Intratumoral delivery of engineered recombinant modified vaccinia virus Ankara expressing Flt3L and OX40L generates potent antitumor immunity through activating the cGAS/STING pathway and depleting tumor-infiltrating regulatory T cells

Abstract

SummaryIntratumoral (IT) delivery of immune-activating viruses can serve as an important strategy to turn “cold” tumors into “hot” tumors, resulting in overcoming resistance to immune checkpoint blockade (ICB). Modified vaccinia virus Ankara (MVA) is a highly attenuated, non-replicative vaccinia virus that has a long history of human use. Here we report that IT recombinant MVA (rMVA), lacking E5R encoding an inhibitor of the DNA sensor cyclic GMP-AMP synthase (cGAS), expressing a dendritic cell growth factor, Fms-like tyrosine kinase 3 ligand (Flt3L), and a T cell co-stimulator, OX40L, generates strong antitumor immunity, which is dependent on CD8+ T cells, the cGAS/STING-mediated cytosolic DNA-sensing pathway, and STAT1/STAT2-mediated type I IFN signaling. Remarkably, IT rMVA depletes OX40hi regulatory T cells via OX40L/OX40 interaction and IFNAR signaling. Taken together, our study provides a proof-of-concept for improving MVA-based cancer immunotherapy, through modulation of both innate and adaptive immunity.One Sentence SummaryIntratumoral delivery of recombinant MVA for cancer immunotherapy

IL-12-expressing highly immunogenic recombinant modified vaccinia virus Ankara reprograms tumor-infiltrating myeloid cells to overcome immune resistance

“…Infection of mouse bone marrow-derived dendritic cells (BMDCs) with MVA lacking the E5R gene (MVAΔE5R) generates high levels of IFN-β production and promotes DC maturation, which is dependent on cGAS 26 . We engineered our first-generation recombinant MVA (rMVA or MQ710), MVAΔE5R-hFlt3L-mOX40L, by inserting an expression cassette containing two transgenes encoding membrane-bound human Flt3L and murine OX40L linked by a P2A self-cleaving sequence into the E5R gene 25 . Flt3L encodes a growth factor for DCs 27 , and OX40L encodes a co-stimulatory ligand for OX40-expressing T cells 28 .…”

“…Flt3L encodes a growth factor for DCs 27 , and OX40L encodes a co-stimulatory ligand for OX40-expressing T cells 28 . Intratumoral (IT) delivery of rMVA (MQ710) generates strong antitumor immunity in multiple tumor models, which is dependent on CD8 + T cells, the cGAS/STING pathway, and IFNAR feedback loop 25 . IT rMVA (MQ710) also leads to depletion of intratumoral OX40 hi regulatory T cells via OX40L/OX40 interaction and IFNAR signaling 25 .…”

“…Intratumoral (IT) delivery of rMVA (MQ710) generates strong antitumor immunity in multiple tumor models, which is dependent on CD8 + T cells, the cGAS/STING pathway, and IFNAR feedback loop 25 . IT rMVA (MQ710) also leads to depletion of intratumoral OX40 hi regulatory T cells via OX40L/OX40 interaction and IFNAR signaling 25 . To further potentiate the antitumor efficacy of the virus, we designed our second-generation recombinant MVA (MQ833) by removing two more viral immune evasion genes, E3L and WR199, and inserting the gene encoding interleukin 12 (IL-12) anchored to extracellular matrix to prevent systemic toxicity.…”

“…Instead, the activation of the innate immune system within the tumor is crucial for antitumor responses. Based on these results, we engineered our first generation non-replicative recombinant MVA (rMVA or MQ710), MVAΔE5R-Flt3L-OX40L, which activates the cGAS/STING pathway and depletes immune-suppressive OX40 hi regulatory T cells via OX40L-OX40 interaction and IFNAR signaling 25 . However, how innate immune sensing and type I IFN pathways contribute to viral-induced antitumor immunity at individual immune cell type is not well understood.…”