# US 11,111,546 B2 · 3.4 KB mitochondrial DNA deletion for use in the detection of cancer

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- Number: US 11,111,546 B2
- Document ID: US11111546B2
- Status: Granted
- Kind: B2
- Application number: 16/429,739
- Filed: 2019-06-03
- Published: 2019-10-24
- Granted: 2021-09-07
- Earliest claimed: 2005-04-18
- Assignees: MDNA Life Sciences Inc.
- Applicants: MDNA Life Sciences Inc.
- Inventors: Ryan L. Parr; Robert Thayer; Gabriel D. Dakubo; Jennifer Creed; Kerry Robinson; Andrea Maggrah; Brian Reguly
- Web page: https://patentgrep.com/patent/US11111546B2

## CPC

- C12Q 1/6886: for cancer (immunoassay for cancer G01N33/575)
- C12Q 1/686: Polymerase chain reaction [PCR]
- G01N 33/57515: of the breast
- G01N 33/57555: of the prostate
- C12Q 2600/112: Disease subtyping, staging or classification
- C12Q 2600/118: Prognosis of disease development
- C12Q 2600/156: Polymorphic or mutational markers
- 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

A method is provided for detecting a cancer in an individual, the method comprising detection of a deletion in the nucleic acid sequence between residues 10743 and 14125 in mitochondrial DNA by extracting the mitochondrial DNA (mtDNA) from the biological sample obtained from the individual and quantifying the amount of mtDNA in the sample having a deletion in the nucleic acid sequence between approximately residues 10743 and 14125 of the mtDNA genome, and comparing the amount of mtDNA in the sample having the deletion to at least one known reference sample. Kits for conducting the method are also provided.

## Claims

### Claim 1

A method of detecting a cancer in an individual comprising;
a) extracting mitochondrial DNA, mtDNA, from a biological sample from the individual;
b) quantifying the amount of a mtDNA deletion in the sample, the deletion having a nucleic acid sequence as set forth in SEQ ID NO: 1, the step of quantifying comprising the use of a pair of amplification primers, wherein one of the pair of amplification primers is adapted to bind to a rejoining site of the deletion after the deletion has re-circularized;
c) comparing the amount of the deletion in the sample to at least one known reference value.

### Claim 2 (depends on 1)

The method of claim 1, wherein the at least one known reference value is the amount of the deletion in a reference sample of mtDNA from known non-cancerous tissue or body fluid.

### Claim 3 (depends on 1)

The method of claim 1, wherein the at least one known reference value is the amount of the deletion in a reference sample of mtDNA from known cancerous tissue or body fluid.

### Claim 4 (depends on 1)

The method of claim 1, wherein the step of quantifying is conducted using real-time PCR.

### Claim 5 (depends on 4)

The method of claim 4, wherein the one of the pair of amplification primers has a nucleic acid sequence as set forth in SEQ ID NO: 9.

### Claim 6 (depends on 1)

The method of claim 1, wherein the one of the pair of amplification primers has a nucleic acid sequence as set forth in SEQ ID NO: 9.

### Claim 7 (depends on 1)

The method of claim 1, wherein the cancer is prostate cancer.

### Claim 8 (depends on 1)

The method of claim 1, wherein the cancer is breast cancer.

### Claim 9 (depends on 1)

The method of claim 1, wherein the biological sample is a body tissue or a body fluid.

### Claim 10 (depends on 1)

The method of claim 1, wherein the biological sample is breast tissue, prostate tissue, prostate massage fluid, or urine.

### Claim 11 (depends on 4)

The method of claim 4, wherein the reference value is a cycle threshold.

### Claim 12

A method of monitoring an individual for the development of a cancer, the method comprising:
a) extracting mitochondrial DNA, mtDNA, from a biological sample obtained from the individual at a first time point;
b) quantifying the amount of a mtDNA deletion in the sample, the deletion having a nucleic acid sequence as set forth in SEQ ID NO: 1, the step of quantifying comprising the use of a pair of amplification primers, wherein one of the pair of amplification primers is adapted to bind to a rejoining site of the deletion after the deletion has re-circularized;
c) repeating steps a) to b) at one or more second time points;
wherein an increasing level of the deletion over time is indicative of the presence or development of the cancer.

### Claim 13 (depends on 12)

The method of claim 12 further comprising: (i) comparing the amount of the deletion in the sample to the amount of the deletion in a reference sample of mtDNA from known non-cancerous tissue or body fluid; and/or (ii) comparing the amount of the deletion in the sample to the amount of the deletion in a reference sample of mtDNA from known cancerous tissue or body fluid.

### Claim 14 (depends on 12)

The method of claim 12, wherein the step of quantifying is conducted using real-time PCR.

### Claim 15 (depends on 12)

The method of claim 12, wherein the one of the pair of amplification primers has the nucleic acid sequence set forth in SEQ ID NO: 9.

### Claim 16 (depends on 12)

The method of claim 12, wherein the cancer is prostate cancer.

### Claim 17 (depends on 12)

The method of claim 12, wherein the cancer is breast cancer.

### Claim 18 (depends on 12)

The method of claim 12, wherein the biological sample is a body tissue or a body fluid.

### Claim 19 (depends on 12)

The method of claim 12, wherein the biological sample is breast tissue, prostate tissue, prostate massage fluid, or urine.

### Claim 20

A kit for detecting a cancer in a subject, the kit comprising a pair of PCR primers for amplifying a target region of mitochondrial DNA, mtDNA, in a biological sample obtained from the subject, said target region comprising a region of the mtDNA having a deletion spanning approximately nucleotides 10744 and 14124 of the mtDNA genome.

### Claim 21 (depends on 20)

The kit of claim 20, wherein one primer of the pair of primers is adapted to bind to a region of the mtDNA comprising a splice joining opposite ends of the mtDNA sequence after removal of the deletion.

### Claim 22 (depends on 20)

The kit of claim 20, wherein one primer of the pair of primers has a nucleic acid sequence as set forth in SEQ ID NO: 2.

### Claim 23 (depends on 20)

The kit of claim 20, wherein the cancer is breast cancer or prostate cancer.

### Claim 24 (depends on 20)

The kit of claim 20, further comprising reagents and/or instructions.

### Claim 25

A kit for detecting a cancer in a subject, the kit comprising a pair of PCR primers for amplifying a mitochondrial DNA, mtDNA, deletion in a biological sample obtained from the subject, said deletion having a nucleic acid sequence as set forth in SEQ ID NO: 1, wherein one primer of the pair of primers is adapted to bind to a region of the deletion comprising a rejoining site of the deletion after the deletion has re-circularized.

### Claim 26 (depends on 25)

The kit of claim 25, wherein the one primer of the pair of primers has a nucleic acid sequence as set forth in SEQ ID NO: 2.

### Claim 27 (depends on 25)

The kit of claim 25, wherein the cancer is breast cancer or prostate cancer.

### Claim 28 (depends on 25)

The kit of claim 25, further comprising reagents and/or instructions.