y Ciencia to J. A. M.; by grants from Comunidad de Madrid and SAF-2004-00534 from Ministerio de Educación y Ciencia to M. D. V.; and by a joint grant from Instituto de Salud Carlos III to D. L., J. A. M. and M. D. V.S

TAP-independent human histocompatibility complex-Cw1 antigen processing of an HIV envelope protein conserved peptide.

Individuals with nonfunctional transporters associated with antigen processing (TAP) complexes are not particularly susceptible to viral infections or neoplasms. Therefore, their immune system must be reasonably efficient, and the present, though reduced, cytolytic CD8 αβ T subpopulation specific for TAP-independent antigens may be sufficient to establish an immune defense protecting against viral infections in these individuals. The objective of the present study was to identify TAP-independent ligands from HIV gp160 protein. An analysis and comparison of complex human histocompatibility complex (HLA)-bound peptide pools isolated from large quantities of healthy or HIV gp160-expressing human cells was performed using mass spectrometry and bioinformatics tools. A conserved TAP-independent HLA peptide ligand endogenously processed and presented in infected human cells was identified. This ligand originates from the envelope protein bound to the HLA-Cw1 class I molecule with high affinity. It was concluded that HLA class I peptides derived from a large fraction of the N-terminal HIV envelope protein could be presented even in the absence of the TAP complex.The present work was supported by grants to D.L. from the Fondo Investigaciones Sanitarias de la Seguridad Social, and the FIPSE Foundation and to A.A. from the Israel Science Foundation ().S