801. Novel Targeting Non-AAV Parvoviral Vectors for Gene Therapy

Abstract

Top of pageAbstract Parvoviruses infect and are able to kill mid- to late-embryo and cancer cells, but are not pathogenic to fully differentiated, nor to early embryonic cells. While their pathogenicity in vivo is highly species specific, they frequently cross the species barrier when infecting oncogenically transformed cells. Hence, the parvovirus minute virus of mice (MVM) can establish lytic infections in a variety of transformed human cells. The tissue specificity of MVM is largely determined intracellularly by a capsid host-range determinant (hrd), which presumably interacts with an unknown cellular receptor-an interaction that facilitates uncoating of the infecting virus. When forced upon non-permissive cells, MVM was shown to evolve by generating variants with altered host-range properties. We have isolated and sequenced a number of such variants by end-point dilution and plaque purification. The isolates varied from the wild type virus by two types of mutations: a base substitution at nt 1955, in the single Sp1 binding site of the viral P38 promoter (which directs the synthesis of the two capsid proteins), and variable combinations of mutations in the codons for 6 amino acids within a 220 amino acid stretch of the viral capsid. Each of the viruses was examined for its ability to grow and induce cell death in a variety of cell lines. The results demonstrated that MVM isolates with different combinations of surface amino acids possessed characteristic host range properties. The examination of MVM host range mutants in tissue culture cells represents only a fraction of what is likely to occur during normal infection. To overcome this limitation we have begun evaluating the host-range of MVM that bear capsid mutations by injecting mouse embryos in utero. The existence in the embryo of hundreds of different cell types in various differentiation states permits the examination, in parallel, of hundreds of different potential MVM/host-cell combinations. The in vivo analysis shows that the variant viruses exhibit striking differences in tissue specificity and pathogenicity compared to the parental strain. Understanding the recognition code between parvoviral capsid and the host cell components is expected to enable the targeting of this unique oncotropic activity to specific tumor cells.