14 research outputs found
Biased small-molecule ligands for selective inhibition of HIV-1 cell entry via CCR5
Since the discovery of HIV's use of CCR5 as the primary coreceptor in fusion, the focus on developing smallâmolecule receptor antagonists for inhibition hereof has only resulted in one single drug, Maraviroc. We therefore investigated the possibility of using smallâmolecule CCR5 agonists as HIVâ1 fusion inhibitors. A virusâfree cellâbased fusion reporter assay, based on mixing âeffector cellsâ (expressing HIV Env and luciferase activator) with âtarget cellsâ (expressing CD4, CCR5 wild type or a selection of wellâdescribed mutations, and luciferase reporter), was used as fusion readout. Receptor expression was evaluated by ELISA and fluorescence microscopy. On CCR5 WT, Maraviroc and Aplaviroc inhibited fusion with high potencies (EC (50) values of 91 and 501Â nM, respectively), whereas removal of key residues for both antagonists (Glu283Ala) or Maraviroc alone (Tyr251Ala) prevented fusion inhibition, establishing this assay as suitable for screening of HIV entry inhibitors. Both ligands inhibited HIV fusion on signalingâdeficient CCR5 mutations (Tyr244Ala and Trp248Ala). Moreover, the steric hindrance CCR5 mutation (Gly286Phe) impaired fusion, presumably by a direct hindrance of gp120 interaction. Finally, the efficacy switch mutation (Leu203Phe) â converting smallâmolecule antagonists/inverse agonists to full agonists biased toward Gâprotein activation â uncovered that also smallâmolecule agonists can function as direct HIVâ1 cell entry inhibitors. Importantly, no agonistâinduced receptor internalization was observed for this mutation. Our studies of the pharmacodynamic requirements for HIVâ1 fusion inhibitors highlight the possibility of future development of biased ligands with selective targeting of the HIVâCCR5 interaction without interfering with the normal functionality of CCR5