Details for Patent: 5,635,207

Title:	Methods for the preparation of blood substitutes for in vivo delivery
Abstract:	In accordance with the present invention, there are provided compositions useful for the in vivo delivery of a biologic, wherein the biologic is associated with a polymeric shell formulated from a biocompatible material. The biologic can be associated with the polymeric shell itself, and/or the biologic, optionally suspended/dispersed in a biocompatible dispersing agent, can be encased by the polymeric shell. In another aspect, the biologic associated with polymeric shell is administered to a subject, optionally dispersed in a suitable biocompatible liquid.
Inventor(s):	Grinstaff; Mark W. (Pasadena, CA), Soon-Shiong; Patrick (Los Angeles, CA), Wong; Michael (Champaign, IL), Sandford; Paul A. (Los Angeles, CA), Suslick; Kenneth S. (Champaign, IL), Desai; Neil P. (Los Angeles, CA)
Assignee:	Vivorx Pharmaceuticals, Inc. (Santa Monica, CA)
Filing Date:	Jun 07, 1995
Application Number:	08/480,621
Claims:	1. A method for the preparation of blood substitutes for in vivo delivery, said method comprising subjecting aqueous medium containing oxygen binding protein to high intensity ultrasound conditions for a time sufficient to promote crosslinking of said oxygen binding protein by disulfide bonds to form a polymeric shell; wherein the largest cross-sectional dimension of shell is no greater than about 10 microns. 2. A method for the delivery of a blood substitute to a subject, said method comprising administering to said subject an effective amount of blood substitute prepared according to the method of claim 1. 3. The method according to claim 1, wherein said oxygen binding protein is selected from hemoglobin, myoglobin, or a mixture thereof. 4. The method according to claim 1, wherein said oxygen binding protein is hemoglobin. 5. A method for the preparation of blood substitutes for in vivo delivery, said method comprising subjecting aqueous medium containing biocompatible material and a blood substitute to high intensity ultrasound conditions for a time sufficient to promote crosslinking of said biocompatible material by disulfide bonds; wherein said blood substitute is substantially completely contained within a polymeric shell, and wherein the largest cross-sectional dimension of said shell is no greater than about 10 microns. 6. The method according to claim 5, wherein said biocompatible material is a naturally occurring polymer, a synthetic polymer, or a combination thereof, wherein said polymer, prior to crosslinking, has covalently attached thereto sulfhydryl groups or disulfide linkages. 7. The method according to claim 6, wherein said naturally occurring polymer is selected from proteins containing sulfhydryl groups and/or disulfide groups, polypeptides containing sulfhydryl groups and/or disulfide groups, lipids containing sulfhydryl groups and/or disulfide groups, polynucleic acids containing sulfhydryl groups and/or disulfide groups, or polysaccharides containing sulfhydryl groups and/or disulfide groups. 8. The method according to claim 7, wherein said protein is selected from hemoglobin, myoglobin, albumin, insulin, lysozyme, immunoglobulins, .alpha.-2-macroglobulin, fibronectin, vitronectin, fibrinogen, or combinations of any two or more thereof. 9. The method according to claim 8, wherein said protein is albumin. 10. The method according to claim 8, wherein said protein is hemoglobin. 11. The method according to claim 8, wherein said protein is a combination of albumin and hemoglobin. 12. A method for the delivery of a blood substitute to a subject, said method comprising administering to said subject an effective amount of blood substitute prepared according to the method of claim 5. 13. The method according to claim 4, wherein said hemoglobin is modified with an allosteric effector. 14. The method according to claim 13, wherein said allosteric effector is selected from inositol hexaphosphate, 2,3-bisphosphoglycerate, a di or monoaspirin ester, or combinations of any two or more thereof. 15. The method according to claim 14, wherein said diaspirin ester is 3,5-bis(dibromosalicyl)fumarate. 16. The method according to claim 3, wherein said protein is myoglobin. 17. The method according to claim 1, wherein said oxygen binding protein is combined with a second protein selected from albumin, insulin, lysozyme, an immunoglobulin, alpha-2-macroglobulin, fibronectin, vitronectin, fibrinogen, or combinations of any two or more thereof. 18. The method according to claim 17, wherein said second protein is albumin. 19. The method according to claim 17 wherein said second protein is albumin and said oxygen binding protein is hemoglobin. 20. The method according to claim 1, wherein said polymeric shell is modified by a suitable agent, wherein said suitable agent is selected from a synthetic polymer, a phospholipid, a protein, a polysaccharide, a surface active agent, a chemical modifying agent, or combination thereof, wherein said suitable agent is associated with said polymeric shell through an optional covalent linkage. 21. The method according to claim 1, wherein said polymeric shell is suspended in a biocompatible medium, and wherein said biocompatible medium is selected from water, buffered aqueous media, saline, buffered saline, a solution of amino acids, a solution of proteins, a solution of sugars, a solution of vitamins, a solution of carbohydrates, a solution of synthetic polymers, a lipid-containing emulsion, or combinations of any two or more thereof. 22. The method according to claim 1, wherein said aqueous medium containing the oxygen binding protein contains a biologic and wherein after crosslinking, said biologic is completely contained within said polymeric shell. 23. The method according to claim 22, wherein said biologic within said shell is dissolved or suspended in a biocompatible dispersing agent. 24. The method according to claim 7, wherein said polysaccharides are selected from an alginate, a polymannuronic acid, a polymannuronate, a hyaluronic acid, a hyaluronate, heparin, dextran, chitosan, chitin, cellulose, starch, glycogen, guar gum, locust bean gum, levan, inulin, cyclodextrin, agarose, xanthan gum, carrageenan, heparin, pectin, gellan gum, scleroglucan, or combinations of any two or more thereof. 25. The method according to claim 24, wherein said alginate is a high M-content alginate. 26. The method according to claim 6, wherein said synthetic polymer is selected from a synthetic polyamino acid containing cysteine residues and/or disulfide groups, a synthetic polypeptide containing sulfhydryl groups and/or disulfide groups, a polyvinyl alcohol modified to contain free sulfhydryl groups and/or disulfide groups, a polyhydroxyethyl methacrylate modified to contain free sulfhydryl groups and/or disulfide groups, a polyacrylic acid modified to contain free sulfhydryl groups and/or disulfide groups, polyethyloxazoline modified to contain free sulfhydryl groups and/or disulfide groups, a polyacrylamide modified to contain free sulfhydryl groups and/or disulfide groups, polyvinyl pyrrolidinone modified to contain free sulfhydryl groups and/or disulfide groups, a polyalkylene glycol modified to contain free sulfhydryl groups and/or disulfide groups, as well as mixtures of any two or more thereof. 27. The method according to claim 5, wherein said polymeric shell is modified by a suitable agent, wherein said suitable agent is selected from a synthetic polymer, a phospholipid, a protein, a polysaccharide, a surface active agent, a chemical modifying agent, or combination thereof, wherein said suitable agent is associated with said polymeric shell through an optional covalent linkage. 28. The method according to claim 5, wherein said blood substitute is selected from hemoglobin, myoglobin, or mixtures thereof. 29. The method according to claim 28, wherein said blood substitute is hemoglobin. 30. The method according to claim 29, wherein said hemoglobin is modified with an allosteric effector. 31. The method according to claim 30, wherein said allosteric effector is selected from inositol hexaphosphate, 2,3-bisphosphoglycerate, a di or monoaspirin ester, or combinations of any two or more thereof. 32. The method according to claim 31, wherein said diaspirin ester is 3,5-bis(dibromosalicyl) fumarate. 33. The method according to claim 28, wherein said blood substitute is myoglobin. 34. The method according to claim 5, wherein said blood substitute is selected from perfluorodecalin or perfluorotripropylamine. 35. The method according to claim 5, wherein said blood substitute is crosslinked. 36. The method according to claim 5, wherein said blood substitute within said shell is dissolved or suspended in a biocompatible dispersing agent. 37. The method according to claim 5, wherein said polymeric shell containing said blood substitute is suspended in a biocompatible medium, and wherein said biocompatible medium is selected from water, buffered aqueous media, saline, buffered saline, a solution of amino acids, a solution of protein, a solution of sugars, a solution of vitamins, a solution of carbohydrates, a solution of synthetic polymers, a lipid-containing emulsion, or combinations of any two or more thereof. 38. The method according to claim 10, wherein a phospholipid bilayer is formed around the crosslinked hemoglobin shell. 39. The method according to claim 38, wherein said phospholipid bilayer is formed from charged lipids, nonionic lipids, polymerizable lipids, or combinations of any two or more thereof. 40. The method according to claim 39, wherein said charged lipid is selected from phosphatidyl choline, phosphatidyl ethanol amine, phosphatidyl serine, phosphatidyl inositol, phosphatidyl glycerol, sphingomyelin, dimyristoylphosphatidic acid, dipalmitoyl phosphatidic acid, a sarcosinate (sarcosinamide), a betaine, a monomeric and dimeric alkyd, or combinations of any two or more thereof. 41. The method according to claim 39, wherein said nonionic lipid is selected from a polyethylene fatty acid ester, a polyethylene fatty acid ether, a diethanolamide, a long chain acyl hexosamide, a long chain acyl amino acid amide, a long chain amino acid amine, a polyoxyethylene sorbitan ester, a polyoxy glycerol mono- and di-ester, a glycerol mono- and di-stearate, a glycerol mono- and di-oleate, a glycerol mono- and di-palmitate, or combinations of any two or more thereof. 42. The method according to claim, 39, wherein said photopolymerizable lipid is selected from phosphatidyl choline, phosphatidyl ethanol amine, phosphatidyl serine, phosphatidyl glycerol, dimyristoylphosphatidic acid, dipalmitoyl phosphatidic acid, a lipid with native polymerizable unsaturation, or combinations of any two or more thereof. 43. The method according to claim 42, wherein said lipid with native polymerizable unsaturation comprises an unsaturated phosphatidyl choline with diacetylene groups, an unsaturated phosphatidyl choline with conjugated diene groups, or combinations of any two or more thereof. 44. The method according to claim 39, wherein said crosslinkable lipid comprises one or more derivatives of a phosphatidyl choline esterified with lipoic acid.