Env-glycoprotein heterogeneity as a source of apparent synergy and enhanced cooperativity in inhibition of HIV-1 infection by neutralizing antibodies and entry inhibitors

We measured the inhibition of infectivity of HIV-1 isolates and derivative clones by combinations of neutralizing antibodies (NAbs) and other entry inhibitors in a single-cycle-replication assay. Synergy was analyzed both by the current linear and a new non-linear method. The new method reduced spurious indications of synergy and antagonism. Synergy between NAbs was overall weaker than between other entry inhibitors, and no stronger where one ligand is known to enhance the binding of another. However, synergy was stronger for a genetically heterogeneous HIV-1 R5 isolate than for its derivative clones. Enhanced cooperativity in inhibition by combinations, compared with individual inhibitors, correlated with increased synergy at higher levels of inhibition, while being less variable. Again, cooperativity enhancement was stronger for isolates than clones. We hypothesize that genetic, post-translational or conformational heterogeneity of the Env protein and of other targets for inhibitors can yield apparent synergy and increased cooperativity between inhibitors.