Comparison of insulin lispro protamine suspension versus insulin glargine once daily added to oral antihyperglycaemic medications and exenatide in type 2 diabetes: a prospective randomized open-label trial.

AIMS: To compare efficacy and safety of two, once-daily basal insulin formulations [insulin lispro protamine suspension (ILPS) vs. insulin glargine (glargine)] added to oral antihyperglycaemic medications (OAMs) and exenatide BID in suboptimally controlled type 2 diabetes (T2D) patients. METHODS: This 24-week, open-label, multicentre trial randomized patients to bedtime ILPS (n = 171) or glargine (n = 168). Non-inferiority of ILPS versus glargine was assessed by comparing the upper limit of 95% confidence intervals (CIs) for change in haemoglobin A1c (HbA1c) from baseline to week 24 (adjusted for baseline HbA1c) with non-inferiority margin 0.4%. RESULTS: Non-inferiority of ILPS versus glargine was demonstrated: least-squares mean between-treatment difference (ILPS minus glargine) (95% CI) was 0.22% (0.06, 0.38). Mean HbA1c reduction was less for ILPS- versus glargine-treated patients (-1.16 ± 0.84 vs. -1.40 ± 0.97%, p = 0.008). Endpoint HbA1c < 7.0% was achieved by 53.7% (ILPS) and 61.7% (glargine) (p = NS). Overall hypoglycaemia rates (p = NS) and severe hypoglycaemia incidence (p = NS) were similar. Nocturnal hypoglycaemia rate was higher in patients treated with ILPS versus glargine (p = 0.004). Weight gain was similar between groups (ILPS: 0.27 ± 3.38 kg; glargine: 0.66 ± 3.93 kg, p = NS). Endpoint total insulin doses were lower in patients treated with ILPS versus glargine (0.30 ± 0.17 vs. 0.37 ± 0.17 IU/kg/day, p < 0.001). CONCLUSIONS: ILPS was non-inferior to glargine for HbA1c change over 24 weeks, but was associated with less HbA1c reduction and more nocturnal hypoglycaemia. Treat-to-target basal insulin therapy improves glycaemic control and is associated with minimal weight gain when added to OAMs and exenatide BID for suboptimally controlled T2D.

A comparison between simplified and intensive dose-titration algorithms using AIR inhaled insulin for insulin-naive patients with type 2 diabetes in a randomized noninferiority trial.

BACKGROUND: Insulin initiation and optimization is a challenge for patients with type 2 diabetes. Our objective was to determine whether safety and efficacy of AIR inhaled insulin (Eli Lilly and Co., Indianapolis, IN) (AIR is a registered trademark of Alkermes, Inc., Cambridge, MA) using a simplified regimen was noninferior to an intensive regimen. METHODS: This was an open-label, randomized study in insulin-naive adults not optimally controlled by oral antihyperglycemic medications. Simplified titration included a 6 U per meal AIR insulin starting dose. Individual doses were adjusted at mealtime in 2-U increments from the previous day's four-point self-monitored blood glucose (SMBG) (total < or =6 U). Starting Air insulin doses for intensive titration were based on fasting blood glucose, gender, height, and weight. Patients conducted four-point SMBG daily for the study duration. Insulin doses were titrated based on the previous 3 days' mean SMBG (total < or =8 U). RESULTS: End point hemoglobin A1C (A1C) was 7.07 +/- 0.09% and 6.87 +/- 0.09% for simplified (n = 178) and intensive (n = 180) algorithms, respectively. Noninferiority between algorithms was not established. The fasting blood glucose (least squares mean +/- standard error) values for the simplified (137.27 +/- 3.42 mg/dL) and intensive (133.13 +/- 3.42 mg/dL) algorithms were comparable. Safety profiles were comparable. The hypoglycemic rate at 4, 8, 12, and 24 weeks was higher in patients receiving intensive titration (all P < .0001). The nocturnal hypoglycemic rate for patients receiving intensive titration was higher than for those receiving simplified titration at 8 (P < 0.015) and 12 weeks (P < 0.001). CONCLUSIONS: Noninferiority between the algorithms, as measured by A1C, was not demonstrated. This finding re-emphasizes the difficulty of identifying optimal, simplified insulin regimens for patients.

Efficient translocation and processing with Xenopus egg extracts of proteins synthesized in rabbit reticulocyte lysate.

Cell-free translation/translocation systems are broadly applied to examine gene expression and characterize the structure-function relationship of gene products. We present the characterization of Xenopus egg extract (XEE) translocation and processing of proteins synthesized in rabbit reticulocyte lysate. The XEE was prepared from eggs laid by adult female frogs that received serial injections of gonadotropins. The eggs were then dejellied in 2% L-cysteine-HCl and the cytoplasm extracted by centrifugation at 10,000 rpm for 15 min. The in vitro translocation and processing of XEE was examined with a cell-free translation system containing reticulocyte lysate, and appropriate messenger ribonucleic acid (RNA) or complementary deoxyribonucleic acid plasmids with RNA polymerase. Cell-free production of the following proteins were used to assess posttranslational modifications: Escherichia coli beta-lactamase for signal sequence cleavage, Saccharomyces cerevisiae alpha-mating factor for translocation and N-linked glycosylation, the soluble protein luciferase for functional activity, and the membrane-bound human insulin receptor for translation efficiency. All translation products were identified by [35S]-methionine labeling, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography. The results demonstrate that (1) XEE produces near-complete signal sequence and N-glycosylation processing of proteins synthesized in reticulocyte lysate, (2) XEE contains endoplasmic reticulum-equivalent microsomes, which allows for protein translocation and protease protection, (3) the addition of XEE in the translation reaction does not affect synthesis and chemiluminescence activity of luciferase, (4) XEE is efficient in processing the nascent 160-kDa human insulin receptor precursor, a transmembrane protein, and (5) as compared to canine pancreatic microsomes, XEE translocation efficiency is minimally decreased with the addition of dimethylsulfoxide. These results are the first description of the combined use of XEE with reticulocyte lysate and clearly demonstrate a higher efficiency of translocation and processing compared to canine pancreatic microsomes. This method of cell-free translation and processing allows for more extensive in vitro examination of posttranslational modifications of secretory and membrane-bound proteins.

Prevalence of glucose intolerance among Native Hawaiians in two rural communities. Native Hawaiian Health Research (NHHR) Project.

OBJECTIVE: To estimate prevalence of type 2 diabetes and impaired glucose tolerance (IGT) among a population of native Hawaiians in two rural communities. RESEARCH DESIGN AND METHODS: Prevalence of glucose intolerance was assessed in two rural communities by history (confirmed by record review) or with a 75-g oral glucose tolerance test according to World Health Organization criteria. Anthropometric and demographic data were also obtained. A short survey was used to estimate the prevalence of known diabetes among nonparticipants. Prevalence rates were adjusted using the standard world population of Segi. RESULTS: A total of 574 native Hawaiians age > or = 30 years participated. The crude prevalence of IGT and type 2 diabetes were 15.5 and 20.4%, respectively. Only IGT prevalence was significantly higher (P = 0.03) among women (18.7%) than among men (10.9%). Prevalence of glucose intolerance was significantly associated with BMI, waist circumference, and waist-to-hip ratio (WHR). After adjusting for age and BMI, waist circumference and WHR were significantly and independently associated with type 2 diabetes prevalence only among women. Prevalence of type 2 diabetes was not significantly associated with the percentage of Hawaiian ancestry after adjusting for age. CONCLUSIONS: This study observed a high prevalence of glucose intolerance associated with being overweight among native Hawaiians. Age-adjusted type 2 diabetes prevalence was four times higher than among the U.S. National Health and Nutrition Examination Survey (NHANES) II population. Prevalence was high despite high rates of admixture with other ethnic groups of Hawaii, suggesting that these other Asian and Pacific Island populations share similar susceptibility to type 2 diabetes risk.

The insulin resistance syndrome in native Hawaiians. Native Hawaiian Health Research (NHHR) Project.

OBJECTIVE: To investigate whether fasting hyperinsulinemia is associated with a clustering of cardiovascular disease (CVD) risk factors, manifesting as the insulin resistance syndrome (IRS), in a population of native Hawaiians. RESEARCH DESIGN AND METHODS: A total of 574 native Hawaiians > or = 30 years of age were examined for blood pressure, waist-to-hip ratio (WHR), BMI, oral glucose tolerance, and fasting lipid, insulin, and C-peptide concentrations. All statistical analyses (n = 384) excluded 190 individuals who had NIDDM or who were taking hypertension medication. Using logistic regression analysis, fasting insulin and C-peptide levels were compared with CVD risk factors (glucose intolerance, hypertension, central adiposity, elevated triglyceride levels, and low HDL cholesterol levels) after adjusting for age and obesity. RESULTS: Sixty-six percent of native Hawaiians were overweight or obese, and 70% were found to have central adiposity. Fasting insulin concentrations were correlated with BMI, WHR, blood pressure, and triglyceride, HDL cholesterol, and glucose concentrations. Fasting insulin was also significantly associated with an increasing number of CVD risk factors in each participant (P < 0.001). Fasting insulin and C-peptide concentrations were independently associated with glucose intolerance, high triglyceride levels, and low HDL cholesterol levels. However, only fasting C-peptide concentrations were independently associated with hypertension and central adiposity. Apparent differences in the correlates of fasting insulin and C-peptide may be related to multiple factors and warrant further evaluation. CONCLUSIONS: This study provides cross-sectional data confirming the existence of the IRS in native Hawaiians. However, further longitudinal studies are needed to examine the relationship of insulin resistance and/or surrogate markers to increased rates of NIDDM and CVD mortality in native Hawaiians.

In vitro translation of the human insulin proreceptor results in N-linked glycosylation without dimer formation.

The heterotetrameric configuration of the cell surface human insulin receptor (hINSR) is important for mediating insulin action. Investigation of proreceptor dimerization, the quaternary processing event during biogenesis, offers the potential to examine interactions between disulfide-linked receptor subunits. Thus, dimer formation of the proreceptor was examined in a cell-free system that utilized a coupled transcription and translation method with rabbit reticulocyte lysate. Translation products were labeled with [35S]methionine and identified by non-reducing SDS-polyacrylamide gel electrophoresis and autoradiography. In vitro synthesis in the presence of oxidized glutathione failed to demonstrate dimerization of the nascent proreceptor. Co-translational processing with the addition of microsomal membranes resulted in N-linked glycosylation of the proreceptor but without dimer formation. Thus, similar to that observed in vivo, insulin proreceptor dimerization does not appear to be a co-translational or early post-translational event.

A prospective examination of octreotide-induced gall-bladder changes in acromegaly.

OBJECTIVE: We wished to determine the effects of octreotide acetate, a somatostatin analogue, on gall-bladder function during treatment of acromegaly. DESIGN: We used a prospective, open label trial of somatostatin analogue. PATIENTS: Seventeen patients with acromegaly took part. MEASUREMENTS: Ultrasonographic evaluation of gall-bladder contents were performed pretreatment, after 1 month, and subsequently at intervals of 3-6 months. RESULTS: Non-shadowing floating echogenic particles were observed in the gall-bladder in 12 of 17 patients after (mean +/- SEM) 2.5 +/- 0.6 months of treatment. During long-term treatment (mean 20.8 +/- 4.3, median 13, range 1-59 months), ultrasound evidence for cholelithiasis was observed in four patients after 20 +/- 4 months (range 4.2-43) months of octreotide therapy. No symptoms of biliary tract disease have been observed. Duration of acromegaly, average GH, average IGF-I, gender, age at entry, dose of analogue, and concurrent use of non-steroidal anti-inflammatory drugs did not affect the occurrence of sludge or gallstones. CONCLUSIONS: Formation of non-shadowing, floating echogenic particles occurs commonly during the first 6 months of treatment with octreotide acetate. Cholelithiasis is a risk of long-term treatment.

Evidence for exogenous substrate phosphorylation and dimer formation by the activated 190 kDa insulin proreceptor in vitro.

The insulin receptor is synthesized as a single chain, 190 kDa glycoprotein precursor, which undergoes proteolytic cleavage, carbohydrate processing, and fatty acylation to generate the mature receptor on the plasma membrane. The relationship of these post-translational modifications to the acquisition of receptor function, i.e. ligand binding and phosphokinase activity, is not fully understood. Therefore, the 190 kDa proreceptor and mature receptor kinase activities were separately examined in vitro, and their phosphorylation properties compared. The solubilized receptor precursor from IM-9 lymphocytes was purified by sequential lectin chromatography and, following site specific anti-receptor antibody immunoprecipitation, phosphokinase studies performed. The isolated proreceptor was activated by insulin and phosphorylated exogenous substrate alpha-casein, as similarly observed for the mature receptor. Structurally, the phosphorylated proreceptor was identified as a 360 kDa homodimer under non-reducing condition.

Effects of castanospermine and 1-deoxynojirimycin on insulin receptor biogenesis. Evidence for a role of glucose removal from core oligosaccharides.

The insulin proreceptor is a 190-kDa glycoprotein that is processed to mature alpha (135-kDa) and beta (95-kDa) subunits. In order to determine the role of carbohydrate chain processing in insulin receptor biogenesis, we investigated the effect of inhibiting glucose removal from core oligosaccharides of the insulin proreceptor with glucosidase inhibitors, castanospermine and 1-deoxynojirimycin. Cultured IM-9 lymphocytes treated with inhibitors had 50% reduction in surface insulin receptors as demonstrated by ligand binding, affinity cross-linking with 125I-insulin, and lactoperoxidase/Na 125I labeling studies. Degradation rates of surface labeled receptors were similar in both control and inhibitor-treated cells (t1/2 = 5 h); thus, accelerated receptor degradation could not account for this reduction. Biosynthetic labeling experiments with [3H]leucine and [3H]mannose identified an apparently higher molecular size proreceptor (approximately 205 kDa) that failed to show the characteristic decline with time as seen in the normal 190-kDa proreceptor. Along with this finding, the biosynthetic label appearing in the mature subunits was reduced in these inhibitor-treated cells. Endoglycosidase H treatment of both precursors produced identical 170-kDa bands. Carbohydrate chains released from the 205-kDa precursor by endoglycosidase H migrated in the same position as the Glc2-3Man9GlcNAc standards when separated by high performance liquid chromatography, whereas the 190-kDa proreceptor oligosaccharides migrated similar to the Man7-9GlcNAc chains. Although the mature subunits of control and inhibitor-treated cells demonstrated equal electrophoretic mobility, the endoglycosidase H-sensitive oligosaccharides of the mature subunits in treated cells also contained residues that migrated similar to the Glc2-3Man9GlcNAc standards. Thus, glucose removal from core oligosaccharides is apparently not necessary for the cleavage of the insulin proreceptor, but does delay processing of this precursor, which probably accounts for the reduction in cell-surface receptors.

The pharmacokinetics of subcutaneous regular insulin in type I diabetic patients: assessment using a glucose clamp technique.

We recently reported that the peak effect and duration of action of regular insulin injected sc were prolonged in diabetic patients and were not related to the presence of insulin antibodies. The results suggested that the ambient level of plasma glucose might be an important factor in determining the pharmacokinetics of regular insulin. In the present study we used a glucose clamp technique, which minimizes interference by counterregulatory phenomena, to study the pharmacokinetics of regular insulin injected sc at 2 different blood glucose concentrations [276 +/- 7 (+/- SEM) and 130 +/- 5 mg/dl] in 10 insulin-dependent diabetic patients. The patient's blood glucose concentration was maintained constant by means of a variable rate iv infusion of 20% dextrose in water after sc injection of regular insulin (0.2 U/kg) in the deltoid region of the arm. The onset of insulin action occurred at similar times at both glucose concentrations (0.6 +/- 0.1 h at 276 mg/dl vs. 0.5 +/- 0.1 h at 130 mg/dl; P greater than 0.05). Peak insulin action (determined from the time of the maximal glucose infusion rate) was delayed in the studies done at 276 mg/dl (4.7 +/- 0.2 h) compared to that in studies done at mean glucose concentrations of 130 mg/dl (4.3 +/- 0.2 h; P less than 0.05). The duration of insulin action was also significantly prolonged in the studies done at the higher glucose concentrations (9.1 +/- 0.3 h at 276 mg/dl vs. 7.7 +/- 0.2 h at 130 mg/dl; P less than 0.01). These results confirm previous reports of prolonged insulin action in diabetic patients, especially in the presence of hyperglycemia.

Prolonged action of regular insulin in diabetic patients: lack of relationship to circulating insulin antibodies.

We previously reported that in insulin-treated diabetic subjects the time course of action of regular insulin injected sc is different from that reported in standard textbooks. The present studies evaluated the role of insulin antibodies (Abs) in the altered pharmacokinetics of regular insulin by comparing the time course of insulin action in 10 patients receiving chronic insulin therapy and having insulin Abs with that in 15 previously untreated patients without detectable Abs. After an overnight fast, the patients were given an infusion of 5% dextrose in water at 100 ml/h. Regular insulin (15 U) was then injected sc in the deltoid region of the arm. The onset of action of sc insulin, as indicated by a 10% fall in serum glucose, was similar in both patient groups [1.9 +/- 0.1 (+/- SEM) hour in Ab-negative and 1.8 +/- 0.1 h in Ab-positive patients]. The peak effect of insulin action, as determined by the nadir of serum glucose, was 4.6 +/- 0.2 h in the previously untreated patients, not significantly different from the value in the diabetic patients with insulin Abs (5.2 +/- 0.4 h). The duration of action of insulin was also similar in both groups (14.7 +/- 0.7 vs. 14.4 +/- 1.0 h). No significant correlations were found between insulin Ab levels and any of these 3 parameters of insulin action. However, the peak effect and total duration of insulin action were significantly correlated with the baseline serum glucose levels. A possible role of insulin Abs was evaluated in these patients by repeating the studies over a 2-year period. During this time, the previously untreated patients were treated with highly purified pork insulin, to which they developed low titers of insulin Abs. The diabetic patients who had been chronically treated with insulin were changed from less purified insulin to highly purified pork insulin, and all had a significant reduction in their Ab titers. No changes in insulin pharmacokinetics were found in either group. These studies demonstrate that the prolonged action of sc injected regular insulin in diabetic patients is not related to the effect of circulating insulin Abs.