Human Insulin Analogues Research Articles

MYL1501D Insulin Glargine: A Review in Diabetes Mellitus.

Subcutaneous MYL1501D insulin glargine 100 U/mL (hereafter referred to as MYL1501D insulin glargine) [Semglee®] is a long-acting human insulin analogue approved as a biosimilar of insulin glargine 100 U/mL (hereafter referred to as reference insulin glargine 100 U/mL) [Lantus®] in various countries, including those of the EU for the treatment of diabetes mellitus in patients aged ≥ 2 years, as well as Japan for diabetes where insulin therapy is indicated. MYL1501D insulin glargine has similar physicochemical characteristics and biological properties to those of EU- and US-sourced reference insulin glargine 100 U/mL, with the bioequivalence of pharmacodynamic and pharmacokinetic parameters between these agents shown in adults with type 1 diabetes. Once-daily MYL1501D insulin glargine demonstrated noninferior glycaemic efficacy to that of once-daily reference insulin glargine 100 U/mL in adults with type 1 or 2 diabetes, with its glycated haemoglobin-lowering benefits maintained over the longer-term (52 weeks) and unaffected by previous insulin exposure. Switching between MYL1501D insulin glargine and reference insulin glargine 100 U/mL did not appear to impact glycaemic efficacy in adults with type 1 diabetes. MYL1501D insulin glargine was well tolerated, demonstrating a safety and immunogenicity profile similar to that of reference insulin glargine 100 U/mL in patients with type 1 and 2 diabetes, and in those with type 1 diabetes switching between the two agents. As expected, hypoglycaemia was the most frequently reported treatment-emergent adverse event. Thus, MYL1501D insulin glargine provides an effective biosimilar alternative for patients requiring insulin glargine therapy.

Clinically Relevant Insulin Degludec and its Interaction with Polysaccharides: A Biophysical Examination.

Protein polysaccharide complexes have been widely studied for multiple industrial applications and are popular due to their biocompatibility. Insulin degludec, an analogue of human insulin, exists as di-hexamer in pharmaceutical formulations and has the potential to form long multi-hexamers in physiological environment, which dissociate into monomers to bind with receptors on the cell membrane. This study involved complexation of two negatively charged bio-polymers xanthan and alginate with clinically-relevant insulin degludec (PIC). The polymeric complexations and interactions were investigated using biophysical methods. Intrinsic viscosity [η] and particle size distribution (PSD) of PIC increased significantly with an increase in temperature, contrary to the individual components indicating possible interactions. [η] trend was X > XA > PIC > A > IDeg. PSD trend was X > A > IDeg > XA > PIC. Zeta (ζ)- potential (with general trend of IDeg < A < XA < X ≈ PIC) revealed stable interaction at lower temperature which gradually changed with an increase in temperature. Likewise, sedimentation velocity indicated stable complexation at lower temperature. With an increase in time and temperature, changes in the number of peaks and area under curve were observed for PIC. Conclusively, stable complexation occurred among the three polymers at 4 °C and 18 °C and the complex dissociated at 37 °C. Therefore, the complex has the potential to be used as a drug delivery vehicle.

Hundred Years of Insulin Therapy: Purified Early Insulins.

The discovery of insulin has changed dramatically the outcome of patients with type 1 diabetes, giving them the possibility to survive. This is of particular concern due to the fact that type 1 diabetes most frequently occurs in children who were destined to die in ketoacidosis coma. From insulin discovery to the availability of human insulin and human insulin analogs to be used in diabetes therapy, a series of problems have arisen as the difficulty of insulin purifications, the animal insulin used by the first researches were in fact contaminated by proteins, fats, and other impurities, and the presence of side effects such as allergy, antibodies generation, and lipoatrophy. Data strictly related to the argument have been searched in Pub Med and used. Starting from insulin discovery in 1921 to nowadays, significant efforts have been made by a series of researches to purify animal insulin, discover the molecular structure of human insulin, and develop methods to synthetize human insulin and then insulin analogs. The history of insulin discovery here reported is fascinating; insulin is a hormone, a product of biotechnology, a field of research that saved and save the life of many diabetic patients.

Аллергическая реакция на инсулин детемир у ребенка на фоне базис-болюсной терапии

Objective: To present a case study demonstrating an atypic side effect of insulin — hepersensitivity reaction to insulin detemir, a long-term human insulin analogues. Key Points. At the inpatient paediatric unit, we had an 11-year old girl with symptoms and clinical signs of diabetes mellitus (DM). Following physical examination, the patient was diagnosed with type 1 diabetes (manifestation). She was prescribed intensified basal-bolus therapy with insulin analogues: insulin detemir and insulin aspart. On day 5, the patient developed a hypersensitivity reaction in the site of insulin detemir injections. Replacement of insulin detemir with insulin degludec resolved the allergic reaction. Conclusion. We described a case of a hypersensitivity reaction to intensified insulin therapy with basal insulin analogue detemir. Hypersensitivity reactions to insulin analogues are rare; however they can be life-threatening. The primary management is the use of another insulin analogue. Keywords: diabetes mellitus, allergy, insulin detemir, insulin degludec.

Severe Lipoatrophy in a Patient With Type 2 Diabetes in Response to Human Insulin Analogs Glargine and Degludec: Possible Involvement of CD4 T Cell-Mediated Tissue Remodeling.

Lipoatrophy in response to insulin therapy is a rare condition resulting in a loss of subcutaneous adipose tissue at the site of injection. Since the introduction of humanized insulins, insulin-induced lipoatrophy (IIL) is almost exclusively associated with T1D. IIL was suggested to be an aspect of autoimmunity (1,2), but its underlying pathophysiology is poorly understood. …

Synthesis and Characterization of an A6-A11 Methylene Thioacetal Human Insulin Analogue with Enhanced Stability.

Insulin has been a life-saving drug for millions of people with diabetes. However, several challenges exist which limit therapeutic benefits and reduce patient convenience. One key challenge is the fibrillation propensity, which necessitates refrigeration for storage. To address this limitation, we chemically synthesized and evaluated a methylene thioacetal human insulin analogue (SCS-Ins). The synthesized SCS-Ins showed enhanced serum stability and aggregation resistance while retaining bioactivity compared with native insulin.

Glycemic control in patients with diabetes mellitus on hemodialysis

Achievement of stabilization of carbohydrate metabolism in patients with diabetes mellitus, receiving renal replacement therapy with hemodialysis, is a significant problem in endocrinology. It has to do with multiple factors of this cohort of patients, which affect the level of glycemia, pharmacokinetic of drugs, the efficiency of glycemic control. At the moment, the most efficiency method of glycemic control in patients with type 2 diabetes mellitus on hemodialysis is insulin therapy in the basis - bolus regime by analogues of human insulin. The use of oral hypoglycemic agents is significantly limited. The hemoglobin A1c (HbA1c) remains the main parameter of glycemic control. The simultaneous use of continuous glucose monitoring allows to reveal the true level of glucose of the blood and to carry out the timely correction of therapy in order to achieve targets for glycemic control and to decrease the risk of hypoglycemic episodes. At the moment other glycemic control markers such as glycated albumin and fructosamine are described. However, in routine practice at the moment these indicators are not used due to the lack of sufficient evidentiary base of their use in this cohort of patients.

Development and Validation of Approach for the Detection of Neutralizing Antibodies Against Insulin (Glargine) in Human Blood Plasma

Introduction. IImmunogenicity identification of therapeutic proteins, such as human insulin analogues, is one of the most relevant and significant area in medicine and pharmaceuticals. Determination the possibility of producing neutralizing antibodies to insulin reducing the therapeutic effect of the drug, is an important step to understand the pharmacological profile of the drug. Applying of cell-based methods one allows to determinate neutralizing antibodies to insulin.Aim. Development and validation methods for detection of neutralizing antibodies against insulin in human plasma.Materials and methods. The method is based on the use of the iLiteTM Insulin Assay Ready Cells [1], in the genome of which the firefly luciferase reporter gene is introduced under the control of an insulin-dependent promoter. As the insulin concentration increases, the firefly luciferase expression (Firefly) increases, allowing one to use this cell line to estimate the number of neutralizing antibodies against insulin. For normalization by the number of cells and considering the matrix effect of studied samples, the second reporter gene luciferase Renilla is used, which is expressed under the control of a constitutive promoter. The activity of both luciferases was measured using the DualGlo Luciferase Assay System (Promega) assay [2].Results and discussion. Optimal insulin concentration and plasma/serum dilution were determined to identify neutralizing antibodies to insulin. The long-term stability of neutralizing antibodies to insulin were shown in human plasma for more than 3 months. The developed method was applied in a comparative research of the safety and immunogenicity of insulin analogues (Glargine). Method for the determination of antibodies to insulin was.Conclusion. A method for determination of neutralizing antibodies to insulin in human K2EDTA plasma was developed and validated using iLiteTM Insulin Assay Ready Cells system; based on the binding of the insulin alpha chain to the high-affinity heterodimeric CD220 receptor.

Physico-chemical stability of co-formulation of PEGylated human amylin with insulin

Since the discovery of amylin no combined formulation with insulin has been made available. Amylin or its triple proline analog pramlintide are not compatible in solution with insulin. The drug candidate hAmy-PEG5k is a novel monoPEGylated amylin derivative with improved physicochemical properties and retained similar pharmacological activity compared to free amylin and pramlintide. We have investigated the short- and long-term physicochemical compatibility of hAmy-PEG5k co-formulated with slow-acting human insulin analogs glargine or detemir. While human amylin promptly aggregates over a large range of pH, and both free and in the presence of regular, glargine or detemir insulin, the hAmy-PEG5k analog is stable at these conditions as shown by Thioflavin T (ThT) binding assay. When hAmy-PEG5k (100 or 500 µg/mL) was added to the commercial formulations of either insulin glargine or detemir (95 IU/mL), the combinations remained stable after 6 months stored at 4 °C, as probed by ThT, dynamic light scattering (DLS) measurements and high performance liquid chromatography (HPLC) analyses, confirming the absence of amyloid fibers, minor aggregation products or loss of material. These results suggest hAmy-PEG5k and the insulin analogs glargine and detemir are physicochemically compatible and are candidate ready-to-use fixed-dose combinations.

Role of NIB in connecting science to patients by assuring safe and quality anti diabetic products

In India burden of diabetes is increasing and expected to cross the 100 million mark by 2030. Therefore, the development of national policies for the prevention, treatment, and care of diabetes in line with the sustainable development of health care systems is the need of the hour. National Institute of Biologicals (NIB) responsibly assures and reviews the quality of Biological products to the end users by weeding out “Not-of-Standard-Quality” Biological Medicines from reaching Indian market, thereby safeguarding the Public Health by testing at our state-of-art-laboratory. NIB receives samples from various sources such as State Port offices, Central Drug Standard Control Organization (CDSCO), Stakeholders of biotherapeutic manufacturers, legal &amp; global tender samples. They are tested by pharmacopoeial methods or manufacturer’s method for Key Quality Attributes (KQA). The heterogeneity of products involves human recombinant biotherapeutic Insulin, Insulin Analogues, and Peptides of nearly 20 different types eg. Biphasic Insulin, Glargine, Aspart, Exenatide, Liraglutide, etc. which have their global presence for the antidiabetic patient care. Testing for quality attributes such as Identification, Potency, Purity, Stability, Related Impurity and Safety parameters along with the physiochemical test is carried out to ensure the safety, quality, and effectiveness of the product. In addition, reference standards are also being developed at NIB which serves as the measurement standards and complements the written standards in India pharmacopeia. All the testing is carried out strictly adhering to the timelines so that the quality product is delivered to meet the ever-growing public health challenge&#x0D; NIB maintains quality systems of ISO 17025 standards for continuous improvement, function as Central Drugs Laboratory(CDL). It focuses on a continuous supply of lifesaving biotherapeutics to ensure the success of the Non-Communicable Disease (NCD) programme for Diabetes.

Identification of recombinant human insulin and biosynthetic insulin analogues by multiplexed targeted unlabeled mass spectrometry of proteotypic tryptic peptides

Direct qualitative methods that allow the rapid screening and identification of insulin products during early stages of the drug development process and those already in the market can be of great utility for manufacturers and regulatory agencies and the recent scientific literature describes several methods. Herein, a qualitative proteomic method is presented for the identification of recombinant human insulin and all marketed biosynthetic analogues -insulin lispro, aspart, glulisine, glargine, detemir and degludec- via tryptic digestion and identification of proteotypic peptides for each insulin. Individual insulins were first denatured under reducing conditions and the cysteine residues blocked by iodoacetamide. The proteins were then digested with trypsin and the peptide products separated by reversed phase liquid chromatography on an Ascentis® Express ES-C18 column and detected by positive polarity ESI-MS/MS. The digestion peptides were characterized using a multiplexed MRM approach that monitors the fragmentation of the doubly charged unlabeled precursor ion of each peptide into a collection of signature y and b ions. The MRM transitions for the individual peptides were optimized to allow maximal ionization on a standard triple quadrupole mass spectrometer. All products of the digestion procedure for all insulins were detected with adequate signal intensity except for the C-terminal B30Thr whenever it was present and cleaved and the tryptic B1-3 tripeptide of insulin glulisine. The unique proteotypic peptides identified for each of the insulin analogues coupled with their signature y and b ions permitted the unambiguous verification of all sequence variations and chemical modifications. The elution of the A polypeptide chain for all insulins and the tryptic peptides of the B chain, with the exception of a very few, occurred around the same time point. This underscores the close similarity in the physicochemical properties between the digestion peptides and is consistent with the subtle variations in amino acid sequence among the various insulins. Therefore, the identification and distinction of the different types of insulin based solely on the chromatographic retention time of their respective proteolytic products can be deceptive without proper mass spectrometric analysis and may result in false positives.

SUN-186 Insulin Antibody Associated Severe Insulin Resistance Leading to Uncontrolled Hyperglycemia

Background: Severe insulin resistance is unusual in young patients with type 1 DM and normal BMI. We report a case of severe insulin resistance attributed to the development of insulin antibodies. Clinical Case: An 18 year old male with uncontrolled Type 1 DM was admitted to psychiatry for management of a delusional disorder. His home insulin regimen consisted of degludec 56 units daily and aspart 30 units before meals. He weighed 75 kg with a BMI of 27.5 kg/m2. Initial blood work revealed hyperglycemia with glucose of 549 mg/dl, HbA1C of 10%, C-peptide of <0.1 ng/ml (0.9-7.1 ng/ml), and no evidence of urinary ketones, acidemia, hyperosmolarity or thyroid dysfunction. He was started on a basal bolus regimen with a total daily dose of 0.9 units/kg. His hyperglycemia persisted despite rising standing insulin doses and metformin 1000 mg twice daily was added with minimal improvement in glycemic control. Due to persistent hyperglycemia despite rising insulin doses, insulin antibody titers were measured and found to be elevated at 4.4 U/ml (<0.4 U/ml.) When his psychiatric symptoms stabilized, he was discharged on a basal bolus regimen with total daily dose of 2.6 units/kg and metformin 1000 mg twice a day with improved glycemic control. Conclusion: Development of insulin antibodies is known to occur in type 1 diabetics using animal preparations of insulin but far less commonly in patients receiving recombinant human insulin or insulin analogues. Factors related to their development include the type of insulin (animal insulin having higher immunogenicity), mode of delivery (increased levels seen with insulin pump use and more frequent injections) and predisposing genetic factors such as certain HLA types. In some individuals, these antibodies may be clinically insignificant while others may develop severe insulin resistance with highly variable glycemic control. Often these patients experience marked hypoglycemia at night yet are hyperglycemic during the day. Treatment options include switching insulin formulations, insulin sensitization with metformin or thiazolidinediones, stimulation of insulin production with sulfonylureas or GLP-1 receptor agonists and immunosuppression with glucocorticoids, mycophenolate or cyclophosphamide. In patients who fail these treatments, plasmapheresis has been described as an alternative. In conclusion, the presence of insulin antibodies should be considered in patients with severe hyperglycemia despite increasing doses of insulin.

Identification of Lysine Misincorporation at Asparagine Position in Recombinant Insulin Analogs Produced in E. coli

PurposeIdentification of human insulin analogs’ impurity with a mass shift +14 Da in comparison to a parent protein.MethodsThe protein sequence variant was detected and identified with the application of peptide mapping, liquid chromatography, tandem mass spectrometric analysis, nuclear magnetic resonance spectroscopy (NMR) and Edman sequencing.ResultsThe misincorporated lysine (Lys) at asparagine (Asn) position A21 was detected in recombinant human insulin and its analogs.ConclusionsAlthough there are three asparagine residues in the insulin derivative, the misincorporation of lysine occurred only at position A21. The process involves G/U or A/U wobble base pairing.

Continuous subcutaneous insulin infusion vs modern multiple injection regimens in type 1 diabetes: an updated meta-analysis of randomized clinical trials.

Meta-analyses of clinical trials comparing CSII with traditional insulin injections usually show a small, but significant advantage of CSII with respect to HbA1c and risk of severe hypoglycemia. On the other hand, CSII is associated with a small, but relevant risk of ketoacidosis, mainly due to malfunction of insulin pump and/or catheter occlusion. During last time, the technology of insulin pumps and infusion sets has improved as the profound evolution in type and schemes with traditional insulin injections. Aim of the present study is to update previous meta-analyses comparing CSII with traditional insulin injections in subjects with type 1 diabetes. Specific subgroup analyses were designed for assessing the effects of CSII in comparison with basal-bolus MDI, with short-acting analogues as bolus and long-acting analogues as basal insulin. In addition, an exploratory analysis was performed to verify the effect of CSII in insulin-naïve patients with type 1 diabetes. The present analysis includes all randomized clinical trials comparing CSII with traditional injections in type 1 diabetes, with a duration of at least 12 weeks. Animal studies were excluded, whereas no language or date restriction was imposed. If duplicate publications of a single trial were present, the paper containing more adequate information was considered as principal publication. In trials comparing CSII with basal-bolus MDI, performed before the introduction of rapid-acting analogues, regular human insulin was used for CSII, and as prandial insulin in control groups. CSII was associated with a significant reduction of A1c, in comparison with MDI, irrespective of the use of either human insulin or rapid-acting analogues. However, in trials with rapid-acting analogue the advantage of CSII was significantly smaller than in trials with regular human insulin (HbA1c difference: - 0.29[- 0.46; - 0.13] vs - 1.93[- 1.84; - 0.42]%; p = 0.02). Different rapid-acting analogues provided similar results (HbA1c reduction vs MDI: - 0.25 [- 0.48; - 0.02]%, p = 0.03, and - 0.29 [- 0.49; - 0.09]%, p = 0.005, for lispro and aspart, respectively). In addition, in trials comparing CSII with basal-bolus MDI, CSII reduced HbA1c to a similar extent irrespective of the use of either NPH or long-acting analogues as basal insulin in the control groups (HbA1c reduction vs MDI: - 0.31 [- 0.55; - 0.06]%, p = 0.01, and - 0.20 [- 0.38; - 0.03]%, p = 0.02, for NPH and long-acting analogues, respectively. With respect to severe hypoglycemia, CSII did not produce a significant reduction of risk in comparison with traditional insulin injections. Conversely, CSII was associated with a significant increase in the incidence of reported diabetic ketoacidosis (DKA). Notably, the increased risk of DKA was significant in trials comparing CSII with conventional insulin therapy, whereas only a nonsignificant trend toward an increased risk was observed in comparisons with basal-bolus MDI. Only two trials comparing CSII with basal-bolus MDI, both using rapid-acting analogues, were performed on insulin-naïve type 1 diabetic patients. When those two trials were analyzed separately, CSII did not produce any relevant effect on HbA1c (difference from control: - 0.10[- 0.38; + 0.17]%; p = 0.46). No meta-analysis could be performed on either severe hypoglycemia or DKA, which were not reported by one of the two trials. CSII seems to produce a small improvement in HbA1c in patients with type 1 diabetes inadequately controlled with MDI. This apparent effect, which could be partly due to publication bias, is smaller when MDI is properly performed using basal-bolus schemes with short-acting insulin analogues. Other outcomes different from HbA1c (such as quality of life) could be relevant for the choice of CSII instead of MDI. In addition, further studies are needed to better define the profile of patients who could benefit most from CSII.

Comparative Study between the Glycemic Control of Human Insulin and Insulin analogue In Sample of Iraqi Type 1 Diabetic Children and Adolescents

Insulin analogue introduced to offer insulin replacement therapy mimic to normal human physiology. The aim of this study is to compare between the glycemic control of insulin analogue and conventional human insulin in a sample of type 1 diabetic Iraqi children and adolescents. Forty type 1 diabetic Iraqi children and adolescents age between (6-18) years enrolled in this study and divided into two groups. Group 1 contains 20 patients switched from human insulin to insulin analogue. Group 2 contain 20 patients continued with conventional human insulin. The results showed that both therapies reduced fasting plasma glucose (FPG) and glycated hemoglobin (HbA1c %) but insulin analogue treated group had highly significant reduction. Both therapies did not affect on blood urea nitrogen and serum creatinine. Human insulin reduced triglyceride (TG) and very low-density lipoprotein (VLDL) significantly. The parameters measures at baseline and after three months of treatments. In conclusion insulin analogue is superior over conventional human insulin in reducing glycemic indices in a sample of type 1 diabetic Iraqi children and adolescents.

Extreme Adhesion Activity of Amyloid Fibrils Induces Subcutaneous Insulin Resistance.

Insulin-derived amyloidoma, also called an insulin ball, is a skin-related complication of insulin therapy caused by repeated insulin injections at the same site, where native folded insulin changes into amyloid fibrils and forms a mass with a granulomatous reaction. Insulin-derived amyloidoma is a clinically important condition because of its association with subcutaneous insulin resistance, but the precise effect and mechanism of the insulin absorption impairment have not been clarified. We generated insulin-derived amyloidomas in mouse skin, with the amyloidomas large enough to perform insulin tolerance tests in the mass by repeated injections of highly concentrated insulin amyloid fibrils. We demonstrated that the insulin-derived amyloidomas inhibit insulin absorption. By simultaneous administration of insulin and insulin amyloid fibrils, we showed that this effect is due to the amyloid fibril itself in the absence of a granulomatous reaction. In vitro studies revealed that insulin amyloid fibrils have extremely strong adhesion to native human insulin and various insulin analogs. Furthermore, we showed that native insulin that had adhered to insulin amyloid forms amyloid fibrils at physiological pH. These results suggest that the extreme adhesion of insulin amyloid to native insulin is the main mechanism of impaired insulin absorption and amyloidoma growth.

Comparative Study between the Glycemic Control of Human Insulin and Insulin analogue In Sample of Iraqi Type 1 Diabetic Children and Adolescents

Insulin analogue introduced to offer insulin replacement therapy mimic to normal human physiology. The aim of this study is to compare between the glycemic control of insulin analogue and conventional human insulin in a sample of type 1 diabetic Iraqi children and adolescents. Forty type 1 diabetic Iraqi children and adolescents age between (6-18) years enrolled in this study and divided into two groups. Group 1 contains 20 patients switched from human insulin to insulin analogue. Group 2 contain 20 patients continued with conventional human insulin. The results showed that both therapies reduced fasting plasma glucose (FPG) and glycated hemoglobin (HbA1c %) but insulin analogue treated group had highly significant reduction. Both therapies did not affect on blood urea nitrogen and serum creatinine. Human insulin reduced triglyceride (TG) and very low-density lipoprotein (VLDL) significantly. The parameters measures at baseline and after three months of treatments. In conclusion insulin analogue is superior over conventional human insulin in reducing glycemic indices in a sample of type 1 diabetic Iraqi children and adolescents.

Production costs and potential prices for biosimilars of human insulin and insulin analogues

IntroductionHigh prices for insulin pose a barrier to treatment for people living with diabetes, with an estimated 50% of 100 million patients needing insulin lacking reliable access. As insulin analogues...

Activation of insulin receptors and IGF-1 receptors in COLO-205 colon cancer xenografts by insulin and insulin analogue X10 does not enhance growth under normo- or hypoglycaemic conditions.

Recent studies with normal rats and mouse allograft models have reported that insulin and insulin analogues do not activate the IGF-1 receptor in vivo, and that this characteristic therefore cannot be responsible for the increased incidence of mammary tumours observed for the insulin analogue X10 in chronic toxicity studies with Sprague Dawley rats. This is in clear contrast to reports of insulin and insulin analogues in vitro. Clarification of this is important for understanding the mechanisms behind possible growth-promoting effects of insulin analogues, and will have implications for the development of novel insulin analogues. We established a xenograft model in BALB/c nude mice with the human colon cancer cell line COLO-205, which expresses human insulin and IGF-1 receptors, and explored the acute and chronic effects of treatment with supra-pharmacological doses of human insulin, insulin analogue X10 and human IGF-1. With a novel antibody, acute IGF-1 receptor activation was also examined in various tissues from normal rats treated with human insulin, insulin analogue X10 or human IGF-1. Finally, the effects of pharmacologically relevant doses of human insulin and insulin analogue X10 on receptor activation and growth of COLO-205 xenograft were explored in BALB/c nude mice with alloxan-induced hyperglycaemia. In normal rats and in BALB/c nude mice bearing a COLO-205 cell xenograft, treatment with supra-pharmacological doses of human insulin, insulin analogue X10 or human IGF-1 resulted in activation of insulin receptors as well as IGF-1 receptors. Treatment of diabetic nude mice with pharmacologically relevant doses of human insulin or insulin analogue X10, which decreased blood glucose from hyperglycaemic levels to the normoglycaemic range, did not increase IGF-1 receptor activation. Furthermore, repeated treatment with supra-pharmacological as well as pharmacological doses of human insulin or insulin analogue X10 did not influence the growth of COLO-205 xenografts. This study demonstrates that activation of IGF-1 receptors in cancer cells by insulin and insulin analogues cannot be considered as a purely in vitro phenomenon. It does occur in vivo in animal models, although only after treatment with supra-pharmacological doses. Furthermore, treatment with insulin or insulin analogue X10 did not influence the growth of COLO-205 xenografts under normo- or hypoglycaemic conditions. Further studies are needed before a conclusion can be reached on whether IGF-1 receptor activation by insulin analogues correlates with increased growth in vivo.

Electron Microscopic Findings of Insulin-Induced Local Amyloidosis

Introduction: Insulin-induced local amyloidosis (ILA) was firstly described in 1983, but still underdiagnosed. Here we describe clinicopathological and electron microscopic findings of a case with ILA. Case: A 52-year-old male with type 2 diabetes mellitus for 14 years, on basal-bolus insulin (i.e., insulin lispro and glargine), had noticed 2x2 cm mass at injection site. MRI showed a fibrous mass at the subcutaneous tissue. Punch biopsy of the lesion revealed amyloid deposition surrounded by giant cells. Immunohistochemistry studies proved that amyloid material is insulin. Electron microscopic evaluation (EM) revealed both extra-, and intracellular amyloid fibrils (Figure 1A,B). Discussion: Local amyloid deposition due to insulin injection could be seen in both types of diabetes. Despite earlier reports were associated with porcine insulin, subsequent publications linked also with human insulin and recombinant insulin analogues. High doses of insulin and inappropriate insulin injections are the major risk factors. Extracellular amyloid deposits in ILA arises primarily from exogenous insulin, but the precise mechanism of pathogenicity remains to be established. There are only few case reports of ILA with EM. To the best of our knowledge, here in this case we have firstly showed intracellular accumulation of those amyloid fibrils as well as extracellular compartment, which might have further clinical implications. Disclosure S. Sendur: None. K. Katipoglu: None. O. Kockara: None. Ö. Gököz: None. F. Kaymaz: None. S. Dagdelen: None.