Using size and number aberrations in plasma DNA for detecting cancer

• US 10,364,467 B2
• Filed: 01/12/2016
• Issued: 07/30/2019
• Est. Priority Date: 01/13/2015
• Status: Active Grant

• Current Assignee: Chinese University of Hong Kong
• Original Assignee: Chinese University of Hong Kong
• Inventors: Lo, Yuk-Ming Dennis, Chiu, Rossa Wai Kwun, Chan, Kwan Chee, Jiang, Peiyong
• Primary Examiner(s): Zeman, Mary K
• Application Number:
  US14/994,053
• Publication Number:
  US 20160201142A1
• Time in Patent Office: 1,295 Days
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6886 for cancer immunoassay for cancer G01N33/574
• CPC Class Codes: C12Q 2535/122 Massive parallel sequencing
• CPC Class Codes: C12Q 2537/16 Assays for determining copy number or wherein the copy number is of special importance
• CPC Class Codes: C12Q 2600/112 Disease subtyping, staging or classification
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: G16B 20/00 ICT specially adapted for functional genomics or proteomics, e.g. genotype-phenotype associations
• CPC Class Codes: G16B 20/10 Ploidy or copy number detection
• CPC Class Codes: G16B 20/20 Allele or variant detection, e.g. single nucleotide polymorphism [SNP] detection
• CPC Class Codes: G16B 40/00 ICT specially adapted for biostatistics; ICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding
• CPC Class Codes: G16B 40/30 Unsupervised data analysis

Abstract

Analysis of tumor-derived circulating cell-free DNA opens up new possibilities for performing liquid biopsies for solid tumor assessment or cancer screening. However, many aspects of the biological characteristics of tumor-derived cell-free DNA remain unclear. Regarding the size profile of plasma DNA molecules, some studies reported increased integrity of tumor-derived plasma DNA while others reported shorter tumor-derived plasma DNA molecules. We performed an analysis of the size profiles of plasma DNA in patients with cancer using massively parallel sequencing at single base resolution and in a genomewide manner. Tumor-derived plasma DNA molecules were further identified using chromosome arm-level z-score analysis (CAZA). We showed that populations of aberrantly short and long DNA molecules co-existed in the plasma of patients with cancer. The short ones preferentially carried the tumor-associated copy number aberrations. These results show the ability to use plasma DNA as a molecular diagnostic tool.

Claims

1. A method of analyzing a biological sample of an organism, the biological sample including nucleic acid molecules originating from normal cells and potentially from cells associated with cancer, wherein at least some of the nucleic acid molecules are cell-free in the biological sample, the method comprising:

identifying a plurality of chromosomal regions of the organism, each chromosomal region including a plurality of loci;

for each of a plurality of the nucleic acid molecules in the biological sample;

measuring a size of the nucleic acid molecule; and

identifying a location of the nucleic acid molecule in a reference genome of the organism;

for each of the plurality of chromosomal regions;

identifying a respective group of nucleic acid molecules as being from the chromosomal region based on the identified locations, the respective group including at least one nucleic acid molecule located at each of the plurality of loci of the chromosomal region;

calculating, with a computer system, a respective amount of the respective group of nucleic acid molecules; and

comparing the respective amount to a count reference value to determine a count classification of whether the chromosomal region exhibits an aberration, the count classification for a first chromosomal region of the plurality of chromosomal regions indicating a first aberration;

identifying a first group of nucleic acid molecules as being from the first chromosomal region based on the identified locations;

calculating, with a computer system, a first statistical value of a first size distribution of the first group of nucleic acid molecules;

comparing the first statistical value to a size reference value to determine a size classification of whether the first chromosomal region exhibits the first aberration;

determining a final classification of whether the first chromosomal region exhibits the first aberration using the count classification and the size classification of the first chromosomal region; and

determining whether cancer exists in the organism using the final classification.

2. The method of claim 1, wherein the final classification is that the first aberration exists only when the count classification and the size classification indicate a same aberration.

3. The method of claim 1, wherein the plurality of chromosomal regions are non-overlapping.

4. The method of claim 1, wherein the final classification is that the first aberration exists, the method further comprising:

obtaining a calibration function that provides a relationship between a size of a tumor and the first statistical value; and

determining the size of the tumor using the calibration function.

5. The method of claim 4, wherein the calibration function is determined from calibration data points of reference samples from organisms with tumors of known size, wherein a calibration data point includes a measurement of the size of the tumor and a corresponding statistical measurement of sizes of nucleic acid molecules from the first chromosomal region.

6. The method of claim 1, wherein the respective value comprises a mean of the first size distribution, a median of the first size distribution, a mode of the of the first size distribution, or a proportion of nucleic acid molecules having a size below a size threshold.

7. The method of claim 1, wherein the size reference value corresponds to a second statistical value of a second size distribution of a second chromosomal region.

8. The method of claim 7, wherein comparing the first statistical value to the size reference value includes:

determining a separation value between the first statistical value and the size reference value; and

comparing the separation value to a threshold value.

9. The method of claim 1, further comprising:

determining a set of size classifications for a set of chromosomal regions identified as aberrant based on corresponding count classifications; and

confirming whether the set of chromosomal regions are aberrant based on the set of size classifications.

11. The method of claim 1, further comprising:

for each of the plurality of chromosomal regions;

identifying a respective group of nucleic acid molecules as being from the chromosomal region based on the identified locations,

calculating, with a computer system, a respective statistical value of a respective size distribution of the respective group of nucleic acid molecules,

comparing the respective statistical value to the size reference value to determine a size classification of whether the chromosomal region exhibits the aberration, and

determining a respective final classification of whether the chromosomal region exhibits the aberration using the respective count classification and the respective size classification of the chromosomal region.

12. The method of claim 11, wherein the count classification for each of the plurality of chromosomal regions is that the chromosomal region exhibits an aberration.

13. The method of claim 11, wherein determining whether cancer exists in the organism comprises:

determining an amount of chromosomal regions exhibiting an aberration; and

comparing the amount of chromosomal regions to a threshold.

10. A computer product comprising a computer readable medium storing a plurality of instructions for controlling a computer system to perform a method, the method comprising:

identifying a plurality of chromosomal regions of the organism, each chromosomal region including a plurality of loci;

for each of a plurality of the nucleic acid molecules in the biological sample;

measuring a size of the nucleic acid molecule; and

identifying a location of the nucleic acid molecule in a reference genome of the organism;

for each of the plurality of chromosomal regions;

identifying a respective group of nucleic acid molecules as being from the chromosomal region based on the identified locations, the respective group including at least one nucleic acid molecule located at each of the plurality of loci of the chromosomal region;

calculating, with a computer system, a respective amount of the respective group of nucleic acid molecules; and

comparing the respective amount to a count reference value to determine a count classification of whether the chromosomal region exhibits an aberration, the count classification for a first chromosomal region of the plurality of chromosomal regions indicating a first aberration;

identifying a first group of nucleic acid molecules as being from the first chromosomal region based on the identified locations;

calculating, with a computer system, a first statistical value of a first size distribution of the first group of nucleic acid molecules;

comparing the first statistical value to a size reference value to determine a size classification of whether the first chromosomal region exhibits the first aberration;

determining a final classification of whether the first chromosomal region exhibits the first aberration using the count classification and the size classification of the first chromosomal region; and

determining whether cancer exists in the organism using the final classification.