Attenuated human parainfluenza virus type 1 (HPIV1) expressing the fusion glycoprotein of human respiratory syncytial virus (RSV) as a bivalent HPIV1/RSV vaccine

Live attenuated recombinant human parainfluenza virus type 1 (rHPIV1) was investigated as a vector to express the respiratory syncytial virus (RSV) fusion (F) glycoprotein, to provide a bivalent vaccine against RSV and HPIV1. The RSV F gene was engineered to include HPIV1 transcription signals and inserted individually into three gene locations in each of the two attenuated rHPIV1 backbones. Each backbone contained a single previously described attenuating mutation that was stabilized against deattenuation, specifically, a non-temperature-sensitive deletion mutation involving 6 nucleotides in the overlapping P/C open reading frames (ORFs) (C^Δ170) or a temperature-sensitive missense mutation in the L ORF (L^Y942A). The insertion sites in the genome were pre-N (F1), N-P (F2), or P-M (F3) and were identical for both backbones. In vitro, the presence of the F insert reduced the rate of virus replication, but the final titers were the same as the final titer of wild-type (wt) HPIV1. High levels of RSV F expression in cultured cells were observed with rHPIV1-C^Δ170-F1, -F2, and -F3 and rHPIV1-L^Y942A-F1. In hamsters, the rHPIV1-C^Δ170-F1, -F2, and -F3 vectors were moderately restricted in the nasal turbinates, highly restricted in lungs, and genetically stable in vivo. Among the C^Δ170 vectors, the F1 virus was the most immunogenic and protective against wt RSV challenge. The rHPIV1-L^Y942A vectors were highly restricted in vivo and were not detectably immunogenic or protective, indicative of overattenuation. The C^Δ170-F1 construct appears to be suitably attenuated and immunogenic for further development as a bivalent intranasal pediatric vaccine.