Treatment intensification without improved HbA1c levels in children and adolescents with Type 1 diabetes mellitus.

AIMS: To examine trends in diabetes treatment in Danish children and adolescents with Type 1 diabetes mellitus, comparing treatment intensity with metabolic outcomes in the population, and to describe the challenges of population-based registries in a clinical setting with rapidly changing treatment methods. METHODS: This observational study is based on the Danish national population registry of childhood diabetes, which includes 99% of children diagnosed with Type 1 diabetes before the age of 15 years. We included 4527 people diagnosed between 2000 and 2012. Self-monitored blood glucose measurements, insulin injections/boluses, treatment method and metabolic control quantifications were analysed and adjusted for the effects of gender and ethnicity, the combined effect of age, visit year and duration, and for the random effects of individual and hospital settings. RESULTS: Treatment was intensified via an increasing number of self-monitored blood glucose measurements and injections/boluses. More than six injections/boluses and an increased number of self-monitored blood glucose measurements were significantly associated with lower metabolic control. No reduction, however, in the overall mean HbA1c concentration was observed between 2005 [66 mmol/mol (8.2%)] and 2012 [65 mmol/mol (8.1%)]. Changed registration practices in 2009 introduced artificial jumps in data. CONCLUSIONS: Intensifying treatment alone does not lead to improved metabolic control in the overall population despite the appearance of lower HbA1c in individuals with a greater number of self-monitored blood glucose measurements and injections/boluses. The contradictory results reflect difficulties in using observational studies to predict results of intervention in the individual. Data collected from population-based registries need to be adjusted continuously to reflect changes in care.

Ultra-long pharmacokinetic properties of insulin degludec are comparable in elderly subjects and younger adults with type 1 diabetes mellitus.

BACKGROUND: Management of diabetes in elderly subjects is complex and careful management of glucose levels is of particular importance in this population because of an increased risk of diabetes-related complications and hypoglycaemia. OBJECTIVE: The aim of this study was to evaluate the pharmacokinetic and pharmacodynamic properties of insulin degludec (IDeg), a basal insulin with an ultra-long duration of action, in elderly subjects with type 1 diabetes compared with younger adults. METHODS: This trial was a randomised, double-blind, two-period, crossover trial conducted in a single centre and included both inpatient and outpatient periods. Subjects were men and women aged 18-35 years inclusive (younger adult group) or ≥65 years (elderly group) with type 1 diabetes who received IDeg (0.4 U/kg) via subcutaneous injection in the thigh once-daily for six days. Following 6-day dosing, a 26-hour euglycaemic glucose clamp procedure was conducted to evaluate the steady-state pharmacodynamic effects of IDeg. Blood samples were taken for pharmacokinetic analysis up to 120 h post-dose. Pharmacokinetic endpoints included the total exposure of IDeg, ie the area under the IDeg serum concentration curve during one dosing interval at steady state (AUC(IDeg,τ,SS)) (τ = 0-24 h, equal to one dosing interval) and the maximum IDeg serum concentration at steady state (C(max,IDeg,SS)). Pharmacodynamic endpoints included the total glucose-lowering effect of IDeg, ie the area under the glucose infusion rate (GIR) curve at steady state (AUC(GIR,τ,SS)), and the maximum GIR at steady state (GIR(max,IDeg,SS)). RESULTS: Total exposure (AUC(IDeg,τ,SS)) and maximum concentration (C(max,IDeg,SS)) of IDeg were comparable between elderly subjects and younger adults. Estimated mean age group ratios (elderly/younger adult) for AUC(IDeg,τ,SS) and C(max,IDeg,SS) and corresponding two-sided 95 % confidence intervals (CIs) were 1.04 (95 % CI 0.73-1.47) and 1.02 (95 % CI 0.74-1.39), respectively. Mean AUC(IDeg,0-12h,SS)/AUC(IDeg,τ,SS) was 53 % in both younger adult and elderly subjects, showing that in both age groups IDeg exposure was evenly distributed across the first and second 12 h of the 24-hour dosing interval. No statistically significant differences were observed between younger adult and elderly subjects with regard to AUC(GIR,τ,SS) (the primary endpoint of this study) and GIR(max,IDeg,SS). Estimated mean age group ratios (elderly/younger adult) for AUC(GIR,τ,SS) and GIR(max,IDeg,SS) and corresponding two-sided 95 % CIs were 0.78 (95 % CI 0.47-1.31) and 0.80 (95 % CI 0.54-1.17), respectively. Duration of action was beyond the clamp duration of 26 h in all subjects. CONCLUSIONS: The exposure of IDeg at steady state during once-daily dosing was similar in younger adult and elderly subjects. The glucose-lowering effect of IDeg was numerically lower in elderly subjects compared with younger adults, but no significant differences were observed between age groups. The ultra-long pharmacokinetic and pharmacodynamic properties of IDeg observed in younger adults were preserved in elderly subjects with type 1 diabetes. Clinical trials.gov number: NCT00964418.

Ultra-long-acting insulin degludec has a flat and stable glucose-lowering effect in type 2 diabetes.

AIMS: Insulin degludec (IDeg) is a new-generation, ultra-long-acting basal insulin that forms soluble multihexamers upon subcutaneous injection, resulting in a depot from which IDeg is absorbed slowly and continuously into circulation. This double-blind, two-period, incomplete block cross-over trial investigated the pharmacodynamic and pharmacokinetic properties of IDeg at steady state (SS) in people with type 2 diabetes. METHODS: Forty-nine subjects treated with insulin without concomitant oral anti-diabetic drugs were given IDeg (0.4, 0.6 and/or 0.8 U/kg) once daily for two 6-day periods, separated by an interval of 13-21 days. Following dosing on Day 6, subjects underwent a 26-h euglycaemic glucose clamp (Biostator®; clamp blood glucose level: 90 mg/dl; 5.0 mmol/l). Pharmacokinetic samples were taken until 120 h after last dosing. RESULTS: For all dose levels, the mean glucose infusion rate (GIR) profiles were flat and stable. The glucose-lowering effect of IDeg was evenly distributed over the dosing interval τ, with area under the curve (AUC) for each of the four 6-h intervals being approximately 25% of the total AUC (AUC(GIR) (,τ,) (SS) ). Total glucose-lowering effect increased linearly with increasing dose. The blood glucose levels of all subjects stayed very close to the clamp target until end of clamp. The terminal half-life of IDeg was approximately 25 h at steady state. IDeg was well tolerated and no safety concerns were identified. No injection site reactions were reported. CONCLUSIONS: IDeg has a flat and consistent glucose-lowering effect in people with type 2 diabetes.

Symmetrical dimer of the human dopamine transporter revealed by cross-linking Cys-306 at the extracellular end of the sixth transmembrane segment.

There is evidence both for and against Na(+)- and Cl(-)-dependent neurotransmitter transporters forming oligomers. We found that cross-linking the human dopamine transporter (DAT), which is heterologously expressed in human embryonic kidney 293 cells, either with copper phenanthroline (CuP) or the bifunctional reagent bis-(2-methanethiosulfonatoethyl)amine hydrochloride (bis-EA) increased the apparent molecular mass determined with nonreducing SDS/PAGE from approximately 85 to approximately 195 kDa. After cross-linking, but not before, coexpressed, differentially epitope-tagged DAT molecules, solubilized in Triton X-100, were coimmunoprecipitated. Thus, the 195-kDa complex was a homodimer. Cross-linking of DAT did not affect tyramine uptake. Replacement of Cys-306 with Ala prevented cross-linking. Replacement of all of the non-disulfide-bonded cysteines in the extracellular and membrane domains, except for Cys-306, did not prevent cross-linking. We conclude that the cross-link is between Cys-306 at the extracellular end of TM6 in each of the two DATs. The motif GVXXGVXXA occurs at the intracellular end of TM6 in DAT and is found in a number of other neurotransmitter transporters. This sequence was originally found at the dimerization interface in glycophorin A, and it promotes dimerization in model systems. Mutation of either glycine disrupted DAT expression and function. The intracellular end of TM6, like the extracellular end, is likely to be part of the dimerization interface.

Two active molecular phenotypes of the tachykinin NK1 receptor revealed by G-protein fusions and mutagenesis.

The NK1 neurokinin receptor presents two non-ideal binding phenomena, two-component binding curves for all agonists and significant differences between agonist affinity determined by homologous versus heterologous competition binding. Whole cell binding with fusion proteins constructed between either Galpha(s) or Galpha(q) and the NK1 receptor with a truncated tail, which secured non-promiscuous G-protein interaction, demonstrated monocomponent agonist binding closely corresponding to either of the two affinity states found in the wild-type receptor. High affinity binding of both substance P and neurokinin A was observed in the tail-truncated Galpha(s) fusion construct, whereas the lower affinity component was displayed by the tail-truncated Galpha(q) fusion. The elusive difference between the affinity determined in heterologous versus homologous binding assays for substance P and especially for neurokinin A was eliminated in the G-protein fusions. An NK1 receptor mutant with a single substitution at the extracellular end of TM-III-(F111S), which totally uncoupled the receptor from Galpha(s) signaling, showed binding properties that were monocomponent and otherwise very similar to those observed in the tail-truncated Galpha(q) fusion construct. Thus, the heterogenous pharmacological phenotype displayed by the NK1 receptor is a reflection of the occurrence of two active conformations or molecular phenotypes representing complexes with the Galpha(s) and Galpha(q) species, respectively. We propose that these molecular forms do not interchange readily, conceivably because of the occurrence of microdomains or "signal-transductosomes" within the cell membrane.

Septide and neurokinin A are high-affinity ligands on the NK-1 receptor: evidence from homologous versus heterologous binding analysis.

The three main tachykinins, substance P, neurokinin A (NKA), and neurokinin B, are believed to be selective ligands for respectively the NK-1, NK-2 and NK-3 receptors. However, NKA also has actions which cannot be mediated through its normal NK-2 receptor and the synthetic peptide [pGlu6,Pro9]-Substance P9-11--called septide--is known to have tachykinin-like actions despite its apparent lack of binding to any known tachykinin receptor. In the cloned NK-1 receptor expressed in COS-7 cells NKA and septide as expected were poor competitors for radiolabeled substance P. However, by using radiolabeled NKA and septide directly, it was found that both peptides in homologous binding assays as well as in competition against each other in fact bound to the NK-1 receptor with high affinity: Kd values of 0.51 +/- 0.15 nM (NKA) and 0.55 +/- 0.03 nM (septide). It is concluded that NKA and septide are high-affinity ligands for the NK-1 receptor but that they are poor competitors for substance P, which in contrast competes very well for binding with both NKA and septide.