HIV protease inhibitors (PIs) have been associated with acute induction of peripheral insulin resistance and impaired insulin secretion.1-3 We previously reported that 4 weeks of indinavir treatment in healthy men caused a blunting of the ability of insulin to suppress endogenous glucose production (EGP) during a euglycemic hyperinsulinemic clamp, in the absence of significant changes in body composition or lipid profiles.4 It is unknown whether other PIs also alter EGP in the hyperinsulinemic state. Therefore, we determined the rates of EGP during a euglycemic hyperinsulinemic clamp using stable isotopic tracer methods in 3 separate studies of identical design with indinavir, full-dose ritonavir, and amprenavir administration to healthy normal volunteers. Previously, we found that a single dose of indinavir or ritonavir, but not amprenavir, acutely block insulin-mediated glucose disposal, which was consistent with an acute blockade of the glucose transporter GLUT4 found in vitro.1,3 Here, we present results on the ability of PIs to acutely alter EGP in the hyperinsulinemic state.
We report data from 3 separate studies of similar protocol design.1,3 The studies were approved by the Committee on Human research at the University of California, San Francisco. Indinavir, full-dose ritonavir, and amprenavir were each studied in randomized, double-blind, placebo-controlled, crossover trials, as described previously.1,3 The timing of drug administration was designed to take into account the pharmacokinetics of each PI to achieve therapeutic plasma levels during the hyperinsulinemic clamp study.
In the indinavir study, 6 subjects randomly received a single dose of indinavir (Merck and Co, Rahway, NJ) 1200 mg or placebo at time 0 at the start of the clamp study.1 In the ritonavir study, 9 subjects randomly received ritonavir (Abbott Laboratories Abbott Park, IL) 800 mg soft gel caps or placebo, 2 hours before the start of the clamp study.3 In the amprenavir study, amprenavir (GlaxoSmithKline, Research Triangle Park, NC) 1200 mg soft gel caps or placebo was given to 6 subjects, 1 hour before the start of the clamp study.3 Indinavir levels averaged 6-9 μmol/L, ritonavir 6-10 μmol/L, and ampremnavir 2-6 μmol/L throughout the clamp.1,3 A 3-hour euglycemic hyperinsulinemic clamp was performed in each case. The subjects underwent the alternative treatment (drug or placebo) within 7-28 days, and the above studies were repeated. As reported previously, 1 subject in the ritonavir study failed to achieve therapeutic drug levels at the beginning of the clamp and was excluded from data analysis.
During the hyperinsulinemic clamp, we used the labeled exogenous glucose infusate technique.4 In the indinavir study, a continuous infusion of [6,6-2H2] glucose (3.6 mg kg−1·h−1) was begun before the start of the clamp. In the ritonavir and amprenavir studies, a continuous infusion of [13C6] glucose (1.2 mg kg−1·h−1) was used. Stable isotopes were purchased from Cambridge Isotope Laboratories (Andover, MA) and Isotec (Miamisburg, OH). Steady state blood sampling was performed every 10 minutes during the final 30 minutes of the study. Rate of appearance of glucose was determined using gas chromatography/mass spectrometry by traditional dilution techniques.4
Paired t tests were used for normally distributed data. Data are presented as mean ± standard error of the mean.
As reported previously,1,3 during these studies, insulin-mediated glucose disposal decreased with indinavir and ritonavir by 34% and 16% respectively, compared with placebo (P < 0.001 and P = 0.008, respectively). There was no statistically significant change in insulin-mediated glucose disposal with amprenavir.3
EGP Measurements During the Euglycemic Hyperinsulinemic Clamp
With indinavir, EGP was higher in the hyperinsulinemic state than with placebo (placebo 2.2 ± 0.8, indinavir 4.1 ± 1.3 μg·kg−1·min−1, P = 0.04, Fig. 1A). With ritonavir, there was a trend toward higher EGP during insulin infusion (placebo 3.0 ± 0.5, ritonavir 3.6 ± 0.3 μg·kg−1·min−1, P = 0.08, Fig. 1B). There was no significant difference in EGP during hyperinsulinemia with amprenavir compared with placebo (Fig. 1C). There was no significant difference in EGP with placebo administration between the indinavir, ritonavir, and amprenavir studies. When the effects of indinavir and ritonavir on insulin suppression of EGP during the clamp are taken into account, they are not of sufficient magnitude to eliminate the previously reported effects of either PI drug on the inhibition of insulin-mediated glucose disposal.1,3
In 3 separate studies, we found that indinavir, ritonavir, and amprenavir had different effects on EGP in the hyperinsulinemic state. Indinavir had the greatest effect on increasing EGP during the euglycemic hyperinsulinemic clamp. The effect of indinavir was approximately half the difference in EGP between type 2 diabetes and controls at this insulin concentration.5,6 Ritonavir, given at the full therapeutic dose rather than boosting dose, had a more modest effect on EGP, whereas amprenavir had no significant effect. These studies were performed in subjects of very similar age, ethnicity, and family history of diabetes.1,3 All subjects in the amprenavir study had participated in the ritonavir study, and 1 subject participated in all 3 studies. Although a direct comparison is not possible, these 3 studies of identical study design in similar populations of healthy normal men suggest these PIs have different effects on hepatic insulin sensitivity.
Both insulin-mediated glucose disposal and insulin inhibition of EGP play a role in determining the levels of glucose after a meal. EGP is also the major determinant of fasting glucose levels. Due to the single-dose design and the pharmacokinetics of the drugs, we did not measure the effects of PIs on fasting EGP in these acute studies. We have previously shown that 4 weeks of indinavir leads to increased fasting glucose and EGP in the fasting and hyperinsulinemic state.4 In this study, EGP in the hyperinsulinemic state was 86% higher on indinavir, whereas it was 46% higher after 4 weeks on indinavir.4 We have previously found that the effects of indinavir on insulin-mediated glucose disposal are also higher acutely than after 4 weeks.1,7
Although the mechanism by which indinavir and ritonavir alter EGP is not known, our findings suggest that it is likely to be an acute process. Indinavir and ritonavir increased EGP in the hyperinsulinemic state within several hours of administration. The degree to which the PIs altered EGP paralleled the previously reported effects of these PIs on acute induction of peripheral insulin resistance in humans.1,3 PIs have been shown to decrease insulin-mediated glucose disposal through an acute blockade of GLUT4 in vitro.8 Mice that have a tissue-specific knockout of GLUT4 in muscle and adipose tissue manifest an impaired ability of insulin to inhibit EGP.9 Thus, it is likely that some of the effects of PIs on EGP during insulin infusion or postprandially may be mediated via an acute blockade of GLUT4.
In summary, we found that indinavir and full-dose ritonavir acutely increase EGP in the hyperinsulinemic state, whereas amprenavir had no significant effect on EGP, indicating that the effects of PI are drug-specific, not class-specific. These findings emphasize the acute and heterogeneous nature by which PIs induce alterations in glucose metabolism.
The authors thank Seongsoo Park, PhD, and the GCRC nursing and dietary staff for technical assistance.
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