Modified heparinase iii and methods of sequencing therewith

Abstract

The invention relates to heparinase III and mutants thereof. Modified forms of heparinase III having reduced enzymatic activity which are useful for a variety of purposes, including sequencing of heparin-like glycosaminoglycans (HLGAGs), removing active heparan sulfate from a solution, inhibition of angiogenesis, etc. have been discovered according to the invention. The invention in other aspects relates to methods of treating cancer and inhibiting tumor cell growth and/or metastasis using heparinase III, or products produced by enzymatic cleavage by heparinase III of HLGAGs.

1 . A method for preventing proliferation of a tumor, comprising: exposing a tumor cell to an effective amount of heparinase III for preventing proliferation of the tumor cells in order to prevent growth of the tumor. 
     
     
         2 . The method of  claim 1 , wherein the heparinase III is a modified heparinase III. 
     
     
         3 . The method of  claim 1 , wherein the heparinase III is a native heparinase III. 
     
     
         4 . The method of  claim 1 , wherein the heparinase III is injected into the tumor in vivo. 
     
     
         5 . The method of  claim 1 , wherein the heparinase III is administered systemically to a subject having a tumor. 
     
     
         6 . The method of  claim 1 , wherein the heparinase III is administered orally to a subject having a tumor. 
     
     
         7 . The method of  claim 1 , wherein the tumor cell is administered the heparinase III in vitro. 
     
     
         8 . The method of  claim 1 , wherein the heparinase III is administered in conjunction with an anti-cancer drug. 
     
     
         9 . The method of  claim 1 , wherein the heparinase III is administered to a subject having a non-metastatic tumor in order to prevent the tumor from becoming metastatic. 
     
     
         10 . The method of  claim 1 , wherein the tumor is selected from the group consisting of a prostate tumor and melanoma. 
     
     
         11 . The method of  claim 1 , wherein the heparinase III is administered to a subject without any additional anti-cancer drugs. 
     
     
         12 - 22 . (canceled) 
     
     
         23 . A composition, comprising: heparinase III or a therapeutic HLGAG fragment in an effective amount for preventing metastasis of a tumor cell and a targeting molecule for targeting the heparinase III to the tumor, in a pharmaceutically acceptable carrier. 
     
     
         24 . The composition of  claim 23 , wherein the heparinase III is a modified heparinase III. 
     
     
         25 . The composition of  claim 23 , wherein the heparinase III is a native heparinase 
     
     
         26 . The composition of  claim 23 , wherein the targeting molecule is a compound which binds specifically to an antigen on the surface of a tumor cell. 
     
     
         27 - 49 . (canceled) 
     
     
         50 . A method comprising treating or preventing a subject having a cancer or at risk of developing a cancer by administering to the subject a therapeutic HLGAG fragment. 
     
     
         51 . The method of  claim 50 , wherein the therapeutic HLGAG fragment is a composition of HLGAG fragments wherein at least 50% of the HLGAG fragments are di-or tri-sulfated disaccharides. 
     
     
         52 . The method of  claim 50 , wherein the therapeutic HLGAG fragment is a composition of HLGAG fragments wherein at least 75% of the HLGAG fragments are di-or tri-sulfated disaccharides. 
     
     
         53 . The method of  claim 50 , wherein the therapeutic HLGAG fragment is a composition of HLGAG fragments wherein at least 90% of the HLGAG fragments are di-or tri-sulfated disaccharides. 
     
     
         54 . The method of  claim 50 , wherein the therapeutic HLGAG fragment is free of mono-or un-sulfated disaccharides. 
     
     
         55 - 60 . (canceled)