RESUMEN
Tumor-targeted immunomodulation using oncolytic viral vectors is currently being investigated as a promising strategy in cancer therapy. In a previous study, we showed that a measles virus Schwarz vaccine strain (MeVac) vector encoding an interleukin-12 fusion protein (FmIL-12) is an effective immunotherapy in the MC38cea murine colon adenocarcinoma model. We hypothesized that MeVac encoding interleukin-15 may mediate enhanced T and NK cell responses and thus increase the therapeutic efficacy, especially in NK cell-controlled tumors. Therefore, we generated MeVac vectors encoding an interleukin-15 superagonist, FmIL-15. Replication and oncolytic capacity, transgene expression, and functionality of MeVac FmIL-15 vectors were validated in vitro. Effects on the tumor immune landscape and therapeutic efficacy of both FmIL-12 and FmIL-15 vectors were studied in the MC38cea and B16hCD46 tumor models. Treatment with MeVac FmIL-15 increased T and NK cell infiltration in both models. However, MeVac FmIL-12 showed more robust viral gene expression and immune activation, resulting in superior anti-tumor efficacy. Based on these results, MeVac encoding a human IL-12 fusion protein was developed for future clinical translation.
Asunto(s)
Regulación Viral de la Expresión Génica , Interleucina-12/agonistas , Interleucina-15/agonistas , Vacuna Antisarampión/inmunología , Adenocarcinoma , Animales , Línea Celular Tumoral , Supervivencia Celular , Colon , Modelos Animales de Enfermedad , Femenino , Genes Virales , Inmunoterapia , Interleucina-12/genética , Interleucina-15/genética , Células Asesinas Naturales/inmunología , Sarampión , Ratones , Ratones Endogámicos C57BL , Virus Oncolíticos , Transcriptoma , Vacunas Sintéticas , Proteínas Virales de Fusión/inmunología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The envelope glycoproteins (Env) of HIV-1 mediate cell entry through fusion of the viral envelope with a target cell membrane. Intramembrane mobility and clustering of Env trimers at the viral budding site are essential for its function. Previous live-cell and super-resolution microscopy studies were limited by lack of a functional fluorescent Env derivative, requiring antibody labeling for detection. Introduction of a bio-orthogonal amino acid by genetic code expansion, combined with click chemistry, offers novel possibilities for site-specific, minimally invasive labeling. Using this approach, we established efficient incorporation of non-canonical amino acids within HIV-1 Env in mammalian cells. The engineered protein retained plasma membrane localization, glycosylation, virion incorporation, and fusogenic activity, and could be rapidly and specifically labeled with synthetic dyes. This strategy allowed us to revisit Env dynamics and nanoscale distribution at the plasma membrane close to its native state, applying fluorescence recovery after photo bleaching and STED nanoscopy, respectively.
Asunto(s)
Proteína gp120 de Envoltorio del VIH/metabolismo , VIH-1/metabolismo , Microscopía Fluorescente/métodos , Movimiento , Nanotecnología/métodos , Membrana Celular/metabolismo , Supervivencia Celular , Química Clic , Colorantes Fluorescentes/química , Células HEK293 , Proteína gp120 de Envoltorio del VIH/química , Proteína gp120 de Envoltorio del VIH/genética , VIH-1/fisiología , Humanos , Ingeniería de Proteínas , Transporte de ProteínasRESUMEN
Norovirus infects different animals, including humans, mice, dogs, and cats. Here, we show an X-ray crystal structure of a feline GIV.2 norovirus capsid-protruding (P) domain to 2.35Å resolution. The feline GIV.2 P domain was reminiscent of human norovirus P domains, except for a novel P2 subdomain α-helix and an extended P1 subdomain interface loop. These new structural features likely obstructed histo-blood group antigens, which are attachment factors for human norovirus, from binding at the equivalent sites on the feline GIV.2 P domain. Additionally, an ELISA showed that the feline GIV.2 was antigenically distinct from a human GII.10 norovirus.
