# US 2021/0172028 A1 · Early lung cancer detection by dna methylation phenotyping of sputum-derived cells > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2021/0172028 A1 - Document ID: US20210172028A1 - Status: Published application - Kind: A1 - Application number: 17/180,433 - Filed: 2021-02-19 - Published: 2021-06-10 - Earliest claimed: 2014-08-28 - Assignees: Cedars-Sinai Medical Center; The Regents of the University of California; The United States Government as represented by the Department of Veterans Affairs - Applicants: Cedars-Sinai Medical Center; The Regents of the University of California; United States Government Represented By the Department of Veterans Affairs - Inventors: Jian Tajbakhsh; Fariborz Mortazavi - Web page: https://patentgrep.com/patent/US20210172028A1 ## CPC - C12Q 1/6886: for cancer (immunoassay for cancer G01N33/575) - C12Q 1/6841: In situ hybridisation - G01N 33/5752: of the lungs - C12Q 2600/154: Methylation markers - G01N 1/30: Staining; Impregnating {; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis} - G01N 2440/12: alkylation, e.g. methylation, (iso-)prenylation, farnesylation - G01N 2800/50: Determining the risk of developing a disease - 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 - 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 In certain embodiments, this application discloses methods for detecting lung cancer. The method includes characterization of cells extracted from human sputum, which is a valuable tissue surrogate and source of upper respiratory cells that become cancerous early in the process of lung cancer development. The method includes the staining of extracted cells with fluorescent reporters that produce a specific pattern in the nuclei of labeled cells, which can be made visible by light microscopy. The pattern is relevant to a type of epigenetic coding of DNA known as DNA methylation, which changes in specific cells of the lung during cancer development, in comparison to normal respiratory cells. ## Claims ### Claim at position 1 1-16. (canceled) ### Claim 17 A method, comprising: obtaining a biological sample from a subject, the biological sample including a plurality of cells; determining (i) a global 5-methylcytosine (5mC) content and (ii) a spatial nuclear co-distribution of 5mC and global DNA (gDNA) in a nucleus of each of the plurality of cells; determining that a cell of the plurality of cells is hypomethylated if at least the global 5mC content or the spatial nuclear co-distribution of 5mC and gDNA in the nucleus of the cell is significantly different from one or more non-cancerous or non-precancerous reference cells; and based at least in part on a percentage of determined hypomethylated cells in the plurality of cells exceeding a threshold, determining that the subject has a high risk of developing clinically verifiable cancer. ### Claim 18 (depends on 17) The method of claim 17, further comprising treating the subject for cancer, based at least in part on the determining that the subject has a high risk for developing clinically verifiable cancer. ### Claim 19 (depends on 18) The method of claim 18, wherein the biological sample comprises sputum. ### Claim 20 (depends on 19) The method of claim 19, wherein the sputum comprises the plurality of cells. ### Claim 21 (depends on 20) The method of claim 20, wherein the plurality of cells is of lung origin. ### Claim 22 (depends on 19) The method of claim 19, wherein the subject has a history of smoking cigarettes. ### Claim 23 (depends on 19) The method of claim 19, wherein the subject does not have a history of smoking cigarettes. ### Claim 24 (depends on 17) The method of claim 17, wherein the global 5mC content is determined with a microscope after the plurality of cells has been subjected to (a) immunofluorescence staining with an antibody specific for 5mC and a dye to delineate the cytoplasm, and (b) counterstaining with 4′,6-diamidino-2-phenylindole (DAPI). ### Claim 25 (depends on 17) The method of claim 17, wherein the spatial nuclear co-distribution of 5mC and gDNA is determined with a microscope after the plurality of cells has been subjected to (a) immunofluorescence staining with an antibody specific for 5mC and a dye to delineate the cytoplasm, and (b) counterstaining with 4′,6-diamidino-2-phenylindole (DAPI). ### Claim 26 (depends on 19) The method of claim 19, wherein the biological sample was obtained from a subject by a method comprising: administering hypertonic saline into the subject's respiratory tract; and collecting a quantity of sputum that is expelled from the subject as the result of inhaling said hypertonic saline. ### Claim 27 (depends on 26) The method of claim 26, wherein the hypertonic saline is administered via a nebulizer. ### Claim 28 (depends on 27) The method of claim 27, wherein the hypertonic saline is 3-5% NaCl. ### Claim 29 (depends on 24) The method of claim 24, wherein the microscope is a confocal scanning microscope with a resolution equal to or less than 500 nanometers. ### Claim 30 (depends on 17) The method of claim 17, wherein the plurality of cells for determining the hypomethylated cells is a plurality of human respiratory cells.