Rhone-poulenc Rorer Pharmaceuticals Inc.

Centeon L.l.c.

David L Farb

Jack Newman

Michael E Hrinda

Ted C Lee

Christopher P Prior

David Weber

Processes for preparing aqueous solutions of cysteine-altered von Willebrand Factor fragment which are substantially free of aggregate and capable of therapeutic use for treating thrombosis are provided. The claimed process comprises providing an aqueous solution of vWF fragment and denaturant and containing undesired contaminants, said solution having an acidic pH; separating said contaminants from said solution by contacting said solution with an affinity chromatography medium to which said vWF fragments adhere; eluting said vWF fragment from said affinity chromatography medium in the presence of the denaturant; and separating the eluted fragment from said denaturant while maintaining the aqueous solution of the fragment at a pH of about 2.5 to less than about 5.5 to increase monomerization of, and decrease aggregation of, said fragment, thereby forming an aqueous solution of vWF fragment which is substantially free of aggregate.

Therapeutic fragments of von Willebrand factor

A method of improving the therapeutic activity of von Willebrand Factor obtained from materials comprising on Willebrand Factor comprising incubating said Factor at a temperature of about 20.degree. C. to about 55.degree. C. for about 1 to about 30 hours, most preferably at from about 45.degree. C. to about 55.degree. C. for about 1 to about 5 hours.

Purification of von Willebrand factor by affinity chromatography

An aqueous solution of cysteine-altered von Willebrand Factor fragment which is substantially free of aggregate and capable of therapeutic use for treating thrombosis and a process for preparing such a solution comprising: (A) providing an aqueous solution of the fragment and denaturant; (B) purifying the solution of fragment and denaturant under conditions which promote conversion of aggregated forms of the fragment to the dimeric and/or monomeric forms thereof to provide purified fragment; (C) separating the dissolved, purified fragment from the denaturant while maintaining the aqueous solution of the fragment at a pH of about 2.5 to less than about 5.5 to increase monomerization of, and decrease aggregation of, said purified fragment, thereby forming an aqueous solution of fragment which is substantially free of aggregate.

A method of improving the therapeutic activity of von Willebrand Factor obtained from materials comprising said Factor comprising incubating said Factor at a temperature of about 20.degree. C. to about 55.degree. C. for about 1 to about 30 hours.

Purification of vonwillebrand factor by affinity chromatography

A method of preparing von Willebrand Factor by disassociating it from a chaotropic agent in solution therewith and preferably treating the same under controlled temperature either in liquid or lyophilized form.

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Chalfont, PA, United States of America

David L Farb