title: Evaluation of the safety and adjuvant effect of a detoxified listeriolysin O mutant on the humoral response to dengue virus antigens.
abstract: Listeriolysin O (LLO) has been proposed as a potential carrier or adjuvant molecule in the vaccination field. However, the cytotoxic and pro-apoptotic effects of LLO are the major limitations for this purpose. Here, we have performed a preclinical safety evaluation and characterized a new potential adjuvant application for a non-cytolytic LLO mutant (dtLLO) to enhance and modulate the immune response against the envelope (E) protein from dengue virus. In addition, we have studied the adjuvant effects of dtLLO on human immune cells and the role of membrane cholesterol for the binding and proinflammatory property of the toxoid. Our in-vivo results in the murine model confirmed that dtLLO is a safer molecule than wild-type LLO (wtLLO), with a significantly increased survival rate for mice challenged with dtLLO compared with mice challenged with wtLLO (P < 0·001). Histopathological analysis showed non-toxic effects in key target organs such as brain, heart, liver, spleen, kidney and lung after challenge with dtLLO. In vitro, dtLLO retained the capacity of binding to plasma membrane cholesterol on the surface of murine and human immune cells. Immunization of 6-8-week-old female BALB/c mice with a combination of dtLLO mixed with E protein elicited a robust specific humoral response with isotype diversification of immunoglobulin (Ig)G antibodies (IgG1 and IgG2a). Finally, we demonstrated that cholesterol and lipid raft integrity are required to induce a proinflammatory response by human cells. Taken together, these findings support a potential use of the dtLLO mutant as a safe and effective adjuvant molecule in vaccination.
authors.affiliation: Instituto de Investigaciones Médico-Biológicas, Universidad Veracruzana, Veracruz City, Veracruz, México.|Doctorado en Ciencias Biomédicas, Centro de Investigaciones Biomédicas, Universidad Veracruzana, Xalapa Veracruz, México.
authors.forename: K G
authors.initials: KG
authors.lastname: Hernández-Flores
authors.affiliation: Instituto de Medicina Forense, Universidad Veracruzana, Boca del Río Veracruz, México.
authors.forename: A L
authors.initials: AL
authors.lastname: Calderón-Garcidueñas
authors.affiliation: Instituto de Investigaciones Médico-Biológicas, Universidad Veracruzana, Veracruz City, Veracruz, México.
authors.forename: G
authors.initials: G
authors.lastname: Mellado-Sánchez
authors.affiliation: Instituto de Medicina Forense, Universidad Veracruzana, Boca del Río Veracruz, México.
authors.forename: R
authors.initials: R
authors.lastname: Ruiz-Ramos
authors.affiliation: Instituto de Investigaciones Médico-Biológicas, Universidad Veracruzana, Veracruz City, Veracruz, México.|Doctorado en Ciencias Biomédicas, Centro de Investigaciones Biomédicas, Universidad Veracruzana, Xalapa Veracruz, México.
authors.forename: L A
authors.initials: LA
authors.lastname: Sánchez-Vargas
authors.affiliation: Instituto de Investigaciones Médico-Biológicas, Universidad Veracruzana, Veracruz City, Veracruz, México.|Doctorado en Ciencias Biomédicas, Centro de Investigaciones Biomédicas, Universidad Veracruzana, Xalapa Veracruz, México.
authors.forename: P
authors.initials: P
authors.lastname: Thomas-Dupont
authors.affiliation: Instituto de Investigaciones Médico-Biológicas, Universidad Veracruzana, Veracruz City, Veracruz, México.|Doctorado en Ciencias Biomédicas, Centro de Investigaciones Biomédicas, Universidad Veracruzana, Xalapa Veracruz, México.
authors.forename: I Y
authors.initials: IY
authors.lastname: Izaguirre-Hernández
authors.affiliation: Departamento de Inmunología, Instituto Nacional de Salud Pública (INSP), Centro de Investigación Sobre Enfermedades Infecciosas (CISEI), Cuernavaca, México.
authors.forename: J
authors.initials: J
authors.lastname: Téllez-Sosa
authors.affiliation: Departamento de Inmunología, Instituto Nacional de Salud Pública (INSP), Centro de Investigación Sobre Enfermedades Infecciosas (CISEI), Cuernavaca, México.
authors.forename: J
authors.initials: J
authors.lastname: Martínez-Barnetche
authors.affiliation: Department of Microbiology, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: L
authors.initials: L
authors.lastname: Wood
authors.affiliation: Department of Microbiology, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Paterson
authors.affiliation: Departamento de Biomedicina Molecular, Centro de Investigación y de Estudios Avanzados IPN, Ciudad de México, México.
authors.forename: L
authors.initials: L
authors.lastname: Cedillo-Barrón
authors.affiliation: Centro de Estudios y Servicios en Salud. Universidad Veracruzana, Veracruz City, Veracruz, México.
authors.forename: O
authors.initials: O
authors.lastname: López-Franco
authors.affiliation: Instituto de Investigaciones Médico-Biológicas, Universidad Veracruzana, Veracruz City, Veracruz, México.
authors.forename: H
authors.initials: H
authors.lastname: Vivanco-Cid
mesh_terms: D000276:Adjuvants, Immunologic; D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology; D000918:Antibody Specificity / Q000276:immunology; D000956:Antigens, Viral / Q000276:immunology*; D001427:Bacterial Toxins / Q000235:genetics / Q000276:immunology*; D002784:Cholesterol / Q000378:metabolism; D016207:Cytokines / Q000378:metabolism; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D003715:Dengue / Q000276:immunology / Q000473:pathology / Q000517:prevention & control; D003716:Dengue Virus / Q000276:immunology*; D004195:Disease Models, Animal; D005260:Female; D006360:Heat-Shock Proteins / Q000235:genetics / Q000276:immunology*; D006460:Hemolysin Proteins / Q000235:genetics / Q000276:immunology*; D006461:Hemolysis / Q000276:immunology; D056724:Immunity, Humoral; D007114:Immunization; D018836:Inflammation Mediators / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000378:metabolism; D008563:Membrane Lipids / Q000378:metabolism; D051379:Mice; D050505:Mutant Proteins / Q000235:genetics / Q000276:immunology*; D011485:Protein Binding / Q000276:immunology
chemical_list: D000276:Adjuvants, Immunologic; D000914:Antibodies, Viral; D000956:Antigens, Viral; D001427:Bacterial Toxins; D016207:Cytokines; D006360:Heat-Shock Proteins; D006460:Hemolysin Proteins; D018836:Inflammation Mediators; D008563:Membrane Lipids; D050505:Mutant Proteins; D002784:Cholesterol; C038705:hlyA protein, Listeria monocytogenes
keywords: adjuvant; cholesterol; dengue; listeriolysin O