Anti-inflammatory approaches that target the chemokine network.

Chemokine-ligand/receptor axes play pivotal roles in a myriad of inflammatory, allergic and autoimmune diseases, as well as in the promotion of tumor growth and metastasis. Upon insult, tissue resident cells (and cancer cells in general) release a defined set of inflammatory chemokines that are responsible for the recruitment of activated pathological leukocytes. Recruited leukocytes synthesize and release a host of inflammatory mediators such as chemokines, cytokines, reactive oxygen and nitrogen species, and proteinases. These agents are responsible for the maintenance and amplification of inflammatory responses, and are directly responsible for secondary tissue damage, promotion of autoimmunity, fibrosis and tissue remodelling. Many cancers are associated with the expression of chemokine ligands that co-opt leukocytes such as tumor associated macrophages which in turn provide mediators including growth factors, chemokines and proteinases that promote angiogenesis, tumor growth, and cancer metastasis. Here, we discuss the relevant patents, development and the mechanism of action of a range of therapeutic and potential therapeutic agents that specifically target the chemokine ligand and receptor network. The main approaches that will be highlighted here are antagonism, cell depletion and the relatively unexplored fields of anti-sense, gene and stem cell therapies.

Selective chemokine receptor-targeted depletion of pathological cells as a therapeutic strategy for inflammatory, allergic and autoimmune diseases.

Targeting cell surface antigens or receptors with lytic monoclonal antibodies and specific ligand-directed fusion proteins in order to eliminate cancer cells has been in development for at least forty years. More recently, leukocyte populations known to drive a host of allergic, autoimmune and inflammatory diseases have been targeted. For fusion protein constructs, a number of different classes of cellular toxins have been fused to a variety of ligands such as monoclonal antibodies, growth factors and cytokines. Although there has been great clinical success using these biologics, there are some limitations. The target antigens are often expressed on normal cells leading to side effects. More recently, several groups have explored the use of chemokine receptor ligands and antibodies to target leukocytes and cancer cells. There are a number of inducible chemokine receptors that are only up-regulated in inflammation and their expression is relatively restricted to pathological cells. This confers another degree of specificity on biologics that are composed of chemokine receptor targeting agents. This review discusses articles, recent patents and patent applications that explore the selective depletion of pathological cells by targeting chemokine receptors with chemokine ligands, monoclonal antibodies and different bispecific constructs as a therapeutic strategy for allergic, autoimmune and inflammatory diseases.

Clinical grade production and characterization of a fusion protein comprised of the chemokine CCL2-ligand genetically fused to a mutated and truncated form of the Shiga A1 subunit.

First generation chemokine ligand-Shiga A1 (SA1) fusion proteins (leukocyte population modulators, LPMs) were previously only obtained in small quantities due to the ribosomal inactivating protein properties of the SA1 moiety which inhibits protein synthesis in host cells. We therefore employed 4-aminopyrazolo[3,4-d]-pyrimidine, an inhibitor of Shiga A1, to allow the growth of these cells prior to induction and during the expression phase post-induction with IPTG. Scale-up allowed the production of gram quantities of clinical grade material of the lead candidate, OPL-CCL2-LPM. A manufacturing cell bank was established and used to produce OPL-CCL2-LPM in a fed-batch fermentation process. Induction of the expression of OPL-CCL2-LPM led to the production of 22.47 mg/L per OD(600) unit. The LPM was purified from inclusion bodies using solubilization, renaturation, refolding and chromatography steps. The identity and purity of the OPL-CCL2-LPM was determined using several analytical techniques. The product retained the ability of the SA1 moiety to inhibit protein synthesis as measured in a rabbit reticulocyte lysate cell-free protein synthesis assay and was cytotoxic to target cells. Binding studies established that the protein exerts its effects via CCR2, the cognate receptor for CCL2. Clinical trials in inflammatory nephropathies are planned.