Programming alphavirus with IFN-γ overrides the adverse effects of macrophages on oncolytic virotherapy

Virus uptake by tumor-associated macrophages may significantly reduce the availability of oncolytic viruses for cancer cell infection and limit therapeutic efficacy. Through a computational model, we hypothesized that oncolytic viruses encoding a T cell-stimulating signal like IFN-γ can enhance efficacy regardless of macrophages. To test this, we engineered an alphavirus-based replicon expressing IFN-γ and studied its effect in various tumor-immune coculture systems. While alphavirus replicons do not replicate in macrophages, macrophages readily take up the virus, limiting tumor infection in a frequency-dependent but phenotype-independent manner. However, virus uptake activates the pro-inflammatory responses, further enhanced by neighboring cancer cells expressing virus-encoded IFN-γ. Consequently, T-cell activation was ensured even when a fraction of infected tumor cells expressed IFN-γ, regardless of macrophage presence, frequency, or phenotype. These findings suggest a strategy for optimizing oncolytic virotherapy in tumors with high macrophage infiltration by designing viruses that can stimulate T-cell activation, ensuring therapeutic efficacy.