Mirac Proteins

• US 20100260739A1
• Filed: 03/09/2010
• Published: 10/14/2010
• Est. Priority Date: 03/09/2009
• Status: Abandoned Application

• Current Assignee: Bioatla, Inc.
• Original Assignee: Bioatla, Inc.
• Inventors: Short, Jay M., Frey, Gerhard, Chang, Hwai Wen
• Application Number:
  US12/719,972
• Publication Number:
  US 20100260739A1
• Time in Patent Office: Days
• US Class Current: 424/94.500
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61P 13/12 of the kidneys
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 17/06 Antipsoriatics
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 29/02 without antiinflammatory effect
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 37/06 Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 5/10 of the posterior pituitary hormones, e.g. oxytocin, ADH
• CPC Class Codes: A61P 7/02 Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: A61P 9/04 Inotropic agents, i.e. stimulants of cardiac contraction; Drugs for heart failure
• CPC Class Codes: A61P 9/10 for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: A61P 9/12 Antihypertensives
• CPC Class Codes: C07K 14/3153 Streptokinase
• CPC Class Codes: C07K 14/47 : from mammals
• CPC Class Codes: C07K 14/575 : Hormones derived from pro-opiomelanocortin, pro-enkephalin or pro-dynorphin C07K14/665, e.g. corticotropin C07K14/695
• CPC Class Codes: C07K 14/57545 : Neuropeptide Y
• CPC Class Codes: C07K 14/57563 : Vasoactive intestinal peptide [VIP]; Related peptides
• CPC Class Codes: C07K 7/06 : having 5 to 11 amino acids
• CPC Class Codes: C12N 15/102 : Mutagenizing nucleic acids
• CPC Class Codes: C12N 15/1058 : Directional evolution of libraries, e.g. evolution of libraries is achieved by mutagenesis and screening or selection of mixed population of organisms
• CPC Class Codes: C12N 9/16 : acting on ester bonds (3.1)
• CPC Class Codes: C12N 9/50 : Proteinases , e.g. Endopeptidases (3.4.21-3.4.25)
• CPC Class Codes: C12N 9/6435 : Plasmin (3.4.21.7), i.e. fibrinolysin
• CPC Class Codes: C12N 9/6459 : t-plasminogen activator (3.4.21.68), i.e. tPA
• CPC Class Codes: C12N 9/6462 : u-Plasminogen activator (3.4.21.73), i.e. urokinase
• CPC Class Codes: C12Y 302/01035 : Hyaluronoglucosaminidase (3.2.1.35), i.e. hyaluronidase
• CPC Class Codes: C12Y 304/21007 : Plasmin (3.4.21.7), i.e. fibrinolysin
• CPC Class Codes: C12Y 304/21069 : Protein C activated (3.4.21.69)
• CPC Class Codes: C12Y 304/21073 : u-Plasminogen activator (3.4.21.73), i.e. urokinase
• CPC Class Codes: C12Y 304/23015 : Renin (3.4.23.15)
• CPC Class Codes: C12Y 304/24029 : Aureolysin (3.4.24.29)
• CPC Class Codes: G01N 33/54306 : Solid-phase reaction mechanisms
• CPC Class Codes: G01N 33/573 : for enzymes or isoenzymes
• CPC Class Codes: G01N 33/6845 : Methods of identifying protein-protein interactions in protein mixtures
• CPC Class Codes: G01N 33/6854 : Immunoglobulins
• CPC Class Codes: Y02A 50/30 : Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

This disclosure relates to a method of generating conditionally active biologic proteins from wild type proteins, in particular therapeutic proteins, which are reversibly or irreversibly inactivated at the wild type normal physiological conditions. For example, evolved proteins are virtually inactive at body temperature, but are active at lower temperatures.

Claims

1. A method of preparing a conditionally active biologic protein, the method comprising:

i. selecting a wild-type biologic protein;

ii. evolving the DNA which encodes the wild-type biologic protein using one or more evolutionary techniques to create a mutant DNA;

iii. expressing the mutant DNA to obtain a mutant protein;

iv. subjecting the mutant protein and the wild-type protein to an assay under a normal physiological condition and to an assay under an aberrant condition; and

v. selecting the conditionally active biologic protein from those mutant proteins which exhibit both (a) a decrease in activity in the assay at the normal physiological condition compared to the wild-type protein, and (b) an increase in activity in the assay under the aberrant condition compared to the wild-type protein.

2. The method of claim 1 wherein the wild-type biologic protein is an enzyme.

3. The method of claim 2 wherein the wild-type biologic protein is selected from the group consisting of tissue plasminogen activator, streptokinase, urokinase, renin, and hyaluronidase.

4. The method of claim 1 wherein the wild-type biologic protein is selected from calcitonin gene-related peptide, substance P, neuropeptide Y, vasoactive intestinal peptide, vasopressin, and angiostatin.

5. The method of claim 1, wherein the normal physiological condition is selected from one or more of normal physiological temperature, pH, osmotic pressure, osmolality, oxidation and electrolyte concentration.

6. The method of claim 5, wherein the normal physiological condition is temperature;

and wherein the conditionally active biologic protein is substantially inactive at the normal physiological temperature, and is active at an aberrant temperature less than the normal physiological temperature.

7. A conditionally active biologic protein prepared by the method of claim 1, wherein the protein is reversibly or irreversibly inactivated at the normal physiological condition.

8. The conditionally active biologic protein of claim 7, wherein the protein is reversibly inactivated at the wild type normal physiological conditions.

9. The conditionally active biologic protein of claim 7, wherein the wild-type biologic protein is selected from the group consisting of tissue plasminogen activator, streptokinase, urokinase, renin, and hyaluronidase.

10. The conditionally active biologic protein of claim 7, wherein the wild-type biologic protein is selected from calcitonin gene-related peptide, substance P, neuropeptide Y, vasoactive intestinal peptide, vasopressin, and angiostatin.

11. A pharmaceutical composition comprising an effective amount of the conditionally active biologic protein of claim 7, and a pharmaceutically acceptable carrier.

12. The pharmaceutical composition of claim 11, wherein the conditionally active biologic protein is selected from the group consisting of a tissue plasminogen activator mutant, a streptokinase mutant, a urokinase mutant, a renin mutant, and a hyaluronidase mutant.

13. The pharmaceutical composition of claim 12, wherein the conditionally active biologic protein is selected from the group consisting of a tissue plasminogen activator mutant, a streptokinase mutant, and a urokinase mutant.

14. The pharmaceutical composition of claim 11, wherein the conditionally active biologic protein is selected from the group consisting of a calcitonin gene-related peptide mutant, a substance P mutant, a neuropeptide Y mutant, a vasoactive intestinal peptide mutant, a vasopressin mutant, and an angiostatin mutant.

15. The pharmaceutical composition of claim 14, wherein the conditionally active biologic protein is selected from the group consisting of a calcitonin gene-related peptide mutant, a vasopressin mutant, and an angiostatin mutant.

16. A method of preparing a conditionally active biological response modifier, the method comprising:

a. selecting an inflammatory response mediator;

b. identifying a wild-type antibody to the mediator;

c. evolving the wild-type antibody;

d. screening differentially for mutants that exhibit decreased binding to the mediator relative to the wild-type antibody at a first condition, and exhibit increased binding affinity to the mediator at a second condition to identify up-mutants; and

e. recombining the heavy chains and the light chains of the up-mutants to create recombined up-mutants; and

f. screening the recombined up-mutants for mutants that exhibit decreased binding to the mediator relative to the wild-type antibody at the first condition, and show increased binding affinity to the mediator at the second condition to identify the conditionally active biological response modifier.

17. The method of claim 16 wherein the inflammatory response mediator is selected from IL-6, IL-6 receptor, TNF-alpha, IL-23 and IL-12.

18. The method of claim 16 wherein the first and second condition are each selected from conditions of pH, osmotic pressure, osmolality, oxidation and electrolyte concentration.