Ex vivo methods for testing toxicity of substances using donated human organs or tissues

• US 8,771,930 B2
• Filed: 05/18/2007
• Issued: 07/08/2014
• Est. Priority Date: 05/18/2007
• Status: Active Grant

• Current Assignee: Lifeline Scientific, Inc. (Shanghai Genext Medical Technology Co., Ltd.)
• Original Assignee: Lifeline Scientific, Inc. (Shanghai Genext Medical Technology Co., Ltd.)
• Inventors: Curtis, Gerald, Brassil, John, Kravitz, David
• Primary Examiner(s): SINGH, SATYENDRA K
• Application Number:
  US11/802,059
• Publication Number:
  US 20080288399A1
• Time in Patent Office: 2,608 Days
• US Class Current: 435/1.2
• CPC Class Codes: G01N 33/5014 for testing toxicity
• CPC Class Codes: G01N 33/5082 Supracellular entities, e.g. tissue, organisms
• CPC Class Codes: G01N 33/566 using specific carrier or receptor proteins as ligand binding reagents where possible specific carrier or receptor proteins are classified with their target compounds
• CPC Class Codes: G01N 33/574 for cancer
• CPC Class Codes: G01N 33/68 involving proteins, peptides or amino acids involving lipoproteins G01N33/92
• CPC Class Codes: G06Q 20/102 Bill distribution or payments

Abstract

Properties such as toxicity of substances may be determined by assaying properties, fates and effects of substances in an ex vivo metabolically active human organ or tissue under normothermic perfusion with a fluid containing a test substance. The data can be used as, for example, part of a submission to a government regulatory organization. Preferred methods use perfused endocrine gland organs or tissues to evaluate hormone or other bodily chemical disruption caused by substances and pre-donation diseased or injured organs or tissues.

Claims

1. A method of testing the toxicity of a substance, comprising:

passing a perfusate comprising the substance under normothermic conditions and physiological oxygenation, osmolality, electrolyte balance and pH conditions through at least one metabolically active ex vivo donated human organ or tissue that was injured or diseased before donation, wherein perfusate comprising the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

passing a perfusate under normothermic conditions and physiological oxygenation, osmolality, electrolyte balance and pH conditions that does not contain the substance through the at least one donated human organ or tissue before and/or after passing the at least one donated human organ or tissue with the perfusate comprising the substance, wherein

the at least one donated human organ or tissue acts as its own control,

the perfusate that does not contain the substance comprises the positive and negative controls, and

the perfusate that does not contain the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

collecting data derived from the perfused organ or tissue, wherein collecting data derived from the perfused organ or tissue comprises;

assaying an effluent from the perfused organ or tissue, and

monitoring one or more parameters selected from the group consisting of cell death, apoptosis, genetic mutation, changes in gene expression, biochemical inhibition, reduction in metabolism, induction and oxidative damage, hormone disruption, and bodily chemical disruption; and

analyzing the data to assess the toxicity of the substance, wherein the substance is selected from the group consisting of agricultural products, industrial products, building materials, tobacco products, cleaning products and cosmetics;

wherein said at least one metabolically active ex vivo donated human organ or tissue is a glandular organ or tissue.

2. The method of claim 1, wherein the data are collected by electrophysiological testing, medical diagnostic imaging, spectroscopic testing, microdialysis, solid state tissue probe testing or a combination thereof.

3. The method of claim 1, wherein the substance is selected from the group consisting of industrial products, building materials and cleaning products.

4. The method of claim 3, wherein the cleaning products are selected from the group consisting of detergents, bleaches, soaps, shampoos and conditioners.

5. The method of claim 1, wherein the substance is selected from the group consisting of tobacco products and cosmetics.

6. A method of testing the toxicity of a substance, comprising:

passing a perfusate comprising the substance under normothermic conditions through at least one metabolically active ex vivo donated human organ or tissue that was injured or diseased before donation, wherein perfusate comprising the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

passing a perfusate that does not contain the substance through the donated human organ or tissue before and/or after perfusing the donated human organ or tissue with the perfusate comprising the substance, wherein

the donated human organ or tissue acts as its own control,

the perfusate that does not contain the substance comprises the positive and negative controls, the perfusate that does not contain the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP); and

but for the presence of the substance, the perfusate comprising the substance is compositionally the same as the perfusate that does not contain the substance;

collecting data derived from the perfused organ or tissue, wherein collecting data derived from the perfused organ or tissue comprises;

assaying an effluent from the perfused organ or tissue, and

monitoring one or more parameters selected from the group consisting of cell death, apoptosis, genetic mutation, changes in gene expression, biochemical inhibition, reduction in metabolism, induction and oxidative damage, hormone disruption, and bodily chemical disruption; and

analyzing the data to assess the toxicity of the substance, wherein the at least one metabolically active ex vivo donated human organ or tissue is an intact organ or tissue, and data are used to analyze at least one of the following;

hormone level, target cell-receptor mediation, glandular secretion function, hormone function, organ function and a combination thereof;

wherein the substance is selected from the group consisting of agricultural products, industrial products, building materials, tobacco products, cleaning products and cosmetics;

wherein said at least one metabolically active ex vivo donated human organ or tissue is a glandular organ or tissue.

7. A method of testing the toxicity of a substance, comprising:

passing a perfusate comprising the substance under normothermic conditions through at least one metabolically active ex vivo donated human organ or tissue that was injured or diseased before donation, wherein perfusate comprising the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

passing a perfusate that does not contain the substance through the donated human organ or tissue before and/or after perfusing the donated human organ or tissue with the perfusate comprising the substance, wherein the donated human organ or tissue acts as its own control, and the perfusate that does not contain the substance comprises the positive and negative controls, wherein the perfusate that does not contain the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

collecting data derived from the perfused organ or tissue, wherein collecting data derived from the perfused organ or tissue comprises assaying an effluent from the perfused organ or tissue, and

analyzing the data to assess the toxicity of the substance, wherein the organ or tissue is selected from the group consisting of endocrine glands and exocrine glands, and analyzing the data comprises determining a concentration of the substance at which the secretion ability of said glands decreases, increases or is otherwise altered;

wherein the substance is selected from the group consisting of agricultural products, industrial products, building materials, tobacco products, cleaning products and cosmetics.

8. A method of testing the toxicity of a substance, comprising:

passing a perfusate comprising the substance under normothermic conditions through at least one metabolically active ex vivo donated human organ or tissue that was injured or diseased before donation, wherein perfusate comprising the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

passing a perfusate that does not contain the substance through the donated human organ or tissue before and/or after perfusing the donated human organ or tissue with the perfusate comprising the substance, wherein the donated human organ or tissue acts as its own control, and the perfusate that does not contain the substance comprises the positive and negative controls, wherein the perfusate that does not contain the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

collecting data derived from the perfused organ or tissue, wherein collecting data derived from the perfused organ or tissue comprises assaying an effluent from the perfused organ or tissue; and

analyzing the data to assess the toxicity of the substance, wherein the organ or tissue is an organ selected from the group consisting of pancreas, thymus, male reproductive organs, female reproductive organs, thyroid gland and adrenal gland, and analyzing the data comprises determining the concentration of substance at which the rate of production of secretory products or concentration of secretory products of the organ decreases, increases or is otherwise altered;

wherein the substance is selected from the group consisting of agricultural products, industrial products, building materials, tobacco products, cleaning products and cosmetics.

9. A method of testing the toxicity of a substance, comprising:

passing a perfusate comprising the substance under normothermic conditions through at least one metabolically active ex vivo donated human organ or tissue that was injured or diseased before donation, wherein perfusate comprising the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

passing a perfusate that does not contain the substance through the donated human organ or tissue before and/or after perfusing the donated human organ or tissue with the perfusate comprising the substance, wherein the donated human organ or tissue acts as its own control, and the perfusate that does not contain the substance comprises the positive and negative controls, wherein the perfusate that does not contain the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

collecting data derived from the perfused organ, or tissue, wherein collecting data derived from the perfused organ or tissue comprises assaying an effluent from the perfused organ or tissue;

analyzing the data to assess the toxicity of the substance; and

quantifying a functional state of the donated human organ or tissue that was injured or diseased before donation, wherein the passing comprises perfusing the organ or tissue with a first perfusate that does not contain the substance, followed by perfusing the organ or tissue with a second perfusate that contains the substance;

wherein the substance is selected from the group consisting of agricultural products, industrial products, building materials, tobacco products, cleaning products and cosmetics;

wherein said at least one metabolically active ex vivo donated human organ or tissue is a glandular organ or tissue.

10. The method of claim 9, wherein the organ or tissue is an entire organ or tissue and has been injured by mechanical injury.

11. The method of claim 9, wherein the organ or tissue is an entire organ or tissue and diseased with a disease that the substance comprised in the perfusate is intended to treat.

12. A method of testing the toxicity of a substance, comprising:

passing a perfusate comprising the substance under normothermic conditions through at least one metabolically active ex vivo donated human organ or tissue that was injured or diseased before donation, wherein perfusate comprising the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

passing a perfusate that does not contain the substance through the donated human organ or tissue before and/or after perfusing the donated human organ or tissue with the perfusate comprising the substance, wherein the donated human organ or tissue acts as its own control, and the perfusate that does not contain the substance comprises the positive and negative controls, wherein the perfusate that does not contain the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

collecting data derived from the perfused organ or tissue, wherein collecting data derived from the perfused organ or tissue comprises assaying an effluent from the perfused organ or tissue;

analyzing the data to assess the toxicity of the substance;

quantifying a functional state of the donated human organ or tissue that was injured or diseased before donation with respect to a hormone level, wherein the data are analyzed to assess whether the hormone level is unaltered or is increased, decreased or otherwise altered in response to passing the perfusate comprising the substance under normothermic conditions through said at least one metabolically active ex vivo donated human organ or tissue;

wherein the substance is selected from the group consisting of agricultural products, industrial products, building materials, tobacco products, cleaning products and cosmetics;

wherein said at least one metabolically active ex vivo donated human organ or tissue is a glandular organ or tissue.

13. The method of claim 12, wherein the substance is selected from the group consisting of industrial products, building materials, tobacco products and cleaning products.

14. A method of testing the toxicity of a substance, comprising:

passing a perfusate comprising the substance under normothermic conditions through at least one metabolically active ex vivo donated human endocrine gland or endocrine gland tissue under normothermic conditions, wherein perfusate comprising the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

passing a perfusate that does not contain the substance through the human endocrine gland or endocrine gland tissue before and/or after perfusing the human endocrine gland or endocrine gland tissue with the perfusate comprising the substance, wherein the human endocrine gland or endocrine gland tissue acts as its own control, and the perfusate that does not contain the substance comprises the positive and negative controls, wherein the perfusate that does not contain the substance further comprises about 15 to about 20% (v/v) matched human erythrocytes, about 2 to about 6% human, serum albumin, N-acetylcysteine, and adenosine monophosphate (AMP);

collecting data derived from the perfused gland or gland tissue, wherein collecting data derived from the perfused gland or gland tissue comprises assaying an effluent from the perfused gland or gland tissue;

quantifying a functional state of the at least one metabolically active ex vivo donated human endocrine gland or endocrine gland tissue with respect to hormone uptake or production; and

analyzing the data to assess effects of the substance on the hormone uptake or production by said gland or gland tissue;

wherein the substance is selected from the group consisting of agricultural products, industrial products, building materials, tobacco products, cleaning products and cosmetics.

15. The method of claim 14, wherein the data are collected by electrophysiological testing, medical diagnostic imaging, spectroscopic testing, microdialysis, solid state tissue probe testing or a combination thereof.

16. The method of claim 14, wherein the gland is selected from the group consisting of pancreas, thymus, male reproductive organs, female reproductive organs, thyroid gland and adrenal gland.

17. The method of claim 14, wherein the gland or gland tissue has been injured by prolonged warm ischemia.

18. The method of claim 14, wherein the gland or gland tissue has been injured by prolonged hypothermic storage.

19. The method of claim 14, wherein the gland or gland tissue has been injured by mechanical injury.

20. The method of claim 14, wherein the gland or gland tissue is diseased.

21. The method of claim 14, wherein the data are analyzed to assess whether a hormone level is unaltered or is increased, decreased or otherwise altered.

22. The method of claim 14, wherein the substance is selected from the group consisting of industrial products, building materials and cleaning products.

23. The method of claim 14, wherein the substance is selected from the group consisting of tobacco products and cosmetics.