METHODS AND SYSTEMS FOR DETECTING RESIDUAL DISEASE
Described herein are methods, devices, and systems for measuring a level of a disease (such as cancer), for example a fraction of nucleic acid molecules (such as cell-free DNA) in a sample from an individual that relate to diseased tissue (such as cancer tissue). Also described are methods, devices, and systems for measuring a presence, recurrence, progression, or regression of the disease in the individual. Certain methods include comparing, using nucleic acid sequencing data associated with the individual, a signal indicative of a rate at which sequenced loci selected from a personalized disease-associated small nucleotide variant (SNV) locus panel are derived from a diseased tissue to a background factor indicative of a sequencing false positive error rate, or a noise factor indicative of a sampling variance, across the selected loci.
1 . A method of measuring a level of a disease in an individual, comprising:
comparing, using nucleic acid sequencing data associated with the individual, a signal indicative of a rate at which sequenced loci selected from a personalized disease-associated small nucleotide variant (SNV) locus panel are derived from a diseased tissue to a background factor indicative of a sequencing false positive error rate across the selected loci; and
determining the level of the disease in the individual based on the comparison of the signal to the background factor.
2 . The method of claim 1 , wherein the level of the disease is a fraction of nucleic acid molecules associated with the disease in a sample from the individual.
3 . The method of claim 1 , wherein comparing comprises subtracting the background factor from the signal.
4 . The method of claim 1 , further comprising determining an error for the measurement of the level of the disease.
5 . The method of claim 4 , wherein the error is a confidence interval for the level of the disease.
6 . The method of claim 4 , wherein the error is proportional to a total number of individual small nucleotide variant reads detected at the selected loci.
7 . (canceled)
8 . The method of claim 1 , wherein the method comprises measuring a recurrence of the disease.
9 . The method of claim 1 , wherein the method comprises measuring a progression or regression of the disease by comparing the measured level of the disease to a previously measured level of the disease.
10 . The method of claim 9 , wherein progression or regression of the disease is based on a statistically significant change in the measured level of the disease.
11 . A method of detecting a disease in an individual, comprising:
comparing, using nucleic acid sequencing data associated with the individual, a signal indicative of a rate at which sequenced loci selected from a personalized disease-associated small nucleotide variant (SNV) locus panel are derived from a diseased tissue to a noise factor indicative of a sampling variance across the selected loci; and
determining whether the individual has the disease based on the comparison of the signal to the noise factor.
12 . The method of claim 11 , wherein the individual is determined to have a disease recurrence or a residual level of the disease if the signal exceeds the noise factor by more than a predetermined threshold.
13 - 16 . (canceled)
17 . The method of claim 11 , wherein the method comprises detecting a recurrence of the disease.
18 . The method of claim 1 , wherein a magnitude of the signal depends on at least a number of selected loci and an average sequencing depth associated with the nucleic acid sequencing data.
19 . A method of detecting a presence, a progression, or a regression, of a disease in an individual, comprising:
measuring at least one of:
(a) a likelihood that a value indicative of a fraction, F, of nucleic acid molecules in a sample that originate from a diseased tissue of the individual is greater than zero, wherein F being greater than zero is indicative of a presence of the disease in the individual, and
(b) a statistically significant change in a value indicative of the fraction, F, of nucleic acid molecules in a sample that originate from a diseased tissue of the individual, wherein the statistically significant change is relative to a previously measured fraction, F prior , and wherein a statistically significant change in F indicates progression or regression of the disease in the individual;
wherein the fraction F is determined by comparing a total number of single nucleotide variants (SNVs) detected in cell-free nucleic acid sequencing data, N total , wherein the SNVs are selected from a personalized disease-associated SNV locus panel, to the number of SNVs selected from the SNV panel, N var , adjusted by a mean sequencing depth, D, and further adjusted by a sequencing false positive error rate, E, across the selected SNVs.
20 . The method of claim 1 , further comprising generating the personalized disease-associated SNV locus panel.
21 . The method of claim 20 , wherein generating the personalized disease-associated SNV locus panel comprises:
sequencing nucleic acid molecules derived from a sample of the diseased tissue to determine a set of disease-associated SNVs; and
filtering the set of disease-associated SNVs to remove germline variants and non-disease related somatic variants.
22 . The method of claim 21 , wherein the sample of the diseased tissue is a tumor biopsy sample obtained from the individual.
23 . The method of claim 21 , wherein the germline variants or the non-disease related somatic variants, or both, are determined by sequencing nucleic acid molecule derived from a sample of non-diseased tissue obtained from the individual.
24 . The method of claim 23 , wherein the sample of non-diseased tissue comprises white blood cells.
25 . The method of claim 24 , wherein the sample of non-diseased tissue is a buffy coat.
26 . The method of claim 21 , further comprising:
filtering the set of diseased-associated SNVs to remove SNVs supported by only one sequencing read;
filtering the set of diseased-associated SNVs to remove SNVs not supported complementary sequencing reads; or
filtering the set of diseased-associated SNVs to remove SNVs present in a general population of individuals at an allele frequency greater than a predetermined threshold.
27 - 29 . (canceled)
30 . The method of claim 21 , further comprising filtering SNVs within a homopolymer region or filtering SNVs within a short tandem repeat.
31 . The method of claim 21 , wherein the nucleic acid sequencing data is obtained by sequencing nucleic acid molecules from a fluidic sample obtained from the individual using non-terminating nucleotides provided in separate nucleotide flows according to a flow-cycle order comprising a plurality of flow positions, wherein the flow positions correspond to the nucleotide flows; and
generating the personalized disease-associated SNV locus panel further comprises filtering the set of disease-associated SNVs to include only those SNVs that result in nucleic acid sequencing data that differs from reference sequencing data associated with a reference sequence at two or more flow positions when the nucleic acid sequencing data and the reference sequencing data are sequenced using non-terminating nucleotides provided in separate nucleotide flows according to the flow-cycle order.
32 . The method of claim 1 , wherein the nucleic acid sequencing data is obtained by sequencing nucleic acid molecules from a fluidic sample obtained from the individual using non-terminating nucleotides provided in separate nucleotide flows according to a flow-cycle order comprising a plurality of flow positions, wherein the flow positions correspond to the nucleotide flows; and
the method further comprises generating the personalized disease-associated SNV locus panel comprising,
sequencing nucleic acid molecules derived from a sample of the diseased tissue to determine a set of disease-associated SNVs; and
generating the personalized disease-associated SNV locus panel further comprises filtering the set of disease-associated SNVs to include only those SNVs that result in nucleic acid sequencing data that differs from reference sequencing data associated with a reference sequence at two or more flow positions when the nucleic acid sequencing data and the reference sequencing data are sequenced using non-terminating nucleotides provided in separate nucleotide flows according to the flow-cycle order.
33 . The method of claim 31 , wherein generating the personalized disease-associated SNV locus panel comprises filtering the set of disease-associated SNVs to include only those SNVs that result in nucleic acid sequencing data that differs from reference sequencing data associated with a reference sequence across one or more flow cycles when the nucleic acid sequencing data and the reference sequencing data are sequenced using non-terminating nucleotides provided in separate nucleotide flows according to the flow-cycle order.
34 - 38 . (canceled)
39 . The method of claim 1 , wherein the disease is cancer.
40 . The method of claim 39 , wherein the cancer is a metastatic cancer.
41 . The method of claim 1 , wherein the method further comprises sequencing nucleic acid molecules to obtain the sequencing data.
42 . The method of claim 1 , wherein the nucleic acid sequencing data is obtained by sequencing nucleic acid molecules according to a predetermined nucleotide sequencing cycle order.
43 . The method of claim 42 , wherein the nucleic acid sequencing data is further obtained by re-sequencing the nucleic acid molecules according to a different predetermined nucleotide sequencing cycle, wherein the different predetermined nucleotide sequencing cycle results in a different false positive variant rate at a subset of the sequencing loci compared to the first predetermined nucleotide sequencing cycle order.
44 . The method of claim 1 , wherein the sequencing data is untargeted sequencing data.
45 - 49 . (canceled)
50 . The method of claim 1 , wherein the disease-associated SNV locus panel comprises passenger mutations or driver mutations.
51 . The method of claim 1 , wherein the disease-associated SNV locus panel comprises driver mutations.
52 . The method of claim 1 , wherein the disease-associated SNV locus panel comprises single nucleotide polymorphism (SNP) loci, indel loci, or both.
53 . (canceled)
54 . The method of claim 1 , wherein the selected loci from the disease-associated SNV locus panel comprise about 300 or more loci.
55 . The method of claim 1 , wherein the loci selected from the disease-associated SNV panel are selected based on a false positive rate of the individual loci.
56 . The method of claim 1 , wherein the loci selected from the disease-associated SNV panel based on unique SNVs associated with a selected sub-clone of the disease.
57 . The method of claim 1 , wherein the disease-associated SNV panel is determined by comparing sequencing data associated with the diseased tissue to sequencing data associated with a non-diseased tissue.
58 . The method of claim 57 , comprising sequencing nucleic acid molecules derived from the diseased tissue to obtain the sequencing data associated with the diseased tissue.
59 . The method of claim 57 , comprising sequencing nucleic acid molecules derived from the non-diseased tissue to obtain the sequencing data associated with the non-diseased tissue.
60 . The method of claim 1 , wherein the nucleic acid sequencing data is obtained using surface-based sequencing of nucleic acid molecules, and wherein the nucleic acid molecules are not amplified prior to attaching the nucleic acid molecules to a surface.
61 . The method of claim 1 , wherein the nucleic acid sequencing data is obtained without the use of unique molecular identifiers (UMIs).
62 . (canceled)
63 . The method of claim 1 , wherein the sequencing false positive error rate is measured using a panel of control loci.
64 - 67 . (canceled)
68 . The method of claim 1 , comprising generating a report that indicates the presence, absence, or level of disease in the individual.
69 . The method or system of claim 68 , comprising providing the report to a patient or a healthcare representative of the patient.
70 . A system, comprising:
one or more processors; and
a non-transitory computer-readable medium that stores one or more programs comprising instructions for implementing the method of claim 1 .