Claims for Patent: 7,919,273

Summary for Patent: 7,919,273

Title:	Structural variants of antibodies for improved therapeutic characteristics
Abstract:	The present invention provides substituted humanized, chimeric or human anti-CD20 antibodies or antigen binding fragments thereof and bispecific antibodies or fusion proteins comprising the substituted antibodies or antigen binding fragments thereof. The antibodies, fusion proteins or fragments are useful for treatment of B-cell disorders, such as B-cell malignancies and autoimmune diseases, as well as GVHD, organ transplant rejection, and hemolytic anemia and cryoglobulinemia. Amino acid substitutions, particularly substitution of an aspartate residue at Kabat position 101 of CDR3 V.sub.H (CDRH3), result in improved therapeutic properties, such as decreased dissociation rates, improved CDC activity, improved apoptosis, improved B-cell depletion and improved therapeutic efficacy at very low dosages. Veltuzumab, a humanized anti-CD20 antibody that incorporates such sequence variations, exhibits improved therapeutic efficacy compared to similar antibodies of different CDRH3 sequence, allowing therapeutic effect at dosages as low as 200 mg or less, more preferably 100 mg or less, more preferably 80 mg or less, more preferably 50 mg or less, most preferably 30 mg or less of naked antibody when administered i.v. or s.c.
Inventor(s):	Goldenberg; David M. (Mendham, NJ), Chang; Chien-Hsing (Downingtown, PA), Hansen; Hans J. (Picayune, MS)
Assignee:	Immunomedics, Inc. (Morris Plains, NJ)
Application Number:	12/506,992
Patent Claims:	1. A method of improving a chimeric or humanized anti-CD20 antibody or antigen-binding fragment thereof comprising making at least one amino acid substitution in the third complementarity determining region (CDR) sequence of the heavy chain of a chimeric or humanized or human anti-CD20 antibody or antigen binding fragment thereof to make a substituted antibody or antigen binding fragment thereof, wherein the substitution results in at least one improved characteristic selected from the group consisting of a slower off-rate, slower antigen dissociation rate, higher CDC activity, higher ADCC activity, higher apoptotic activity, greater ability to induce cell death in vitro in the absence of cross-linking and greater ability to kill or inhibit the growth of CD20-positive cells in vivo when administered to a subject with CD20-positive cells, wherein the CDR sequences of the unsubstituted anti-CD20 antibody or antigen-binding fragment thereof comprise the light chain CDR sequences CDRL1 (RASSSVSYIH; SEQ ID NO:1), CDRL2 (ATSNLAS; SEQ ID NO:2) and CDRL3 (QQWTSNPPT; SEQ ID NO:3) and heavy chain CDR sequences CDRH1 (SYNMH; SEQ ID NO:4), CDRH2(AIYPGNGDTSYNQKFKG; SEQ ID NO:5) and CDRH3(STYYGGDWYFNV; SEQ ID NO:32) and wherein the substitution comprises replacing the asparagine residue at Kabat position 101 of the heavy chain CDR3 (CDRH3) of the antibody with an aspartate residue. 2. The method of claim 1 wherein the CD20-positive cells are B-cells. 3. The method of claim 1, wherein the substituted antibody comprises an arginine residue at Kabat position 94 and the aspartate residue at Kabat position 101 forms an ionic bond with the arginine residue at Kabat position 94. 4. The method of claim 1, wherein the substituted antibody or fragment thereof inhibits binding to CD20 of rituximab. 5. The method of claim 1, wherein the substituted antibody is veltuzumab. 6. The method of claim 1, wherein substitution of aspartate for asparagine at Kabat position 101 results in at least a two-fold slower dissociation rate of the anti-CD20 antibody from CD20. 7. A method of improving a chimeric or humanized anti-CD20 antibody or antigen-binding fragment thereof comprising making at least one amino acid substitution in the third complementarity determining region (CDR) sequence of the heavy chain of a chimeric or humanized anti-CD20 antibody or antigen binding fragment thereof to make a substituted antibody or antigen binding fragment thereof, wherein the substitution results in at least one improved characteristic selected from the group consisting of a slower off-rate, slower antigen dissociation rate, higher CDC activity, higher ADCC activity, higher apoptotic activity, greater ability to induce cell death in vitro in the absence of cross-linking and greater ability to kill or inhibit the growth of CD20-positive cells in vivo when administered to a subject with CD20-positive cells, wherein the substituted anti-CD20 antibody or fragment thereof comprises the light chain CDR sequences CDRL1 (RASSSVSYIH; SEQ ID NO:1), CDRL2 (ATSNLAS; SEQ ID NO:2) and CDRL3 (QQWTSNPPT; SEQ ID NO:3) and heavy chain CDR sequences CDRH1 (SYNMH; SEQ ID NO:4), CDRH2 (AIYPGNGDTSYNQKFKG; SEQ ID NO:5) and CDRH3 (STYYGGDWYFDV; SEQ ID NO:6). 8. The method of claim 1, wherein the humanized anti-CD20 antibody comprises the framework region sequences of the humanized anti-CD22 antibody, epratuzumab, or the framework region sequences of the humanized anti-CD20 antibody, veltuzumab. 9. The method of claim 1, wherein substitution of aspartate for the asparagine residue at Kabat position 101 results in an increase in complement-dependent cytotoxicity of Daudi cells exposed to the antibody. 10. The method of claim 1, wherein substitution of aspartate for the asparagine residue at Kabat position 101 results in a 30 to 40% reduction in EC.sub.50 in complement-dependent cytotoxicity of Daudi cells exposed to the antibody. 11. The method of claim 7, wherein the CD20-positive cells are B-cells. 12. The method of claim 7, wherein the substituted antibody comprises an arginine residue at Kabat position 94 and the aspartate residue at Kabat position 101 forms an ionic bond with the arginine residue at Kabat position 94. 13. The method of claim 7, wherein the substituted antibody or fragment thereof inhibits binding to CD20 of rituximab. 14. The method of claim 7, wherein the substituted antibody is veltuzumab. 15. The method of claim 7, wherein substitution of aspartate for asparagine at Kabat position 101 results in at least a two-fold slower dissociation rate of the anti-CD20 antibody from CD20. 16. The method of claim 7, wherein the CDR sequences of the unsubstituted antibody are identical to the CDR sequences of a murine anti-CD20 antibody. 17. The method of claim 7, wherein the humanized anti-CD20 antibody comprises the framework region sequences of the humanized anti-CD22 antibody, epratuzumab, or the framework region sequences of the humanized anti-CD20 antibody, veltuzumab. 18. The method of claim 7, wherein substitution of aspartate for the asparagine residue at Kabat position 101 results in an increase in complement-dependent cytotoxicity of Daudi cells exposed to the antibody. 19. The method of claim 7, wherein substitution of aspartate for the asparagine residue at Kabat position 101 results in a 30 to 40% reduction in EC.sub.50 in complement-dependent cytotoxicity of Daudi cells exposed to the antibody.

Details for Patent 7,919,273

Applicant	Tradename	Biologic Ingredient	Dosage Form	BLA	Approval Date	Patent No.	Expiredate
Genentech, Inc.	RITUXAN	rituximab	Injection	103705	11/26/1997		2028-07-21
Idec Pharmaceuticals Corp.	RITUXAN	rituximab	Injection	103737	02/19/2002		2028-07-21
Genentech, Inc.	RITUXAN HYCELA	rituximab and hyaluronidase human	Injection	761064	06/22/2017		2028-07-21