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Identifying bias in CCR 1 antagonists using radiolabelled binding, receptor internalization, β‐arrestin translocation and chemotaxis assays
Author(s) -
Gilchrist A,
Gauntner T D,
Fazzini A,
Alley K M,
Pyen D S,
Ahn J,
Ha S J,
Willett A,
Sansom S E,
Yarfi J L,
Bachovchin K A,
Mazzoni M R,
Merritt J R
Publication year - 2014
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/bph.12835
Subject(s) - internalization , ccr1 , chemotaxis , chemokine receptor , microbiology and biotechnology , receptor , biology , antagonist , arrestin , chemokine , cancer research , g protein coupled receptor , signal transduction , biochemistry
Background and Purpose Investigators have suggested that the chemokine receptor CCR 1 plays a role in multiple myeloma. Studies using antisense and neutralizing antibodies to CCR 1 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 CCR 1 antagonist BX 471 reduced osteolytic lesions, while the CCR 1 antagonist MLN‐ 3897 prevented myeloma cell adhesion to osteoclasts. However, information is limited regarding the pharmacology of CCR 1 antagonists in myeloma cells. Experimental Approach We compared several well‐studied CCR 1 antagonists including AZD 4818, BX 471, CCX 354, CP ‐481715, MLN ‐3897 and PS899877 for their ability to inhibit binding of [ 125 I]‐ CCL 3 in vitro using membranes prepared from RPMI 8226 cells, a human multiple myeloma cell line that endogenously expresses CCR 1. In addition, antagonists were assessed for their ability to modulate CCL 3‐mediated internalization of CCR 1 and CCL 3‐mediated cell migration using RPMI 8226 cells. As many GPCR s 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. Key Results There were clear differences between the CCR 1 antagonists in their ability to inhibit CCL 3 binding to myeloma cells, as well as in their ability to inhibit G –protein‐dependent and ‐independent functional responses. Conclusions and Implications Our studies demonstrate that tissue phenotype seems to be relevant with regards to CCR 1. Moreover, it appears that for CCR 1 antagonists, inhibition of β ‐arrestin translocation is not necessarily linked to chemotaxis or receptor internalization.