In vitro transcribed mRNA constitutes a versatile platform to encode antigens and to evoke CD8 T-cell responses. Systemic delivery of mRNA packaged into cationic liposomes (lipoplexes) has proven particularly powerful in achieving effective antitumor immunity in animal models. Yet, T-cell responses to mRNA lipoplexes critically depend on the induction of type I interferons (IFN), potent pro-inflammatory cytokines, which inflict dose-limiting toxicities. Here, we explored an advanced hybrid lipid polymer shell mRNA nanoparticle (lipopolyplex) endowed with a trimannose sugar tree as an alternative delivery vehicle for systemic mRNA vaccination. Like mRNA lipoplexes, mRNA lipopolyplexes were extremely effective in conferring antitumor T-cell immunity upon systemic administration. Conversely to mRNA lipoplexes, mRNA lipopolyplexes did not rely on type I IFN for effective T-cell immunity. This differential mode of action of mRNA lipopolyplexes enabled the incorporation of N1 methyl pseudouridine nucleoside modified mRNA to reduce inflammatory responses without hampering T-cell immunity. This feature was attributed to mRNA lipopolyplexes, as the incorporation of thus modified mRNA into lipoplexes resulted in strongly weakened T-cell immunity. Taken together, we have identified lipopolyplexes containing N1 methyl pseudouridine nucleoside modified mRNA as potent yet low-inflammatory alternatives to the mRNA lipoplexes currently explored in early phase clinical trials.

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Improved understanding of cancer immunology has provided insight into the phenomenon of frequent tumor recurrence after initially successful immunotherapy. A delicate balance exists between the capacity of the immune system to control tumor growth and various resistance mechanisms that arise to avoid or even counteract the host's immune system. These resistance mechanisms include but are not limited to (i) adaptive expression of inhibitory checkpoint molecules in response to the proinflammatory environment and (ii) amplification of cancer stem cells, a small fraction of tumor cells possessing the capacity for self-renewal and mediating treatment resistance and formation of metastases after long periods of clinical remission. Several individual therapeutic agents have so far been developed to revert T-cell exhaustion or disrupt the cross-talk between cancer stem cells and the tumor-promoting microenvironment. Here, we demonstrate that a three-arm combination therapy-consisting of an mRNA-based vaccine to induce antigen-specific T-cell responses, monoclonal antibodies blocking inhibitory checkpoint molecules (PD-1, TIM-3, LAG-3), and antibodies targeting IL-6 and TGF-β-improves the therapeutic outcome in subcutaneous TC-1 tumors and significantly prolongs survival of treated mice. Our findings point to a need for a rational development of multidimensional anticancer therapies, aiming at the induction of tumor-specific immunity and simultaneously targeting multiple resistance mechanisms.

Over the past two decades, several cell types with fibroblast-like morphology, including mesenchymal stem/stromal cells, but also other adult, embryonic and extra-embryonic fibroblast-like cells, have been brought forward in the search for cellular therapies to treat severe brain injuries and/or diseases. Although current views in regenerative medicine are highly focused on the immune modulating and regenerative properties of stromal cell transplantation in vivo, many open questions remain regarding their true mode of action. In this perspective, this study integrates insights gathered over the past 10 years to formulate a unifying model of the cellular events that accompany fibroblast-like cell grafting in the rodent brain. Cellular interactions are discussed step-by-step, starting from the day of implantation up to 10 days after transplantation. During the short period that precedes stable settlement of autologous/syngeneic stromal cell grafts, there is a complex interplay between hypoxia-mediated cell death of grafted cells, neutrophil invasion, microglia and macrophage recruitment, astrocyte activation and neo-angiogenesis within the stromal cell graft site. Consequently, it is speculated that regenerative processes following cell therapeutic intervention in the CNS are not only modulated by soluble factors secreted by grafted stromal cells (bystander hypothesis), but also by in vivo inflammatory processes following stromal cell grafting. Copyright © 2016 John Wiley & Sons, Ltd.

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