Small Neutralizing Molecules to Inhibit Actions of the Chemokine CXCL12

Abstract

The chemokine CXCL12 and the receptor CXCR4 play pivotal roles in normal vascular and neuronal development, in inflammatory responses, and in infectious diseases and cancer. For instance, CXCL12 has been shown to mediate human immunodeficiency virus-induced neurotoxicity, proliferative retinopathy and chronic inflammation, whereas its receptor CXCR4 is involved in human immunodeficiency virus infection, cancer metastasis and in the rare disease known as the warts, hypogammaglobulinemia, immunodeficiency, and myelokathexis (WHIM) syndrome. As we screened chemical libraries to find inhibitors of the interaction between CXCL12 and the receptor CXCR4, we identified synthetic compounds from the family of chalcones that reduce binding of CXCL12 to CXCR4, inhibit calcium responses mediated by the receptor, and prevent CXCR4 internalization in response to CXCL12. We found that the chemical compounds display an original mechanism of action as they bind to the chemokine but not to CXCR4. The highest affinity molecule blocked chemotaxis of human peripheral blood lymphocytes ex vivo. It was also active in vivo in a mouse model of allergic eosinophilic airway inflammation in which we detected inhibition of the inflammatory infiltrate. The compound showed selectivity for CXCL12 and not for CCL5 and CXCL8 chemokines and blocked CXCL12 binding to its second receptor, CXCR7. By analogy to the effect of neutralizing antibodies, this molecule behaves as a small organic neutralizing compound that may prove to have valuable pharmacological and therapeutic potential.Chemokines are small (8 -10-kDa) secreted proteins that play roles in the normal physiology of the immune system as well as in orchestrating leukocyte recruitment and activation in the context of inflammatory and infectious diseases (1). Most of them belong to one of two major subfamilies: the ␤ or CC chemokines in which two conserved cysteines from the amino terminus are adjacent to each other and the ␣ or CXC chemokines in which these two cysteines are separated by one residue. Chemokine receptors are members of the superfamily of G proteincoupled receptors characterized by seven transmembranespanning regions and coupling to heterotrimeric G proteins.The CXC chemokine stromal cell-derived factor-1 (SDF1), 5 now named CXCL12, binds to and activates the chemokine receptor CXCR4 as well as the more recently identified CXCR7 receptor (19). CXCL12 stimulates a rapid receptor-mediated intracellular calcium mobilization and signaling through a Pertussis toxin-sensitive G i protein. The response to CXCL12 and expression of the CXCR4 receptor occur at a very early stage of embryonic development and appear to be widely used whenever cell migration is required (2). Indeed mice lacking either CXCL12 or CXCR4 die prenatally and exhibit defects in vascular development, neuronal development, hematopoiesis, and cardiogenesis (3-6).Besides the regulation of homeostatic processes, the CXCR4 receptor is implicated in tumor metastasis (7) as well as in infectious and inflammatory diseases....

Leading and trailing cells cooperate in collective migration of the zebrafish posterior lateral line primordium

“…To test these predictions, we repeated these experiments in the presence of chalcone-4-hydrate, an inhibitor of Cxcl12a activity (Hachet-Haas et al, 2008). We confirmed the efficacy of this inhibitor by first demonstrating that treatment with 10 μM of chalcone-4-hydrate resulted in slowing and eventual stalling of PLLp migration (data not shown).…”

Prodrugs of a CXC Chemokine-12 (CXCL12) Neutraligand Prevent Inflammatory Reactions in an Asthma Model in Vivo

“…1 In screening a proprietary chemical library, 2 we recently identified compounds belonging to the chalcone family and preventing CXCL12 from binding to its CXCR4 or CXCR7 receptors. 3 These compounds have an original mechanism of action: they bind to the chemokine rather than to the receptors. The highest affinity molecule, chalcone 4 3 (compound 1 in Scheme 1, now commercialized by Sigma-Aldrich, C7870), inhibits both chemotaxis of human peripheral blood lymphocytes and eosinophil infiltration in a mouse model of airway hypereosinophilia.…”

“…3−5 Because of its mechanism of action, compound 1 does not affect the resting/basal level of the receptors for either calcium or chemotactic responses. 3 By analogy to the activity of neutralizing antibodies, we named compound 1 a "neutraligand". 4 It is a tool that complements traditional antagonists of the CXCR4 receptor, such as AMD3100, AMD3465, ALX40-4C, and T22/T140, 6−8 but has the disadvantage of poor solubility in aqueous buffer (9 μM).…”

A Dissociated Glucocorticoid Receptor Modulator Reduces Airway Hyperresponsiveness and Inflammation in a Mouse Model of Asthma

“…Determination of total and differential cell counts in bronchoalveolar lavage fluids Bronchoalveolar lavage (BAL; 24 h after the last OVA challenge) was performed as described previously (16,17 …”