Methods and processes for non-invasive assessment of genetic variations

• US 10,622,094 B2
• Filed: 06/20/2014
• Issued: 04/14/2020
• Est. Priority Date: 06/21/2013
• Status: Active Grant

• Current Assignee: Sequenom Incorporated (Laboratory Corporation Of America Holdings)
• Original Assignee: Sequenom Incorporated (Laboratory Corporation Of America Holdings)
• Inventors: Kim, Sung K., Hannum, Gregory, Geis, Jennifer, Deciu, Cosmin
• Primary Examiner(s): Brusca, John S
• Application Number:
  US14/311,070
• Publication Number:
  US 20150005176A1
• Time in Patent Office: 2,125 Days
• Field of Search: None
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6872 involving mass spectrometry
• CPC Class Codes: C12Q 2537/165 Mathematical modelling, e.g. logarithm, ratio
• 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 30/00 ICT specially adapted for sequence analysis involving nucleotides or amino acids
• CPC Class Codes: G16B 30/10 Sequence alignment; Homology search
• CPC Class Codes: G16H 10/40 for data related to laboratory analysis, e.g. patient specimen analysis
• CPC Class Codes: G16H 50/20 for computer-aided diagnosis, e.g. based on medical expert systems
• CPC Class Codes: Y02A 90/10 Information and communication technologies [ICT] supporting adaptation to climate change, e.g. for weather forecasting or climate simulation

Abstract

Provided herein are methods, processes, systems, machines and apparatuses for non-invasive assessment of genetic variations.

Claims

1. A computer-implemented, fetal gender-independent, fetal ploidy independent and fetal polymorphism-independent method for determining a fraction of fetal nucleic acid in a test sample from sequenced test sample nucleic acid, the method comprising:

(a) receiving into a memory counts of sequence reads of sequenced test sample nucleic acid that are mapped to corresponding portions of a reference genome, which test sample nucleic acid comprises circulating cell-free (CCF) nucleic acid from blood of a pregnant female bearing a fetus;

(b) receiving into a memory a set of weighting factors that have been calculated for the corresponding portions of the reference genome from a fitted relation between (i) a fraction of fetal nucleic acid for each of multiple training samples, and (ii) counts of sequence reads mapped to each of the multiple training samples, wherein at least one weighting factor corresponds to each of the corresponding portions and is indicative of the fraction of fetal nucleic acid in each of the corresponding portions;

(c) assigning, using a microprocessor, the set of weighting factors to the corresponding portions of the reference genome, wherein each of the corresponding portions is independently associated with a corresponding weighting factor;

(d) generating, using a microprocessor, portion-specific fetal fraction calculations for the corresponding portions according to (i) the count of the sequence reads of the sequenced test sample nucleic acid mapped to the portion of the reference genome for each portion of the sequenced test sample, and (ii) the assigned weighting factor for each portion of the sequenced test sample; and

(e) determining, using a microprocessor, a fraction of fetal nucleic acid for the test sample based on the portion-specific fetal fraction calculations.

2. The method of claim 1, wherein the weighting factors are associated with portions in a plurality of portions corresponding to autosomes.

3. The method of claim 2, wherein the weighting factors are associated with portions that do not correspond to any one or more of chromosomes 13, 18, and 21.

4. The method of claim 1, wherein the counts in (a) or (b)(ii) are normalized counts.

5. The method of claim 4, wherein the normalized counts have reduced guanine-cytosine (GC) bias with respect to raw counts.

6. The method of claim 1, wherein determining the fraction of fetal nucleic acid for the test sample comprises averaging or summing the portion-specific fetal fraction calculations.

7. The method of claim 1, wherein the weighting factor for each corresponding portion is related to the average ratio for the corresponding portion for the multiple training samples.

8. The method of claim 1, wherein the weighting factor for each corresponding portion is proportional to the average number of reads from CCF fetal nucleic acid fragments mapped to the corresponding portion for the multiple samples.

9. The method of claim 1, wherein the corresponding portions are chosen from discrete genomic bins, genomic bins having sequential sequences of predetermined length, variable-size bins, point-based views of a smoothed coverage map, or a combination thereof.

10. The method of claim 1, wherein each of the portions are about 10 contiguous kilobases to about 75 contiguous kilobases from the reference genome.

11. The method of claim 1, wherein about 75% or more of the weighting factors are greater than zero.

12. The method of claim 1, wherein the weighting factors are estimated coefficients from the fitted relations in (b).

13. The method of claim 1, which further comprises estimating coefficients for each portion from the fitted relations in (b).

14. The method of claim 12, wherein each of the fitted relations in (b) is a regression model and the weighting factors are, or are based on, regression coefficients from the fitted relations.

15. The method of claim 1, wherein the fitted relations in (b) are fitted by an estimation chosen from least squares, ordinary least squares, linear, partial, total, generalized, weighted, non-linear, iteratively reweighted, ridge regression, least absolute deviations, Bayesian, Bayesian multivariate, reduced-rank, LASSO, elastic net estimator, and any combination thereof.

16. The method of claim 1, further comprising, prior to (a), sequencing the test sample nucleic acid by a massively parallel sequencing process, thereby generating sequence reads;

and mapping at least some of the sequence reads to a subset of the portions of the reference genome and counting the sequence reads mapped to the subset of the portions, thereby obtaining counts of sequence reads mapped to the subset of the portions,

wherein each portion in the subset of the portions corresponds to a portion of the reference genome based on an observed proportion of fetal nucleic acid, a specific fragment length ratio, or a predetermined sequence length of the each portion, and

wherein the number of portions in the subset of the portions is less than the number of portions of the reference genome.

17. The method of claim 1, further comprising outputting a classification of the presence or absence of a fetal chromosome copy number variation for the test sample based on the fraction of fetal nucleic acid determined in (e).

18. The method of claim 17, wherein the fetal chromosome copy number variation is an aneuploidy.

19. The method of claim 18, wherein the aneuploidy is a trisomy that is determined with a sensitivity of 95% or greater or a specificity of 95% or greater, or a sensitivity of 95% or greater and a specificity of 95% or greater.

20. The method of claim 16, wherein the test sample is sequenced at a fold coverage less than 1.