Breast cancer associated circulating nucleic acid biomarkers
The invention provides methods and reagents for diagnosing breast cancer that are based on the detection of biomarkers in the circulating nucleic acids from a patient to be evaluated.
1. A method of analyzing circulating cell-free DNA in a patient sample to determine copy number of cell-free DNA present in the sample, comprising
(a) isolating cell-free DNA from a blood, serum, or plasma sample from a human patient;
(b) preparing a sequencing library from the cell-free DNA obtained in (a) and performing whole-genome sequencing;
(c) aligning sequences of cell-free DNA molecules to a human genome database;
(d) determining the normalized copy number in the sample of at least one cell-free DNA that has a sequence, free of repetitive elements, that is unambiguously assigned to a first chromosomal region Hs3, region 26316904-26317098, corresponding to nucleotide sequence SEQ ID NO:3023 as determined by human genome build 36; a second chromosomal region Hs5, region 138546637-138546876, corresponding to nucleotide sequence SEQ ID NO:3031 as determined by human genome build 36; or a third chromosomal region Hs5, region 1895609-1895744, corresponding to nucleotide sequence SEQ ID NO:3032 as determined by human genome build 36; and
(e) identifying in the sample at least one of the cell-free DNA sequences unambiguously assigned to the first chromosomal region, the second chromosomal region, or the third chromosomal region that is increased in copy number by at least 1.5-fold relative to a reference value from normal controls.
2. The method of claim 1 , wherein said blood, serum, or plasma sample is from a human patient who has been diagnosed with breast cancer.
3. The method of claim 2 , further comprising administering a treatment for breast cancer to the human patient having an increase in copy number of at least one region by at least 1.5-fold relative to the reference value obtained from normal controls.
4. A method of analyzing circulating cell-free DNA in a patient sample to determine copy number of cell-free DNA present in the sample, comprising
(a) isolating cell-free DNA from a blood, serum, or plasma sample from a human patient;
(b) performing sequencing of the cell-free DNA obtained in (a) using an Illumina or SOLiD next generation sequencing platform;
(c) aligning sequences of cell-free DNA molecules to a human genome database;
(d) determining the normalized copy number in the sample of at least one cell-free DNA that has a sequence, free of repetitive elements, that is unambiguously assigned to a first chromosomal region Hs3, region 26316904-26317098, corresponding to nucleotide sequence SEQ ID NO:3023 as determined by human genome build 36; a second chromosomal region Hs5, region 138546637-138546876, corresponding to nucleotide sequence SEQ ID NO:3031 as determined by human genome build 36; or a third chromosomal region Hs5, region 1895609-1895744, corresponding to nucleotide sequence SEQ ID NO:3032 as determined by human genome build 36; and
(e) identifying in the sample at least one of the cell-free DNA sequences unambiguously assigned to the first chromosomal region, the second chromosomal region, or the third chromosomal region that is increased in copy number by at least 1.5-fold relative to a reference value from normal controls.