RESUMO
A limited number of sites on the HIV-1 Envelope protein are vulnerable to broadly neutralizing antibodies (bnAbs). One of these sites, the membrane proximal external region (MPER), is located at the C-terminus of the gp41 ectodomain (gp41ecto). This highly conserved sequence is bound by several well-characterized bnAbs. Efforts to produce a gp41 immunogen are in part hampered by the MPER's hydrophobicity and propensity to induce aggregation. We sought to produce a DNA vaccine expressing a gp41ecto that is both secreted from mammalian cells and maintains binding by bnAbs to the MPER. Through in silico analysis, we predicted regions of gp41ecto that could induce aggregation and possibly hinder secretion. We generated deletion mutants of gp41ecto and tested their ability to be secreted by mammalian cells. Upon deletion of regions in either the fusion peptide (FP) or MPER, secretion of the gp41ecto increased. In an effort to both augment secretion and maintain binding by bnAbs, we developed constructs with the FP deletion and introduced point mutations in the MPER. Two constructs (gp41 ΔFP and gp41 ΔFP+I682E) maintained binding by gp41 MPER-specific bnAbs (4E10, Z13e1 and 10E8). These were evaluated as DNA vaccines in a mouse model. Both vaccines proved to be immunogenic and appeared to elicit some MPER-specific antibodies that bound gp41 ectodomain-derived proteins but not short peptides spanning the MPER. No neutralizing capacity was detected against a clade C virus containing a homologous MPER.
Assuntos
Vacinas contra a AIDS/imunologia , Proteína gp41 do Envelope de HIV/imunologia , HIV-1/imunologia , Vacinas de DNA/imunologia , Vacinas contra a AIDS/administração & dosagem , Vacinas contra a AIDS/genética , Animais , Feminino , Anticorpos Anti-HIV/sangue , Proteína gp41 do Envelope de HIV/genética , HIV-1/genética , Desenvolvimento Humano , Humanos , Camundongos Endogâmicos BALB C , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologiaRESUMO
In 54/64 subjects with nosocomial diarrhea, fecal calprotectin levels correlated with the results of stool samples tested for Clostridium difficile toxin gene by PCR. Fecal calprotectin levels can be used as an adjunctive measure to PCR to support the diagnosis of C. difficile infection.
