# US 2021/0239686 A1 · Biosensors for Measuring Metastatic Potential and Chemoresistance of Single Cancer Cells

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- Number: US 2021/0239686 A1
- Document ID: US20210239686A1
- Status: Published application
- Kind: A1
- Application number: 17/051,293
- Filed: 2019-05-13
- Published: 2021-08-05
- Earliest claimed: 2018-05-14
- Assignees: The Regents of the University of California
- Applicants: The Regents of the University of California
- Inventors: Pradipta Ghosh
- Web page: https://patentgrep.com/patent/US20210239686A1

## CPC

- G01N 33/542: with steric inhibition or signal modification, e.g. fluorescent quenching
- G01N 33/575: for cancer
- G01N 33/57595: {involving intracellular compounds}
- G01N 2800/52: Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis
- G01N 2800/56: Staging of a disease; Further complications associated with the disease
- 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

The present invention provides a Fluorescence Resonance Energy Transfer (FRET) biosensor that measures the metastatic potential and/or chemoresistance of single living cancer cells, and/or cells associated with fibrogenesis, diabetes, aging, and/or infertility.

## Claims

### Claim 1

A method for detecting metastatic potential and/or chemoresistance in single cells, comprising;
providing one or more multi-modular biosensors;
providing a sample comprising a cell;
contacting said sample with the one or more biosensors under conditions that allow folding of probe donor and acceptor fluorescent proteins into close proximity; and
detecting fluorescence resonance energy transfer (FRET) signal to detect said interacting molecules.

### Claim 2 (depends on 1)

The method of claim 1, wherein the one or more multi-modular biosensors target short stretches of the C-terminus of GIV flanking one or both of tyrosines, Y1764 and Y1798, and N-SH2 domain of p85α(PI3K).

### Claim 3 (depends on 1)

The method of claim 1, wherein the multi-modular biosensor consists of two modules.

### Claim 4 (depends on 3)

The method of claim 3, wherein the two modules are separated by a flexible linker and sandwiched by eCFP (donor) and eYFP (acceptor) proteins, such that binding of the N-SH2-p85α(PI3K) to phosphotyrosine (pY) GIV ligand leads to folding of the probe bringing donor and acceptor in proximity sufficient to generate a detectable FRET signal.

### Claim 5 (depends on 1)

The method of claim 1, wherein the single cells, wherein metastatic potential and chemoresistance are being detected, are cancer cells.

### Claim 6 (depends on 1)

The method of claim 1, wherein the single cells, wherein metastatic potential and chemoresistance are being detected, are cells associated with fibrogenesis, diabetes, aging, and/or infertility.

### Claim 7

A method for identifying agents specifically targeting a GNAI:GIV interface, comprising;
providing one or more multi-modular biosensors;
providing a sample comprising a cell;
contacting said sample with the one or more biosensors to form a first mixture under conditions that allow the folding of probe donor and acceptor fluorescent proteins into close proximity;
detecting a first FRET signal to detect said interacting molecules;
contacting said first mixture with an agent under said conditions that allow the folding of probe donor and acceptor fluorescent proteins into close proximity;
detecting a second FRET signal to detect said interacting molecules; and
determining the difference between said first and said second FRET signal.

### Claim 8 (depends on 7)

The method of claim 7, wherein the one or more biosensors target short stretches of the C-terminus of GIV flanking one or both of tyrosines, Y1764 and Y1798, and the N-SH2 domain of p85α(PI3K).

### Claim 9 (depends on 7)

The method of claim 7, wherein the multi-modular biosensor consists of two modules.

### Claim 10 (depends on 9)

The method of claim 9, wherein the two modules are separated by a flexible linker and sandwiched by eCFP (donor) and eYFP (acceptor) proteins, such that binding of N-SH2-p85α(PI3K) to phosphotyrosine (pY) GIV ligand leads to folding of the probe bringing donor and acceptor in proximity sufficient to generate a detectable FRET signal.

### Claim 11 (depends on 7)

The method of claim 7, wherein the sample comprises cancer cells.

### Claim 12 (depends on 7)

The method of claim 7, wherein sample comprises cells associated with fibrogenesis, diabetes, aging, and/or infertility.

### Claim 13

A biosensor device for detecting metastatic potential and/or chemoresistancy of single cells comprising a biosensor that targets short stretches wherein the one or more multi-modular biosensors target short stretches of the C-terminus of GIV flanking one or both of tyrosines, Y1764 and Y1798, and N-SH2 domain of p85α(PI3K).

### Claim 14 (depends on 13)

The biosensor device of claim 13, wherein the multi-modular biosensor consists of two modules.

### Claim 15 (depends on 14)

The biosensor device of claim 14, wherein the two modules are separated by a flexible linker and sandwiched by eCFP (donor) and eYFP (acceptor) proteins, such that binding of the N-SH2-p85α(PI3K) to phosphotyrosine (pY) GIV ligand leads to folding of the probe bringing donor and acceptor in proximity sufficient to generate a detectable FRET signal.