METHODS OF DETECTING AND ENRICHING CIRCULATING TUMOR DNA
The present disclosure relates to a laboratory execution system that provides for automation of laboratory processes. A centralized data management system may be dynamically updated and used to facilitate management of components of the laboratory execution system, such as an automation system and an analytics results management system that may facilitate complex analytical functions, such as synthesizing raw test data. Potential workflows include the detection of specific molecules of interest.
1 - 30 . (canceled)
31 . A method for preparing a deoxyribonucleic acid (DNA) fraction enriched with circulating tumor DNA (ctDNA) from a cancer patient, comprising:
(a) obtaining a fluid sample taken from the cancer patient following completion of a cancer treatment, wherein the fluid sample comprises blood plasma or blood serum;
(b) extracting cell-free DNA (cfDNA) from the fluid sample;
(c) enriching a fraction of ctDNA by contacting the cfDNA with a plurality of oligonucleotides, wherein the plurality of oligonucleotides comprises at least 10 distinct oligonucleotides and each of the at least 10 distinct oligonucleotides comprises a nucleic acid sequence that hybridizes to a DNA fragment comprising a corresponding tumor-specific somatic mutation sequence selected from a predetermined set of tumor-specific somatic mutation sequences, wherein the predetermined set of tumor-specific somatic mutation sequences comprises somatic mutations that are specific to the subject based on a comparison of sequence reads from sequencing DNA from a tumor sample from the subject and sequence reads from sequencing DNA from a non-tumor sample from the subject.
32 . The method of claim 31 , wherein enriching the fraction of cfDNA corresponding to ctDNA comprises:
(i) hybrid capture-based enrichment;
(ii) PCR-target enrichment; or
(iii) on-sequencer enrichment.
33 . The method of claim 31 , wherein sequencing genomic DNA from the tumor sample of the patient and sequencing genomic DNA from the non-tumor sample of the patient comprises whole genome sequencing or targeted sequencing of exomes.
34 . The method of claim 31 , further comprising:
sequencing the ctDNA, thereby obtaining a plurality of sequence reads; and
detecting the presence or absence of a DNA fragment comprising any one of the set of tumor-specific somatic mutation sequences.
35 . The method of claim 31 , further comprising repeating (a)-(c) one or more times.
36 . The method of claim 31 , wherein the comparison of sequence reads from sequencing DNA from the tumor sample from the subject and sequence reads from sequencing DNA from the non-tumor sample from the subject does not comprise genotyping the DNA from the tumor sample and non-tumor sample.
37 . A method for preparing a deoxyribonucleic acid (DNA) fraction enriched with circulating tumor DNA (ctDNA) from a cancer patient, comprising:
(a) obtaining a fluid sample taken from the cancer patient while the cancer patient is in remission, wherein the fluid sample comprises blood plasma or blood serum;
(b) extracting cell-free DNA (cfDNA) from the fluid sample;
(c) enriching a fraction of ctDNA by contacting the cfDNA with a plurality of oligonucleotides, wherein the plurality of oligonucleotides and each oligonucleotide in the plurality comprises a nucleic acid sequence that hybridizes to a DNA fragment comprising a corresponding tumor-specific somatic mutation sequence selected from a predetermined set of tumor-specific somatic mutation sequences, wherein the predetermined set of tumor-specific somatic mutation sequences comprises somatic mutations that are specific to the subject based on a comparison of sequence reads from sequencing DNA from a tumor sample from the subject and sequence reads from sequencing DNA from a non-tumor sample from the subject.
38 . The method of claim 37 , wherein enriching the fraction of cfDNA corresponding to ctDNA comprises:
(i) hybrid capture-based enrichment;
(ii) PCR-target enrichment; or
(iii) on-sequencer enrichment.
39 . The method of claim 37 , wherein sequencing genomic DNA from the tumor sample of the patient and sequencing genomic DNA from the non-tumor sample of the patient comprises whole genome sequencing or targeted sequencing of exomes.
40 . The method of claim 37 , further comprising:
sequencing the ctDNA, thereby obtaining a plurality of sequence reads; and
detecting the presence or absence of a DNA fragment comprising any one of the set of tumor-specific somatic mutation sequences.
41 . The method of claim 37 , further comprising repeating (a)-(c) one or more times.
42 . The method of claim 37 , wherein the plurality of oligonucleotides comprises at least 10 distinct oligonucleotides capable of detecting at least 50 different tumor-specific somatic mutations.
43 . A method for preparing a deoxyribonucleic acid (DNA) fraction enriched with circulating tumor DNA (ctDNA) from a cancer patient, comprising:
(a) obtaining a fluid sample taken from the cancer patient at a time coinciding with or prior to surgery; following, during, or prior to administration of chemotherapy; or following, during, or prior to radiation therapy; wherein the fluid sample comprises blood plasma or blood serum;
(b) extracting cell-free DNA (cfDNA) from the fluid sample;
(c) enriching a fraction of ctDNA by contacting the cfDNA with a plurality of oligonucleotides, wherein the plurality of oligonucleotides and each oligonucleotide in the plurality comprises a nucleic acid sequence that hybridizes to a DNA fragment comprising a corresponding tumor-specific somatic mutation sequence selected from a predetermined set of tumor-specific somatic mutation sequences, wherein the predetermined set of tumor-specific somatic mutation sequences comprises somatic mutations that are specific to the subject based on a comparison of sequence reads from sequencing DNA from a tumor sample from the subject and sequence reads from sequencing DNA from a non-tumor sample from the subject.
44 . The method of claim 43 , wherein enriching the fraction of cfDNA corresponding to ctDNA comprises:
(i) hybrid capture-based enrichment;
(ii) PCR-target enrichment; or
(iii) on-sequencer enrichment.
45 . The method of claim 43 , wherein sequencing genomic DNA from the tumor sample of the patient and sequencing genomic DNA from the non-tumor sample of the patient comprises whole genome sequencing or targeted sequencing of exomes.
46 . The method of claim 43 , further comprising:
sequencing the ctDNA, thereby obtaining a plurality of sequence reads; and
detecting the presence or absence of a DNA fragment comprising any one of the set of tumor-specific somatic mutation sequences.
47 . The method of claim 43 , wherein the comparison of sequence reads from sequencing DNA from the tumor sample from the subject and sequence reads from sequencing DNA from the non-tumor sample from the subject does not comprise genotyping the DNA from the tumor sample and non-tumor sample.
48 . The method of claim 43 , wherein the fluid sample is taken from the cancer patient at a time coinciding with or prior to surgery.
49 . The method of claim 43 , wherein the fluid sample is taken from the cancer patient at a time following, during, or prior to administration of chemotherapy or radiation therapy.
50 . The method of claim 43 , further comprising repeating (a)-(c) one or more times.