DETECTION OF LUNG NEOPLASIA BY ANALYSIS OF METHYLATED DNA
First Claim
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1. A method of processing a sample, the method comprising:
- a) assaying a sample from a subject for an amount of at least one methylation marker selected from the group consisting of BARX1, LOC100129726, SPOCK2, TSC22D4, MAX.chr8.124, RASSF1, ZNF671, ST8SIA1, NKX6_2, FAM59B, DIDO1, MAX_Chr1.110, AGRN, SOBP, MAX_chr10.226, ZMIZ1, MAX_chr8.145, MAX_chr10.225, PRDM14, ANGPT1, MAX.chr16.50, PTGDR_9, ANKRD13B, DOCK2, MAX_chr19.163, ZNF132, MAX chr19.372, HOXA9, TRH, SP9, DMRTA2, ARHGEF4, CYP26C1, ZNF781, PTGDR, GRIN2D, MATK, BCAT1, PRKCB_28, ST8SIA_22, FLJ45983, DLX4, SHOX2, EMX1, HOXB2, MAX.chr12.526, BCL2L11, OPLAH, PARP15, KLHDC7B, SLC12A8, BHLHE23, CAPN2, FGF14, FLJ34208, B3GALT6, BIN2_Z, DNMT3A, FERMT3, NFIX, S1PR4, SKI, SUCLG2, TBX15, ZDHHC1 and ZNF329;
b) assaying said sample for an amount of a reference marker;
c) comparing the amount of said at least one methylation marker to the amount of reference marker in said sample to determine a methylation state for said at least one methylation marker in said sample; and
optionallyd) generating a record reporting the methylation state for said at least one methylation marker in said sample.
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Abstract
Provided herein is technology for lung neoplasia screening and particularly, but not exclusively, to methods, compositions, and related uses for detecting the presence of lung cancer.
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Citations
47 Claims
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1. A method of processing a sample, the method comprising:
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a) assaying a sample from a subject for an amount of at least one methylation marker selected from the group consisting of BARX1, LOC100129726, SPOCK2, TSC22D4, MAX.chr8.124, RASSF1, ZNF671, ST8SIA1, NKX6_2, FAM59B, DIDO1, MAX_Chr1.110, AGRN, SOBP, MAX_chr10.226, ZMIZ1, MAX_chr8.145, MAX_chr10.225, PRDM14, ANGPT1, MAX.chr16.50, PTGDR_9, ANKRD13B, DOCK2, MAX_chr19.163, ZNF132, MAX chr19.372, HOXA9, TRH, SP9, DMRTA2, ARHGEF4, CYP26C1, ZNF781, PTGDR, GRIN2D, MATK, BCAT1, PRKCB_28, ST8SIA_22, FLJ45983, DLX4, SHOX2, EMX1, HOXB2, MAX.chr12.526, BCL2L11, OPLAH, PARP15, KLHDC7B, SLC12A8, BHLHE23, CAPN2, FGF14, FLJ34208, B3GALT6, BIN2_Z, DNMT3A, FERMT3, NFIX, S1PR4, SKI, SUCLG2, TBX15, ZDHHC1 and ZNF329; b) assaying said sample for an amount of a reference marker; c) comparing the amount of said at least one methylation marker to the amount of reference marker in said sample to determine a methylation state for said at least one methylation marker in said sample; and
optionallyd) generating a record reporting the methylation state for said at least one methylation marker in said sample. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 19, 20, 21, 22)
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17. A method of analyzing a plasma sample obtained from a subject suspected of having a lung neoplasm, the method comprising:
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a) combining the plasma sample with; i) protease; and ii) a first lysis reagent, said first lysis reagent comprising guanidine thiocyanate; and non-ionic detergent; to form a mixture wherein proteins are digested by said protease; b) to the mixture of step a) adding iii) silica particles, and iv) a second lysis reagent, said second lysis reagent comprising; guanidine thiocyanate; non-ionic detergent; and isopropyl alcohol; under conditions wherein DNA is bound to said silica particles; c) separating silica particles with bound DNA from the mixture of b); d) to the separated silica particles with bound DNA adding a first wash solution, said first wash solution comprising A) guanidine hydrochloride or guanidine thiocyanate, and B) ethyl alcohol; e) separating the silica particles with bound DNA from said first wash solution; f) to the separated silica particles with bound DNA adding a second wash solution, said second wash solution comprising a buffer and ethyl alcohol; g) separating washed silica particles with bound DNA from said second wash solution; and h) eluting DNA from the washed silica particles with bound DNA separated in step g); i) assaying eluted DNA for an amount of at least one methylated methylation marker and for an amount of reference marker in said eluted DNA; j) comparing the amount of said at least one methylated methylation marker to the amount of reference marker in said DNA to determine a methylation state for said at least one methylation marker in said plasma sample; and
optionallyd) generating a record reporting the methylation state for said at least one methylation marker in said plasma sample. - View Dependent Claims (18, 23, 24)
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25. A kit, comprising:
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a) at least one oligonucleotide, wherein at least a portion of said oligonucleotide specifically hybridizes to a methylation marker selected from the group consisting of BARX1, LOC100129726, SPOCK2, TSC22D4, MAX.chr8.124, RASSF1, ZNF671, ST8SIA1, NKX6_2, FAM59B, DIDO1, MAX_Chr1.110, AGRN, SOBP, MAX_chr10.226, ZMIZ1, MAX_chr8.145, MAX_chr10.225, PRDM14, ANGPT1, MAX.chr16.50, PTGDR_9, ANKRD13B, DOCK2, MAX_chr19.163, ZNF132, MAX_chr19.372, HOXA9, TRH, SP9, DMRTA2, ARHGEF4, CYP26C1, ZNF781, PTGDR, GRIN2D, MATK, BCAT1, PRKCB_28, ST8SIA_22, FLJ45983, DLX4, SHOX2, EMX1, HOXB2, MAX.chr12.526, BCL2L11, OPLAH, PARP15, KLHDC7B, SLC12A8, BHLHE23, CAPN2, FGF14, FLJ34208, B3GALT6, BIN2_Z, DNMT3A, FERMT3, NFIX, S1PR4, SKI, SUCLG2, TBX15, ZDHHC1, and ZNF329, and b) at least one additional oligonucleotide, wherein at least a portion of said additional oligonucleotide specifically hybridizes to a reference nucleic acid. - View Dependent Claims (26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37)
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- 38. A composition comprising a reaction mixture comprising a complex of a target nucleic acid selected from the group consisting of BARX1, LOC100129726, SPOCK2, TSC22D4, MAX.chr8.124, RASSF1, ZNF671, ST8SIA1, NKX6_2, FAM59B, DIDO1, MAX_Chr1.110, AGRN, SOBP, MAX_chr10.226, ZMIZ1, MAX_chr8.145, MAX_chr10.225, PRDM14, ANGPT1, MAX.chr16.50, PTGDR_9, ANKRD13B, DOCK2, MAX_chr19.163, ZNF132, MAX_chr19.372, HOXA9, TRH, SP9, DMRTA2, ARHGEF4, CYP26C1, ZNF781, PTGDR, GRIN2D, MATK, BCAT1, PRKCB_28, ST8SIA_22, FLJ45983, DLX4, SHOX2, MIXT HOXB2, MAX.chr12.526, BCL2L11, OPLAH, PARP15, KLHDC7B, SLC12a, BHLHE23, CAPN2, FGF14, FLJ34208, B3GALT6, BIN2_Z, DNMT3A, FERMT3, NFIX, S1PR4, SKI, SUCLG2, TBX15, ZDHHC1, and ZNF329, and an oligonucleotide that specifically hybridizes to said target nucleic acid.
Specification