The Journal of Experimental Medicine
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© The Rockefeller University Press, 0022-1007/1997/10/1373/ $5.00
The Journal of Experimental Medicine, Volume 186, Number 8, October 20, 1997 1373-1381

Articles

Interaction of Chemokine Receptor CCR5 with its Ligands: Multiple Domains for HIV-1 gp120 Binding and a Single Domain for Chemokine Binding

Lijun Wu*, Greg LaRosa*, Nasim Kassam*, Cynthia J. Gordon{ddagger}, Heidi Heath*, Nancy Ruffing*, Howard Chen*, Jason Humblias*, Michel Samson§, Marc Parmentier§, John P. Moore{ddagger}, and Charles R. Mackay*

From * LeukoSite, Inc., Cambridge, Massachusetts 02142; the {ddagger} Aaron Diamond AIDS Research Center, The Rockefeller University, New York 10016; and § IRIBHN, Universite Libre de Bruxelles, Campus Erasme, B-1070 Bruxelles, Belgium

CCR5 is a chemokine receptor expressed by T cells and macrophages, which also functions as the principal coreceptor for macrophage (M)-tropic strains of HIV-1. To understand the molecular basis of the binding of chemokines and HIV-1 to CCR5, we developed a number of mAbs that inhibit the various interactions of CCR5, and mapped the binding sites of these mAbs using a panel of CCR5/CCR2b chimeras. One mAb termed 2D7 completely blocked the binding and chemotaxis of the three natural chemokine ligands of CCR5, RANTES (regulated on activation normal T cell expressed and secreted), macrophage inflammatory protein (MIP)-1{alpha}, and MIP-1β, to CCR5 transfectants. This mAb was a genuine antagonist of CCR5, since it failed to stimulate an increase in intracellular calcium concentration in the CCR5 transfectants, but blocked calcium responses elicited by RANTES, MIP-1{alpha}, or MIP-1β. This mAb inhibited most of the RANTES and MIP-1{alpha} chemotactic responses of activated T cells, but not of monocytes, suggesting differential usage of chemokine receptors by these two cell types. The 2D7 binding site mapped to the second extracellular loop of CCR5, whereas a group of mAbs that failed to block chemokine binding all mapped to the NH2-terminal region of CCR5. Efficient inhibition of an M-tropic HIV-1–derived envelope glycoprotein gp120 binding to CCR5 could be achieved with mAbs recognizing either the second extracellular loop or the NH2-terminal region, although the former showed superior inhibition. Additionally, 2D7 efficiently blocked the infectivity of several M-tropic and dual-tropic HIV-1 strains in vitro. These results suggest a complicated pattern of HIV-1 gp120 binding to different regions of CCR5, but a relatively simple pattern for chemokine binding. We conclude that the second extracellular loop of CCR5 is an ideal target site for the development of inhibitors of either chemokine or HIV-1 binding to CCR5.

Address correspondence to Dr. Lijun Wu, LeukoSite, Inc., 215 First St., Cambridge, MA 02142. Phone: 617-621-9350, ext. 1380; FAX: 617-621-9349; E-mail: lijun_wu{at}leukosite.com. C.R. Mackay's current address is Millennium BioTherapeutics, Cambridge, MA 02139.

M. Samson and M. Parmentier at the I.R.I.B.H.N. were supported by the Belgian programme on Interdisciplinary Poles of Attraction and the French Agence Nationale de Recherche contre le SIDA. J.P. Moore was supported by National Institutes of Health grant AI-41420 and by the Pediatric AIDS foundation, of which he is an Elizabeth Glaser Scientist.

1 Abbreviations used in this paper: M, macrophage; MCP, monocyte chemotactic protein; MIP, macrophage inflammatory protein; RANTES, regulated on activation normal T cell expressed and secreted.


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