Development, optimization, and scale‐up of suspension Vero cell culture process for high titer production of oncolytic herpes simplex virus‐1

Oncolytic viruses (OVs) have emerged as a novel cancer treatment modality, and four OVs have been approved for cancer immunotherapy. However, high-yield and cost-effective production processes remain to be developed for most OVs. Here suspension-adapted Vero cell culture processes were developed for high titer production of an OV model, herpes simplex virus type 1 (HSV-1). Our study showed the HSV-1 productivity was significantly affected by multiplicity of infection, cell density, and nutritional supplies. Cell culture conditions were first optimized in shake flask experiments and then scaled up to 3 L bioreactors for virus production under batch and perfusion modes. A titer of 2.7 × 10⁸ TCID50 mL^-1 was obtained in 3 L batch culture infected at a cell density of 1.4 × 10⁶ cells mL^-1 , and was further improved to 1.1 × 10⁹ TCID50 mL^-1 in perfusion culture infected at 4.6 × 10⁶ cells mL^-1 . These titers are similar to or better than the previously reported best titer of 8.6 × 10⁷ TCID50 mL^-1 and 8.1 × 10⁸ TCID50 mL^-1 respectively obtained in labor-intensive adherent Vero batch and perfusion cultures. HSV-1 production in batch culture was successfully scaled up to 60 L pilot-scale bioreactor to demonstrate the scalability. The work reported here is the first study demonstrating high titer production of HSV-1 in suspension Vero cell culture under different bioreactor operating modes.