SBIR-STTR Award

We seek to develop an ultra-fast, zinc-free insulin analog formulation for the treatment of diabetes mellitus. An ultra-fast pharmacokinetic-dynamic (PK/PD) profile promises to enable superior performance of pump therapy (continuous subcutaneous insulin infusion) with enhanced safety and more robust integration with glucose- sensing technologies. Ultra-fast kinetics would also facilitate post-prandial glycemic control following single mealtime injections. The barrier to subcutaneous absorption is insulin self-assembly. The key component of an ultra-fast formulation would, thus, be an engineered insulin monomer with sufficient intrinsic chemical and physical stability to render zinc-mediated self-assembly unnecessary. Such an analog was developed 20 years ago (AspB10-insulin), but failed preclinical testing due to its potential tumorigenicity and increased in vitro mitogenicity relative to native insulin. These properties are thought to reflect increased binding to the IGF-I receptor (IGF-1R) and prolonged residence time at the insulin receptor (IR). We have discovered that a strategic fluoro-modification of an AspB10-insulin analog (a) eliminates undesirable binding to IGF-1R and prolonged IR residence time and, at the same time, (b) enhances the stabilizing effects of AspB10. The fluoro-modified residue is ortho-F-PheB24, which is amenable to insertion by chemical synthesis or by novel genetic engineering. Phase-I support is, therefore, requested to achieve milestones related to the stability, potency, mitogenicity, and PK/PD of fluoro-protected AspB10 analogs containing rapid-acting B-chain substitutions at positions B28 and B29 (derived from current products Humalog(R) and Novolog(R)). This proposal makes innovative use of fluorine (a mainstay of medicinal chemistry) in protein biotechnology to enhance the safety and efficacy of insulin replacement therapy.

Public Health Relevance:
Diabetes is increasing in global prevalence. To provide greater convenience, improved glycemic control, and fewer adverse side-effects (all of which result in greater compliance and lower healthcare costs), we have invented novel ultra-stable and ultra-rapid-acting insulin analogs (designated Fluorolog-1 and Fluorolog-2) that exhibit optimized receptor binding profiles attenuating unwanted effects of AspB10 on binding to the insulin receptor (IR) and the IGF-I receptor (IGF-1R), in principle reducing cancer risk. The innovative design of these analogs exploits fluorine-based electrostatic engineering to ""tune"" the stability, mitogenicity, and potency of an engineered Zn-free insulin monomer. This project will complete feasibility testing on Fluorolog-1 and Fluorolog-2.

Thesaurus Terms:
2-Deoxy-2-((Methylnitrosoamino)Carbonyl)Amino-D-Glucose;2-Deoxy-2-(3-Methyl-3-Nitrosoureido)-D-Glucopyranose;2-Deoxy-2-[[(Methylnitrosamino)-Carbonyl]amino]-D-Glucopyranose;28(B)-Lys-29(B)-Pro-Insulin;28(B)-Lysine-29(B)-Prolineinsulin;A21-Gly-B31-Arg-B32-Arg-Insulin;Adverse Effects;Assay;Attenuated;B23;Binding;Binding (Molecular Function);Bioassay;Biologic Assays;Biological Assay;Biotechnology;Blood Glucose;Blood Serum;Blood Sugar;Brittle Diabetes Mellitus;Budgets;Buffers;Cancer Cell Line;Cell Communication And Signaling;Cell Signaling;Chemicals;Clinical;Clinical Research;Clinical Study;Clinical Trials;Common Rat Strains;D-Glucose;Dextrose;Diabetes Mellitus;Dose;Drug Formulations;Drug Kinetics;Elisa;Electrostatics;Engineering;Enzyme-Linked Immunosorbent Assay;Event;Exhibits;F Element;Family Suidae;Fermentation;Fluorine;Formulation;Generations;Genetic Engineering;Genetic Engineering Biotechnology;Genetic Engineering Molecular Biology;Glucose;Glycohemoglobin A;Glycosylated Hemoglobin A;Group 17 Elements;Guidelines;Halogens;Hb A1;Hb A1a+B;Hb A1c;Hba1;Hba1c;Health Care Costs;Health Costs;Health Sciences;Healthcare Costs;Hemoglobin A(1);Humalog;Human;Humulin R;Hypoglycemia;Iddm;Igf Receptor;Igf-1;Igf-I;Igf-I-Smc;In Vitro;Infusion;Infusion Procedures;Injection Of Therapeutic Agent;Injections;Insulin;Insulin (Ox), 8a-L-Threonine-10a-L-Isoleucine-30b-L-Threonine-;Insulin Receptor;Insulin Receptor Protein-Tyrosine Kinase;Insulin, Glargine, Human;Insulin, Lispro, Human;Insulin-Dependent Diabetes Mellitus;Insulin-Dependent Tyrosine Protein Kinase;Insulin-Like Growth Factor 1;Insulin-Like Growth Factor I;Insulin-Like Growth Factor Receptor;Insulin-Like Somatomedin Peptide I;Intracellular Communication And Signaling;Juvenile-Onset Diabetes Mellitus;Ketosis-Prone Diabetes Mellitus;Kinetics;Loinc Axis 2 Property;Life;Lispro Insulin;Mcf-7;Mcf-7 Cell;Mcf7;Mcf7 Cell;Man (Taxonomy);Mediating;Medicinal Chemistry;Modeling;Modern Man;Modification;Molecular Interaction;Niddk;Nih Rfa;Npm;Npm1;Npm1 Gene;National Institute Of Diabetes And Digestive And Kidney Diseases;Novolog;Novolin R;Octreotide;Oregon;Peer Review;Performance;Pharmaceutic Chemistry;Pharmaceutical Chemistry;Pharmacodynamics;Pharmacokinetics;Phase;Physical Condensation;Physiologic;Physiological;Pigs;Position;Positioning Attribute;Preclinical Testing;Prevalence;Production;Proinsulin;Property;Protein Analysis;Proteins;Pump;Rat;Rats Mammals;Rattus;Receptor Signaling;Recombinant Dna Technology;Recombinants;Regular Insulin;Relative;Relative (Related Person);Replacement Therapy;Request For Applications;Risk;Sbir;Sbirs (R43/44);Stz;Safety;Sampling;Serum;Signal Pathway;Signal Transduction;Signal Transduction Systems;Signaling;Small Business Innovation Research;Small Business Innovation Research Grant;Solubility;Somatomedin C;Somatomedin Receptors;Streptozocin;Streptozotocin;Structure;Sudden-Onset Diabetes Mellitus;Suidae;Swine;T1 Dm;T1 Diabetes;T1d;T1dm;Technology;Testing;Thermodynamic;Thermodynamics;Time;Timeline;Treatment Side Effects;Tripcellim;Trypsin;Tumorigenicity;Type 1 Diabetes Mellitus;Type 1 Diabetes;Type I Diabetes Mellitus;Universities;Zanosar;Zinc;Zn Element;Absorption;Analog;Base;Biological Signal Transduction;Blood Glucose Regulation;Cancer Risk;Chemical Stability;Chemical Synthesis;Clinical Investigation;Condensation;Design;Designing;Diabetes;Gene Product;Glargine;Glucose Control;Glucose Homeostasis;Glucose Regulation;Glycemic Control;Hemoglobin A1c;Hypoglycemic;Hypoglycemic Episodes;Improved;In Vivo;Innovate;Innovation;Innovative;Insulin Dependent Diabetes;Insulin, Gly(A21)-Arg(B31,B32)-;Insulin, Lys(28b)-Pro(29b)-;Insulin, Glycyl(A21)-Arginyl(B31,B32)-;Insulin, Lysyl(28b)-Prolyl(28b)-;Juvenile Diabetes;Juvenile Diabetes Mellitus;Ketosis Prone Diabetes;Lispro;Monomer;Non-Diabetic;Nondiabetic;Novel;Physical Property;Porcine;Pre-Clinical;Preclinical;Programs;Receptor Binding;Residence;Response;Self Assembly;Side Effect;Subcutaneous;Suid;Therapy Adverse Effect;Treatment Adverse Effect;Type I Diabetes

In this Phase 2 SBIR application we seek to advance toward human clinical trials a rapid-acting and ultra- concentrated monomeric insulin analog of novel halogen-based protein design. Designated Fluorolog, this analog was found in Phase I studies to exhibit rapid-acting PK/PD properties irrespective of protein concentration in the range 0.6 - 3.0 mM, i.e., from U-100 to U-500 strengths. We anticipate that a U-500 formulation of Fluorolog will be of particular benefit to the treatment of Type 2 diabetes mellitus (T2DM) in the setting of marked insulin resistance. Such patients are disproportionately members of underprivileged minority communities, including African-Americans, Hispanic Americans, and Indigenous Americans. T2DM represents a major component of health-care disparities in American society. Our Phase 1 studies validated a key hypothesis underlying design of Fluorolog: that introduction of a single fluorine atom at the receptor-binding surface of insulin (para-F-PheB24) can at the same time (i) protect the insulin monomer from degradation and (ii) modulate mitogenicity such that untoward effects of AspB10 on cellular proliferation in culture and on cross-binding to the IGF receptor are each mitigated. In Phase 2 we seek to extend these studies to obtain data required in an IND application. To this end, pilot stability data willbe enlarged to include formal testing of the individual aspects of chemical and physical degradation (such as disulfide cleavage, covalent polymer formation, and fibrillation) by procedures and under conditions customarily provided in an IND application. Similarly, pilot cell-culture data wil be extended to analysis of tumor xenograft growth rates in nude mice and by formal toxicity studies in Sprague-Dawley rats. The overarching goal of this Phase 2 application is thus the submission of an appropriate and well-documented IND application.

Thesaurus Terms:
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