USOO8623419B2 (12) United States Patent (10) Patent No.: US 8,623,419 B2 Malakhov et al. (45) Date of Patent: Jan. 7, 2014 (54) TECHNOLOGY FOR PREPARATION OF WO 2004/047735 6, 2004 MACROMOLECULAR MICROSPHERES WO WO2005/035088 4/2005 WO 2006/031291 3, 2006 (75) Inventors: Michael P. Malakhov, San Francisco, CA (US); Fang Fang, Rancho Santa Fe, OTHER PUBLICATIONS CA (US) US 5,849,884, 11/09/99, Woiszwillo et al (withdrawn). Agrawal et al., “Basis of rise in intracellular sodium in airway hyper (73) Assignee: Ansun Biopharma, Inc., San Diego, CA responsiveness and asthma.” Lung, 183:375-387. (2005). 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Microspheres are produced by contacting an aqueous solu tion of a protein or other macromolecule with an organic FOREIGN PATENT DOCUMENTS Solvent and a counterion, and chilling the solution. The CN 1729013 2, 2006 microspheres are useful for preparing pharmaceuticals of EP O542314 5, 1993 defined dimensions. WO WO9420856 9, 1994 WO 98.06.279 2, 1998 WO O1,28524 4/2001 11 Claims, No Drawings US 8,623.419 B2 Page 2 (56) References Cited Sellers, S.P. et al., “Drypowders of stable protein formulations from aqueous solutions prepared using Supercritical CO(2)-assisted OTHER PUBLICATIONS aerosolization.” Journal of Pharmaceutical Sciences, 90(6):785-797. (2001). Lee, M.K. and A.D. 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Potier, M., et al., “Fluorometric assay of neuraminidase with a Zhirnov, O.P. et al., “Protective effect of protease inhibitors in influ sodium (4-methylumbelliferyl-alpha-D-N-acetylneuraminate) Sub enza virus infected animals.” Archives of Virology, 73:263-272, strate.” Analytical Biochemistry, 94:287-296, (1979). (1982). US 8,623,419 B2 1. 2 TECHNOLOGY FOR PREPARATION OF temperature and that do not contain a significant amount of MACROMOLECULAR MICROSPHERES denatured protein. There also is a need for a method of pro ducing microparticles of proteins and other macromolecules RELATED APPLICATIONS wherein substantially all of the protein or macromolecule in the starting material is recovered in the microparticle formu This application is a continuation of U.S. application Ser. lation, with minimal loss. There also is a need for micropar No. 12/633,773, filed Dec. 8, 2009, now abandoned, which ticles of proteins or other macromolecules containing these claims priority to U.S. application Ser. No. 1 1/657,812, filed properties for administration, for example, as a therapeutic or Jan. 24, 2007, now abandoned, which claims priority to U.S. nutritional Supplement. provisional application No. 60/762.002, filed Jan. 24, 2006. 10 The prior applications are hereby incorporated by reference SUMMARY in their entireties. This application also is related to International PCT appli Provided herein are methods for producing protein and cation Application Serial No.