Growth hormone receptor antagonism with pegvisomant in insulin resistant non-diabetic men: A phase II pilot study

Participants and study design

Four men, aged 52–57 years, with a BMI between 18–35 kg/m² and insulin resistance, defined as a HOMA-IR score >2.77¹⁶, were enrolled. The first subject was enrolled on March 3, 2014. The sample size was determined after a power analysis based on prior work and the effect size of the primary outcome. Participants were recruited through a combination of advertising, word of mouth, and doctor’s appointments. Participants were required to be on a stable medication regimen for any lipid disorders. Participants were excluded if they had type 1 or type 2 diabetes, fasting blood glucose >126 mg/dl, hemoglobin A1c >6.5, unstable hypertension, human immunodeficiency virus infection, hepatitis B or hepatitis C infection, evidence of chronic kidney disease, major gastrointestinal surgery, history of pancreatitis, pancreatic disease, liver or biliary disorders, or fasting plasma triglyceride >500 mg/dl. The study was completed in August 2015, and the final follow-up was completed in December 2015. The Committee on Human Research of the University of California, San Francisco approved the study (IRB Number: 13-10982; full protocol available as Supplementary File 1). Written, informed consent was obtained from each individual.

Participants were admitted to the Clinical Research Center (CRC) at San Francisco General Hospital the evening before baseline testing and consumed a controlled metabolic diet with fixed proportions of macronutrients (10 kcal/kg; 15% protein, 30% fat, 55% carbohydrate with <20% from simple sugars). All meals were prepared in the metabolic kitchen of the CRC under the supervision of CRC bionutritionists. In the morning, the participants underwent metabolic assessments, including measurements of body composition by dual-energy X-ray absorptiometry (DXA), resting energy expenditure by indirect calorimetry, and hyperinsulinemic-euglycemic clamp with stable isotope tracer infusions. After baseline testing, participants were discharged and self-administered pegvisomant 20 mg subcutaneously nightly, after supervised instruction from a CRC nurse. Participants were instructed to maintain their usual diets and activity levels and attended weekly follow up visits. During these visits, the participants had updated medical history and a brief physical exam, including weight and vital signs. All reported signs and symptoms were recorded. Safety laboratory studies (i.e. fasting glucose, lipids, electrolytes, and renal and hepatic function), as well as insulin, GH and IGF1 levels, were obtained. At these visits they also returned unused vials of drug and received their next week’s supply of drug and supplies. After four weeks of treatment, the inpatient metabolic assessments were repeated, as at baseline.

Euglycemic, hyperinsulinemic clamp

Whole-body insulin sensitivity was measured by the euglycemic hyperinsulinemic clamp technique¹⁷. After the baseline measurements were completed, insulin (Humulin®, Eli Lilly & Co., Indianapolis, IN, USA; 40 mU/m² min) was infused for 180 minutes and blood samples were collected at 5-minute intervals from a retrograde intravenous line placed in a hand that was warmed in a heated box at 50–55^oC. Whole-blood glucose concentrations were determined by the glucose oxidase method (YSI Stat glucose analyzer, Yellow Springs, OH, USA). A variable infusion of 20% dextrose (labeled with [U-¹³C] glucose, as described below) was adjusted to maintain blood glucose concentrations at 90 mg/dL. Blood samples were collected at 30-minute intervals during the final hour of the clamp and the serum was frozen and batched for measurement of insulin. Insulin sensitivity was calculated as a measure of whole-body glucose uptake during the final hour of the clamp (M) divided by steady-state serum insulin level (I)¹⁸.

Stable isotope tracer methods

EGP (R_a glucose) and whole body lipolysis (R_a glycerol) were measured under fasting conditions using primed constant infusions of [U-¹³C] glucose (0.96 mg/kg/h, prime 0.096 mg/kg/min for 10 min) and [²H₅]-glycerol (0.67 mg/kg/h, prime 0.067 mg/kg/min for 10 min) started at 430 h. Blood samples were obtained every 10 minutes between 0800 and 0830 h for steady-state fasting measurements. The isotope infusions continued, and a 180-minute euglycemic-hyperinsulinemic clamp was started at 0900 h, as described above. The constant glycerol infusion continued, while the enriched glucose became a variable infusion of 0.6% [U-¹³C] glucose within the 20% dextrose infusion used for the clamp. During the final 30 minutes of the clamp (1130 to 1200 h), blood samples were collected every 10 minutes for determination of EGP and lipolysis under conditions of steady-state hyperinsulinemia.

Isotope enrichments were measured by Metabolic Solutions (Nashua, NH, USA). [²H₅]-glycerol was determined by gas chromatography-mass spectrometry (GC-MS), using the trimethylsilyl (TMS) derivative¹⁹. Plasma ¹³C₆-glucose was determined using the aldonitrile penta-acetate derivative²⁰.

Ra glucose and Ra glycerol were calculated by the dilution technique using the average of the last 4 samples during the fasting state and during the clamp²¹.

Body composition

Total and regional fat and lean body mass were measured by DXA (Hologic, Marlborough, MA, USA) and subsequently analyzed using Apex 5.5™ software (Hologic, Marlborough, MA, USA) to provide estimates of visceral adipose tissue.

Indirect calorimetry

Resting energy expenditure was measured by indirect calorimetry under fasting conditions and during the clamp using a Deltatrac II Metabolic Monitor (Sensormedics, Yorba Linda, CA, USA). Respiratory quotient (RQ), and index of substrate utilization, was calculated as carbon dioxide production divided by oxygen production rates.

Other laboratory measures

Free insulin like growth factor (IGF)-1 and IGF-BP3 were determined by ELISA (GenWay Biotech, San Diego, CA, USA). The San Francisco General Hospital Clinical Laboratory measured total and high-density lipoprotein (HDL) cholesterol, fasting triglycerides (TG), and calculated low-density lipoprotein (LDL) cholesterol, serum insulin, and fasting serum glucose. Serum insulin was measured by chemiluminescent sandwich assay.

Statistical methods

The primary outcome was specified to be changes in insulin sensitivity (M/I). Key secondary outcomes were changes in endogenous glucose production (%EGP suppression) and changes in lipolysis (% Ra glycerol suppression). Analyses were performed using Graph Pad Prism 7.0. Student’s paired t-test was used to compare baseline values to values after one month of treatment, using two tailed p-values. Values are represented as mean ± SD.

ClinicalTrials.gov identifier: NCT02023918

Changes from ClinicalTrials.gov protocol

The following deviations from the protocol were made. A total of 6 male participants were recruited, enrolled, and completed the entire study. There was a modification to the protocol after the first 4 participants to change the insulin infusion rate during the clamp. This was done because there was near complete suppression of endogenous glucose production with the original insulin dose. However, there did not appear to be a significant effect of lowering the insulin dose in the final 2 participants. To maintain consistency, the final analysis included only the data from the first 4 participants who were treated according to the original clamp protocol. The summary data published on the ClinicalTrials.gov website include all 6 participants, while the data presented here include on the first 4. Despite enrolling fewer than the expected number of participants, the investigators felt that there was an extremely low likelihood that additional participants would change the outcomes, so the study was terminated at this point.

Consort checklist and flowchart are available as Supplementary File 2 and Supplementary File 3.

Participants and clinical course

The mean age of the participants was 54.5 ± 2.1 years (Table 1). Three of the participants carried a diagnosis of hyperlipidemia and were on statin medications. Three of the participants carried a diagnosis of hypertension and two were on antihypertensive medications (amlodipine and metoprolol). One participant carried a diagnosis of gout, but did not require medication during the study.

Participants were adherent to the daily self-infections of pegvisomant based on weekly medication reconciliation and measurement of IGF-1 levels.

IGF-1 and binding proteins

As shown in Figure 1, total IGF-1 levels decreased in all participants (134.0 ± 41.5 vs. 72 ± 11.7 ng/mL, p = 0.04). There was no significant change in IGF-BP3 levels (Table 1).

Figure 1. Effects of pegvisomant treatment on total insulin-like growth factor (IGF)-1 levels.

IGF-1 decreased as expected over the four-week treatment period. Circles indicate individual baseline and post-treatment values. P values are derived from paired t-tests.

Glucose metabolism

There was no significant change in fasting blood glucose, fasting insulin, or HOMA-IR following four weeks of pegvisomant treatment (Table 1). There was no significant difference in the serum glucose level or the glucose infusion rate during the clamp (Figure 2). There was no difference in clamped insulin levels pre- and post-treatment (Figure 3). There was no difference in basal EGP pre- or post-pegvisomant treatment or in the percent suppression of EGP by insulin (Figure 4). There was a small increase in Ra glucose post-treatment, but there was no significant difference in suppression of EGP during the clamp (Table 1). There was no significant change in whole body insulin sensitivity as assessed by M/I (3.2 ± 1.3 vs. 3.4 ± 2.4 p = 0.82).

Figure 2. Glucose infusion rate and plasma glucose during hyperinsulinemic euglycemic clamp.

There was no difference between the glucose infusion rate (GIR) after treatment with pegvisomant. Blue symbols indicate GIR at baseline. Red symbols indicate GIR post-treatment.

Figure 3. Clamped insulin values.

Insulin was measured at various times during the clamp. Data represent the mean insulin levels +/- SD for the pre- (closed circles) and post- (dark squares) during the steady state portion of the clamp.

Figure 4. Endogenous glucose production (EGP) suppression.

Data represent the percent suppression of endogenous glucose production during the hyperinsulinemic euglycemic clamp at 140 minutes at baseline and after treatment with pegvisomant for one month.

Lipid metabolism

There was no significant difference in fasting TG, HDL, or LDL following pegvisomant treatment. Whole body lipolysis did not change in either the fasting state or during hyperinsulinemia.

Body composition

Lean body mass did not change significantly during the treatment period. There was no change in total fat mass, nor was there a change in visceral adipose tissue mass, but there was a small but statistically significant decrease in appendicular fat (decrease of 0.4 kg, p <0.01). While truncal fat also decreased by 0.5 kg, this did not reach statistical significance (p = 0.11).

Indirect calorimetry

There was no significant change in resting energy expenditure or RQ measured during the clamp. Fasting RQ declined significantly (p = 0.04).

Safety measures

One participant, who had prior aspartate aminotransferase (AST) and abnormal liver function (ALT), had a mild increase of his transaminases during weeks 2 and 3 of monitoring, but these remained less than twice the upper limits of normal and decreased back to his baseline while on drug treatment. Side effects for all participants were limited to injection site discomfort. No participants discontinued the drug as a consequence of side effects or laboratory abnormalities.