Methods for modeling protein stability

Patent metadata

Field	Value
Publication number	US-9388452-B2
Application number	US-201113638919-A
Country	US
Kind code	B2
Filing date	Apr 8, 2011
Priority date	Apr 8, 2010
Publication date	Jul 12, 2016
Grant date	Jul 12, 2016

Abstract

The present invention relates to a method for determining the highest temperature that is suitable for performing accelerated protein stability studies, as well as to a method for modeling real-time protein stability from accelerated stability data generated at said temperature.

First claim

The invention claimed is:

1. A method for modeling protein stability over time, comprising the steps:
(a) determining a stability of a protein at a plurality of different temperatures within a specific temperature range, wherein the protein stability comprises a measure of retention of enzymatic activity over time;
(b) creating an Arrhenius plot comprising the stability of the protein at the plurality of temperatures determined in step (a), the Arrhenius plot having a linear portion at lower temperatures and a non-linear portion at higher temperatures, the linear and non-linear portions joining at an inflection point on the Arrhenius plot;
(c) identifying the inflection point at the junction between the linear portion and the non-linear portion of the Arrhenius plot created in step (b);
(d) determining an assay temperature consisting of the temperature at the inflection point of the Arrhenius plot created in step (b);
(e) performing an accelerated stability study of the protein at the assay temperature determined in step (d), wherein the stability study comprises determining a retention of enzymatic activity over time; and
(f) determining a stability of the protein at a second temperature based on the accelerated stability data obtained in step (e), wherein the second temperature is lower than the assay temperature.

2. The method of claim 1 , wherein the protein stability determined in step (a) is determined over a time course of from 1 to 12 weeks.

3. The method of claim 1 , wherein each respective temperature in the plurality of temperatures used in step (a) is in the temperature range of from 4° C. to 60° C.

4. The method of claim 1 , wherein the method further comprises determining an expected shelf-life of the protein at a predetermined temperature below the assay temperature, wherein the expected shelf-life is the period of time a formulation comprising the protein is expected to maintain a predetermined level of stability.

5. The method of claim 4 , wherein the predetermined level of stability is a threshold level of enzymatic activity.

6. The method of claim 4 , wherein the predetermined level of stability is a threshold level of protein aggregation.

7. The method of claim 4 , wherein the predetermined level of stability is a threshold level of protein degradation.

8. The method of claim 4 , wherein the predetermined temperature is between 2° C. and 8° C.

9. The method of claim 4 , wherein the protein is an antibody.

10. The method of claim 9 , wherein the antibody is a recombinant antibody.

11. The method of claim 4 , wherein the protein is a plasma-derived protein.

12. The method of claim 4 , wherein the protein is Factor VIII.

13. The method of claim 4 , wherein the protein is antithrombin III (AT III).

14. A method for modeling protein stability over time, comprising the steps:
(a) determining a stability of a protein at a plurality of different temperatures within a specific temperature range, wherein the protein stability comprises a measure of retention of enzymatic activity over time;
(b) creating an Arrhenius plot comprising the stability of the protein at the plurality of temperatures determined in step (a), the Arrhenius plot having a linear portion at lower temperatures and a non-linear portion at higher temperatures, the linear and non-linear portions joining at an inflection point on the Arrhenius plot;
(c) identifying the inflection point at the junction between the linear portion and the non-linear portion of the Arrhenius plot created in step (b);
(d) determining an assay temperature consisting of the temperature at the inflection point of the Arrhenius plot created in step (b);
(e) performing an accelerated stability study of the protein at the assay temperature determined in step (d), wherein the stability study comprises determining a retention of enzymatic activity over time; and
(f) determining a stability of the protein at a second temperature based on the accelerated stability data obtained in step (e), wherein the second temperature is lower than the assay temperature,
wherein the respective temperatures in the plurality of temperatures used in step (a) are at 5° C. intervals.

15. A method for determining the shelf-life of Factor VIII formulation, comprising the steps:
(a) performing an accelerated stability study of the Factor VII formulation at a temperature equal to or less 30° C.; and
(b) determining the expected shelf-life of the formulation based on the accelerated stability study.

16. The method of claim 15 , wherein the accelerated stability study is performed at a temperature equal to or less 25° C.

17. A method for determining the shelf-life of an antithrombin III (AT III) formulation, comprising the steps:
(a) performing an accelerated stability study of the AT III formulation at a temperature equal to or less 20° C.; and
(b) determining the expected shelf-life of the formulation based on the accelerated stability study.

18. The method of claim 17 , wherein the accelerated stability study is performed at a temperature equal to or less 15° C.

19. A method for modeling protein stability over time, comprising the steps:
(a) determining a stability of a protein at a plurality of different temperatures within a specific temperature range, wherein the protein stability comprises a measure of protein aggregation over time;
(b) creating an Arrhenius plot comprising the stability of the protein at the plurality of temperatures determined in step (a), the Arrhenius plot having a linear portion at lower temperatures and a non-linear portion at higher temperatures, the linear and non-linear portions joining at an inflection point on the Arrhenius plot;
(c) identifying the inflection point at the junction between the linear portion and the non-linear portion of the Arrhenius plot created in step (b);
(d) determining an assay temperature consisting of the temperature at the inflection point of the Arrhenius plot created in step (b);
(e) performing an accelerated stability study of the protein at the assay temperature determined in step (d), wherein the stability study comprises determining protein aggregation over time; and
(f) determining a stability of the protein at a second temperature based on the accelerated stability data obtained in step (e), wherein the second temperature is lower than the assay temperature.

20. A method for modeling protein stability over time, comprising the steps:
(a) determining a stability of a protein at a plurality of different temperatures within a specific temperature range, wherein the protein stability comprises a measure of protein degradation over time;
(b) creating an Arrhenius plot comprising the stability of the protein at the plurality of temperatures determined in step (a), the Arrhenius plot having a linear portion at lower temperatures and a non-linear portion at higher temperatures, the linear and non-linear portions joining at an inflection point on the Arrhenius plot;
(c) identifying the inflection point at the junction between the linear portion and the non-linear portion of the Arrhenius plot created in step (b);
(d) determining an assay temperature consisting of the temperature at the inflection point of the Arrhenius plot created in step (b);
(e) performing an accelerated stability study of the protein at the assay temperature determined in step (d), wherein the stability study comprises determining protein degradation over time; and
(f) determining a stability of the protein at a second temperature based

Assignees

• Matthiessen Peter
• Baxalta Inc
• Baxalta GmbH

Inventors

• Matthiessen Peter

Classifications

• G01N2333/755

  Factors VIII, e.g. factor VIII C [AHF], factor VIII Ag [VWF] · CPC title

• C12Q1/56 Primary

  involving blood clotting factors, e.g. involving thrombin, thromboplastin, fibrinogen · CPC title

• G01N33/6803

  General methods of protein analysis not limited to specific proteins or families of proteins · CPC title

• G06F17/11

  for solving equations {, e.g. nonlinear equations, general mathematical optimization problems (optimization specially adapted for a specific administrative, business or logistic context G06Q10/04)} · CPC title

• G06F19/12

  Physics · mapped topic