Functional characterization of a competitive peptide antagonist of p65 in human macrophage-like cells suggests therapeutic potential for chronic inflammation

Abstract

Glucocorticoid-induced leucine zipper (GILZ) is a glucocorticoid responsive protein that links the nuclear factor-kappa B (NFκB) and the glucocorticoid signaling pathways. Functional and binding studies suggest that the proline-rich region at the carboxy terminus of GILZ binds the p65 subunit of NFκB and suppresses the immunoinflammatory response. A widely-used strategy in the discovery of peptide drugs involves exploitation of the complementary surfaces of naturally occurring binding partners. Previously, we observed that a synthetic peptide (GILZ-P) derived from the proline-rich region of GILZ bound activated p65 and ameliorated experimental encephalomyelitis. Here we characterize the secondary structure of GILZ-P by circular dichroic analysis. GILZ-P adopts an extended polyproline type II helical conformation consistent with the structural conformation commonly observed in interfaces of transient intermolecular interactions. To determine the potential application of GILZ-P in humans, we evaluated the toxicity and efficacy of the peptide drug in mature human macrophage-like THP-1 cells. Treatment with GILZ-P at a wide range of concentrations commonly used for peptide drugs was nontoxic as determined by cell viability and apoptosis assays. Functionally, GILZ-P suppressed proliferation and glutamate secretion by activated macrophages by inhibiting nuclear translocation of p65. Collectively, our data suggest that the GILZ-P has therapeutic potential in chronic CNS diseases where persistent inflammation leads to neurodegeneration such as multiple sclerosis and Alzheimer’s disease.

Keywords:

• glucocorticoid-induced leucine zipper
• therapeutic potential
• translational impact
• chronic inflammation

Acknowledgments

We sincerely appreciate the grant supports from the Indiana CTSI to MS and DKL. This work was also partly supported by grants from National Institutes of Health (NIA-R01 and -R21) and Alzheimer’s Association (IIRG) to DKL. We sincerely appreciate assistance from Mr. Nipun Chopra in some of the procedures.

Disclosure

DKL: Scientific advisory board – QR Pharma, Yuma Therapeutics, Entia Biosciences; International advisory board – Drug Discovery and Therapy World Congress; stock options, QR Pharma; patent involving AIT-082, memantine and pending on acamprosate; Editor in Chief, Current Alzheimer Research; research grant support from Baxter Healthcare. MS: cofounder, Provaidya LLC., Indianapolis. CB reports no conflicts of interest in this work.

These disclosures and the funding sources noted in the Acknowledgment section had no role in study design, in the collection, analysis, and interpretation of data, in the writing of the report, or in the decision to submit the article for publication.