REISHI F3 SUB FRACTION POLYSACCHARIDES AND METHODS OF USING SAME

• US 20090263897A1
• Filed: 10/02/2008
• Published: 10/22/2009
• Est. Priority Date: 04/21/2008
• Status: Active Grant

• Current Assignee: Academia Sinica (Government of The Republic of China)
• Original Assignee: Academia Sinica (Government of The Republic of China)
• Inventors: Chen, Yen-Po, Yang, Wen-Bin, Wong, Chi-Huey, Liang, Shu-Mei
• Granted Patent: US 8,071,105 B2
• Time in Patent Office: Days
• US Class Current: 435/377
• CPC Class Codes: A61K 2039/55588 Adjuvants of undefined constitution
• CPC Class Codes: A61K 39/0002 Fungal antigens, e.g. Trichophyton, Aspergillus, Candida

Abstract

The present disclosure relates to the discovery of methods of isolating subfractions of an F3 Reishi extract, and of administration of these novel isolates to eukaryotic cells in order to induce certain immumodulatory, hematopoeitic and tumor-inhibiting phenotypic changes in those eukaryotic cells, mediated through particular toll-like receptor (TLR) and other transmembrane receptors. F3 subfractions F301 and F331 have demonstrated that F331 is capable of activating at least TLR-2 while F301 is capable of activating at least TLR-2, TLR-4, and TLR-5.

Claims

1. A method comprising:

administering a sufficient amount of F301 to at least one member of the group consisting of eukaryotic cells and eukaryotic cell-based organisms; and

wherein the administered F301, increases toll-like receptor mediated activation of intracellular nuclear factor-kappa B by at least 5% when assayed by Reporter Gene Assay.

2. The method of claim 1, wherein the group consisting of eukaryotic cells and eukaryotic cell-based organisms includes dendritic, epithelial, and monocyte cells and cellular precursors.

3. The method of claim 1, wherein the toll-like receptor mediated activation of intracellular nuclear factor-kappa B is mediated at least through the TLR-5 receptor.

4. The method of claim 1, wherein the toll-like receptor mediated activation of intracellular nuclear factor-kappa B is mediated at least through the TLR-2 receptor.

5. A method comprising:

administering a sufficient amount of F301 to at least one member of the group consisting of eukaryotic cells and eukaryotic cell-based organisms; and

wherein the administered F301, increases toll-like receptor mediated transcription of messenger RNA coding for cytokines by at least 5% when assayed by RT-PCR.

6. The method of claim 5, wherein the toll-like receptor increased transcription of messenger RNA coding for cytokines is mediated at least through the TLR-5 receptor.

7. The method of claim 5, wherein the toll-like receptor increased transcription of messenger RNA coding for cytokines is mediated at least through the TLR-2 receptor.

8. The method of any of claims 5, 6 or 7, wherein the increased toll-like receptor mediated transcription of messenger RNA coding results in an increase of mRNA coding for cytokines selected from the group consisting of interleukin-6, interleukin-8, and tumor necrosis factor-alpha.

9. A method comprising:

administering a sufficient amount of F301 to at least one member of the group consisting of eukaryotic cells and eukaryotic cell-based organisms; and

wherein the administered F301, increases toll-like receptor mediated translation of messenger RNA to cytokines by at least 5% when assayed by ELISA.

10. The method of claim 9, wherein the toll-like receptor mediated increased translation of messenger RNA to cytokines is mediated at least through the TLR-5 receptor.

11. The method of claim 9, wherein the toll-like receptor mediated increased translation of messenger RNA to cytokines is mediated at least through the TLR-2 receptor.

12. The method of any of claims 9, 10 or 11, wherein the increased toll-like receptor mediated translation of messenger RNA to cytokines results in an increase in production of intracellular and/or extracellular cytokines selected from the group consisting of interleukin-6, interleukin-8, and tumor necrosis factor-alpha.

13. A product formed by the process of:

homogenizing tissue of a Ganoderma lucidum;

extracting the homogenized tissue with 0.1 N NaOH aqueous alkaline solution to form a crude extract;

subjecting the crude extract to gel filtration chromatography using a Sephacryl S-500 column and eluting with an aqueous Tris buffer solution to form at least one fraction comprising a polysaccharide or glycopeptide component having terminal fucose residues;

dialyzing at least one of the fractions containing a glycopeptide or polysaccharide component having a terminal fucose residue;

precipitating the dialyzed fraction by adding ethanol to the dialyzed fraction; and

resuspending the precipitate in an aqueous solution.

14. A method comprising:

administering a sufficient amount of F331 to at least one member of the group consisting of eukaryotic cells and eukaryotic cell-based organisms; and

wherein the administered F331, increases toll-like receptor mediated activation of intracellular nuclear factor-kappa beta by at least 5% when assayed by Reporter Gene Assay.

15. The method of claim 14, wherein the group consisting of eukaryotic cells and eukaryotic cell-based organisms includes dendritic, epithelial, and monocyte cells and cellular precursors.

16. The method of claim 14, wherein the toll-like receptor mediated activation of intracellular nuclear factor-kappa beta is mediated at least through the TLR-2 receptor.

17. A method comprising:

administering a sufficient amount of F331 to at least one member of the group consisting of eukaryotic cells and eukaryotic cell-based organisms; and

wherein the administered F331, increases toll-like receptor mediated transcription of messenger RNA coding for cytokines by at least 5% when assayed by RT-pCR.

18. The method of claim 17, wherein the toll-like receptor increased transcription of messenger RNA coding for cytokines is mediated at least through the TLR-2 receptor.

19. The method of any of claims 17 or 18, wherein the increased toll-like receptor mediated transcription of messenger RNA coding results in an increase of mRNA coding for cytokines selected from the group consisting of interleukin-6, interleukin-8, and tumor necrosis factor-alpha.

20. A method comprising:

administering a sufficient amount of F331 to at least one member of the group consisting of eukaryotic cells and eukaryotic cell-based organisms; and

wherein the administered F331, increases toll-like receptor mediated translation of messenger RNA to cytokines by at least 5% when assayed by ELISA.

21. The method of claim 20, wherein the toll-like receptor mediated increased translation of messenger RNA to cytokines is mediated at least through the TLR-2 receptor.

22. The method of any of claims 20 or 21, wherein the increased toll-like receptor mediated translation of messenger RNA to cytokines results in an increase in production of intracellular and/or extracellular cytokines selected from the group consisting of interleukin-6, interleukin-8, and tumor necrosis factor-alpha.

23. A product formed by the process of:

homogenizing tissue of a Ganoderma lucidum;

extracting the homogenized tissue with 0.1 N NaOH aqueous alkaline solution to form a crude extract;

subjecting the crude extract to gel filtration chromatography using a Sephacryl S-500 column and eluting with an aqueous Tris buffer solution to form at least one fraction comprising a polysaccharide or glycopeptide component having terminal fucose residues;

dialyzing at least one of the fractions containing a glycopeptide or polysaccharide component having a terminal fucose residue;

precipitating the dialyzed fraction by adding ethanol to the dialyzed fraction; and

evaporating the ethanol/aqueous solution.

24. A composition of matter comprising an subfraction of a F3 fraction of Reishi, the composition comprising:

a total sugar content of substantially 37 percent;

fucose content of substantially 4 percent;

glucosamine content of substantially 5 percent;

galactose content of substantially 9 percent;

Glucose content of substantially 74 percent; and

,

and mannose content of substantially 9 percent;

25. A composition of matter comprising a subfraction of an F3 fraction of Reishi, the composition comprising:

a total sugar content of substantially 19 percent;

fucose content of substantially 3 percent;

glucosamine content of substantially 18 percent;

galactose content of substantially 7 percent;

Glucose content of substantially 59 percent; and

,

and mannose content of substantially 13 percent;