# US 10,501,472 B2 · Method to isolate inoscavin a from Fulviformes fastuosus and medicinal preparation thereof to treat rhabdomyosarcoma cancer conditions

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- Number: US 10,501,472 B2
- Document ID: US10501472B2
- Status: Granted
- Kind: B2
- Application number: 15/775,007
- Filed: 2015-11-23
- Published: 2018-12-27
- Granted: 2019-12-10
- Earliest claimed: 2015-07-22
- Applicants: University of Colombo; National Research Council
- Inventors: Dilusha Fernando; Ravi Wijesundera; Preethi Soysa; Dilip de Silva; Chandrika Nanayakkara; Achyut Adhikari
- Web page: https://patentgrep.com/patent/US10501472B2

## CPC

- C07D 493/20: Spiro-condensed systems
- A61K 31/366: having six-membered rings, e.g. delta-lactones
- A61K 36/07: Basidiomycota, e.g. Cryptococcus
- A61P 35/00: Antineoplastic agents
- C07D: HETEROCYCLIC COMPOUNDS (macromolecular compounds C08)
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS

## Abstract

The invention relates to a novel method to isolate a natural product, a flavonoid namely inoscavin A from a Sri Lankan mushroom, medicinal preparation thereof to treat rhabdomyosarcoma cancer condition. This compound was isolated from the terrestrial basidiomycete Fulviformes fastuosus for the first time. The isolation method of the inoscavin A comprises the following steps of: (1) carrying out sonication extraction of pulverized specimen by methanol; (2) partitioning in to hexane, dichloromethane and ethyl acetate respectively by liquid-liquid extraction 3) fractionation using normal phase silica gel column chromatography 4) purification using preparative high performance liquid chromatography (HPLC) to obtain flavonoid compound inoscavin A in substantially purified form (≥98%). In vitro cytotoxicity experiments demonstrated that the composition comprising inoscavin A strongly induced the apoptosis of human rhabdomyosarcoma (RD) cells compared to the cytotoxicity produced on normal epithelial cells (CC-1). Therefore, isolated flavonoid compound can be used for manufacturing of a medicament to treat rhabdomyosarcoma cancer condition. A method of treating rhabdomyosarcoma using medicinal preparation comprising inoscavin A can be achieved by administering said composition to a subject in need of such treatment.

## Claims

### Claim 1

A method to isolate inoscavin A comprising:
(a) providing Fulviformes fastuosus fruiting bodies;
(b) pulverization and sonication extraction of the Fulviformes fastuosus fruiting bodies to obtain an extract;
(c) partitioning of the extract of step (b) using liquid-liquid extraction;
(d) fractionation of extract from step (c) using column chromatography; and
(e) purification of product from step (d) using high performance liquid chromatography.

### Claim 2 (depends on 1)

The pulverization and sonication in claim 1 wherein, the Fulviformes fastuous fruiting bodies is pulverized into a fine powder.

### Claim 3 (depends on 1)

The pulverization and sonication in claim 1, step (b), wherein the extraction is performed into a polar organic solvent comprising methanol and obtaining a dried extract.

### Claim 4 (depends on 3)

The liquid -liquid extraction in claim 3 comprising dissolving the dried extract in water and transferring the dissolved extract into a separatory funnel containing hexane and obtaining an aqueous layer containing the extract.

### Claim 5 (depends on 4)

The liquid aqueous layer of claim 4 wherein the aqueous layer containing the extract was further extracted using ethyl acetate and dried.