# US 12,760,374 B2 · Cell-free DNA hydroxymethylation profiles in the evaluation of pancreatic lesions

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- Number: US 12,760,374 B2
- Document ID: US12760374B2
- Status: Granted
- Kind: B2
- Application number: 18/105,167
- Filed: 2023-02-02
- Published: 2024-01-25
- Granted: 2026-10-13
- Earliest claimed: 2018-09-19
- Assignees: ClearNote Health, Inc.
- Applicants: ClearNote Health, Inc.
- Inventors: Samuel Levy; Patrick A. Arensdorf; Chin-Jen Ku; Francois Collin
- Web page: https://patentgrep.com/patent/US12760374B2

## CPC

- C12Q 1/6886: for cancer (immunoassay for cancer G01N33/575)
- G06F 17/18: for evaluating statistical data {, e.g. average values, frequency distributions, probability functions, regression analysis (forecasting specially adapted for a specific administrative, business or logistic context G06Q10/04)}
- G16H 50/30: for calculating health indices; for individual health risk assessment
- C12Q 2537/164: Methylation detection other then bisulfite or methylation sensitive restriction endonucleases
- C12Q 2600/154: Methylation markers
- C12Q 2600/158: Expression markers
- G01N 2800/60: Complex ways of combining multiple protein biomarkers for diagnosis
- G01N 2800/7028: Cancer
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- G06F: ELECTRIC DIGITAL DATA PROCESSING (computer systems based on specific computational models G06N)
- G16H: HEALTHCARE INFORMATICS, i.e. INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR THE HANDLING OR PROCESSING OF MEDICAL OR HEALTHCARE DATA
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

Disclosed herein are methods for identifying patients with pancreatic cancer and subjects at risk for developing pancreatic cancer, methods for monitoring a patient with an identified pancreatic lesion, methods for evaluating the effectiveness of a treatment used for a patient with pancreatic cancer, and methods for selecting a therapy for treating pancreatic cancer in a particular patient. The invention makes use of hydroxymethylation biomarkers, which in combination with one or more clinical parameters and optionally one or more additional types of biomarkers and/or patient-specific risk factors, exhibit a hydroxymethylation level that correlates with pancreatic cancer. Kits and other methods of use are also provided.

## Claims

### Claim 1

A method for monitoring the effectiveness of treatment in a patient with a pancreatic lesion identified on an imaging scan, the method comprising:
(a) obtaining an initial cell-free DNA sample that comprises genes relevant to PDAC and its variants from a patient who is being treated for a pancreatic lesion;
(b) enriching for hydroxymethylated DNA in the sample;
(c) quantifying the hydroxymethylated DNA in the enriched initial sample that map to each of a plurality of selected loci in a reference hydroxymethylation profile, wherein the selected loci comprise hydroxymethylation biomarkers related to the presence, absence, or risk of pancreatic PDAC and its variants comprising ADARB2-AS1, ANKRD36B, ASAH2B, ATG4B, ATP8B1, BOLA1, C11orf88, C17orf97, C1orf170, C3orf36, C8orf74, CAMSAP2, CCDC54, CCDC59, CKAP2, CLK2P, CRTC1, CSRP2, CYB5D1, DNAJC27, DYNAP, FAM166A, FAM188B, FAM196A, FAM86JP, FAT4, FBXO5, FGF2, FUT2, GAS2L2, GAS6, GGACT, GLRX5, GPX1, GPX5, HBD, HLA-A, HTR1F, IL36G, KANSL1, KCNH6, KCTD15, KLHL38, KLK2, KRT6B, LAMC1, LGALS14, LGALS8-AS1, LIFR, LINC00266-1, LINC00310, LOC100130452, LOC100130557, LOC100130894, LOC100288778, LOC100505633, LOC100505648, LOC100505738, LOC100652909, LOC389033, LOC90784, LRRC37A2, MED11, MRPL23-AS1, NAT8L, NEUROD1, NEUROG2, NME5, NOMO3, NPRL2, NXN, ODF3L1, ODF3L2, OSCP1, PARD6G, PGAM1, PLA2G2E, PLSCR4, PPAP2A, PPP1R15A, PPP1R3E, RASL10B, REXO1L1, RIMBP3, RNF126P1, RNU6-76, RPP25, RPS27, SH3PXD2B, SHISA4, SLC25A38, SLC4A1, SLCO5A1, SPDEF, SRSF6, STRA6, SYNM, TBCB, TDRD6, TEX26, TMEM253, TNFSF13B, TTC14, TUBA4A, UBB, VAMP8, VGLL2, WASH2P, WNT9B, XBP1, and ZNF789;
(d) comparing, at each selected locus, the hydroxymethylation level of the enriched cell-free DNA in the initial sample with the hydroxymethylation level in the reference profile, to ascertain differences in hydroxymethylation levels between the sample and the reference profile for each biomarker;
(e) generating an initial hydroxymethylation profile for the patient comprising the hydroxymethylation level of the enriched cell-free DNA in the initial sample, at each selected locus;
(f) repeating steps (a) through (c) at a later time with a subsequent cell-free DNA sample obtained from the patient;
(g) generating a subsequent hydroxymethylation profile for the patient comprising the hydroxymethylation level of the enriched cell-free DNA in the subsequent sample, at each selected locus;
(h) comparing, at each selected locus, the hydroxymethylation level of the enriched cell-free DNA in the subsequent hydroxymethylation profile to the hydroxymethylation level of the enriched cell-free DNA in the initial hydroxymethylation profile, to ascertain a change in the pancreatic lesion; and
(i) identifying a change in the patient's hydroxymethylation profile that correlate with a progression toward cancer; and
(j) commencing the adminstration of treatment for PDAC and its variants after a change has been identified in step (i).

### Claim 2 (depends on 1)

The method of claim 1, wherein the cell-free DNA sample is extracted from a blood sample.

### Claim 3 (depends on 1)

The method of claim 1, wherein the cell-free DNA sample is extracted from pancreatic cyst fluid.

### Claim 4 (depends on 1)

The method of claim 1, wherein the hydroxymethylation biomarkers additionally comprise loci that are associated with one or more of the following genes: GATA4, GATA6, PROX1, ONECUT2, YAP1, TEAD1, ONECUT2/ONECUT1-TCGA, IGF1, and IGF2.

### Claim 5 (depends on 1)

The method of claim 1 wherein the treatment of step (j) comprises radiation therapy, chemotherapy, pharmacotherapy, surgical resection of the lesion, or a combination thereof.

### Claim 6 (depends on 1)

The method of claim 1 wherein the sample comprises whole blood, serum, plasma, urine, cyst fluid, or another body fluid comprising hydroxymethylation biomarkers related to the presence, absence, or risk of PDAC and its variants.