DIAGNOSING FETAL CHROMOSOMAL ANEUPLOIDY USING MASSIVELY PARALLEL GENOMIC SEQUENCING

• US 20090029377A1
• Filed: 07/23/2008
• Published: 01/29/2009
• Est. Priority Date: 07/23/2007
• 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
• Granted Patent: US 12,054,776 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6827 for detection of mutation or polymorphism
• CPC Class Codes: C12Q 1/6883 for diseases caused by alterations of genetic material
• CPC Class Codes: C12Q 1/6888 for detection or identification of organisms
• CPC Class Codes: C12Q 2521/331 Methylation site specific nuclease
• CPC Class Codes: C12Q 2535/122 Massive parallel sequencing
• CPC Class Codes: C12Q 2537/159 Reduction of complexity, e.g. amplification of subsets, removing duplicated genomic regions
• CPC Class Codes: C12Q 2537/16 Assays for determining copy number or wherein the copy number is of special importance
• CPC Class Codes: C12Q 2565/501 being an array of oligonucleotides
• CPC Class Codes: C12Q 2600/112 Disease subtyping, staging or classification
• CPC Class Codes: C12Q 2600/154 Methylation markers
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: G01N 2800/387 Down syndrome; Trisomy 18; Trisomy 13
• 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: Y02A 90/10 Information and communication technologies [ICT] supporting adaptation to climate change, e.g. for weather forecasting or climate simulation

Abstract

Embodiments of this invention provide methods, systems, and apparatus for determining whether a fetal chromosomal aneuploidy exists from a biological sample obtained from a pregnant female. Nucleic acid molecules of the biological sample are sequenced, such that a fraction of the genome is sequenced. Respective amounts of a clinically-relevant chromosome and of background chromosomes are determined from results of the sequencing. A parameter derived from these amounts (e.g. a ratio) is compared to one or more cutoff values, thereby determining a classification of whether a fetal chromosomal aneuploidy exists.

Claims

1. A method for performing prenatal diagnosis of a fetal chromosomal aneuploidy in a biological sample obtained from a pregnant female subject, wherein the biological sample includes nucleic acid molecules, the method comprising:

receiving the biological sample;

sequencing at least a portion of a plurality of the nucleic acid molecules contained in the biological sample, wherein the sequenced portion represents a fraction of the human genome;

based on the sequencing;

determining a first amount of a first chromosome from sequences identified as originating from the first chromosome;

determining a second amount of one or more second chromosomes from sequences identified as originating from one of the second chromosomes;

determining a parameter from the first amount and the second amount;

comparing the parameter to one or more cutoff values; and

based on the comparison, determining a classification of whether a fetal chromosomal aneuploidy exists for the first chromosome.

2. The method of claim 1, wherein the sequencing is performed randomly on a portion of the nucleic acid molecules contained in the biological sample.

3. The method of claim 1, wherein the biological sample is maternal blood, plasma, serum, urine or saliva

4. The method of claim 1, wherein the biological sample is transcervical lavage fluid.

5. The method of claim 1, wherein the first chromosome is chromosome 21, chromosome 18, chromosome 13, chromosome X, or chromosome Y.

6. The method of claim 1, wherein the parameter is a ratio of sequences that originate from the first chromosome.

7. The method of claim 6, wherein the ratio is obtained from any one or more of a fractional count of the number of sequenced tags, a fractional number of sequenced nucleotides, and a fractional length of accumulated sequences.

8. The method of claim 6, wherein the sequences that originate from the first chromosome are selected to be less than a specified number of base pairs.

9. The method of claim 8, wherein the specified number of base pairs is 300 bp, 200 bp, or 100 bp.

10. The method of claim 1, wherein the nucleic acid molecules of the biological sample have been enriched for sequences originating from at least one particular chromosome.

11. The method of claim 1, wherein the nucleic acid molecules of the biological sample have been enriched for sequences less than 300 bp.

12. The method of claim 1, wherein the nucleic acid molecules of the biological sample have been enriched for sequences less than 200 bp.

13. The method of claim 1, wherein the nucleic acid molecules of the biological sample have been amplified using a polymerase chain reaction.

14. The method of claim 1, wherein the sequenced portion represents at least a pre-determined fraction of the human genome.

15. The method of claim 1, wherein the fraction represents at least 0.1% of the human genome.

16. The method of claim 1, wherein the fraction represents at least 0.5% of the human genome.

17. The method of claim 1, wherein at least one of the cutoff values is related to the fractional concentration of fetal DNA in the biological sample.

18. The method of claim 17, wherein the fractional concentration of fetal DNA in the biological sample is determined by any one or more of a proportion of Y chromosome sequences, a fetal epigenetic marker, or using single nucleotide polymorphism analysis.

19. The method of claim 1, wherein a cutoff value is a reference value established in a normal biological sample.

20. The method of claim 1, further comprising:

identifying an amount of fetal DNA in the biological sample; and

calculating a number N of sequences to be analyzed based on a desired accuracy.

21. A computer program product comprising a computer readable medium encoded with a plurality of instructions for controlling a computing system to perform an operation for performing prenatal diagnosis of a fetal chromosomal aneuploidy in a biological sample obtained from a pregnant female subject, wherein the biological sample includes nucleic acid molecules, the operation comprising the steps of:

receiving data from a random sequencing of a portion of the nucleic acid molecules contained in the biological sample obtained from a pregnant female subject, wherein the biological sample includes nucleic acid molecules, wherein the portion represents a fraction of the human genome;

based on the data from the random sequencing;

determining a first amount of a first chromosome from sequences identified as originating from the first chromosome;

determining a second amount of one or more second chromosomes from sequences identified as originating from one of the second chromosomes;

determining a parameter from the first amount and the second amount;

comparing the parameter to one or more cutoff values; and

based on the comparison, determining a classification of whether a fetal chromosomal aneuploidy exists for the first chromosome.

22. A method for performing prenatal diagnosis of a fetal chromosomal aneuploidy in a biological sample obtained from a pregnant female subject, wherein the biological sample includes nucleic acid molecules, the method comprising:

receiving the biological sample;

calculating a number N of sequences to be analyzed based on a desired accuracy;

randomly sequencing at least N of the nucleic acid molecules contained in the biological sample, wherein the portion represents a fraction of the human genome;

based on the random sequencing;

determining a first amount of a first chromosome from sequences identified as originating from the first chromosome;

determining a second amount of one or more second chromosomes from sequences identified as originating from one of the second chromosomes;

determining a parameter from the first amount and the second amount;

comparing the parameter to one or more cutoff values; and

based on the comparison, determining a classification of whether a fetal chromosomal aneuploidy exists for the first chromosome.

23. The method of claim 22, further comprising:

identifying a percentage of fetal DNA in the biological sample, wherein calculating a number N of sequences to be analyzed based on a desired accuracy is based on the percentage.