# US 7,408,095 B2 · Methods for introducing heterologous cells into fish to determine if the heterologous cells contain cancer cells or a pathogen

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- Number: US 7,408,095 B2
- Document ID: US7408095B2
- Status: Granted
- Kind: B2
- Application number: 10/766,134
- Filed: 2004-01-27
- Published: 2004-09-23
- Granted: 2008-08-05
- Earliest claimed: 1998-02-22
- Assignees: Phylonix Pharmaceuticals, Inc.
- Inventors: George N. Serbedzija; Patricia McGrath
- Web page: https://patentgrep.com/patent/US7408095B2

## CPC

- A61K 49/0058: {Antibodies}
- A01K 67/0271: Chimeric vertebrates, e.g. comprising exogenous cells
- A61K 49/00: Preparations for testing in vivo
- A61K 49/0008: {Screening agents using (non-human) animal models or transgenic animal models or chimeric hosts, e.g. Alzheimer disease animal model, transgenic model for heart failure}
- A61K 49/0032: {Methine dyes, e.g. cyanine dyes}
- A61K 49/0043: {Fluorescein, used in vivo}
- A61P 35/00: Antineoplastic agents
- C12Q 1/68: involving nucleic acids
- G01N 33/5082: {Supracellular entities, e.g. tissue, organisms}
- G01N 33/5088: {of vertebrates}
- A01K 2227/40: Fish
- G01N 2333/4603: from fish
- 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)
- A01K: ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

Methods of introducing heterologous cells into fish are provided. After introduction cells remain viable, and in some instances proliferate, for sufficient time to conduct a variety of analyses on the heterologous cells or the fish or both. Such methods are also useful, for example, for diagnosing the presence of small quantities of cancerous cells or pathogens in patient tissue samples.

## Claims

### Claim 1

A method of analyzing a sample for presence of a cancer cell or pathogen, comprising:
(a) obtaining a sample from a patient containing a cell or population of cells;
(b) introducing the cell or the population of cells into a zebrafish embryo that has not yet developed an immune system that would subject the cell or population of cells to immune rejection; and
(c) detecting a property of the cell or the population of cells to indicate whether the cell or the population of cells comprises a cancer cell or pathogen.

### Claim 2 (depends on 1)

The method of claim 1, wherein the sample is obtained from bone marrow, peripheral blood, a body fluid or a transplanted tissue.

### Claim 3 (depends on 1)

The method of claim 1, wherein the sample is from a biopsy.

### Claim 4 (depends on 1)

The method of claim 1, wherein the cell or the population of cells is a human cell or a population of human cells.

### Claim 5 (depends on 1)

The method of claim 1, wherein introducing comprises microinjecting the cell or the population of cells into the zebrafish embryo.

### Claim 6 (depends on 1)

The method of claim 1, wherein the cell or the population of cells are microinjected into a blastula stage embryo.

### Claim 7 (depends on 1)

The method of claim 1, wherein the cell or the population of cells are microinjected into the yolk of the zebrafish embryo.

### Claim 8 (depends on 1)

The method of claim 1, wherein a population of cells are introduced into the zebrafish embryo.

### Claim 9 (depends on 8)

The method of claim 8, further comprising culturing the zebrafish embryo after the population of cells is introduced, whereby cancer cells present in the cell population proliferate.

### Claim 10 (depends on 8)

The method of claim 8, further comprising culturing the zebrafish embryo after the population of cells is introduced, whereby a pathogen present in the cell population is amplified.

### Claim 11 (depends on 8)

The method of claim 8, wherein the property detected is cell proliferation or metastasis, detection of proliferation or metastasis being an indication that the population of cells contains cancer cells.

### Claim 12 (depends on 8)

The method of claim 8, wherein the property detected is the rate of cell proliferation or metastasis.

### Claim 13 (depends on 11)

The method of claim 11, wherein cell proliferation is detected by immunostaining with an antibody specific for cancer cells.

### Claim 14 (depends on 11)

The method of claim 11, wherein metastasis is detected by detecting movement of cells from the cell population relative to an initial site at which the cell population was introduced.

### Claim 15 (depends on 1)

The method of claim 1, wherein the pathogen is a bacterium, a virus or a fungus.