Pharmacological characterization of the chemokine receptor, CCR5

Abstract

1 We investigated the e ects of a number of naturally occurring chemokines (MIP-1a, MIP-1b, RANTES, MCP-2, MCP-3, MCP-4) on di erent processes linked to the chemokine receptor CCR5 in recombinant CHO cells expressing the receptor at di erent levels. 2 Internalization of CCR5 following chemokine treatment was studied and MIP-1a, MIP-1b and RANTES (50 nM) were able to induce internalization (*50%) of the receptor. Internalization due to MCP-2, MCP-3 and MCP-4 was less (*20%). 3 Phosphorylation of CCR5 following chemokine treatment was studied and MIP-1a, MIP-1b and RANTES (50 nM) were able to induce phosphorylation of CCR5 whereas the other chemokines did not induce CCR5 phosphorylation. 4 MIP-1a, MIP-1b, RANTES and MCP-2 were able to stimulate [ 35 S]-GTPgS binding, an index of receptor/G protein activation, whereas MCP-3 and MCP-4 had no e ect in this assay. MCP-2 was a partial agonist (*80%) compared to MIP-1a, MIP-1b and RANTES, which gave similar maximal stimulations in this assay. 5 MIP-1a, MIP-1b, RANTES, MCP-2 and MCP-4 were able to stimulate increases in intracellular calcium ions via activation of CCR5 whereas MCP-3 was without e ect. 6 It is concluded that di erent chemokines interacting with CCR5 mediate di erent patterns of cellular responses.

RANTES Modulates the Release of Glutamate in Human Neocortex

“…Several authors reported the ability of RANTES to mobilize Ca 2ϩ ions in cultured neurons (Bolin et al, 1998;Meucci et al, 1998;Boutet et al, 2001;Oh et al, 2001;Gillard et al, 2002;Mueller et al, 2002;Watson et al, 2005;Ignatov et al, 2006), an observation consistent with release of glutamate evoked by RANTES. The facilitation by RANTES of glutamate release may, by one side, explain the pathogenic effects that the chemokine can produce under some pathological conditions associated with neuroinflammation (i.e., HIV-1 infection, multiple sclerosis), including the regulation of T-cell chemotactic migration in CNS.…”

“…Activation of these receptors usually triggers G-protein-coupled PTx-sensitive intracellular pathways, leading to facilitation or inhibition of cellular responses (Zhao et al, 1998;Klein et al, 1999;Boutet et al, 2001;Mueller et al, 2002). Because the effects of hRANTES on [ 3 H]D-ASP release are PTx sensitive, CCR1, CCR3, and CCR5 might have mediated the observed effects on basal and depolarization-evoked release.…”

Lack of Prophylactic Efficacy of Oral Maraviroc in Macaques despite High Drug Concentrations in Rectal Tissues

“…This observation is consistent with clinical studies showing that almost complete CCR5 occupancy is required to significantly reduce plasma viremia (29) and suggests that only a few unoccupied receptors may be sufficient to initiate and sustain a spreading infection. However, as with human CCR5, rhesus CCR5 was not fully internalized by MIP-1␤ in PBMCs or rectal leukocytes in the in vitro assay, thus limiting our analysis to the proportion of CCR5 that was downregulated by MIP-1␤ ex vivo (31). Also, our analysis of MVC binding to rectal CCR5 was performed in SHIV-infected macaques, which might have overestimated occupancy since the number of high CCR5-expressing activated memory CD4 ϩ T cells are expected to be significantly reduced with SHIV infection (39).…”

Constitutive Activation of CCR5 and CCR2 Induced by Conformational Changes in the Conserved TXP Motif in Transmembrane Helix 2

“…[ 35 (28,29). Briefly, 7 g of cell membranes were preincubated with 40 M GDP (Sigma) in the absence (basal binding) or presence of an agonist for 30 min at 30°C before addition of [ Calcium Mobilization-CHO cells were loaded with 2 g/ml fura-2 acetoxymethyl ester (Molecular Probes).…”