Chemokine signaling plays a critical role in the pathogenesis of asthma through the activation of eosinophil, Th2 lymphocyte, basophil, and neutrophil migration and recruitment into the lung. Recent studies indicate that targeting eosinophils in patients with properly diagnosed eosinophilic asthma can provide a significant health benefits. However, patients with the non-eosinophilic endotype of asthma exhibit more prevalent neutrophilic inflammation, but there are currently no successful treatments for this endotype. There is a growing realization that to efficiently interfere with the chemotaxis of inflammatory cells in asthma, more than one chemokine receptor needs to be inhibited. Currently, there are no drugs capable of multi-faceted interference with chemoattraction of eosinophils and neutrophils. The CCR3 chemokine receptor binds its preferred chemokines, the eotaxins, and facilitates recruitment of eosinophils to the lungs. In addition to eotaxins, CCR3 also binds chemokines responsible for the chemotaxis of Th2 lymphocytes, basophils, mast cells and neutrophils. This binding is mediated by the receptor's extracellular loops. We propose to design novel peptide- based nanoparticles with CCR3 extracellular loop presentation to act as decoy CCR3 receptors or to directly inhibit receptor function. These nanoparticles would sequester chemokines that target both eosinophils and potentially other inflammatory cells such as neutrophils. Unlike commonly available nanoscale passive carriers used for drug delivery, our novel CCR3 nanoparticles will act as drugs themselves. We used this approach to develop peptide-based nanoparticles that antagonize CXCR4, a chemokine receptor similar to CCR3. X4-2-6, a transmembrane helical peptide analog that includes the extracellular loop 1, targets CXCR4 receptors for inhibition of HIV entry and breast cancer metastasis. Based on the ability of x4-2-6 to assemble into nanoparticles, and to sequester SDF-1, we similarly propose to develop peptides containing transmembrane and extracellular loop portions of CCR3. These peptide sequences consist of hydrophobic amino acids with a high helical propensity and include N- and C-terminal charges. These peptide properties are responsible for the peptide's assembly into spherical nanoparticles in aqueous solution. We hypothesize that peptide analogs of CCR3 can be designed that will display properties similar to the CXCR4 peptide x4-2-6, namely, they will be able to form nanoparticles and sequester chemokines (i.e. eotaxins) that are involved in eosinophil and possibly neutrophil migration/recruitment into tissues or directly inhibit CCR3 function. To address this hypothesis, two specific aims are proposed: (1) Determine the ability of CCR3 transmembrane peptides to assemble into nanoparticles and to sequester chemokines involved in eosinophil and neutrophil recruitment;and (2) Demonstrate the ability of CCR3 peptide nanoparticles to inhibit chemokine-dependent inflammatory responses in vitro and in vivo in the mouse triple antigen allergic asthma model. The proposed project is highly likely to significantly impact the general approach to the development of novel asthma therapeutics.
Asthma continues to increase in incidence and severity in the United States and globally, and new treatments are sorely needed. White blood cells (eosinophils and neutrophils) are attracted into the lung in asthma where they cause significant inflammation that includes tissue damage, remodeling, airway contraction and overproduction of mucus. Proteins called chemokines that are responsible for attracting these inflammatory cells into the lung in response to allergic reactions, work through specific chemokine receptors on the inflammatory cells. The overall goal of this project is to develop novel small protein (peptide) based nanoparticle inhibitors of these chemokine receptors in order to block allergen-induced influx of inflammatory cells into the lung in patients with asthma. This project may result in the development of novel nanotechnology- based treatments that could significantly improve the health and quality of life of asthmatics by blocking this inflammation and returning their lung function to normal;it is therefore innovative and of high biomedical significance and potential impact in this disease.
|Chavan, Tanmay S; Meyer, James O; Chisholm, Luke et al. (2014) A novel method for the production of fully modified K-Ras 4B. Methods Mol Biol 1120:19-32|