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Boosting Dose Ritonavir Does Not Alter Peripheral Insulin Sensitivity in Healthy HIV-Seronegative Volunteers
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JAIDS Journal of Acquired Immune Deficiency Syndromes:
1 November 2010
"Here, we show that boosting dose levels of ritonavir do not acutely alter peripheral insulin sensitivity as measured by insulin-mediated glucose disposal during a euglycemic hyperinsulinemic clamp. Given that the effects of PI are greater in short-term administration7,9 than after 4 weeks,18,19 it is unlikely that there will be effects of boosting dose ritonavir with longer exposure. We studied HIV-seronegative subjects to avoid potential confounding effects on glucose metabolism introduced by HIV-related factors.....In summary, we show a single boosting dose of ritonavir does not alter insulin-mediated glucose disposal. Given our previous finding that a single full dose of ritonavir (which yielded significantly higher plasma levels than those achieved here) acutely induced insulin resistance, we also demonstrate a dose-dependent effect of ritonavir on peripheral insulin sensitivity. Further, our data suggest that lopinavir likely makes the major contribution to the induction of insulin resistance that has been observed in short-term studies of lopinavir/ritonavir. Consideration should be given to these findings when assessing the metabolic effects of antiretroviral regimens and advising patients."
Taylor, Steven A MD*; Lee, Grace A MD*; Pao, Vivian Y MD*; Anthonypillai, Jayaranjan MSc; Aweeka, Francesca T PharmD; Schwarz, Jean-Marc PhD*; Mulligan, Kathleen PhD*; Schambelan, Morris MD*; Grunfeld, Carl MD, PhD*
From the *Department of Medicine, University of California, San Francisco, CA; Division of Endocrinology and Metabolism, Department of Veterans Affairs Medical Center, San Francisco, CA; Drug Research Unit, Department of Clinical Pharmacy, San Francisco General Hospital, University of California, San Francisco, CA; Division of Endocrinology, San Francisco General Hospital, San Francisco, CA; and Department of Basic Sciences, College of Osteopathic Medicine, Touro University, Vallejo, CA.
Abstract
Background: Some HIV protease inhibitors (PIs), including full-dose ritonavir (800 mg) and ritonavir-boosted lopinavir, acutely induce insulin resistance in the absence of HIV infection and changes in body composition. Boosting dose ritonavir (100-200 mg) is the most commonly prescribed PI, yet its effects on glucose metabolism have not been described in the absence of another PI.
Methods: In this randomized, double-blind, cross-over study, a single dose of ritonavir 200 mg or placebo was given to healthy HIV-seronegative volunteers before assessment of insulin sensitivity by euglycemic hyperinsulinemic clamp.
Results: Boosting dose ritonavir had no effect on insulin-mediated glucose disposal (M/I, placebo: 8.59 ± 0.83 vs. ritonavir: 8.51 ± 0.64 mg/kg per minute per µU/mL insulin, P = 0.89).
Conclusions: A single boosting dose of ritonavir does not alter insulin sensitivity, suggesting lopinavir is likely responsible for the induction of insulin resistance demonstrated in prior short-term studies of lopinavir/ritonavir. There is a dose-dependent effect of ritonavir on insulin sensitivity.
INTRODUCTION
After the introduction of highly active antiretroviral therapy with HIV protease inhibitors (PIs), an increase in abnormalities of glucose metabolism was reported.1-6 These included induction of insulin resistance, impaired glucose tolerance, and frank diabetes. Studies have since established that individual PIs have varied effects on glucose metabolism, especially the acute induction of insulin resistance.7-10 Previously, we showed that a single dose of ritonavir at levels used to suppress HIV ("full" dose = 800 mg) acutely induces insulin resistance in healthy HIV-seronegative volunteers.8 However, ritonavir is generally prescribed at lower "boosting" doses (100 or 200 mg) in combination with a second PI, to achieve therapeutic levels of the other PI. Boosting dose ritonavir is the most commonly prescribed PI, yet its effects on glucose metabolism have not been described in the absence of another PI. A single standard dose of ritonavir-boosted lopinavir acutely induces insulin resistance in healthy human subjects.9 Short-term administration of ritonavir-boosted lopinavir induces more insulin resistance than ritonavir-boosted atazanavir.11 It is unclear whether the induction of insulin resistance observed in these studies occurs as a direct effect of ritonavir, lopinavir, or both.
Here, utilizing the euglycemic hyperinsulinemic clamp technique, we compare the effects of a single boosting dose of ritonavir to placebo in a cross-over study. We studied healthy normal volunteers to eliminate contributions of HIV infection itself, treatment-induced restoration of health, and other concomitantly prescribed antiretroviral medications that might alter insulin sensitivity.
RESULTS
Eight men and 2 women, ages 38-65 years (54 ± 3 years), completed the study; 6 were white, 2 were Hispanic, 1 African-American, and 1 Asian. No serious adverse clinical events occurred.
Weight, body mass index, resting energy expenditure and fasting serum insulin, plasma glucose, homeostasis model assessment-insulin resistance index, and lipids measured after administration of active drug or placebo but before the clamp did not differ between the study arms (Table 1). Plasma ritonavir levels reached 2.5 ± 0.5 µmol/L at the start of the clamp and the mean level remained >1.2 µmol/L until the end of the study (Fig. 1A). The 3-hour time averaged area under the concentration time curve for ritonavir was 1.66 µmol/L per hour. All study subjects reached ritonavir concentrations above 0.7 µmol/L during the clamp.
Similar steady-state insulin and glucose levels were maintained during the final hour of the clamp (Fig. 1B). Glucose levels were 4.39 ± 0.01 and 4.40 ± 0.02 mmol/L (P = 0.42) for ritonavir and placebo, respectively. Insulin levels were 447 ± 22 and 456 ± 23 pmol/L (P = 0.46) for ritonavir and placebo, respectively.
M/I measured after administration of boosting dose ritonavir and placebo did not differ during the last hour of the clamp study (Fig. 1C). M/I was 8.51 ± 0.64 and 8.59 ± 0.83 mg/kg per minute per µU/mL insulin for boosting dose ritonavir and placebo, respectively (P = 0.89).
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DISCUSSION
Boosting dose ritonavir is the most commonly prescribed PI. Its potent inhibition of the enzyme responsible for the metabolism of most PIs (CYP3A4) enhances the antiretroviral activity of a concomitantly-dosed PI by allowing it to achieve higher plasma concentrations for more sustained periods of time. Despite its prevalent use, the effects of boosting dose ritonavir on glucose metabolism had only been studied concomitantly with a second PI. Here, we show that boosting dose levels of ritonavir do not acutely alter peripheral insulin sensitivity as measured by insulin-mediated glucose disposal during a euglycemic hyperinsulinemic clamp. Given that the effects of PI are greater in short-term administration7,9 than after 4 weeks,18,19 it is unlikely that there will be effects of boosting dose ritonavir with longer exposure. We studied HIV-seronegative subjects to avoid potential confounding effects on glucose metabolism introduced by HIV-related factors.
In the past, ritonavir was prescribed at full doses to suppress HIV. Previously, we showed that a single full dose of ritonavir caused a 15% reduction in insulin-mediated glucose disposal, also measured by the euglycemic hyperinsulinemic clamp.8 Plasma ritonavir concentrations achieved during determination of M/I in that study (12.5 ± 1.5 µmol/L) were substantially higher than those observed here (1.5 ± 0.3 µmol/L). Thus, our studies demonstrate that boosting dose ritonavir has no effect on M/I and that there is a dose-dependent effect of ritonavir on peripheral insulin sensitivity.
Ritonavir-boosted lopinavir is one of the most common PI combinations used in the treatment of HIV infection.20 Lopinavir cannot be studied in isolation due to its low bioavailability and rapid metabolism. Previously, we showed that a single dose of lopinavir/ritonavir (533 mg/133 mg) acutely decreased M/I by 13%.9 Noor et al10,11 also demonstrated significant reductions in insulin-mediated glucose disposal measured by the clamp after short-term administration of lopinavir/ritonavir (400 mg/100 mg twice daily for 5 or 10 days). In all 3 studies, plasma lopinavir levels reached therapeutic concentrations. However, ritonavir levels achieved during determination of M/I in those 3 studies were one quarter of those found here. Given that we did not detect an effect of boosting dose ritonavir on insulin sensitivity at the higher levels achieved here, it is likely that lopinavir was responsible for the induction of insulin resistance observed in those studies.
Noor et al11 found less effect on M/I of short-term administration of atazanavir/ritonavir (300 mg/100 mg daily for 10 days) in HIV-seronegative subjects. Clamp ritonavir levels achieved in that study were lower than those found here but were more than 2 times greater than those reached in prior short-term studies of lopinavir/ritonavir. Thus, although previous data from our group and Noor et al9-11 suggested that boosting dose ritonavir was unlikely to be a major contributor to the acute induction of peripheral insulin resistance observed in prior short-term studies of lopinavir/ritonavir, the data presented here directly demonstrate the lack of an effect of boosting dose ritonavir on insulin resistance. The major effect is likely due to lopinavir, although we cannot rule out some additive effects.
In summary, we show a single boosting dose of ritonavir does not alter insulin-mediated glucose disposal. Given our previous finding that a single full dose of ritonavir (which yielded significantly higher plasma levels than those achieved here) acutely induced insulin resistance, we also demonstrate a dose-dependent effect of ritonavir on peripheral insulin sensitivity. Further, our data suggest that lopinavir likely makes the major contribution to the induction of insulin resistance that has been observed in short-term studies of lopinavir/ritonavir. Consideration should be given to these findings when assessing the metabolic effects of antiretroviral regimens and advising patients.
METHODS
Ten healthy volunteers were recruited. They had no history of medical illness, a normal screening physical examination, normal hematology and chemistry laboratory results, and negative HIV-1 antibody tests before study. The protocol was approved by the Committee on Human Research at the University of California, San Francisco. Informed consent was obtained from each subject.
Exclusion criteria included a fasting glucose >5.6 mmol/L; body mass index >30 kg/m2; fasting triglyceride level >2.13 mmol/L; low-density lipoprotein-cholesterol >4.87 mmol/L; blood pressure >140/90 mm Hg; taking antihypertensive medications; creatinine >140.8 µmol/L; liver function tests >the upper limits of normal; pregnancy; >3 alcoholic drinks per day for men or 2 per day for women; use of amphetamines, cocaine, or heroin; or use of drugs affecting glucose or lipid metabolism within the previous 30 days.
Study Design
This was a randomized, double-blind, placebo-controlled, cross-over study. Subjects were admitted to the University of California, San Francisco Clinical and Translational Science Institute Clinical Research Center at San Francisco General Hospital the day before the study. After an overnight (10 hours) fast, subjects received either a single dose of ritonavir (Abbott Laboratories, Abbott Park, IL) 200 mg or placebo 3 hours before the start of the euglycemic hyperinsulinemic clamp. Subjects were re-admitted to the Clinical Research Center within 14-28 days, and studies were repeated using the alternative treatment (active drug or placebo). All subjects were examined by a study investigator between admissions to assess for complications or intercurrent illnesses.
Ritonavir plasma concentrations are highly variable; the half-life is 3-5 hours when administered in the fasting state.12 Therefore, a single dose of ritonavir 200 mg was administered 3 hours before the clamp to achieve plasma ritonavir levels of 0.7-1.8 µmol/L during the clamp, the range achieved in patients on boosting doses of ritonavir.9,11,13-15
Euglycemic Hyperinsulinemic Clamp
A 3-hour euglycemic hyperinsulinemic clamp was performed as described by DeFronzo et al16 and employed by us previously.7-9 A cannula was inserted into an antecubital vein for infusion. A vein in the dorsum of the contralateral hand was cannulated and warmed to 50-55°C for arterialized blood sampling. At the start of the clamp (t = 0), insulin (Humulin R, Eli Lilly, Indianapolis, IN) was administered as a primed continuous intravenous infusion for 10 minutes, followed by a constant infusion at 40 mU/m2 per minute for 180 minutes. Glucose was measured every 5 minutes throughout the clamp. A primed 20% glucose infusion was begun and adjusted every 5 minutes to maintain plasma glucose at 4.5 mmol/L (coefficient of variation < 5%). Blood samples were collected during the clamp for serum insulin levels. Insulin-mediated glucose disposal (M/I) was calculated using data from the last hour of the clamp (t = 120-180 minutes).
Resting Energy Expenditure
Oxygen consumption and carbon dioxide production were measured by indirect calorimetry (DeltaTrac II metabolic monitor, Yorba Linda, CA) to calculate resting energy expenditure.
Laboratory Measurements
Glucose and lactate were measured using a YSI 2300 STAT-Plus Glucose and Lactate Analyzer (YSI Inc, Yellow Springs, OH). Serum insulin levels were measured by radioimmunoassay (Millipore Bioscience, St. Charles, MO). Homeostasis model assessment-insulin resistance index was calculated from fasting plasma glucose and serum insulin at the start of the clamp (3 hours after administration of ritonavir or placebo).17 Fasting total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, and triglycerides were measured by spectrophotometric methods (Thermo Scientific, Middletown, VA; Wako Diagnostics, Richmond, VA; and Sigma, St. Louis, MO).
Ritonavir levels were measured by liquid chromatography, tandem mass spectrometry by the Drug Research Unit, San Francisco General Hospital (lower limit of quantitation 50 ng/mL, interassay and intra-assay coefficients of variation 4.2%-10.6% and 1.1%-9.1%, respectively). Area under the concentration time curve from t = 0 to 180 minutes was estimated using the linear-linear trapezoidal rule.
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