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  10th International Workshop on Clinical Pharmacology of HIV Therapy
April 15-17, 2009
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Merck Offers Unique Perspective on Second-Generation Integrase Inhibitor
  10th International Workshop on Clinical Pharmacology of HIV Therapy, April 15-17, 2009, Amsterdam
Mark Mascolini
Invoking terms like "functionally irreversible" inhibition and "one-shot kill," Merck's Jay Grobler offered a fresh perspective on why raltegravir controls HIV well and--perhaps more importantly--why the company's second-generation integrase inhibitor may do better [1]. At the same time, clinicians will probably want to grasp the science behind Merck's compelling locutions before they start turning up in the lay press.
Earlier work by Merck suggested that raltegravir efficacy appears to be independent of its minimum concentration, unlike every other marketed antiretroviral [2]. This analysis of 332 people taking raltegravir in a salvage regimen found that 16 had a minimum (12-hour) concentration below 33 nM, the drug's 95% inhibitory concentration. But this sub-33-nM group responded as well to raltegravir salvage as people with higher Cmins.
If Cmin doesn't explain why raltegravir works or fails, what does? To find out, Grobler and coworkers looked for ways to test the hypothesis that integrase inhibitors like raltegravir need to exert their activity during only a fraction of the "integration window" to block replication. To test the hypothesis, they set out to map the integration window in cell culture and to measure raltegravir binding kinetics in cell studies.
To map the integration window, they added raltegravir to infected cells at different intervals in the 24 hours after infection. At 48 hours they counted the number of HIV-infected cells (labeled with a marker) for each different raltegravir addition time. The Merck team complemented this time-of-addition study with a washout study in which raltegravir is flushed from infected cells at specified intervals after infection. When they plotted curves showing the number of infected cells per well, the time-of-addition curve was the reverse image of the washout curve, with an intersection around 8 hours. Reading the inflections of each curve led Merck to conclude that raltegravir's inhibition window starts 4 hours after infection of a cell and lasts to 12 hours after infection.
Grobler and colleagues calculated raltegravir binding kinetics by measuring raltegravir residence time on integrase/DNA complexes with a scintillation proximity assay, then by measuring dissociation rates from those complexes. These experiments indicated that raltegravir has a residence time on the integrase/DNA complex that equals or exceeds the half-life of the preintegration complex that viral DNA forms to integrate with host-cell DNA. To be precise, the dissociation half-life of raltegravir in wild-type (nonmutated) virus was 7.3 hours.
That lengthy residence time could explain why raltegravir efficacy in clinical trials did not appear to depend of Cmin. Grobler suggested that a residence time on the integrase/DNA complex "comparable to or exceeding the half-life of the pre-integration complex in the cell results in [a] functionally irreversible, 'one-shot kill' of integration."
If the infecting virus bears the N155H resistance mutation, the dissociation half-life shrinks 10-fold to 0.7 hour. Merck proposed that this greatly curtailed residence times explains why raltegravir can't control replication of virus bearing N155H and other mutations.
In contrast, the experimental Merck integrase inhibitor MK-2048 has a dissociation half-life of 32 hours on wild-type integrase--more than four times that of raltegravir. That half-life fades almost 10-fold, to 4 hours, in the face of N155H mutant virus. But that 4-hour residence time of MK-2048 on mutant virus begins to approach the 7.3 hours of raltegravir on wild-type virus. Those findings led Grobler to suggest that "longer-binding strand-transfer integrase inhibitors may have greater durability and forgiveness, as well as reduced susceptibility to resistance mutations." He did not say when MK-2048, or related compounds, will be ready for clinical trials.
1. Grobler JA, McKenna PM, Ly S, et al. Functionally irreversible inhibition of integration by slowly dissociating strand transfer inhibitors. 10th International Workshop on Clinical Pharmacology of HIV Therapy. April 15-17, 2009. Amsterdam. Abstract O-10.
2. Wenning L, Nguyen B, Teppler H, et al. Pharmacokinetic/pharmacodynamic analyses for raltegravir in phase II and III studies in treatment experienced HIV-infected patients. 9th International Workshop on Clinical Pharmacology of HIV Therapy. April 7-9, 2008. New Orleans. Abstract O_21.