A Bivalent Ligand Targeting the Putative Mu Opioid Receptor and Chemokine Receptor CCR5 Heterodimers: Binding Affinity versus Functional Activities

Abstract

Opioid substitution and antiretroviral therapies have steadily increased the life spans of AIDS patients with opioid addiction, while the adverse drug-drug interactions and persistence of HIV-associated neurocognitive disorders still require new strategies to target opioid abuse and HIV-1 comorbidities. A bivalent ligand 1 with a 21-atom spacer was thus synthesized and explicitly characterized as a novel pharmacological probe to study the underlying mechanism of opioid-enhanced NeuroAIDS. The steric hindrance generated from the spacer affected the binding affinity and Ca²⁺ flux inhibition function activity of bivalent ligand 1 at the chemokine receptor CCR5 more profoundly than it did at the mu opioid receptor (MOR). However, the CCR5 radioligand binding affinity and the Ca²⁺ flux inhibition function of the ligand seemed not necessarily to correlate with its antiviral activity given that it was at least two times more potent than maraviroc alone in reducing Tat expression upon HIV-1 infection in human astrocytes. Furthermore, the ligand was also about two times more potent than the simple mixture of maraviroc and naltrexone in the same viral entry inhibition assay. Therefore bivalent ligand 1 seemed to function more effectively by targeting specifically the putative MOR-CCR5 heterodimer in the viral invasion process. The results reported here suggest that a properly designed bivalent ligand may serve as a useful chemical probe to study the potential MOR-CCR5 interaction during the progression of NeuroAIDS.

Keywords: CCR5; MOR; NeuroAIDS; bivalent ligand; functional selectivity.

Figure 1

Chemical structures of bivalent ligand (1), monovalent ligands (2, 3, 6), and maraviroc analogues, 4 and 5.

Figure 2

HIV-1_SF162 infectivity in human astrocytes (HA) was determined based on the relative amount of Tat protein expressed by the virus using a luciferase based assay. Dose response effect was first studied for each compound (See Supporting Information). Values are absorbance ± SEM (x-axis) of three independent experiments at 18 h post-infection [Maraviroc (MVC), 100 nM; naltrexone (NTX), 1.5 μM; bivalent ligand 1, 100 nM; *p < 0.005 vs. uninfected cells (control); ^$p < 0.05 vs. R5 HIV-1; ^§p < 0.05 vs. R5 + MVC; ^¶p < 0.05 vs. R5 + NTX].

Figure 3

Schematic illustration of the proposed mechanism for the “functional selectivity” of bivalent ligand 1 and maraviroc in the radioligand binding assay, Ca²⁺ flux inhibition assay, and HIV-1 invasion assay. The CCR5 receptor may exist as monomer or form homodimers in the monocloned receptor expressed assays while the CCR5 and the MOR may exist as heterodimers in the human astrocytes and the CCR5 binding pocket in the heterodimer might accommodate the bivalent ligand preferably.