Human Insulin Formulations Research Articles

Less is more: Validating a single method for comprehensive rh-insulin analysis

The objective of this current study is to establish a single method for potency and related proteins analysis of human insulin formulations using reverse-phase high performance liquid (RP-HPLC) chromatography technique which was validated and verified for the potency analysis in insulin formulations. Chromatographic separation was achieved using an octadecylsilane (C-18) stationary phase and a mobile phase composed of 55% (v/v) buffer (0.2 M sodium sulfate in water, {pH 2.3}) and 45% (v/v) acetonitrile. Detection was performed by UV detector at 214 nm with a flow rate of 1 ml/min and an injection volume of 20 µL, at 40°C. Currently there are separate methods available in Indian Pharmacopoeia for analysis of Potency and Related proteins in human insulin. We have validated a single method where quantitation of potency and related proteins can be performed in the same run. The method validation exhibited linearity over the concentration range of 0.08–4.5 mg/ml (r2=0.999) with limit of detection of 0.094 mg/ml The accuracy of the method was 99–102.8%. Thus, it is proposed that both potency and related proteins in insulin formulations can be precisely evaluated using a single run thus saving the time and cost for quality analysis of insulin preparations both at manufacturing and regulatory laboratories which in turn will increase the market availability of such standard quality insulin preparations for public health use.

L’insulina alla luce delle nuove linee guida italiane per il trattamento del diabete di tipo 2: quando, in chi e perché?

Aim: this editorial is aimed at providing a reference for the insulin treatment of type 2 diabetes in adults. Methods and results: the present paper revised the recently published Italian guidelines for the treatment of type 2 diabetes (outpatients), either in primary care or specialist referral. The guideline has been developed following the methods described in the Manual of the National Guideline System (http://www.snlg-iss.it) by a panel nominated by the Società Italiana di Diabetologia (SID) and Associazione Medici Diabetologi (AMD). Prior cardiovascular events, heart failure, hypoglycemic risk, and other conditions have been considered as factors capable of modifying treatment strategies. The following areas have been assessed: 1) position of the insulin therapy in the new algorithms; 2) type of formulations (insulin analogues or human insulin); 3) use of insulin infusion pumps. Conclusions: insulin should be used only as a second/thirdline therapy due to its lower efficacy on long-term HbA1c, higher risk of hypoglycemia, and neutral effects on cardiorenal outcomes. Human insulin formulations should be avoided for safety concerns (higher hypoglycemic risk) with no preference among different insulin analogues. Insulin pumps are not recommended for the treatment of type 2 diabetes.

Simultaneous characterization of insulin HMWP and protamine sulphate in complex formulations through SEC-coupled mass spectrometry

High molecular weight protein aggregates present in a recombinant human insulin and analogs are conventionally quantified by SEC-HPLC and identified by SEC-MALS as oligomeric population which lacks precise identification of species. The limitation of these two techniques is vanquished though simultaneous separation and identification by SEC coupled with MS. The identification was established with organic solvent based isocratic elution followed by MS for parallel separation and identification of HMWP species. The developed SEC-MS method is validated to establish the method capability and variability. Further investigations under stress conditions of Insulin analogues revealed the capability of the method to separate higher order oligomeric (Trimeric, and Tetrameric) species alongside covalent dimeric species. Additionally, the method holds good in separating and sequencing protamine peptides used in suspension (Neutral Protamine Hagedorn) and biphasic/mixed (70/30) formulations of Human insulin using ETD-MSMS. The data presented here shows insight towards utilization of state-of-the-art SEC-MS technique in the biopharmaceutical field as a tool to establish the structural comparability of higher order species in biosimilars and to investigate the lot to lot batch variability for protamine sulphate in-terms of sequence confirmation.

1023-P: Dose Titration and Clinical Effects of Inhaled Technosphere Insulin Compared with Mealtime Subcutaneous (SC) Analog Insulin Therapy in Type 1 Diabetes (T1D)

Insulin dosing has been a clinical challenge since the introduction of “islet extract” in the 1920’s. Route of administration, insulin preparation and peptide modifications alter clinical effects. With newer insulins, patients and clinicians must clearly understand effective dosing. Technosphere Insulin (TI) is a dry-powder formulation of recombinant human insulin adsorbed onto Technosphere microparticles administered by pulmonary inhalation. TI cartridges are labeled as insulin “units” (4, 8, 12 unit cartridges). With proper dose titration, TI achieves comparable glucose control, improves mealtime glucose, and lowers hypoglycemia risk vs. insulin analogs in T1D. Methods: These analyses compare doses of TI achieved after active titration in 4 randomized trials in individuals with T1D. The ratio of TI inhalation units (based on labeled cartridge content) to SC prandial analog in the comparator group was calculated for each trial at study endpoint. Results: Conclusion: The current analyses across 4 unique studies support the titration of individual patients to TI doses that are approximately 1.3 - 2.5x those of their comparable SC analog insulin doses. These data show that greater “unit doses” of TI should be used to achieve glycemic control in T1D and thus, insulins should be dosed based on glycemic responses rather than “insulin units.” Disclosure D.M. Kendall: Employee; Self; MannKind Corporation. Stock/Shareholder; Self; Eli Lilly and Company. J. Manoukian: Employee; Self; MannKind Corporation. J.A. Krueger: Employee; Self; MannKind Corporation. Stock/Shareholder; Self; Eli Lilly and Company, MannKind Corporation. M. Jones: Employee; Self; MannKind Corporation. R. Abaniel: None. P.M. Morey: Employee; Self; MannKind Corporation. M.L. Grant: Employee; Self; MannKind Corporation. Stock/Shareholder; Self; MannKind Corporation. B.J. Hoogwerf: Consultant; Self; MannKind Corporation. Employee; Spouse/Partner; MannKind Corporation. Stock/Shareholder; Spouse/Partner; Eli Lilly and Company. Stock/Shareholder; Self; Eli Lilly and Company.

1024-P: Technosphere Insulin Added to Basal Insulin Is Associated with Less Weight Gain than Basal Insulin plus Insulin Aspart or Insulin Analog (Aspart) 70/30 Mixture in Type 2 DM

Technosphere Insulin (TI) is a dry-powder formulation of recombinant human insulin adsorbed onto Technosphere microparticles for oral inhalation. Data from two 52-wk randomized phase 3 trials in type 2 DM were combined to analyze the effects of TI on body weight. In one trial, patients on basal insulin were randomized to prandial TI or insulin aspart. In the other trial, basal insulin plus prandial TI was compared to 70/30 insulin analog mixture. Primary efficacy end point in each trial was change in A1C. Weight data were captured in each study. Weight and A1C data were analyzed based on modified Intention to Treat Analyses and LS mean changes from baseline. In these analyses, the TI arms were combined and comparisons were made between TI and each of the comparator arms. Aspart and 70/30 arms were not compared to each other. Data for 26 and 52 wk time points are reported. Results At both 26 and 52 weeks, TI was associated with less weight gain than either aspart or premixed insulin. Differences were significant for TI vs. each comparator. A1C differences among treatment arms were small and only statistically significant for TI vs. aspart at 52 weeks. Conclusion: In insulin treated T2DM persons, TI is associated with less weight gain than Insulin aspart or premixed 70/30 analog insulin. These weight differences appear to be independent of changes in A1C. Disclosure J. Manoukian: Employee; Self; MannKind Corporation. D.M. Kendall: Employee; Self; MannKind Corporation. Stock/Shareholder; Self; Eli Lilly and Company. P.M. Morey: Employee; Self; MannKind Corporation. B.J. Hoogwerf: Consultant; Self; MannKind Corporation. Employee; Spouse/Partner; MannKind Corporation. Stock/Shareholder; Spouse/Partner; Eli Lilly and Company. Stock/Shareholder; Self; Eli Lilly and Company.

650-P: Safety and Effectiveness of Recombinant Human Insulin Insugen in Real-World Clinical Setting

Background: As the burden of diabetes is increasing dramatically, the demand of insulin is also on rise, affordable access to insulin remains a challenge, particularly in Low and Mid income countries. For that reason, recombinant human insulin (rHI) formulations are considered to be effective alternatives to insulin analogues in countries with limited healthcare systems and resources. Continuing patient education and support programmes with rHI are critical role players in improving the access and overall disease management. Objective: This is a non-interventional study to observe the safety and effectiveness of Recombinant Human insulin in a real world population with type 2 (T2DM) diabetes mellitus from Indian perspective. Methodology: Out of 10426 diabetes patients who were enrolled in patient support program i.e “Insulin Therapy Assistance Program (iTAP)”, 5691 patients were initiated with various formulations of rHI (Insugen R, N, 30/70 and 50/50). Insulin therapy was initiated in T2DM patients with A1C > 7%. HbA1c, FPG and PPG values were recorded at baseline and follow up of 3, 6 and 9 months. Results: The Figure 1, 2, 3 depict the baseline and follow up of FBG, PPG and HbA1c respectively. Figure 1. Change in FPG (*p<0.05). Figure 2. Change in PPG (*p<0.05). Figure 3. Change in HbA1c (*p<0.05). Conclusion: In the current study, various formulations of rHI were well tolerated and resulted in significant reduction of HbA1c, FPG and PPG when aided with patient support program. Considering the economic impact of diabetes patients in taking insulins, human insulin become an alternative option in comparison to analogues which are expensive. Early initiation of insulin therapy can prevent or delay diabetes complications. Access to affordable insulin therapy is the key success factor while considering Insulin Therapy. Disclosure P. Raj: Employee; Self; Biocon. P. J: None. D.C. Mathew: None. C. Biswas: None. S. Nethra: None.

1019-P: Variability of 501 Inhaled Insulin Absorption and Action

501 inhaled insulin (INH) is a novel liquid formulation of human insulin for inhalation with faster onset of action than s.c. insulin lispro (LIS). This randomized, replicated cross-over trial investigated the variability of INH vs. LIS during 10-hour glucose clamps in non-smoking subjects with T1D. Fourteen subjects (age [mean±SD] 32.5±7.9 years, BMI 26.5±2.1 kg/m2, HbA1c 7.5±0.8 %) received 2 single doses of 92.2 IU INH via the Afina inhaler (n=13 completing subjects) and 2 injections of an equivalent 12 U LIS dose (n=12). The inhalations were performed by the subjects after a brief training on the correct inhalation technique; injections were given by trained staff. Repeated dosing of INH by the same subject resulted in reproducible insulin absorption and insulin action, with lower or similar variability vs. LIS (Table 1). The response to INH between subjects was more variable than for LIS. The 13.8±8.1% mean relative biopotency of INH was consistent with previous findings. INH was well tolerated, adverse events (8 with INH, 13 with LIS) were mild to moderate in severity. No cough and no acute changes in spirometry were observed with any inhalation. In conclusion, the intra-individual response to self-administration of 501 inhaled insulin is as reproducible as that of s.c. insulin lispro injected by experienced staff. This is a clinically meaningful outcome and enables individual dose titration of 501 insulin in future treatment. Disclosure E. Zijlstra: Employee; Self; Profil. Speaker’s Bureau; Self; Aerami Therapeutics, Eli Lilly and Company, Novo Nordisk A/S. G. Andersen: Employee; Self; Profil Institute for Clinical Research. L. Plum-Moerschel: Other Relationship; Self; Eli Lilly and Company, Novo Nordisk A/S. M. Rhodes: None. R. Patton: None. B. Bueche: None. M. Kuo: None. T.H. Le: None. B.J. Stedman: None. J.S. Patton: Board Member; Self; Dance Biopharm Holdings, Inc.

1004-P: Oral Insulin (ORMD-0801) Effects on Glucose Parameters in Uncontrolled T2DM on OADs

ORMD-0801 is a novel oral human insulin formulation which can potentially improve acceptance and adherence, as well as glycemic control by increasing hepatic insulin exposure, without increasing hypoglycemia risk. This randomized, placebo-controlled, multi-center, phase 2b 12-week study, in 273 T2DM subjects, on metformin ±1-2 oral antidiabetic drugs (OADs), including sulfonylurea, compared placebo vs. 32 mg ORMD-0801 capsules administered once daily (QD) at bedtime, or two (BID) or three (TID) times daily at bedtime, and 30-40 min before breakfast/lunch. Blinded continuous glucose monitoring (CGM) was performed during a two-week run-in period and over the last two weeks of treatment. HbA1c levels improved from baseline for all active treatment regimens (Table). Glucose AUC improved from baseline following QD and BID treatment. Hypoglycemia rates were low and relatively similar to the placebo arm, and body weight was unchanged in all treatment arms. Frequency of drug-related adverse events was similar across treatment groups. In conclusion, ORMD-0801 elicited clinically significant HbA1c reductions among T2DM patients inadequately controlled on standard therapies, without increasing hypoglycemia rates, or inducing severe or serious adverse effects or significant weight gain. Disclosure R. Eldor: Advisory Panel; Self; Oramed. Speaker’s Bureau; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Medtronic, Merck Sharp & Dohme Corp., Novo Nordisk Inc., Sanofi. Stock/Shareholder; Self; Oramed. G. Fleming: Consultant; Self; ADOCIA, Biocon, Caladrius Biosciences, Inc., Dance Biopharm Holdings, Inc., Diasome Pharmaceuticals, Inc., Intarcia Therapeutics, MannKind Corporation, NuSirt Biopharma, Inc., Oramed Pharmaceuticals, Sanofi, Tolerion, Inc. J. Neutel: None. K.E. Homer: None. M. Kidron: Employee; Self; Oramed Pharmaceuticals. Stock/Shareholder; Self; Oramed Pharmaceuticals. J. Rosenstock: Research Support; Self; AstraZeneca, Bristol-Myers Squibb, Genentech, Inc., GlaxoSmithKline plc., Lexicon Pharmaceuticals, Inc., Oramed Pharmaceuticals, PegBio Co., Ltd., Pfizer Inc., REMD Biotherapeutics. Other Relationship; Self; Applied Therapeutics, Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company, Intarcia Therapeutics, Janssen Pharmaceuticals, Inc., Novo Nordisk Inc., Sanofi.

Exploring the complex map of insulin polymorphism: a novel crystalline form in the presence of m-cresol.

In this study, the first crystal structure of a novel crystal form of human insulin bound to meta-cresol in an acidic environment is reported. The combination of single-crystal and powder X-ray diffraction crystallography led to the detection of a previously unknown monoclinic phase (P21). The structure was identified from the powder patterns and was solved using single-crystal diffraction data at 2.2 Å resolution. The unit-cell parameters at pH 6.1 are a = 47.66, b=70.36, c=84.75 Å, β = 105.21°. The structure consists of two insulin hexamers per asymmetric unit. The potential use of this insulin form in microcrystalline drugs is discussed.

COMPUTATIONAL DESIGN OF NOVEL ORAL INSULIN CONJUGATES FOR THE DEVELOPMENT OF SOLID ORAL INSULIN DOSAGE FORMS

Objective: Insulin is used to manage diabetes mellitus by subcutaneous injections regulary, that has become a part of patients life and is distressing for patients. Hence, we anticipate to get rid of the usage of subcutaneous injections completely from the medicine world and would make the diabetic people to say as “farewell to painful injections.”
 Methods: Virtual screening method (for selection of carriers), Toxtree tool (for toxicity evaluation of carriers), and discovery studio tool (for pharmacophore designing, absorption, distribution, metabolism, excretion, and toxicity [ADMET] analysis, designing oral insulin conjugates (OICs), and interaction studies between insulin receptor and OICs) were used for proposed study.
 Results: We have screened 14 competent drug delivering molecules (DDMs) from seven chemical compound databases. The ADMET and pharmacophoric properties of DDMs were analyzed by drug-informatics’ tools. Consequently, the DDMs were mono, di, and poly conjugated by covalent bonding with various binding sites of monomeric and hexameric form of human insulin and insulin-lispro (Humalog®) individually; and novel OICs were generated. Its binding efficiency and biological activity with insulin-receptor were determined.
 Conclusion: Inulin and Vitamin-B1 are the novel, safe, and proficient carriers for oral delivery of insulin. Insulin lispro is the remarkable option for oral delivery than other insulin forms.

1085-P: Dance 501 Inhaled Human Insulin (INH): Linear Dose Response, Earlier Onset of Action, and Higher Early Effect than s.c. Insulin Lispro (LIS)

Dance 501 is a novel liquid formulation of human insulin for inhalation with the Dance 501 inhaler. This randomized, cross-over trial investigated the pharmacodynamic (PD) action and safety of INH in its final formulation vs. LIS during 10-hour glucose clamps in non-smoking subjects with T2D. LIS was injected s.c. at low (12 U), medium (24 U) and high (48 U) doses, INH was inhaled at equivalent doses, assuming a 13% relative biopotency. Twenty-two subjects completed all 6 single dose administrations. INH and LIS demonstrated a linear dose-response relationship and comparable total PD action (AUC_GIR0-10h). Median relative biopotency of INH was 12.3 to 13.0%. PD profiles (Figure) show a more rapid onset of action for equivalent doses of INH vs. LIS (median differences 6.5 to 20 min, p<0.02) and greater action in the first hour after administration (median differences 19.7 to 48.2 mg/kg, p<0.05). No safety issues and no cough were observed with any inhalation. These results nicely confirm and extend to higher doses the results of a previous PK/PD study in type 2 patients with an earlier version of the Dance 501 device and formulation (Zijlstra et al. Diabetes 2015; 64(Suppl 1): A248). In conclusion, Dance 501 showed favorable glucodynamic properties vs. LIS and excellent tolerability and may therefore become a clinically meaningful alternative to rapid-acting insulin injectables. Disclosure E. Zijlstra: Speaker's Bureau; Self; Novo Nordisk A/S. L. Plum-Moerschel: None. M. Ermer: None. O. Klein: None. L. Porter: Other Relationship; Self; Dance Biopharm Holdings Inc., Eiger BioPharmaceuticals. B. Bueche: Employee; Self; Dance Biopharm Holdings Inc. M. Kuo: Employee; Self; Dance Biopharm Holdings Inc. T. Le: None. B.J. Stedman: None. J.S. Patton: Other Relationship; Self; Dance Biopharm Holdings Inc. Funding Dance Biopharm Holdings Inc.

Identification of N-Terminally Truncated Derivatives of Insulin Analogs Formed in Pharmaceutical Formulations

PurposeIsolation and identification of unknown impurities of recombinant insulin lispro (produced at IBA) formed during accelerated stability testing of pharmaceutical solutions. For comparative purposes also commercially available formulations of recombinant human insulin (Humulin S®; Lilly), recombinant insulin lispro (Humalog®; Lilly), recombinant insulin aspart (NovoRapid® Penfill®; Novo Nordisk), recombinant insulin detemir (Levemir®; Novo Nordisk) and recombinant insulin glargine (Lantus®; Sanofi-Aventis) were analyzed.MethodsThe impurities of insulin analogs were isolated by RP-HPLC and identified with peptide mass fingerprinting using MALDI-TOF/TOF mass spectrometry.ResultsThe identified derivatives were N-terminally truncated insulin analog impurities of decreased molecular mass of 119, 147 and 377 Da related to the original protein. The modifications resulting in a mass decrease were detected at the N-terminus of B chains of insulin lispro, insulin aspart, human insulin, insulin glargine, insulin detemir in all tested formulations. To our knowledge it is the first time that these impurities are reported.ConclusionsThe following derivatives formed by truncation of the B chain in insulin analogs were identified in pharmaceutical formulations: desPheB1-N-formyl-ValB2 derivative, desPheB1 derivative, pyroGluB4 derivative.

Porous reduced graphene oxide modified electrodes for the analysis of protein aggregation. Part 1: Lysozyme aggregation at pH 2 and 7.4

Protein instability due to misfolding and aggregation is of big concern for protein based therapeutics because it impacts the bioavailability and immunogenicity of such drugs. The development of simple and cost-effective methods for the analysis of pharmaceutical formulations, indicating the presence or absence of protein aggregates, is consequently of high importance. This work proposes a novel electrochemical interface based on porous reduced graphene oxide coated glassy carbon electrode (GC/prGO) allowing for the early and sensitive identification of protein aggregation by following the change in the oxidative current of the proteins. The novelty of this work lies in the exploration of the ability of GC/prGO interfaces to capture different aggregation behaviors. Lysozyme is used as a model to follow by electrochemistry its aggregation at two pH values, pH 2 and pH 7.4, leading to the formation of amyloid and amorphous aggregates, respectively. Comparing the oxidation peak of lysozyme by differential pulse voltammetry (DPV) for different electrode architectures allowed validating the higher sensitivity of the GC/prGO interface versus bare glassy electrodes or electrodes coated with non-porous reduced graphene oxide. Parallel experiments were performed by fluorescence with thioflavin T, size exclusion chromatography and Atomic Force Microscopy (AFM) imaging. These tests further highlighted the usefulness of GC/prGO electrode to visualize in a fast and reliable manner the changes in the protein structure and the differences between the processes occurring at pH 2 and pH 7.4. In particular, the ability to emphasize changes related to the first steps in aggregation that could be indicative of the aggregation course, recommend the GC/prGO electrode in combination with DPV as a new analytical tool for aggregation studies of biopharmaceuticals. Part 2 of this work will demonstrate later the utility of this approach for the analysis of a fast acting injectable human insulin formulation, Humulin R, used for diabetes treatment as well as for calcitonin.

Poor glycemic control attributable to insulin antibody in a fulminant type 1 diabetes patient under hemodialysis: successful treatment with double filtration plasmapheresis and prednisolone

BackgroundInsulin antibody appears in approximately 40% of insulin users using human insulin and insulin analog formulations. Insulin antibody, characterized by high affinity and low insulin antibody binding capacity (% bound), rarely causes clinical problems including poor glycemic control. Recently, the insulin antibody against human insulin causing deterioration of glycemic control, characterized by low affinity and high insulin antibody binding capacity similar to those observed in insulin autoimmune syndrome, has been reported.Case presentationA 62-year-old male developing fulminant type 1 diabetes showing rapid hyperglycemia (602 mg/dL) and normal range of glycoalbumin (GA) (18.4%) in the introductory stage of hemodialysis (HD) was transferred to our hospital 3 weeks after the onset of diabetes. He had been administered insulin intensification therapy in other medical clinic; however, his glycemic control remained poor, showing ketoacidosis. After hospitalization, he was initially administered intravenous insulin for 2 days and then switched to subcutaneous intensification therapy (24 U/day); the dose was gradually increased maximum of 72 U/day within 2 months. His glycemic control deteriorated despite high-dose insulin administration and change in the insulin type. Insulin antibody was detected, insulin antibody binding capacity (% bound) was of a high value (89.5%), GA was 32.9%, and casual blood glucose widely changed in continuous glucose monitor, suggesting that unstable blood glucose fluctuations caused by insulin antibody; therefore, treatment with double filtration plasmapheresis (DFPP) and prednisolone (PSL) was performed. The insulin dose reduced, and glycemic control improved from 1 month after the commencement of treatment. Subsequently, insulin antibody level remained high; however, after 6 months, % bound declined (20.9%), and glycemic control improved (GA 23.7%) by PSL (gradually reduced to 5 mg/day). The Scatchard plots also indicated the effectiveness with DFPP and PSL therapy.ConclusionsThe authors present a unique case of fulminant type 1 diabetes developing during the HD introduction period. Although this is a rare complication, physicians specializing in dialysis therapy should also keep in mind that this is a life-threatening complication if the treatment is delayed and that concomitant therapy with apheresis such as DFPP and long-term steroids is effective in some cases when glycemic control deteriorates attributable to insulin antibody.

Structure, Aggregation, and Activity of a Covalent Insulin Dimer Formed During Storage of Neutral Formulation of Human Insulin

A specific covalently linked dimeric species of insulin high molecular weight products (HMWPs), formed during prolonged incubation of a neutral pharmaceutical formulation of human insulin, were characterized in terms of tertiary structure, self-association, biological activity, and fibrillation properties. The dimer was formed by a covalent link between A21Asn and B29Lys. It was analyzed using static and dynamic light scattering and small-angle X-ray scattering to evaluate its self-association behavior. The tertiary structure was obtained using nuclear magnetic resonance and X-ray crystallography. The biological activity of HMWP was determined using 2 in vitro assays, and its influence on fibrillation was investigated using Thioflavin T assays. The dimer's tertiary structure was nearly identical to that of the noncovalent insulin dimer, and it was able to form hexamers in the presence of zinc. The dimer exhibited reduced propensity for self-association in the absence of zinc but significantly postponed the onset of fibrillation in insulin formulations. Consistent with its dimeric state, the tested species of HMWP showed little to no biological activity in the used assays. This study is the first detailed characterization of a specific type of human insulin HMWP formed during storage of a marketed pharmaceutical formulation. These results indicate that this specific type of HMWP is unlikely to antagonize the physical stability of the formulation, as HMWP retained a tertiary structure similar to the noncovalent dimer and participated in hexamer assembly in the presence of zinc. In addition, increasing amounts of HMWP reduce the rate of insulin fibrillation.

A case of type 1 diabetes mellitus with which localized insulin allergy was markedly alleviated by switching to insulin glulisine.

There have been numerous reports of allergies with causes such as differences in the amino acid sequences between human insulin and insulin extracted from porcine and bovine pancreas. Insulin allergy leads to deterioration of blood glucose control and decrease in quality of life. However, the availability of genetically recombinant human insulin formulations has led to insulin allergy becoming uncommon [1], and such allergies also occur much more rarely when insulin analog formulations are administered. The present report is about a patient who showed allergic reactions to various insulin formulations, but showed only a weak allergic reaction to insulin glulisine, so the symptoms of insulin allergy ceased when treatment was switched to insulin glulisine.

AFREZZA® (insulin human) Inhalation Powder: A Review in Diabetes Mellitus.

Afrezza® (insulin human) inhalation powder is a rapid-acting Technosphere® insulin (TI) administered via a breath-powered oral inhaler to patients with diabetes requiring prandial insulin. TI, a dry powdered formulation of recombinant human insulin adsorbed onto a proprietary carrier, is designed to deliver insulin to the deep lung, at the level of the alveoli, where it is absorbed into the systemic circulation. In a randomized, open-label, multinational, phase III trial (trial 171) in type 1 diabetes (T1DM) patients, prandial TI via the Gen2 inhaler provided noninferior glycated haemoglobin (HbA1c) lowering compared with prandial subcutaneous insulin aspart. Although TI was associated with less HbA1c lowering, it provided significantly lower fasting plasma glucose levels and significantly less hypoglycaemia and bodyweight gain compared with insulin aspart. In a randomized, double-blind, placebo-controlled, multinational, phase III trial (trial 175) in type 2 diabetes (T2DM) patients, prandial TI via the Gen2 inhaler provided superior HbA1c lowering compared with inhaled placebo. Cough was the most commonly occurring non-hypoglycaemia adverse event across both studies. In a pooled analysis of tolerability data from phase II and III studies, the most commonly occurring non-hypoglycaemia adverse events in T1DM and T2DM patients were cough and throat pain/irritation. However, cough was generally mild, dry and decreased over time. In addition, treatment with TI was associated with positive patient-reported outcomes. Insulin human inhalation powder is an effective and generally well-tolerated agent for the prandial treatment of hyperglycaemia in T1DM and T2DM patients and may provide a solution to insulin initiation barriers such as injection phobia, concerns of bodyweight gain and concerns of hypoglycaemia.

Supercritical Fluid Particle Design of DPI Formulations (Review).

Pulmonary drug delivery (PDD) with dry powder inhaler (DPI) has rapidly developed for the treatment of local and systemic diseases, which targets the delivery of fine drug particles into the deep lung surface by combining technologies of fine drug particle formulation, small dose packaging and suitable inhaler, where by each contributes to the overall aerodynamic performance. The basic requirements of DPI formulation are an excellent aerodynamic performance, including particle size distribution within 1-5 μm, suitable morphology and electrostatic charge, low surface energy, high deposition rate and long shelf life stability. The strategy of DPI formulation is shifting from carrier-based to carrier free, from single drug to drug combination, from microparticles to nanoparticles and from small molecules to biomacromolecules. Making such DPI formulation is a big challenge for conventional pharmaceutical techniques. Fortunately, an emerging technology of supercritical fluid particle design (SCF PD) provides a powerful platform for DPI formulation since it runs single step operation at near ambient temperature to minimize the potential damage of delicate active ingredients and to ensure the consistency of the DPI formulation. Combining with our research experiences in DPI formulation of budesonide and recombinant human insulin, this review focus on the most recent development of DPI formulation using SCF PD technology, which can well control and tune the particle size, morphology and surface properties through different design routes (nanoparticles or microparticles, polymorphic particles, composite particles and bio-drug particles), and hence enable prominent enhancement aerodynamic performance and pulmonary deposition of such inhaled dry powders. Also considered within this review is the progress of the industrialization of SCF PD processes for DPI formulation.

New insulins and insulin therapy.

Aims Prandial treatment with human regular insulin for diabetes may result in early postprandial hyperglycaemia and late hypoglycaemia due to its slow onset and long duration of action. This study compared injections of recombinant human insulin (rHI) formulated with recombinant human hyaluronidase [rHuPH20] (INSULIN-PH20) to insulin lispro for prandial treatment in subjects with type 1 diabetes (T1D). Methods After a 1-month run-in period using twice-daily insulin glargine (or usual basal insulin therapy for pump users) with prandial lispro, 46 subjects with T1D (42 ± 13 years; body mass index: 26 ± 4 kg/m2; A1c: 6.8 ± 0.5%) were assigned to INSULIN-PH20 or lispro in a random sequence for two consecutive, 12-week periods as the prandial insulin in an intensive treatment regimen. Results The mean glycaemic excursion for INSULIN-PH20 (0.96 ± 2.00 mmol/l) was comparable (p = 0.322) to lispro (0.80 ± 1.95 mmol/l). The 8-point self-monitored blood glucose profiles were also comparable in the two groups. Good glycaemic control (A1c) was maintained for both treatments at 12 weeks (INSULIN-PH20: 7.0 ± 0.5%; lispro: 6.9 ± 0.6%). Overall rates of hypoglycaemia (≤3.9 mmol/l) were 24 events per patient per 4 weeks for INSULIN-PH20 and 22 events for lispro. There were no significant differences in adverse events or immunogenicity between treatments and both treatments were well tolerated. Conclusions Unlike commercially available formulations of regular human insulin, a formulation of rHI with rHuPH20 was comparable to lispro for postprandial glucose excursions in a basal-bolus treatment regimen for T1D patients. Glycaemic control, safety and tolerability profiles were comparable for both treatments.

Purification and identification of high molecular weight products formed during storage of neutral formulation of human insulin.

To identify High Molecular Weight Products (HMWP) formed in human insulin formulation during storage. Commercial formulation of human insulin was stored at 37°C for 1year and HMWP was isolated using preparative size exclusion chromatography (SEC) and reverse phase (RP) chromatography. The primary structure of the isolated species was analysed using liquid chromatography mass spectrometry (LC-MS) and tandem mass spectrometry (MS/MS). To test the hypothesis that amino groups of insulin are involved in HMWP formation, the HMWP content of various formulations spiked with amine compounds or formulations of insulin with modified amino groups was measured. More than 20 species of HMWP were observed and 16 species were identified using LC-MS. All identified species were covalent dimers of human insulin linked via A21Asn and B29Lys, formed via the formation of an anhydride intermediate at A21Asn. Two types of HMWP were identified, with the covalent link in the open or closed (succinimidyl) form. Some species also contained single deamidation at B3 or the desPhe(B1)-N-oxalyl-Val(B2) modification. Reduced rate of HMWP formation was observed after addition of L-lysine, L-arginine or piperazine or when insulin analogues with methylated N-terminals and side chain amines and A21Gly mutation were used. Formulations of human insulin without zinc and m-cresol were found to contain a different pool of HMWP. HMWP formed in formulation of human insulin at pH 7.4 with zinc and m-cresol consists primarily of covalent dimers linked via A21Asn and B29Lys. Insulin formulation properties determine the amount and identity of formed HMWP.