As with HIV, effective pharmacologic control of HCV will likely best be achieved by a cocktail of drugs against independent virus-specific targets. Our results demonstrate that the antiviral effect of the recently discovered NS4B RNA binding inhibitor, clemizole [6
] is highly synergistic with HCV PIs, and additive with interferon, ribavirin or HCV polymerase inhibitors. Importantly, combining clemizole with PIs doesn’t increase host cell toxicity. Moreover, the clemizole-SCH503034 combination decreases emergence of resistance without conferring cross-resistance.
Two major models, Loewe Additivity and Bliss Independence theory are used for analyzing interactions between drugs in combinations. While usually concordant, discordant results may be obtained when analyzing data in these two models [30
]. Both models defined the antiviral effect of the clemizole-SCH503034 combination as synergistic, excluding potential bias and validating the results. Furthermore, the magnitude of the clemizole-SCH503034 combination’s synergy was characterized as strong [17
] further emphasizing its potential relevance in vivo. Clinical trials are needed to determine the maximally tolerated dose of clemizole (based on preclinical animal data, a no observed adverse events level (NOAEL) is estimated to be 100mg/kg/day [32
], and the dose used for antihistamine therapy in humans was typically only ~2mg/kg/day) and pharmacokinetics in HCV patients in order to estimate what are the achievable serum and liver concentrations.
In addition to synergistically increasing the antiviral effect, the SCH503034-clemizole combination decreases the emergence of resistance. This is likely a result of the distinct mechanisms of action of these two drugs and the resulting different resistance profiles. The increased antiviral effect likely contributes as well. Moreover, each of these drugs can select for resistant mutants that have decreased fitness [6
]. Importantly, there was no evidence of cross-resistance between the two drugs. Last, the described synergistic antiviral effects may also permit a reduction in the dose or dosing frequency of individual agents, thereby minimizing potential toxicity and adverse effects. Taken together, these major advantages further emphasize clemizole’s potential as an important component of future combination regimens.
Synergy between antimicrobial drugs often implies a mechanistic interaction between the two targets [33
]. NS4B and NS3 have been shown to bind each other biochemically [35
]. Furthermore, there is genetic evidence for their interaction [36
]. We thus hypothesize that the interaction between these proteins, perhaps involving conformational changes, is the mechanism for the observed synergistic interaction between NS4B RNA binding inhibitors and PIs. Alternatively, it is possible that inhibiting both of these proteins affects two steps in a common pathway that is critical for viral replication thus resulting in synergy by a sequential blocking mechanism [37
In summary, these results suggest that combination of even a moderate NS4B RNA binding inhibitor with PIs represents an attractive paradigm for increasing virologic response rates. Although we hypothesize that more potent inhibitors than clemizole can be obtained, because clemizole has already been extensively used in humans (albeit for a different indication) it may find immediate use as a critical component of next generation anti-HCV strategies.