Background
Modulation of the immune system by checkpoint inhibitors (ICI) has become an important strategy for the treatment of cancer. However, the majority of patients does not respond to the available therapies, indicating a need for innovative targets and new treatment modalities. The membrane-bound multi domain protein neuropilin-1 (NRP1) is involved in cell migration, cell survival and neoangiogenesis. It furthermore can mediate suppression of anti-tumor immune responses by attracting suppressive myeloid cells into the tumor microenvironment (TME), stabilizing the phenotype of Tregs, and by contributing to the exhaustion of effector T cells. NRP1 is therefore an attractive target for cancer therapies. Considering that these effects are mediated by multiple domains of NRP1, there is a conceptual advantage to down-regulate the expression of the whole protein using antisense oligonucleotides over the functional or steric blockade of individual domains.
Materials and Methods
We used our in-house Oligofyer™ bioinformatics system to design NRP1-specific locked nucleic acid (LNA)-modified antisense oligonucleotides. NRP1-specific ASOs (NRP1 ASO 1 and NRP1 ASO 2) were injected intraperitoneally into mice bearing syngeneic tumors (MC38 and EMT6 model) either as monotherapy or in combination with ICIs. NRP1 expression and downstream effects of NRP1 downregulation were investigated on the mRNA and protein level by hybridization-based RNA expression analysis, flow cytometry and ELISA.
Results
Systemic administration of unformulated NRP1-specific ASOs resulted in potent downregulation of NRP1 expression in different cell types in tumors, including myeloid cells, different T cell subsets and non-immune cells. We furthermore observed strong reduction of soluble NRP1 plasma levels. Downregulation of NRP1 resulted in delayed tumor growth and prolonged survival as a monotherapy. Strikingly, we observed prolonged survival in combination with ICI as compared to ICI monotherapy in the MC-38 and EMT6 model. In the EMT6 breast cancer model, median survival time in the ICI monotherapy group was 30 days, whereas median survival was not reached at day 103 in the groups that had been treated with the combination of NRP1 ASO 1 or 2 and ICI. Furthermore, rechallenge with EMT6 tumor cells on day 103 did not result in growth of tumors in animals that responded to the therapy.
Conclusions
Taken together, these encouraging results show that the ASO-mediated down-regulation of NRP1 expression has the potential to overcome resistance to ICI. We are currently performing additional experiments to fully elucidate the mechanisms that underlie the observed anti-tumor efficacy. Furthermore, preparations for a clinical trial have started.
A. Maaske: A. Employment (full or part-time); Significant; Secarna Pharmaceuticals GmbH & Co. KG. N. Kirchhammer: None. J. Festag: A. Employment (full or part-time); Significant; Secarna Pharmaceuticals GmbH & Co. KG. L. Rodriguez: None. M. Buchi: None. S. Michel: A. Employment (full or part-time); Significant; Secarna Pharmaceuticals GmbH & Co. KG. A. Zippelius: B. Research Grant (principal investigator, collaborator or consultant and pending grants as well as grants already received); Significant; Secarna Pharmaceuticals GmbH & Co. KG. F. Jaschinski: A. Employment (full or part-time); Significant; Secarna Pharmaceuticals GmbH & Co. KG. R. Klar: A. Employment (full or part-time); Significant; Secarna Pharmaceuticals GmbH & Co. KG.