Claims for Patent: 7,803,565

Summary for Patent: 7,803,565

Title:	Use of lymphocytes to measure anthrax lethal toxin activity
Abstract:	It is disclosed herein that isolated lymphocytes, such as human B-cells and CD4.sup.+ T-cell can be used to determine an amount of lymphocyte-associated anthrax lethal toxin activity present. Methods of using isolated lymphocytes to identify anthrax therapeutic agents and to determine the efficacy of a potential anthrax therapeutic are disclosed. Methods are also provided for diagnosing and treating anthrax infections.
Inventor(s):	Frucht; David M. (Vienna, VA), Fang; Hui (Germantown, MD)
Assignee:	The United States of America as represented by the Department of Health and Human Services (Washington, DC) N/A (N/A)
Application Number:	11/399,003
Patent Claims:	1. A method of identifying an agent that decreases pathogenicity of anthrax, comprising: exposing isolated lymphocytes to anthrax lethal toxin (LT); contacting the lymphocytes with a test agent; and detecting lymphocyte-associated LT activity, wherein detecting lymphocyte-associated LT activity comprises: detecting a mitogen-activated protein kinase kinase (MAPKK)-dependent cytokine response of the lymphocytes to the LT, wherein an increased MAPKK-dependent cytokine response in the presence of the test agent indicates that the lymphocyte-associated LT activity is decreased and that the test agent decreases pathogenicity of anthrax; or determining whether there is an increase in proliferation of the lymphocytes, wherein an increase in lymphocyte proliferation in the presence of the test agent indicates that the lymphocyte-associated LT activity is decreased and that the test agent decreases pathogenicity of anthrax. 2. The method of claim 1, wherein exposing the isolated lymphocytes to LT comprises exposing the lymphocytes to purified LT. 3. The method of claim 1, wherein exposing the isolated lymphocytes to LT comprises exposing the lymphocytes to an organism that produces LT. 4. The method of claim 3, wherein exposing the isolated lymphocytes to an organism that produces LT comprises exposing the lymphocytes to Bacillus anthracis (B. anthracis). 5. The method of claim 1, wherein exposing the isolated lymphocytes to LT comprises exposing the lymphocytes to B. anthracis spores. 6. The method of claim 1, wherein the isolated lymphocytes are stimulated lymphocytes. 7. The method of claim 1, wherein the method further comprises stimulating the isolated lymphocytes following exposing the lymphocytes to anthrax LT. 8. The method of claim 7, wherein the isolated lymphocytes are isolated T-lymphocytes, and stimulating the T-lymphocytes comprises contacting the T-lymphocytes with an agent that activates a T-cell receptor (TCR). 9. The method of claim 7, wherein the isolated lymphocytes are isolated T-lymphocytes, and stimulating the T-lymphocytes comprises contacting the T-lymphocytes with 12-O tetradecanoylphorbol 13-acetate (PMA) and ionomycin. 10. The method of claim 7, wherein the isolated lymphocytes are isolated B-lymphocytes, and stimulating the B-lymphocytes comprises contacting the B-lymphocytes with an agent that activates a B-cell receptor (BCR). 11. The method of claim 7, wherein the isolated lymphocytes are first contacted with the LT, and subsequently the lymphocytes are contacted with the test agent and stimulated, wherein a decrease in lymphocyte-associated LT activity indicates the test agent is a therapeutic agent that can be used to treat an anthrax infection. 12. The method of claim 11, wherein the isolated lymphocytes are contacted with the test agent during or before stimulating the lymphocytes, wherein a decrease in lymphocyte-associated LT activity indicates the test agent is a therapeutic agent that can be used to treat an anthrax infection. 13. The method of claim 1, wherein the method further comprises comparing the detected lymphocyte-associated LT activity to a baseline or a control. 14. The method of claim 1, wherein the method further comprises comparing the lymphocyte-associated LT activity in the lymphocyte to a standard curve, wherein the standard curve represents an amount of lymphocyte-associated LT activity versus an amount of biologically active LT present. 15. The method of claim 1, wherein the method further comprises: selecting an agent that decreased lymphocyte-associated LT activity; administering the agent that decreased lymphocyte-associated LT activity to a subject; and detecting lymphocyte-associated LT activity in the subject, wherein decreased lymphocyte associated LT activity indicates the test agent decreases pathogenicity of anthrax. 16. The method of claim 15, wherein detecting lymphocyte-associated LT activity in the subject comprises determining whether lymphocyte-associated LT activity is decreased in a sample obtained from the subject. 17. The method of claim 1, wherein the test agent comprises an inhibitor of lymphocyte-associated LT activity. 18. The method of claim 1, wherein the agent that decreases pathogenicity of anthrax is an anti-anthrax therapeutic agent. 19. The method of claim 1, wherein the isolated lymphocytes are isolated human lymphocytes. 20. The method of claim 1, wherein the lymphocytes comprise stimulated human lymphocytes; and wherein an increase in human lymphocyte proliferation or secretion of a MAPKK-dependent cytokine relative to a negative control indicates the test agent decreases pathogenicity of anthrax. 21. The method of claim 1, wherein detecting a MAPKK-dependent cytokine response of the lymphocyte to the LT comprises detecting IL-2. 22. The method of claim 1, wherein detecting a MAPKK-dependent cytokine response of the lymphocyte to the LT comprises detecting IL-4. 23. The method of claim 1, wherein detecting a MAPKK-dependent cytokine response of the lymphocyte to the LT comprises detecting IFN-.gamma.. 24. The method of claim 1, wherein detecting a MAPKK-dependent cytokine response of the lymphocyte to the LT comprises detecting TNF-.alpha., IL-1.alpha., IL-1.beta., IL-2, IL-4, IL-6, IL-12, IL-18, IFN-.gamma., or combinations thereof. 25. The method of claim 7, wherein determining whether there is an increase in proliferation of the lymphocytes, comprises measuring proliferation of a stimulated B-cell or T-cell. 26. The method of claim 1, wherein the method is conducted in vitro. 27. The method of claim 19, wherein the isolated human lymphocytes comprise isolated human T-cells. 28. The method of claim 27, wherein the isolated human T-cells comprise isolated human CD4.sup.+ T-cells. 29. The method of claim 19, wherein the isolated human lymphocytes comprise isolated human B-cells. 30. The method of claim 1, wherein the isolated lymphocytes comprise lymphocytes isolated from a human blood sample. 31. The method of claim 1, wherein the isolated lymphocytes comprise a human lymphocyte cell line. 32. The method of claim 31, wherein the human lymphocyte cell line comprises human Jurkat cells.

Details for Patent 7,803,565

Applicant	Tradename	Biologic Ingredient	Dosage Form	BLA	Approval Date	Patent No.	Expiredate
Emergent Biodefense Operations Lansing Llc	BIOTHRAX	anthrax vaccine adsorbed	Injection	103821	11/12/1998		2025-04-05