Background Vasoactive intestinal peptide (VIP) is an immunosuppressive neuropeptide that significantly affect proliferation and anti-tumor properties of T cells.1-3 VIP overexpression is a potential mechanism of immune escape in solid tumors with paracrine VIP production. Our published work shows that inhibiting VIP receptor (VIP-R) signaling via VIPhyb, an antagonistic fusion peptide between neurotensin and VIP, improves T cell dependent anti-tumor response in mouse models of acute myeloid leukemia (AML) and T lymphoblastic leukemia (TLL).4 In this study, we developed novel VIP-R antagonists with enhanced efficacy when compared to VIPhyb, to generate a significantly more robust anti-tumor response in mouse models of AML.
Methods We created a combinatorial library of 300 peptide sequences that contain the six charged N-terminal residues of the neurotensin present in VIPhyb (first-generation VIP antagonist) with two or more amino acid substitutions within the C-terminal amino acid sequence of VIP (table 1). We performed in-silico screening to identify 10 novel VIP-R antagonists that were predicted to have increased binding affinity to VIP receptors VPAC1 and VPAC2 when compared to VIP or VIPhyb. The efficacy of these peptides where tested in-vitro using T cells from luciferase transgenic mice seeded and expanded on anti-CD3 monoclonal antibody coated plates for three days. Enhanced potency of the novel antagonists in vivo, was tested in a mouse AML model, by treating C1498-bearing mice with subcutaneous administration of VIP, VIPhyb, scrambled peptide or the second-generation VIP-R antagonists (labeled as ‘ANT’) from day 6-12 after tumor implantation.
Results T cell proliferation using 0.3 µM of a novel VIP-R antagonist was increased up to 216% + 20% of control cultures without added peptides versus 197% + 38% in cultures with VIPhyb at 1 uM (table 1). Furthermore, the novel VIP-R antagonists increased median survival times (MST) by up to 57 days and rendered 40% of mice leukemia-free at 60 days compared to MST of 34 days and 5% long-term survival with VIPhyb (figure 1).
Conclusions In this study, for the first time, we have identified novel and more potent VIP-R antagonists when compared to VIPhyb, with enhanced potency to activate and proliferate T cells and generate an effective anti-tumor response in mouse models of leukemia. These novel antagonists can lead to peptide-based immunotherapy for the treatment of various solid and liquid cancers, such as the cancer of the colon and pancreas, that overexpress VIP intratumorally.
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