General Strategy for Decoration of Enveloped Viruses with Functionally Active Lipid-Modified Cytokines

Abstract

Viral particles preferentially incorporate extra-and intracellular constituents of host cell lipid rafts, a phenomenon central to pseudotyping. Based on this mechanism, we have developed a system for the predictable decoration of enveloped viruses with functionally active cytokines that circumvents the need to modify viral proteins themselves. Human interleukin-2 (hIL-2), hIL-4, human granulocyte-macrophage colony-stimulating factor (hGM-CSF), and murine IL-2 (mIL-2) were used as model cytokines and fused at their C terminus to the glycosylphosphatidylinositol (

Surface Modification of Retroviral Vectors for Gene Therapy

“…for tissue engineering purposes. Proof of principle was shown through differentiation of monocytes to dendritic cells (Kueng et al 2007). Again, the GPI-anchored cytokines used were functional and elicited cellular responses such as differentiation and proliferation with similar efficiency as their soluble counterparts when co-cultured with the appropriate target cells.…”

“…These results suggested for the first time that incorporation of recombinantly expressed GPIanchored proteins into the envelopes of viral vectors is possible and that these modifications can be useful for gene therapy approaches. In two more recent studies, co-transfection approaches successfully produced virus-like particles (VLPs) containing glypiated proteins from mammalian (Kueng et al 2007) or insect cells (Skountzou et al 2007). In both cases, recombinant GPI-anchored different cytokine species were generated i.e.…”

“…In the first study, it was demonstrated that the GPI-anchored cytokines are functional and elicit cellular responses such as differentiation and proliferation with similar efficiency as their soluble counterparts when co-cultured with the appropriate target cells (Kueng et al 2007); and in the second it was described that GPI-anchored cytokines engineered onto VLPs based on simian immunodeficiency virus (SIV) can enhance immunogenicity of the VLPs. In both cases a modulation of the immune responses was achieved by displaying GPI-anchored cytokine (Kueng et al 2007;Skountzou et al 2007). The major advantage of this approach is that stable transfection of RV/LV producer cell lines co-or super-transfected with GPI-anchored proteins can provide a reproducible long-term source of modified viral particles.…”

Human TCR Transgenic Bet v 1-Specific Th1 Cells Suppress the Effector Function of Bet v 1-Specific Th2 Cells

“…Intracellular cytokine production was analyzed after pretreatment for 6 h with GolgiStop (1:1500; Becton Dickinson, Palo Alto, CA) using Fix and Perm (An der Grub) and using Abs specific for IFN-g (25723-PerCP; R&D Systems, Minneapolis, MN), IL-2 (MQ1-17H12-allophycocyanin; eBioscience, San Diego, CA), IL-4 (MP4-25D2-PE; Invitrogen, Camarillo, CA), IL-5 (JES1-39D10-PE; BioLegend, San Diego, CA), IL-10 (JES3-9D7; eBioscience), and IL-13 (JES10-5A2-allophycocyanin; BioLegend). [25][26][27][28][29][30][31][32][33][34] and used 72 h after transfection. For polarization experiments, aAPCs were variably transfected with single-chain IL-12::GPI.…”

“…Monocyte-derived dendritic cells (MDDCs) were differentiated as described previously (27) [25][26][27][28][29][30][31][32][33][34][35][36] , or medium alone (control) for 3 h were generated. Subsequently, Bet v 1 142-153 -specific or Art v 1 [25][26][27][28][29][30][31][32][33][34][35][36] -specific TCRtg Jurkat cells (1 3 10 5 ) were added to 5 3 10 4 of the above-described aAPCs and cocultured for 6 h. PMA (10 27 M) plus PHA (5 mg/ml) served as positive and medium alone as negative control.…”