Human ERAP2 blocking peptide

We have developed an optimized array-based approach for customizable allele-specific gene expression (ASE) analysis. The central features of the approach are the ability to select SNPs at will for detection, and the absence of need to PCR amplify the target. A surprisingly long probe length (39-49 nt) was needed for allelic discrimination. Reconstitution experiments demonstrate linearity of ASE over a broad range. Using this approach, we have discovered at least two novel imprinted genes, NLRP2, which encodes a member of the inflammasome, and OSBPL1A, which encodes a presumed oxysterol-binding protein, were both preferentially expressed from the maternal allele. In contrast, ERAP2, which encodes an aminopeptidase, did not show preferential parent-of-origin expression, but rather, cis-acting nonimprinted differential allelic control. The approach is scalable to the whole genome and can be used for discovery of functional epigenetic modifications in patient samples.

Intracellular aminopeptidases endoplasmic reticulum aminopeptidases 1 and 2 (ERAP1 and ERAP2), and as well as insulin-regulated aminopeptidase (IRAP) process antigenic epitope precursors for loading onto MHC class I molecules and regulate the adaptive immune response. Their activity greatly affects the antigenic peptide repertoire presented to cytotoxic T lymphocytes and as a result can regulate cytotoxic cellular responses contributing to autoimmunity or immune evasion by viruses and cancer cells. Therefore, pharmacological regulation of their activity is a promising avenue for modulating the adaptive immune response with possible applications in controlling autoimmunity, in boosting immune responses to pathogens, and in cancer immunotherapy. In this study we exploited recent structural and biochemical analysis of ERAP1 and ERAP2 to design and develop phosphinic pseudopeptide transition state analogs that can inhibit this family of enzymes with nM affinity. X-ray crystallographic analysis of one such inhibitor in complex with ERAP2 validated our design, revealing a canonical mode of binding in the active site of the enzyme, and highlighted the importance of the S2' pocket for achieving inhibitor potency. Antigen processing and presentation assays in HeLa and murine colon carcinoma (CT26) cells showed that these inhibitors induce increased cell-surface antigen presentation of transfected and endogenous antigens and enhance cytotoxic T-cell responses, indicating that these enzymes primarily destroy epitopes in those systems. This class of inhibitors constitutes a promising tool for controlling the cellular adaptive immune response in humans by modulating the antigen processing and presentation pathway.