Modified polypeptides with high activity and reduced allergenicity

• US 6,284,246 B1
• Filed: 07/30/1997
• Issued: 09/04/2001
• Est. Priority Date: 07/30/1997
• Status: Expired due to Fees

• Current Assignee: The Procter & Gamble Company
• Original Assignee: The Procter & Gamble Company
• Inventors: Weisgerber, David J., Rubingh, Donn N.
• Primary Examiner(s): Page, Thurman K.
• Assistant Examiner(s): HOWARD, SHARON LEE
• Application Number:
  US08/903,298
• Time in Patent Office: 1,497 Days
• Field of Search: 435/180, 424/49, 424/94.6, 424/400, 424/94.1, 424/401, 424/402, 514/844, 514/944, 514/947, 514/847
• US Class Current: 424/94.1
• CPC Class Codes: A61K 2800/57 Compounds covalently linked to a(n inert) carrier molecule, e.g. conjugates, pro-fragrances
• CPC Class Codes: A61K 2800/70 Biological properties of the composition as a whole
• CPC Class Codes: A61K 47/60 the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
• CPC Class Codes: A61K 8/0208 Tissues; Wipes; Patches
• CPC Class Codes: A61K 8/66 Enzymes
• CPC Class Codes: A61K 8/86 Polyethers
• CPC Class Codes: A61Q 11/00 Preparations for care of the teeth, of the oral cavity or of dentures; Dentifrices, e.g. toothpastes; Mouth rinses
• CPC Class Codes: A61Q 19/00 Preparations for care of the skin
• CPC Class Codes: A61Q 19/10 Washing or bathing preparations
• CPC Class Codes: A61Q 5/02 Preparations for cleaning the hair
• CPC Class Codes: C11D 3/386 Preparations containing enzymes , e.g. protease or amylase
• CPC Class Codes: C11D 3/38627 containing lipase

Abstract

The present invention relates to a modified polypeptide which has an enzymatic activity level of greater than about 70% of the parent polypeptide and an allergenic response level of less than about 33% of the parent polypeptide. Embodiments of the present invention relate to modified polypeptides with reduced allergenicity and high enzymatice activity comprising the formula:

A-B_n

wherein A is an enzyme, and mixtures thereof; B is a twin polymer moiety, having a total molecular weight of from about 0.5 kilodaltons (KD) to about 40 KD, having the formula

conjugated to the enzyme; wherein R₁ and R₂ are essentially straight chain polymers, having a molecular weight ranging from about 0.25 KD to about 20 KD; wherein the ratio of the molecular weights of R₁ and R₂ is from about 1:10 to about 10:1; wherein X is a linking moiety which links the twin moeity to a single site on the enzyme; and n is the number of twin polymer moietis conjugated to the enzyme, and represents an integer from about 1 to about 15.

Claims

1. A modified polypeptide which has an enzymatic activity level of greater than about 70% of a parent polypeptide and an allergenic response level of less than about 33% of the parent polypeptide, wherein the parent polypeptide comprises an enzyme with no additional conjugation of polymer moieties.

2. A modified polypeptide according to claim 1 wherein the enzymatic activity level is greater than about 80% of a parent polypeptide and an allergenic response level of less than about 20% of the parent polypeptide.

3. A modified polypeptide with reduced allergenicity and high activity comprising the formula:

4. A modified polypeptide according to claim 3 wherein:

the enzyme, A, is selected from the group consisting of lipase enzymes and protease enzymes, and mixtures thereof; and

the modified polypeptide has an enzymatic activity level of greater than about 70% of a parent polypeptide and an allergenic response level of less than about 33% of the parent polypeptide.

5. A modified polypeptide according to claim 4, wherein the enzyme, A, is selected from the group consisting of lipase enzymes and protease enzymes selected from the group consisting of subtilisin, chymotrypsin, and elastase enzymes, and mixtures thereof.

6. A modified polypeptide according to claim 5, wherein the enzyme is a subtilisin protease enzyme.

7. A modified polypeptide according to claim 6, wherein the enzyme is selected from the group consisting of bacterial serine protease enzyme, subtilisin BPN'"'"', Protease A, Protease B, Protease D, Protease F, and mixtures thereof.

8. A modified polypeptide according to claim 7, wherein the enzyme is Protease F.

9. A modified polypeptide according to claim 3 wherein the total molecular weight of the twin polymer moiety is from about 1 KD to about 10 KD and the individual polymer moieties, R₁ and R₂, have a molecular weight ranging from about 0.5 KD to about 5 KD.

10. A modified polypeptide according to claim 3 wherein the ratio of molecular weights of R₁ and R₂, is from about 1:

5 to about 5;

1.

11. A modified polypeptide according to claim 3 wherein the polymers, R₁ and R₂, comprise polyethylene glycol.

12. A modified polypeptide according to claim 3 wherein n is from about 1 to about 10.

14. A modified polypeptide according to claim 12, which exhibits enzymatic activity levels greater than 90% of the activity of Protease F and exhibits allergenic response levels of less than 5% of the allergenic response of Protease F.

15. A personal cleansing composition comprising greater than about 0.001% and less than about 20% of the modified polypeptide according to claim 3.

16. A laundry cleansing composition comprising greater than about 0.001% and less than about 20% of the modified polypeptide according to claim 3.

17. An oral cleansing composition comprising greater than about 0.001% and less than about 20% of the modified polypeptide according to claim 3.

18. A topical skin care composition comprising greater than about 0.001% and less than about 20% of the modified polypeptide according to claim 3.

19. A shower gel comprising greater than about 0.001% and less than about 10% of the modified polypeptide according to claim 3.

20. A leave-on skin moisturizer composition comprising greater than about 0.001% and less than about 10% of the modified polypeptide according to claim 3.

21. A cosmetic composition comprising greater than about 0.001% and less than about 10% of the modified polypeptide according to claim 3.

22. A cleansing wipe composition comprising a cleansing composition comprising greater than about 0.001% and less than about 10% of the modified polypeptide according to claim 3.

13. A modified polypeptide with reduced allergenicity and high activity comprising the formula: