Identification of amino acids of Sindbis virus E2 protein involved in targeting tumor metastases in vivo.

Previous studies conducted in our laboratory with Sindbis viral vectors in animal models demonstrated excellent in vivo targeting of tumor cells and significant reduction of metastatic implant size. To explore the influence of Sindbis strain on these factors, we constructed new plasmids from the wild-type Ar-339 Sindbis virus strain and compared their sequences. We found differences in the replicase and envelope proteins between JT, HRSP, and Ar-339 sequences. We made chimeras combining both strains and studied their efficiency in SCID mice bearing tumor xenograft using IVIS in vivo imaging techniques. We found that JT envelope proteins targeted tumors more efficiently than those of Ar-339, while the Ar-339 replicase showed increased efficacy in tumor reduction. To determine which residues are responsible for tumor targeting, we made mutants of Ar-339 E2 envelope protein and tested them by IVIS imaging in ES-2 tumor-bearing and tumor-free mice. The change of only one amino acid from E70 to K70 in Ar-339 E2 suppressed the ability to target metastatic tumor implants in mice. A K70 and V251 double E2 mutant did not reverse the loss of targeting capability. Only the mutant with JT E2 and Ar-339 helper targeted tumor, though with less intensity.