Galectin-1 fosters an immunosuppressive microenvironment in colorectal cancer by reprogramming CD8⁺ regulatory T cells

Colorectal cancer (CRC) represents the third most common malignancy and the second leading cause of cancer-related deaths worldwide. Although immunotherapy has taken center stage in mainstream oncology, it has shown limited clinical efficacy in CRC, generating an urgent need for discovery of new biomarkers and potential therapeutic targets. Galectin-1 (Gal-1), an endogenous glycan-binding protein, induces tolerogenic programs and contributes to tumor cell evasion of immune responses. Here, we investigated the relevance of Gal-1 in CRC and explored its modulatory activity within the CD8⁺ regulatory T cell (Treg) compartment. Mice lacking Gal-1 (Lgals1 -/- ) developed a lower number of tumors and showed a decreased frequency of a particular population of CD8⁺CD122⁺PD-1⁺ Tregs in the azoxymethane-dextran sodium sulfate model of colitis-associated CRC. Moreover, silencing of tumor-derived Gal-1 in the syngeneic CT26 CRC model resulted in reduced number and attenuated immunosuppressive capacity of CD8⁺CD122⁺PD-1⁺ Tregs, leading to slower tumor growth. Moreover, stromal Gal-1 also influenced the fitness of CD8⁺ Tregs, highlighting the contribution of both tumor and stromal-derived Gal-1 to this immunoregulatory effect. Finally, bioinformatic analysis of a colorectal adenocarcinoma from The Cancer Genome Atlas dataset revealed a particular signature characterized by high CD8⁺ Treg score and elevated Gal-1 expression, which delineates poor prognosis in human CRC. Our findings identify CD8⁺CD122⁺PD-1⁺ Tregs as a target of the immunoregulatory activity of Gal-1, suggesting a potential immunotherapeutic strategy for the treatment of CRC.Centro de Investigaciones Inmunológicas Básicas y Aplicada

Circulating galectin-1 delineates response to bevacizumab in melanoma patients and reprograms endothelial cell biology

Blockade of vascular endothelial growth factor (VEGF) signaling with bevacizumab, a humanized anti-VEGF monoclonal antibody (mAb), or with receptor tyrosine kinase inhibitors, has improved progression-free survival and, in some indications, overall survival across several types of cancers by interrupting tumor angiogenesis. However, the clinical benefit conferred by these therapies is variable, and tumors from treated patients eventually reinitiate growth. Previously we demonstrated, in mouse tumor models, that galectin-1 (Gal1), an endogenous glycan-binding protein, preserves angiogenesis in anti-VEGF–resistant tumors by co-opting the VEGF receptor (VEGFR)2 signaling pathway in the absence of VEGF. However, the relevance of these findings in clinical settings is uncertain. Here, we explored, in a cohort of melanoma patients from AVAST-M, a multicenter, open-label, randomized controlled phase 3 trial of adjuvant bevacizumab versus standard surveillance, the role of circulating plasma Gal1 as part of a compensatory mechanism that orchestrates endothelial cell programs in bevacizumab-treated melanoma patients. We found that increasing Gal1 levels over time in patients in the bevacizumab arm, but not in the observation arm, significantly increased their risks of recurrence and death. Remarkably, plasma Gal1 was functionally active as it was able to reprogram endothelial cell biology, promoting migration, tubulogenesis, and VEGFR2 phosphorylation. These effects were prevented by blockade of Gal1 using a newly developed fully human anti-Gal1 neutralizing mAb. Thus, using samples from a large-scale clinical trial from stage II and III melanoma patients, we validated the clinical relevance of Gal1 as a potential mechanism of resistance to bevacizumab treatment

Multivalent ligands bearing thiosugars. Synthesis and biological activity

Las interacciones específicas entre hidratos de carbono y proteínas regulan una gran diversidad de fenómenos biológicos, puesto que desencadenan numerosos procesos de transferencia de información. Para lograr interacciones de alta afinidad, la Naturaleza ha diseñado biomoléculas que presentan múltiples copias de un determinado residuo oligosacarídico, que actúa como elemento de reconocimiento de una proteína. Esto conlleva al efecto cluster glicosídico o efecto multivalente, en el cual la afinidad del proceso global resulta mucho mayor que la simple suma de las interacciones individuales. Asimismo, estas interacciones entre ligandos multivalentes y sus proteínas receptoras (en la mayoría de los casos, lectinas) conducen a redes entrecruzadas que desencadenan procesos biológicos específicos. En búsqueda de compuestos de alta afinidad por lectinas que pudieran interferir con este tipo de procesos, y que además resultaran estables en medios biológicos sin sufrir degradación por glicosidasas endógenas, se planteó como objetivo principal de este trabajo de tesis la síntesis de ligandos multivalentes con unidades de tioazúcares específicamente diseñados para estudiar fenómenos biológicos regulados por lectinas. Estos ligandos se diseñaron para ser resistentes a la degradación enzimática, y a la vez, afines a lectinas que reconocen ß-galactósidos (lectina de maní y galectinas). Para ello, se sintetizaron cuatro elementos de reconocimiento distintos derivados de ß-galactósidos: ß-tiogalactósidos, ß- tiolactósidos, análogos isostéricos de la 3-desoxilactosa y análogos de tiodigalactósido. Su preparación requirió el desarrollo de metodología de síntesis específicas en cada caso. Estos fragmentos se conectaron a plataformas biocompatibles derivadas de hidratos de carbono a través de la reacción click, la cicloadición de azidas y alquinos catalizada por Cu(I) o Ru(II). Se evaluó la afinidad de los ligandos sintetizados por lectinas así como también su actividad enzimática como inhibidores de glicosidasas. Asimismo, se realizaron estudios conformacionales de los tioazúcares y análisis de la interacción enzima-inhibidor mediante técnicas de RMN y de modelado molecular.The specific interactions between carbohydrates and proteins are determinant in a number of biological phenomena, since they trigger a myriad of biological processes related with communication processes. To achieve high affinity interactions, Nature has designed biomolecules presenting several copies of specific oligosaccharidic moieties recognized by a protein. This fact leads to the cluster glycosidic effect, also called multivalent effect, described as a high affinity process which is stronger than the simple sum of monovalent interactions. Moreover, these interactions between multivalent ligands and their protein receptors (mainly lectins), produce complex lattices which trigger specific biological responses. In order to obtain compounds with high affinity for lectins, being at the same time stable in biological fluids and resistant to endogenous glycosylhydrolases, the aim of this work was the synthesis of multivalent ligands bearing thiosugars, specifically selected to study lectinmediated biological events. These ligands have been designed to interact with lectins that recognize ß-galactosides (peanut lectin and galectins), and to be resistant to enzymatic degradation. Thus, four different recognition elements derived from ß-galactosides have been synthesized: ß-thiogalactosides, ß-thiolactosides, isosteric analogues of 3-deoxylactose and thiodigalactoside analogues. Specific methodology has been developed in each case to achieve their preparation. These fragments were connected to biocompatible carbohydrate-derived scaffolds through the “click” reaction, the cycloaddition between azides and alkynes catalyzed by Cu(I) or Ru(II). The affinity of the synthesized ligands for lectins, as well as their activity as glycosidase inhibitors was determined. In addition, conformational studies of the thiosugars and the analysis of the enzyme-inhibitor interactions were performed using NMR techniques and molecular modeling.Fil:Cagnoni, Alejandro J.. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

Design and Synthesis of Hydrolytically Stable Multivalent Ligands Bearing Thiodigalactoside Analogues for Peanut Lectin and Human Galectin‑3 Binding

Herein, we describe the design and synthesis of a novel family of hydrolytically stable glycoclusters bearing thiodigalactoside (TDG) analogues as recognition elements of β-galactoside binding lectins. The TDG analogue was synthesized by thioglycosylation of a 6-<i>S</i>-acetyl-α-d-glucosyl bromide with the isothiouronium salt of 2,3,4,6-tetra-<i>O</i>-acetyl-β-d-galactose. Further propargylation of the TDG analogue allowed the coupling to azido-functionalized oligosaccharide scaffolds through copper­(I)-catalyzed azide–alkyne cycloaddition (CuAAC) under microwave activation. The final mono-, di-, and tetravalent ligands were resistant to enzymatic hydrolisis by Escherichia coli β-galactosidase. Binding affinities to peanut agglutinin and human galectin-3 were measured by isothermal titration calorimetry which showed <i>K</i>_a constants in the micromolar range as well as a multivalent effect. Monovalent ligand exhibited a binding affinity higher than that of thiodigalactoside. Docking studies performed with a model ligand on both β-galactoside binding lectins showed additional interactions between the triazole ring and lectin amino acid residues, suggesting a positive effect of this aromatic residue on the biological activity

Multivalent sialylation of beta-thio-glycoclusters by Trypanosoma cruzi trans sialidase and analysis by high performance anion exchange chromatography

International audienceThe synthesis of multivalent sialylated glycoclusters is herein addressed by a chemoenzymatic approach using the trans-sialidase of Trypanosoma cruzi (TcTS). Multivalent beta-thio-galactopyranosides and beta-thio-lactosides were used as acceptor substrates and 3'-sialyllactose as the sialic acid donor. High performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) was shown to be an excellent technique for the analysis of the reaction products. Different eluting conditions were optimized to allow the simultaneous resolution of the sialylated species, as well as their neutral precursors. The TcTS efficiently transferred sialyl residues to di, tri, tetra and octa beta-thio-galactosides. In the case of an octavalent thiolactoside, up to six polysialylated compounds could be resolved. Preparative sialylation reactions were performed using the tetravalent and octavalent acceptor substrates. The main sialylated derivatives could be unequivocally assigned by MALDI mass spectrometry. Inhibition of the transfer to the natural substrate, N-acetyllactosamine, was also studied. The octalactoside caused 82 % inhibition of sialic acid transfer when we used equimolar concentrations of donor, acceptor and inhibitor

Synthesis of divalent ligands of β-thio- and β-N-galactopyranosides and related lactosides and their evaluation as substrates and inhibitors of Trypanosoma cruzi trans-sialidase

In this work we describe the synthesis of mono- and divalent β-N- and β-S-galactopyranosides and related lactosides built on sugar scaffolds and their evaluation as substrates and inhibitors of the Trypanosoma cruzi trans-sialidase (TcTS). This enzyme catalyzes the transfer of sialic acid from an oligosaccharidic donor in the host, to parasite βGalp terminal units and it has been demonstrated that it plays an important role in the infection. Herein, the enzyme was also tested as a tool for the chemoenzymatic synthesis of sialic acid containing glycoclusters. The transfer reaction of sialic acid was performed using a recombinant TcTS and 3’-sialyllactose as sialic acid donor, in the presence of the acceptor having βGalp non reducing ends. The products were analyzed by high performance anion exchange chromatography with pulse amperometric detection (HPAEC-PAD). The ability of the different S-linked and N-linked glycosides to inhibit the sialic acid transfer reaction from 3’-sialyllactose to the natural substrate N-acetyllactosamine, was also studied. Most of the substrates behaved as good acceptors and moderate competitive inhibitors. A di-N-lactoside showed to be the strongest competitive inhibitor among the compounds tested (70% inhibition at equimolar concentration). The usefulness of the enzymatic trans-sialylation for the preparation of sialylated ligands was assessed by performing a preparative sialylation of a divalent substrate, which afforded the monosialylated compound as main product, together with the disialylated glycocluster

Emergence of PER-2 and VEB-1a in Acinetobacter baumannii strains in the Americas

Fil: Pasteran, Fernando. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Bacteriología. Servicio de Antimicrobianos; Argentina.Fil: Rapoport, Melina. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Bacteriología. Servicio de Antimicrobianos; Argentina.Fil: Petroni, Alejandro. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Bacteriología. Servicio de Antimicrobianos; Argentina.Fil: Faccone, Diego. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Bacteriología. Servicio de Antimicrobianos; Argentina.Fil: Corso, Alejandra. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Bacteriología. Servicio de Antimicrobianos; Argentina.Fil: Galas, Marcelo. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Bacteriología. Servicio de Antimicrobianos; Argentina.Fil: Vázquez, Miryam. Hospital de Niños Ricardo Gutiérrez. Sección Microbiología; Argentina.Fil: Procopio, Adriana. Hospital de Niños Ricardo Gutiérrez. Sección Microbiología; Argentina.Fil: Tokumoto, Marta. Instituto de Cardiología y Cirugía Cardiovascular Fundación Favaloro; Argentina.Fil: Cagnoni, Viviana. Instituto de Cardiología y Cirugía Cardiovascular Fundación Favaloro; Argentina