bx-471 and Multiple-Myeloma

Investigators have suggested that the chemokine receptor CCR1 plays a role in multiple myeloma. Studies using antisense and neutralizing antibodies to CCR1 showed that down-regulation of the receptor altered disease progression in a mouse model. More recently, experiments utilizing scid mice injected with human myeloma cells demonstrated that the CCR1 antagonist BX471 reduced osteolytic lesions, while the CCR1 antagonist MLN-3897 prevented myeloma cell adhesion to osteoclasts. However, information is limited regarding the pharmacology of CCR1 antagonists in myeloma cells.. We compared several well-studied CCR1 antagonists including AZD4818, BX471, CCX354, CP-481715, MLN-3897 and PS899877 for their ability to inhibit binding of [(125)I]-CCL3 in vitro using membranes prepared from RPMI 8226 cells, a human multiple myeloma cell line that endogenously expresses CCR1. In addition, antagonists were assessed for their ability to modulate CCL3-mediated internalization of CCR1 and CCL3-mediated cell migration using RPMI 8226 cells. As many GPCRs signal through β-arrestin-dependent pathways that are separate and distinct from those driven by G-proteins, we also evaluated the compounds for their ability to alter β-arrestin translocation.. There were clear differences between the CCR1 antagonists in their ability to inhibit CCL3 binding to myeloma cells, as well as in their ability to inhibit G-protein-dependent and -independent functional responses.. Our studies demonstrate that tissue phenotype seems to be relevant with regards to CCR1. Moreover, it appears that for CCR1 antagonists, inhibition of β-arrestin translocation is not necessarily linked to chemotaxis or receptor internalization.

Topics: Animals; Arrestins; beta-Arrestins; Cell Line, Tumor; Chemokine CCL3; Chemotaxis; CHO Cells; Cricetulus; HEK293 Cells; Humans; Multiple Myeloma; Phenylurea Compounds; Piperidines; Quinoxalines; Radioligand Assay; Receptors, CCR1; Spiro Compounds

Multiple myeloma (MM) is a plasma cell malignancy, characterized by the localization of the MM cells in the bone marrow (BM), where they proliferate and induce osteolysis. The MM cells first need to home or migrate to the BM to receive necessary survival signals. In this work, we studied the role of CCR1 and CCR5, two known chemokine receptors, in both chemotaxis and osteolysis in the experimental 5TMM mouse model. A CCR1-specific (BX471) and a CCR5-specific (TAK779) antagonist were used to identify the function of both receptors. We could detect by RT-PCR and flow cytometric analyses the expression of both CCR1 and CCR5 on the cells and their major ligand, macrophage inflammatory protein 1alpha (MIP1alpha) could be detected by ELISA. In vitro migration assays showed that MIP1alpha induced a 2-fold increase in migration of 5TMM cells, which could only be blocked by TAK779. In vivo homing kinetics showed a 30% inhibition in BM homing when 5TMM cells were pre-treated with TAK779. We found, in vitro, that both inhibitors were able to reduce osteoclastogenesis and osteoclastic resorption. In vivo end-term treatment of 5T2MM mice with BX471 resulted in a reduction of the osteolytic lesions by 40%; while TAK779 treatment led to a 20% decrease in lesions. Furthermore, assessment of the microvessel density demonstrated a role for both receptors in MM induced angiogenesis. These data demonstrate the differential role of CCR1 and CCR5 in MM chemotaxis and MM associated osteolysis and angiogenesis.

Topics: Amides; Animals; Bone Marrow; Bone Resorption; CCR5 Receptor Antagonists; Cell Division; Cell Line, Tumor; Cell Movement; Chemokine CCL3; Chemokine CCL4; Chemokines, CC; Chemotaxis; Female; Macrophage Inflammatory Proteins; Mice; Mice, Inbred C57BL; Multiple Myeloma; Neoplasm Proteins; Neovascularization, Pathologic; Osteoclasts; Osteolysis; Phenylurea Compounds; Piperidines; Quaternary Ammonium Compounds; Receptors, CCR1; Receptors, CCR5; Receptors, Chemokine; Recombinant Fusion Proteins; Stromal Cells; Tumor Burden