Triatoma virus-VLPs: a potential new platform for chimeric vaccine design.

SANCHEZ- EUGENIA, R; ROMANO-MORENO, M; GOIKOLEA-REGIA, J; QUERIDO, JFB; DURANA, A; MARTI, G.A; SOUSA SILVA, M; RODRÍGUEZ, JF; GUERIN, D.M.A.

Whashington

Congreso; Worldvaccine Congress; 2014

Triatoma virus (TrV) belongs to the Dicistroviridae family, a viral family of bicistronic+ssRNA viruses pathogenic to arthropods. The capsid structure of TrV is icosahedralof about 30nm in diameter and is made of three structural proteins (ranging from 28to 37kDa) folded with a standard jelly-roll topology. TrV-VLPs can be obtained eitherby purification of infected insects or by a recombinant method using a baculovirusinsectcell expression system. In this study we analyze some of the physicochemical and immunogenic properties of the VLPs. The results presented here provide a preliminary view of the potency of the TrV-VLPs as epitope carrier for vaccine design. TrV Purification Stress Treatments IgG production Stability at acidic pH Squires et. al., 2013 Top: Continuous 5%-30% sucrose gradient of T.infestans (clarified) insect feces. The first peak corresponds to the elution of RNA-full and infective TrV particles (fractions 6-16), and the second peak (fractions 21-25) contains empty TrV capsids or natural VLPs (Agirre et. al., 2011). Bottom: Negative Stained-TEM images of infective (A) and empty or natural VLPs (B) TrV particles. Magnification 88,000X. TrV-VLPs after one week treatments under normal and stress conditions. A-C. Negative Stained-TEM images (88,000X) of VLPs stored in buffer at 4oC (A), at 37oC (B), and lyophilized and then resuspended (C). These images show that structural integrity is better maintained in the lyophilized sample. D. The SDS-PAGE image (12.5%) shows that no protein degradation occurs after these treatments. VP0 (VP4+VP3; 37.3kDa); VP3 (31.8kDa); VP1 (29.7kDa); VP2 (28.4kDa). Levels of total IgG anti TrV proteins antibodies produced by intraperitoneal inoculation (3μg protein) in Mus musculus (BALB/c, 3 groups of 6 to 8 weeks age). Serum samples were taken 30days after injection. VP1-3, the RNA-dependent RNA polymerase (RdRp), the helicase (Hel), and the protease (Prot) are recombinant proteins produced in E. coli (unpublished results). Ctrl corresponds to IgGs' level before inoculation. Values in arbitrary units. Stability of the capsid structure under different pH conditions is a key factor. Human rhinovirus serotype 2 (HRV2) is known to be labile under acidic pH (A), but TrV capsid (B) is stable along a very wide acidic pH range. This TrV stability was observed by intrinsic fluorescence emission experiments (C) and by Static Light Scattering measurements (D) (Agirre et al., 2011).