	US 11,939,636 B2
	Methods and systems for improving patient monitoring after surgery
Ariel Jaimovich, Redwood City, CA (US); Yupeng He, Redwood City, CA (US); Oscar Westesson, Berkeley, CA (US); and William J. Greenleaf, Menlo Park, CA (US)
Assigned to Guardant Health, Inc., Palo Alto, CA (US)
Filed by GUARDANT HEALTH, INC., Redwood City, CA (US)
Filed on Jun. 1, 2020, as Appl. No. 16/889,659.
Claims priority of provisional application 62/855,736, filed on May 31, 2019.
Prior Publication US 2021/0017605 A1, Jan. 21, 2021
Int. Cl. C12Q 1/6886 (2018.01)

CPC C12Q 1/6886 (2013.01) [C12Q 2600/112 (2013.01); C12Q 2600/156 (2013.01); C12Q 2600/158 (2013.01)]

15 Claims

1. A method for detecting a presence or absence of cancer in a subject, comprising:

a) obtaining at least one sample from the subject from at least two different time points to provide at least two samples, wherein a first sample of the at least two samples is obtained at a first time point of the at least two different time points and a second sample of the at least two samples is obtained at a second time point of the at least two different time points, wherein the first time point is prior to or during a cancer treatment and the second time point follows the cancer treatment, and wherein the subject is a human;

b) processing polynucleotides from each of the at least two samples, wherein the processing comprises:

i. partitioning at least a portion of the polynucleotides from the first sample into at least two partitions based on DNA methylation and optionally one or more additional epigenetic features, and partitioning at least a portion of the polynucleotides from the second sample into at least two partitions based on DNA methylation and optionally one or more additional epigenetic features, wherein a first partition from the first sample comprises hypomethylated DNA of the first sample, a first partition from the second sample comprises hypomethylated DNA of the second sample, a second partition from the first sample comprises hypermethylated DNA of the first sample, and a second partition from the second sample comprises hypermethylated DNA of the second sample, thereby generating partitioned polynucleotides;

ii. tagging at least a portion of the partitioned polynucleotides from each of the first and second partitions from the first sample and each of the first and second partitions from the second sample, thereby generating tagged parent polynucleotides;

iii. amplifying at least a portion of the tagged parent polynucleotides from each of the first and second partitions from the first sample and each of the first and second partitions from the second sample to generate progeny polynucleotides; and

iv. sequencing at least a portion of the progeny polynucleotides generated from each of the first and second partitions from the first sample and each of the first and second partitions from the second sample to generate a set of sequencing reads; and

c) analyzing a plurality of genomic regions for DNA methylation and optionally one or more additional epigenetic features from the set of sequencing reads of the at least two samples, to detect the presence or absence of the cancer in the subject at the second time point, wherein the plurality of genomic regions comprise differentially methylated regions that are perturbed by the cancer, the differentially methylated regions comprising hypomethylation variable target regions and/or hypermethylation variable target regions, and wherein the analyzing comprises

i) mapping the set of sequencing reads to a reference sequence;

ii) determining a plurality of epigenetic rates for the plurality of genomic regions of the at least two samples;

iii) adjusting an epigenetic rate threshold based on the epigenetic rate of at least one of the plurality of genomic regions of the first sample; and

iv) comparing the epigenetic rate for the plurality of genomic regions in the second sample with the adjusted epigenetic rate threshold to detect the presence or absence of the cancer; or

i) mapping the set of sequencing reads to a reference sequence;

ii) determining a plurality of epigenetic rates for the plurality of genomic regions of the first sample;

iii) determining a likelihood of a tumor fraction for one or more of the plurality of genomic regions in the second sample based on a predetermined set of epigenetic rates of the plurality of genomic regions of the second sample, a set of epigenetic characteristics for a set of cell-free polynucleotides in the second sample mapped to the plurality of genomic regions, and the epigenetic rates of the plurality of genomic regions of the first sample;

iv) combining the plurality of likelihoods for the one or more of the plurality of genomic regions to determine an overall posterior probability for the presence of the cancer in the subject; and

v) comparing the overall posterior probability for the presence of the cancer in the subject with a predetermined threshold to detect the presence or absence of the cancer.