Anti-melanoma vaccines engineered to simultaneously modulate cytokine priming and silence PD-L1 characterized usingex vivomyeloid-derived suppressor cells as a readout of therapeutic efficacy

Abstract

Efficacious antitumor vaccines strongly stimulate cancer-specific effector T cells and counteract the activity of tumor-infiltrating immunosuppressive cells. We hypothesised that combining cytokine expression with silencing programmed cell death ligand 1 (PD-L1) could potentiate anticancer immune responses of lentivector vaccines. Thus, we engineered a collection of lentivectors that simultaneously co-expressed an antigen, a PD-L1-silencing shRNA, and various T cell-polarising cytokines, including interferon γ (IFNγ), transforming growth factor β (TGFβ) or interleukins (IL12, IL15, IL23, IL17A, IL6, IL10, IL4). In a syngeneic B16F0 melanoma model and using tyrosinase related protein 1 (TRP1) as a vaccine antigen, we found that simultaneous delivery of IL12 and a PD-L1-silencing shRNA was the only combination that exhibited therapeutically relevant anti-melanoma activities. Mechanistically, we found that delivery of the PD-L1 silencing construct boosted T cell numbers, inhibited tumor growth and strongly cooperated with IL12 cytokine priming and antitumor activities. Finally, we tested the capacities of our vaccines to counteract tumor-infiltrating myeloid-derived suppressor cell (MDSC) activities . Interestingly, the lentivector co-expressing IL12 and the PD-L1 silencing shRNA was the only one that counteracted MDSC suppressive activities, potentially underlying the observed anti-melanoma therapeutic benefit. We conclude that (1) evaluation of vaccines in healthy mice has no significant predictive value for the selection of anticancer treatments; (2) B16 cells expressing xenoantigens as a tumor model are of limited value; and (3) vaccines which inhibit the suppressive effect of MDSC on T cells in our assay show promising and relevant antitumor activities.

Signal transducer and activator of transcription 3 in myeloid-derived suppressor cells: an opportunity for cancer therapy

“…However instead of combining therapies, it would be more elegant if one drug could lead to activation of tumor-specific CTLs, while modulating MDSCs. Importantly, various studies have demonstrated that MDSCs can be reverted into stimulatory APCs under the influence of cytokines such as IL-12 [174], [175] or TLR ligands like CpG oligonucleotides [176], [177]. The latter has offered an opportunity to design a drug consisting of CpG oligonucleotides conjugated to STAT3 specific small interfering RNA (referred to as CpG-siSTAT3 conjugates) [178–180].…”

“…These range from the use of cell lines to the differentiation of bone marrow cells [77], [80], [100], [101], [181–194]. In particular, ex vivo differentiation of bone marrow cells using conditioned media from GM-CSF secreting tumor cells has proven to be a successful approach [80], [175], [181], [184]. A proof-of-concept on the value of this strategy to obtain large amounts of MDSCs that resemble those found within various cancer types, including multiple myeloma, melanoma and colorectal cancer was delivered [80], [175], [181].…”

“…In particular, ex vivo differentiation of bone marrow cells using conditioned media from GM-CSF secreting tumor cells has proven to be a successful approach [80], [175], [181], [184]. A proof-of-concept on the value of this strategy to obtain large amounts of MDSCs that resemble those found within various cancer types, including multiple myeloma, melanoma and colorectal cancer was delivered [80], [175], [181]. Since 50 to 60 million MDSCs are obtained from a single mouse without the necessity of inducing cancer, this system allows systematic and high throughput in vitro testing of anti-neoplastic treatments.…”

Molecular mechanisms of programmed cell death-1 dependent T cell suppression: relevance for immunotherapy

“…Since PDL1/PD1 blockade therapies are administrated systematically, such disruption must have a range of effects in the immunological synapse in peripheral tissues. When PDL1 is silenced in the peripheral tissues there is in fact an expansion of polyclonal CD8 T cells (43). Moreover, disruption of PDL1-PD1 interactions abrogates TCR down-modulation leading to the expansion of hyperactivated TCR high T cells.…”

“…For this reason, CD8 T cell infiltration can be used as a biomarker for therapeutic efficacy. Actually, tumor infiltration with PD1high and CTLA4high exhausted CD8+ T cells has been proposed as an accurate predictor of responses to anti-PD1 therapy in melanoma (43).…”