Methylation pattern analysis of tissues in a dna mixture

Patent metadata

Field	Value
Publication number	US-2016017419-A1
Application number	US-201514803692-A
Country	US
Kind code	A1
Filing date	Jul 20, 2015
Priority date	Jul 18, 2014
Publication date	Jan 21, 2016
Grant date	—

Abstract

The contributions of different tissues to a DNA mixture are determined using methylation levels at particular genomic sites. Tissue-specific methylation levels of M tissue types can be used to deconvolve mixture methylation levels measured in the DNA mixture, to determine fraction contributions of each of the M tissue types. Various types of genomic sites can be chosen to have particular properties across tissue types and across individuals, so as to provide increased accuracy in determining contributions of the various tissue types. The fractional contributions can be used to detect abnormal contributions of a particular tissue, indicating a disease state for the tissue. A differential in fractional contributions for different sizes of DNA fragments can also be used to identify a diseased state of a particular tissue. A sequence imbalance for a particular chromosomal region can be detected in a particular tissue, e.g., identifying a location of a tumor.

First claim

What is claimed is:

1 . A method of analyzing a biological sample of an organism, the biological sample including a mixture of cell-free DNA molecules from a plurality of tissues types, including a first tissue type, the method comprising:
analyzing, by a computer system, a plurality of cell-free DNA molecules from the biological sample, the plurality of cell-free DNA molecules being at least 1,000 cell-free DNA molecules, wherein analyzing a cell-free DNA molecule includes:
identifying a location of the cell-free DNA molecule in a reference genome corresponding to the organism;
identifying a first set of the plurality of cell-free DNA molecules that are each located at any one of N genomic sites of a first chromosomal region of the reference genome corresponding to the organism, N being an integer greater than or equal to 10; measuring N first mixture methylation levels at the N genomic sites using the first set of the plurality of cell-free DNA molecules; determining, by the computer system, a first fractional contribution of the first tissue type in the mixture using the N first methylation levels; identifying a second set of the plurality of cell-free DNA molecules that are each located at any one of K genomic sites of a second chromosomal region of the reference genome corresponding to the organism, K being an integer greater than or equal to 10, wherein the second chromosomal region is different from the first chromosomal region; measuring K second mixture methylation levels at the K genomic sites using the second set of the plurality of cell-free DNA molecules; determining, by the computer system, a second fractional contribution of the first tissue type in the mixture using the K second methylation levels; computing a first separation value between the first fractional contribution and the second fractional contribution; and comparing the first separation value to a threshold value to determine a classification of whether the first tissue type has a sequence imbalance for the first chromosomal region.

2 . The method of claim 1 , wherein measuring N first mixture methylation levels at the N genomic sites includes analyzing methylation-aware sequencing results, and wherein the locations of the plurality of cell-free DNA molecules are determined using the methylation-aware sequencing results.

3 . The method of claim 1 , wherein the organism is pregnant with a fetus, wherein the first tissue type is placental tissue, the method further comprising:
detecting whether the fetus has an aneuploidy in the first chromosomal region based on the classification.

4 . The method of claim 1 , wherein the organism is pregnant with a fetus, and wherein the first tissue type is not placental tissue.

5 . The method of claim 1 , wherein the classification is that the first tissue type has the sequence imbalance for the first chromosomal region when the first separation value exceeds the threshold value.

6 . The method of claim 5 , further comprising:
computing a plurality of separation values for the first tissue type, each of the plurality of separation values corresponding to a different chromosomal region; and determining a number of the different chromosomal regions having a corresponding separation value that exceeds the threshold value.

7 . The method of claim 6 , further comprising:
determining the classification of whether the first tissue type has the sequence imbalance for the first chromosomal region based on the number of the different chromosomal regions having a corresponding separation value that exceeds the threshold value.

8 . The method of claim 7 , wherein the first tissue type is determined to have the sequence imbalance for the first chromosomal region when the number of the different chromosomal regions exceeds a cutoff value.

9 . The method of claim 8 , wherein the threshold value is dependent on the cutoff value.

10 . The method of claim 6 , wherein each of the plurality of separation values are computed using the second fractional contribution.

11 . The method of claim 6 , further comprising:
determining a level of cancer for the first tissue type based on the number of the different chromosomal regions.

12 . The method of claim 11 , further comprising:
for each of the number of the different chromosomal regions:
determining an extent that the corresponding separation value exceeds the threshold value,
wherein determining the level of cancer for the first tissue type is further based on the extents that the corresponding separation value exceeds the threshold value.

13 . The method of claim 12 , wherein determining the extent that the corresponding separation value exceeds the threshold value includes:
comparing corresponding separation value to a plurality of threshold values.

14 . The method of claim 1 , further comprising:
identifying the first chromosomal region as exhibiting a copy number aberration based on a first amount of cell-free DNA molecules that are located in the first chromosomal region; and determining which of M tissue types is associated with the copy number aberration by testing corresponding separation values in fractional contributions for each of the M tissue types, the first tissue type being one of the M tissue types.

15 . The method of claim 14 , wherein determining which of M tissue types is associated with the copy number aberration identifies at least two tissue types as being associated with the copy number aberration by the at least two tissue types each having a corresponding separation value that exceeds the threshold value.

16 . The method of claim 15 , wherein the first tissue type is one of the at least two tissue types, the method further comprising:
identifying the first tissue type as an origin of primary cancer when the first separation value has a highest value among the corresponding separation values.

17 . The method of claim 14 , wherein the first chromosomal region exhibits an amplification, the method further comprising:
identifying the second chromosomal region as exhibiting a deletion based on a second amount of cell-free DNA molecules that are located in the second chromosomal region; and using the second chromosomal region to determine the first separation value based on the second chromosomal region exhibiting of the deletion.

18 . The method of claim 1 , wherein the first chromosomal region and the second chromosomal region are different chromosomes.

19 . The method of claim 1 , wherein K does not equal N.

20 . The method of claim 1 , wherein the first chromosomal region is composed of non-contiguous subregions of a first chromosome for which the sequence imbalance is being tested.

21 . The method of claim 1 , wherein the second chromosomal region is composed of non-contiguous subregions of one or more reference chromosomes that do not include the first chromosomal region.

22 . The method of claim 1 , wherein the first chromosomal region exhibits an amplification and wherein the second chromosomal region exhibits a deletion.

23 . The method of claim 1 , wherein analyzing the plurality of cell-free DNA molecules comprises:
sequencing the plurality of cell-free DNA molecules to obtain sequence reads; and aligning the sequence reads to the reference genome, wherein the N mixture methylation levels are measured using sequence reads that each aligns to at least one of the N genomic sites of the reference genome.

Assignees

• Univ Hong Kong Chinese

Inventors

• Chiu Rossa Wai Kwun
• Chan Kwan Chee
• Lo Yuk-Ming Dennis
• Jiang Peiyong
• Sun Kun

Classifications

• C12Q1/6809

  Methods for determination or identification of nucleic acids involving differential detection · CPC title

• G16B30/00

  ICT specially adapted for sequence analysis involving nucleotides or amino acids · CPC title

• G16B20/00 Primary

  ICT specially adapted for functional genomics or proteomics, e.g. genotype-phenotype associations · CPC title

• C12Q1/689

  for bacteria · CPC title

• G06F19/22

  Physics · mapped topic