THE MAIN TEA ETA USANTON 20170335401A1 MARTUA AMAT MALAMATAN ( 19) United States ( 12) Patent Application Publication ( 10 ) Pub. No. : US 2017 /0335401 A1 Allawi et al. (43 ) Pub . Date : Nov . 23 , 2017 (54 ) DETECTION OF LUNG NEOPLASIA BY ( 22 ) Filed : Mar. 28, 2017 ANALYSIS OF METHYLATED DNA Related U . S . Application Data (71 ) Applicants: Exact Sciences Corporation , Madision , (60 ) Provisional application No . 62/ 462 ,677 , filed on Feb . WI (US ) ; Mayo Foundation for 23 , 2017 , provisional application No. 62 /332 , 295 , Medical Education and Research , filed on May 5, 2016 . Rochester, MN (US ) Publication Classification ( 72 ) Inventors : Hatim Allawi, Middleton , WI (US ) ; (51 ) Int . Cl. Graham P . Lidgard , Middleton , WI C12Q 1/ 68 ( 2006 .01 ) (US ) ; Maria Giakoumopoulos, (52 ) U . S . CI. Middleton , WI (US ) ; David A . CPC ...... C12Q 1 /6886 (2013 .01 ); C12Q 1 /6806 Ahlquist, Rochester , MN (US ) ; ( 2013 .01 ) ; C12Q 2600 / 16 ( 2013 .01 ) ; C12Q William R . Taylor , Lake City , MN 2600 / 154 ( 2013 .01 ) ( US) ; Douglas Mahoney , Rochester, (57 ) ABSTRACT MN (US ) Provided herein is technology for lung neoplasia screening and particularly , but not exclusively , to methods, composi tions, and related uses for detecting the presence of lung (21 ) Appl. No. : 15 /471 , 337 cancer. Patent Application Publication Nov . 23, 2017 Sheet 1 of 155 US 2017 /0335401 A1
TIT 5'ACAAATAAACCCCATACAAATTACGAC3(SEQIDNO:4) 5'GGCGTTTTAGTTTTACGGATTCG3(SEQIDNO:3) 5'CGCCGAGGCGAAAACTCCCT/3C6(SEQIDNO:)
FIG.1
Bisulfite-convertedTargetDNA:(SEQIDNO2)
AGRNTargetDNA(SEQIDNO:1) TTTATTTGTATGATTTCGTTTCGCGCGGGAGTCGGGGGCGT3" ????T?TGCAT'GACCCCGCCCCGCGCGGGAGTCGGGGGCGC3? PCRandFlapAssayoligonucleotides: AGRNForwardPrimer: AGRNReversePrimer: AGRNFlapoligo.: 5'GTTCCCGGAACGGCCTCTTGGGGGCGTTCCAGCCCCACGGACCCGCAGGGAGTCCCCGCCGCAATTTGCATGGGG1 5'GTTTTCGGAACGGTTTTTTGGGGGCGTTTTAGTTTTACGGATTCGTAGGGAGTTTTCGTCGTAATTTGTATGGGG Patent Application Publication Nov . 23 , 2017 Sheet 2 of 155 US 2017 /0335401 A1
ANGPT1ReversePrimer:5'ACATCTCTCTACAAAACGTTTATATTATACTAATC3(SEQIDNO9) ANGPT1ForwardPrimer:5'TTTTAGAATACGATGGTAATTGTCGT3(SEQIDNO:8)11 FIG.1(cont'd) I ANGPT1Flapoligo.:5'CGCCGAGGCTATCGTACTCT/3C67(SEQIDNO10)
I
G
Bisulfite-convertedTargetDNA:(SEQIDNO7) ?|Il11 I GACCAGTACAACACAAACGCTCTGCAGAGAGATGCTCCACACGTGGAACCG3" NO6)SEQ:IDDNA(TargetANGPT1 PCRandFlapAssayOligonucleotides: 5'CGGATTCAACATGGGCAATGTGCCTACACTTTCATTCTTCCAGAACACGATGGCAACTGTCGTGAGAGTACGACA 5'CGGATTTAATATGGGTAATGTGTTTATATTTTTATTTTTTTAGAATACGATGGTAATTGTCGTGAGAGTACGATA GATTAGTATAATATAAACGTTTTGTAGAGAGATGTTTTATACGTGGAATCG3T Patent Application Publication Nov . 23 , 2017 Sheet 3 of 155 US 2017 /0335401 A1
JAG 5'CCACGGACGCGACAATTCCAT/3C6(SEQIDNO:15) 5'AGTTACGACGAGTAGTTGCG3(SEQIDNO:13) 5'TCCTCCTACTCCTACGCC3(SEQIDNO:14)
FIG.1(cont'd) ITI TLI 7
17 Bisulfite-convertedTargetDNA:(SEQIDNO12) LO
17
ANKRD13BTargetDNA:(SEOIDNO11) U N GTTAGGAGGAGGAGGAGTTGGAGCGTATTTTGAG3'. nyt GCCAGGAGGAGGAGGAGCTGGAGCGCATCCTGAG3' ANKRD13BFlapoligo. PCRandFlapAssayOligonucleotides: ANKRD13BForwardPrimer: ANKRD13BReversePrimer: 5'GGAGCTACGACGAGCAGCTGCGGCTGGCGATGGAACTGTCGGCGCAGGAGCAGGAGGAGAGGCGGCGGCGCGCGC 5'GGAGTTACGACGAGTAGTTGCGGTTGGCGATGGAATTGTCGGCGTAGGAGTAGGAGGAGAGGCGGCGGCGCGCGCArr: Patent Application Publication Nov . 23, 2017 Sheet 4 of 155 US 2017 /0335401 A1
GTTAGTAGGAAGATTAGTACGACGTTGATGAGAATTAGGAGCGTTAGTACGGCGGAGATTATTACGCG3'. 5'CGTTCGCGTTATTTATTTCGGCG3(SEQIDNO:18) 5'GCTCCTAATTCTCATCAACGTCGT3(SEQIDNO:19) 5'CGCCGAGGGCGGCGTTTTGC/306(SEQIDNO:20) FIG.1(cont'd)
GCCAGCAGGAAGATCAGTACGACGTTGATGAGAACCAGGAGCGCCAGCACGGCGGAGACCACCACGCG3' Bisulfite-convertedTargetDNA:(SEQIDNO17) ARHGEF4TargetDNA:(SEQIDNO16) PCRandFlapAssayoligonucleotides: 5'GGTGGCAACGGCTGGAGTcccGTccccccccccACTCAccccccccccGcccccccccGGccGcTCAccGGAAG 5'GGTGGTAACGGTTGGAGTGTCGTCGTTCGCGTTATTTATTTCGGCGCGGCGTTTTGCGCGGTCGTTTAGCGGAAG ARHGEF4ForwardPrimer: ARHGEF4ReversePrimer: ARHGEF4Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 5 of 155 US 2017 /0335401 A1 mmGIGI mmIII 777177mir 5'GGTTTATTTTGGTTTTTTGAGTTTTCGG3(SEQIDNO:386) TamanILIA 5'TCCAACCTACTATATTTACGCGAA3(SEQIDNO:387) 5'CCACGGACGGCGGATTTAGGG/306(SEQIDNO:388)
d. Bisulfite-convertedtargetDNAwithprimerandFlapoligo.sites:(SEQID ammm 1 FIG.1(cont'd)
mo
GCGCAGGCCTCCGCGCAAACACAGCAGGCTGGAAGTGGCGCTCATCACCGGCACGTCTTCCCAG3' 1 GCGTAGGTTTTCGCGTAAATATAGTAGGTTGGAAGTGGCGTTTATTATCGGTACGTTTTTTTAG3" 384)NO:SEQIDDNA(TargetB3GALT6 AALL PCRandFlapAssayoligonucleotides: B3GALT6ForwardPrimer: B3GALT6ReversePrimer: B3GALT6Flapoligo.i 101 5'GGCCACACAGGCCCACTcTGGCCCTCTGAGccccccccGGAC?CAGGGCATICAAGGAdcGGC???GGGCTGCCA NO:385) 5'GGTTATATAGGTTTATTTTGGTTTTTTGAGTTTTCGGCGGATTTAGGGTATTTAAGGAGCGGTTTTGGGTTGTTA Patent Application Publication Nov . 23 , 2017 Sheet 6 of 155 US 2017 /0335401 A1
I
TI 1411 GIU 5'CGTTAATTTGTTAGATAGAGGGCG3SEQIDNO:23) 5'CGTTAATTTGTTAGATAGAGGGCG3(SEQIDNO:23) 5'CCACGGACGCGCCTACGAAAA/3C6(SEQIDNO:25) AnLU2. 5'ACGATCGTCCGAACAACC3(SEQIDNO:24) manminiIT BARX1ReversePrimerUniversal:57TCCGAACAACCGCCTAC3'(SEQIDNO26) BARX1Flapoligo.Universal:5'CCACGGACGCGAAAAATCCCA/3061(SEQIDNO:27) FIG.1(cont'd) IIAATTIGI 22)SEQIDNO:DNA(TargetconvertedBisulfite-
T veS BARX1TargetDNA:(SEQIDNO21) UIJ PCRandFlapAssayOligonucleotides: GC????cGGACGACCGCATCCGGAG3? GTTGTTCGGACGATCGTATTCGGAG3' BARX1ForwardPrimer: BARX1ReversePrimer: BARX1Flapoligo.i BARX1ForwardPrimer: 5'GGCCCGGGGCCGCCTGGGCCCCTAGGGGCTGGACGTCAACCTGTTAGATAGAGGGCGTGGGACCCCCCGCAGGCG 5'GGTTCGGGGTCGTTTGGGTTTTTAGGGGTTGGACGTTAATTTGTTAGATAGAGGGCGTGGGATTTTTCGTAGGCGMYT WO Patent Application Publication Nov . 23, 2017 Sheet 7 of 155 US 2017 /0335401 A1 MAT
7AMMATTITudIL 5'GTGTTATTGTCGTTTTTTGTAGTTTCG3(SEQIDNO:30) 5'CGCCGAGGGCGTTTTCGTAG/3C6(SEQIDNO:32) mm 5'CGCAACGAAACGAAACGA3(SEQIDNO:31)
T mmIII FIG.1(cont'd)
1 Bisulfite-convertedTargetDNA:(SEQIDNO29)
L63 BCAT1TargetDNA:(SEQIDNO28) G???cGc?ccGCTGcGGC?CTIGcccGCCAGGTACCTCGAACcc3? GTTTCGTTTCGTTGCGGTTTTTGTTCGTTAGGTATTTCGAATTT3' Oligonucleotides:AssayandFlapPCR
I I BCAT1ForwardPrimer: BCAT1ReversePrimer: BCAT1Flapoligo.i 5'GCTTCCAGCCGCGCGCTCCGTGCCACTGCCGCTCTCTGCAGCCCCGCGTccccGCAGCCTCC?CATGGCCAGCCC 5'GTTTTTAGTCGCGCGTTTCGTGTTATTGTCGTTTTTTGTAGTTTCGCGTTTTCGTAGTTTTTTTATGGTTAGTTCB01 free Patent Application Publication Nov . 23, 2017 Sheet 8 of 155 US 2017 /0335401 A1
2 na 5'ACTTTCTTCTACGTAATTCTTTTCCGA3(SEQIDNO:36) Coty 5'CGTAATGTTTCGCGTTTTTCG3(SEQIDNO:35) 5'CGCCGAGGGCGGGGTCGGGC/306!(SEQIDNO:37)
4 IT113 FIG.1(cont'd)
1
YA I ET? 34)SEQIDNO:DNA(TargetBisulfiteconverted- IT TAGAAGAAAGTTTTATTTTTTGTCGTTTGTTTTTGTGTAGTTTTGTAGTTTCGTCGTTTTCGCGT3' CAGAAGAAAGCCCTATTTCTTGTCGTCTGTTCCTGTGCAGCCTTGCAGCCTCGCCGCCCCCGCGT3' 1
o BCL2L11TargetDNA:(SEQIDNO33) W WO
TUIT to
A PCRandFlapAssayoligonucleotides: YAO? BCL2L11Flapoligo.: 5'GCCCGCCGCACGCCGCAATGCTCCGCGCTCCCCGCGGGGTCGGGCGACTCAGACAGGGACCGGAAAAGAACCACG 5'GTTCGTCGTACGTCGTAATGTTTCGCGTTTTTCGCGGGGTCGGGCGATTTAGATAGGGATCGGAAAAGAATTACG N BCL2L11ForwardPrimer: BCL2L11ReversePrimer: Patent Application Publication Nov . 23, 2017 Sheet 9 of 155 US 2017 /0335401 A1
5'AGTATTAGCGTTTTAGGATCGCG3(SEQIDNO:40) 5'ACTCTAAACCGAAAAACGACG3(SEQIDNO:41) 5'CCACGGACGGCGAAACGACGC/3061(SEQIDNO:42)
FIG.1(contd)
Bisulfite-convertedTargetDNA:(SEQIDNO39)
BHLHE23TargetDNA(SEQIDNO:38) PCRandFlapAssayoligonucleotides: BHLHE23ForwardPrimer: BHLHE23ReversePrimer: BHLHE23_Flapoligo.i 5'GCCGGGGAGTCGAGAAGCAAGTACTAGCGCTCCAGGACCGCGCGCGCCGCCCCGCGCCGCCCCGCGCCGCCCCTC GGTCCAGAGC3' 5'GTCGGGGAGTCGAGAAGTAAGTATTAGCGTTTTAGGATCGCGCGCGTCGTTTCGCGTCGTTTCGCGTCGTTTTTC GGTTTAGAGT3' Patent Application Publication Nov . 23 , 2017 Sheet 10 of 155 US 2017 /0335401 A1
5'TCGGGAGTTCGTATTTTTTTTTCGG3(SEQIDNO:45) 5'CGCCGAGGCCCCGCGCCCTA/3C6(SEQIDNO:47) 5'AAAACCGCCGCCCTAAC3(SEQIDNO:46)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO44)
43)SEQNO:IDDNA(TargetBIN2HM_ PCRandFlapAssayOligonucleotides: BIN2HMForwardPrimer: BIN2HMReversePrimer: BIN2_HMFlapoligo.: 5'GCCGGGAGCCCGCACTTCCTCCTCGGGGGCCTCAGAAAACCACAGGGCGCGGGGCCAGGGCGGCGGCCCC CAGG3' 5'GTCGGGAGTTCGTATTTTTTTTTCGGGGGTTTTAGAAAATTATAGGGCGCGGGGTTAGGGCGGCGGTTTT TAGG37 Patent Application Publication Nov . 23 , 2017 Sheet 11 of 155 US 2017 /0335401 A1
1 I. 5'CGAAATTTCGAACAAAAATTAAAACTCGA3(SEOIDNO:51) 5'GGGTTTATTTTTAGGTAGCGTTCG3(SEQIDNO:50) 5'CCACGGACGGTTCGAGGTTAG/3C6(SEQIDNO:52)
FIG.1(cont'd)
II1 103 Bisulfite-convertedTargetDNA:(SEQIDNO49)
BIN2_ZTargetDNA:(SEQIDNO48) 17 III PCRandFlapAssayOligonucleotides: 7 IL BIN2ZForwardPrimer: BIN2ZReversePrimer: BIN2_ZFlapoligo.:
1. 5'cGGGGCCTACCCTCAGGCAGCGCTcGc?cGAGGCCAGC??ccGAGC??????cccTGcccGAAAC??cGGCc??? CTG3' 5'CGGGGTTTATTTTTAGGTAGCGTTCGTTCGAGGTTAGTTTTCGAGTTTTAATTTTTGTTCGAAATTTCGGTTTTA TTG3' Patent Application Publication Nov . 23 , 2017 Sheet 12 of 155 US 2017 /0335401 A1
5'CGAAACTTCAAAAATACGAAACCCGA3(SEQIDNO:56) 5'CGCCGAGGGCGGTTTTACGG/3C6(SEQIDNO:57) 5'TGATGTTTAGGGGTTCGCG3(SEQIDNO:55)
FIG.1(cont'd)
TACACTCGGGCCCCGCACTCCTGAAGTTCCGCGCGGGAGGAGAAGGGCGTCCCTTTCGCAGCTCGG3' Bisulfite-convertedTargetDNA:(SEQIDNO54) TATATTCGGGTTTCGTATTTTTGAAGTTTCGCGCGGGAGGAGAAGGGCGTTTTTTTCGTAGTTCGG3' 53)NO:SEQIDDNA(TargetCAPN2 PCRandFlapAssayoligonucleotides: 5'TGTCCTGACACGATGGCCACAGGCACAGTTTGTGGTGATGCCCAGGGGCCCGCGCGGCCCCACGGTGGTCCAGTT 5'TGTTTTGATACGATGGTTATAGGTATAGTTTGTGGTGATGTTTAGGGGTTCGCGCGGTTTTACGGTGGTTTAGTT CAPN2ForwardPrimer: CAPN2ReversePrimer: CAPN2Flapoligo.: Patent Application Publication Nov . 23, 2017 Sheet 13 of 155 US 2017 /0335401 A1
5'CCACGGACGCGCCAACGACTT/3C6(SEQIDNO:62) 5'GTATTCGTCGTTGCGCG3(SEQIDNO:60) 5'CCTCGAAAACCGCCTACC3(SEQIDNO:61)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO59)
chr5132TargetDNA:(SEQIDNO58) GGCAGCCTGCGCCGCGGCCTCCGCCTCGGCCGCCAGC3' GGTAGTTTGCGTCGCGGTTTTCGTTTCGGTCGTTAGT3' PCRandFlapAssayoligonucleotides: chr5132ForwardPrimer: chr5132ReversePrimer: chr5_132Flapoligo.: 5'CCGGAGCACTcGccGCTGcccccccTGAAGccccTGGCGGTAGGcGccccTCGAGGccGGCGGGCTGGGCGGCTC 5'TCGGAGTATTCGTCGTTGCGCGTTTTGAAGTCGTTGGCGGTAGGCGGTTTTCGAGGTCGGCGGGTTGGGCGGTTC be Patent Application Publication Nov . 23 , 2017 Sheet 14 of 155 US 2017 /0335401 A1
197TI 5'CCACGGACGCGAAACCACGAA/306)(SEQIDNO:67 5'TAAAGGCGGCGTGTTCG3(SEQIDNO:65). 5'CAACTICACTCCTAAACCGAC3(SEQIDNO:66)
A
I
I
FIG.1(contd) Le
Bisulfite-convertedTargetDNA:(SEQIDNO64)
GCTCAGGAGTGAAGCTGCAGATCTTCGCGGTGAGTGTTACAGCTCCTAAGGCGGCGCAT3! GTTTAGGAGTGAAGTTGTAGATTTTCGCGGTGAGTGTTATAGTTTTTAAGGCGGCGTAT3'I chr7_636TargetDNA:(SEQIDNO63) PCRandFlapAssayOligonucleotides: 5'CGCCGTGAGTGTTATAGTTCTTAAAGGCGGCGTGTCCGGAGTTTCTTCCTTCTGGTGGGGTTCGTGGTCTCGCCG 5'CGTCGTGAGTGTTATAGTTTTTAAAGGCGGCGTGTTCGGAGTTTTTTTTTTTTGGTGGGGTTCGTGGTTTCGTCG chr7636HMForwardPrimer: chr7636HMReversePrimer: chr7_636HMFlapoligo.: Patent Application Publication Nov . 23, 2017 Sheet 15 of 155 US 2017 /0335401 A1
5'TGGTTTTTTGGTTATTTCGGAATCGT3(SEQIDNO:70) 5'CGCCGAGGCGACGATCTAAC/306!(SEQIDNO:72) 5'GCGCGTAATCAACGCTAAC3(SEQIDNO:71)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO69) 17
CYP26C1TargetDNA:(SEQIDNO68) ITI CCACAGGTCCTGGCGCGCGTGTTCAGCCGCGC3' TTATAGGTTTTGGCGCGCGTGTTTAGTCGCGT3' PCRandFlapAssayoligonucleotides: CYP26C1ForwardPrimer: CYP26C1ReversePrimer: CYP26C1Flapoligo.: 5'AACTGGCCTTCTGGCTACTCCGGAATCGCCAAGCAGATGAGGCCAGACCGCCGCCAGCGCTGATCACGCGCGCTC 5'AATTGGTTTTTTGGTTATTTCGGAATCGTTAAGTAGATGAGGTTAGATCGTCGTTAGCGTTGATTACGCGCGTTT 2017 Patent Application Publication Nov . 23, 2017 Sheet 16 of 155 US 2017 /0335401 A1
5'CACGAAAAAAAACGAAAACGAAAC3(SEQIDNO:76) 77)SEQIDNO:3061(CGCCGAGGCGCGCCTAAACC/5' 5'GAGGAGGAGTTTAGCGTCG3(SEQIDNO:75)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO74)
DIDO1TargetDNA:(SEQIDNO73) PCRandFlapAssayOligonucleotides: DIDO1Flapoligo.i 5'GGAGCGGGCAGAGGAGGAGCCCAGCGCCGAGGCCCAGGCGCGCCCCGCCCTCGCCCCTCCCCGTGCCCCTCCCCC GCTGCTCCCC3' 5'GGAGCGGGTAGAGGAGGAGTTTAGCGTCGAGGTTTAGGCGCGTTTCGTTTTCGTTTTTTTTCGTGTTTTTTTTTC GTTGTTTTTT3' DIDO1ForwardPrimer: DIDO1ReversePrimer: Patent Application Publication Nov . 23 , 2017 Sheet 17 of 155 US 2017 /0335401 A1
I
7
TTTTT"TAIA TOITAGITIL 5'CTCCTCTACTAAAACGTACGATAAACA3(SEQIDNO:81) TTTTO 5'TGAGTGCGTAGTGTTTTCGG3(SEQIDNO:80) 5'CGCCGAGGATCGTATAAAAC/3C6(SEQIDNO:82) DLX4ForwardPrimerUniversal:5'ATATTTGGTGAGTGCGTAGTG3(SEQIDNO83) DLX4ReversePrimerUniversal:5'ACGTACGATAAACATCGTATAAAACC3(SEQIDNO84) 5'CGCCGAGGGTTTTCGGTAGT/3C6(SEQIDNO:85)DLX4Flapoligo.Universal FIG.1(cont'd) UUGIAU
3
G Bisulfite-convertedTargetDNA:(SEQIDNO79) G 777 17
T DLX4TargetDNA:(SEQIDNO78)
CTACCGCACGCCCTAGCAGAGGAGGTCTCTGT3' TTATCGTACGTTTTAGTAGAGGAGGTTTTTGT3'A PCRandFlapAssayOligonucleotides: ATTAAV TiriIIII DLX4ForwardPrimer: DLX4ReversePrimer: DLX4Flapoligo.i 5'GCGGTCTATCACGGGCAccccTAACACTIGGTGAGTGCGCAGTGc???cGGCAGTCTICTGGGCTCCATACGATGC pored Patent Application Publication Nov . 23 , 2017 Sheet 18 of 155 US 2017 /0335401 A1
5'TGGTGTTTACGTTCGGTTTTCGT3(SEQIDNO:88) 5'CGCCGAGGCGAACGATCACG/3C6(SEQIDNO:90) 5'CCGCAACAACGACGACC3(SEQIDNO:89)
FIG.1(cont'd)
Bisulfiteconverted-TargetDNA:(SEQIDNO87) )
DMRTA2TargetDNA:(SEQIDNO86) GACCGCTCGGCCGCCGCTGCTGCGGCGGTGCACAAGGAGCCGACCT3' GATCGTTCGGTCGTCGTTGTTGCGGCGGTGTATAAGGAGTCGATTT3' PCRandFlapAssayOligonucleotides: 5'TACTCCACTGCCGGCTTGGTGccCACGCTcGGC??ccGC?CACCCATGG?????cccTTTAGCGATCTCATGCGT 5'TATTTTATTGTCGGTTTGGTGTTTACGTTCGGTTTTCGTTTATTTATGGATTACGTTTTTAGCGATTTTATGCGT DMRTA2ForwardPrimer: DMRTA2ReversePrimer: DMRTA2Flapoligo.: Patent Application Publication Nov . 23, 2017 Sheet 19 of 155 US 2017 /0335401 A1
5'CCACGGACGGAGTGCGCGTTC/3C6(SEQIDNO:95) 5'GTTACGAATAAAGCGTTGGCG3(SEQIDNO:93) 5'AACGAAACGTCTTATCGCGA3(SEQIDNO:94)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO92)
DNMT3ATargetDNA:(SEQIDNO91) ccccccCCACGCGGcccccGcGGGCTGAGcc3" TTTCGTTTACGCGGTTTTCGCGGGTTGAGTT3' PCRandFlapAssayOligonucleotides: 5'AGGCCGGTCACGAACAAAGCGCTGGCGAGTGCGCGC?CGCCCACGCGCACAGGTGC?CGCGACAAGACGCCCCGT 5'AGGTCGGTTACGAATAAAGCGTTGGCGAGTGCGCGTTCGTTTACGCGTATAGGTGTTCGCGATAAGACGTTTCGT DNMT3AForwardPrimer: DNMT3AReversePrimer: DNMT3AFlapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 20 of 155 US 2017 /0335401 A1
IT .
5'CGGTTTCGTAGTATTTTTTTGTTCG3(SEQIDNO:98) 5'CGCCGAGGGCGGTTTTTTCG/3C6(SEQIDNO:100) 5'GAACCCCAAAACGCGAC3(SEQIDNO:99)
.
.
I. 117UTI177579 .
L
J IIIIII
FIG.1(cont'd) VU
J.
.) Bisulfite-convertedTargetDNA:(SEQIDNO97) 3
DOCK2TargetDNA:(SEQIDNO96) nir PCRandFlapAssayoligonucleotides: 5'GCCGGCCCCGCAGCATCCTCCTGCTCGCGGCTCTCCCGCCACCTGTCCCGCTCCCTGCCGCGCCCTGGGGCCCGC ACCTAC?CAC3 5'GTCGGTTTCGTAGTATTTTTTTGTTCGCGGTTTTTTCGTTATTTGTTTCGTTTTTTGTCGCGTTTTGGGGTTCGT ATTTATTTAT3'. DOCK2ForwardPrimer: DOCK2ReversePrimer: DOCK2Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 21 of 155 US 2017 /0335401 A1
I TIT
? mmIII yr 5'CGCCGAGGCGCGTTCGTTTT/3C6(SEQIDNO:105) 5.GAGTCGCGGTTTCGTTTTC3'(SEQIDNO:103) 5'GACGCGACGACCGAAAAAC3(SEQIDNO:104)
FIG.1(cont'd) GCCGCCGCGCCGAGCTTTCCGCGCGTGGACAGACTGCCCGGCCGACGGACGGACGCAGG3' Bisulfite-convertedTargetDNA:(SEQIDNO102) GTCGTCGCGTCGAGTTTTTCGCGCGTGGATAGATTGTTCGGTCGACGGACGGACGTAGG3' DTX1TargetDNA:(SEQIDNO101) PCRandFlapAssayoligonucleotides: DTX1Flapoligo.:_S49 I DTX1ForwardPrimer:49 DTX1ReversePrimer:Ver2 5'cGcc?ccTGGGCTccccccGGAGTGGGAGGGAGccGcGGTccccccccccccccGTTcccccCAGGccccTCG 5'CGTTTTTTGGGTTTTTTTCGGAGTGGGAGGGAGTCGCGGTTTCGTTTTCGCGTTCGTTTTTTTTTAGGTTTTTCGTyrir Patent Application Publication Nov . 23 , 2017 Sheet 22 of 155 US 2017 /0335401 A1
FIG.1(cont'd) EMX1ReversePrimer:5'TTCCTTTTCGTTCGTATAAAATTTCGTT3(SEQIDNO109) EMX1ForwardPrimer:5'GGCGTCGCGTTTTTTAGAGAA3(SEQIDNO108) EMX1Flapoligo,:5'CCACGGACGATCGGGTTTTAG/3C6(SEQIDNO110) Bisulfite-convertedTargetDNA:(SEQIDNO107) EMX1TargetDNA:(SEQIDNO106) CCCCACACGAACGAAAAGGAACATGTCTGCGCT3' 5'TCCGGCGCCGCGTTTTCTAGAGAACCGGGTCTCAGCGATGCTCATTTCAGCCCCGTCTTAATGCAACAAACGAAA 5'TCGGCGTCGCGTTTTTTAGAGAATCGGGTTTTAGCGATGTTTATTTTAGTTTCGTTTTAATGTAATAAACGAAAT TTTATACGAACGAAAAGGAATATGTTTGCGTT3' PCRandFlapAssayOligonucleotides: Patent Application Publication Nov . 23, 2017 Sheet 23 of 155 US 2017 /0335401 A1
intir I
Ti12TITI.ILIGI I 5'CGATAGCGTTTTTTATTGTCGCG3(SEQIDNO:113) 5'GCACGACCGTAAAATACTCGTC3(SEQIDNO:114) 5'CCACGGACGCGAAATCGAAAC/3C61(SEQIDNO:115) Tan r17
fummond
1 FIG.1(contd)
Bisulfite-convertedTargetDNA:(SEQIDNO112) A CCCCGACGAGTACTCCACGGCCGTGCGCGAGGCGCCAGCGGAGCTCGCCGAAG3' TTTCGACGAGTATTTTACGGTCGTGCGCGAGGCGTTAGCGGAGTTCGTCGAAG3' FAM59BTargetDNA:(SEQIDNO111) PCRandFlapAssayOligonucleotides: FAM59BForwardPrimer: FAM59BReversePrimer: FAM59BFlapoligo.i 5'GGGCCTGCTGGCCGGGGACCCGCGCGTCGAGCGCCTGGTGCGCGACAGCGCCTCCTACTGCCGCGAGCGCTTCGA 5'GGGTTTGTTGGTCGGGGATTCGCGCGTCGAGCGTTTGGTGCGCGATAGCGTTTTTTATTGTCGCGAGCGTTTCGATiri Patent Application Publication Nov . 23, 2017 Sheet 24 of 155 US 2017 /0335401 A1
1 177 NITI HUIV 5'GTTTTCGGGGATTATATCGATTCG3(SEQIDNO:118) 5'CCCAATAACCCGCAAAATAACC3(SEQIDNO:119) 5'CGCCGAGGCGACTCGACCTC/3C6(SEQIDNO:120)
ITI7 77JULI m
1
FIG.1(cont'd) Y17 117)SEQIDNO:(TargetDNABisulfiteconverted- TGTTTGTGGGAGAGGAGGACCCAGAGGCCGAGTCGGTCACCCTGCGGGTCACTGGGGAGTCGCAC3' my I FERMT3TargetDNA:(SEQIDNO116) ATT PCRandFlapAssayOligonucleotides: FERMT3ForwardPrimer: FERMT3ReversePrimer: FERMT3Flapoligo.: 5'TAGCAGCAGCCGCAGCCATGGCGGGGATGAAGACAGCCTCCGGGGACTACATCGACTCGTCATGGGAGCTGCGGG TI 5'TAGTAGTAGTCGTAGTTATGGCGGGGATGAAGATAGTTTTCGGGGATTATATCGATTCGTTATGGGAGTTGCGGG yTTTILII Patent Application Publication Nov . 23, 2017 Sheet 25 of 155 US 2017 /0335401 A1
17 WTI IIII
ITIMII 5'TTTCGTGGTAACGGAAATTTTTCG3(SEOIDNO:123) 5'CGCCGAGGGCGTTACGGCGG/3061(SEQIDNO:125) TITI I.1 5'CGACGAAAACGACCCGT3(SEQIDNO:124)
FIG.1(cont'd)
TOTI21 III. Bisulfite-convertedTargetDNA:(SEQIDNO122)
FGF14TargetDNA:(SEQIDNO121) AM 01 GccGC??ccGCCGCATTGCGTAGCGAAGC3? GTCGTTTTCGTCGTATTGCGTAGCGAAGT3' PCRandFlapAssayoligonucleotides: 5'GTCCCAGAGACGCCCTAGGGTCAGAGGTCATCTCCGTGGCAACGGAAACTTCCCGCGCTACGGCGGCTCCAACGG 5?GTTTTAGAGACGTTTTAGGGTTAGAGGTTATTTTCGTGGTAACGGAAATTTTTCGCGTTACGGCGGTTTTAACGG FGF14ForwardPrimer: FGF14ReversePrimer: FGF14Flapoligo.: Patent Application Publication Nov . 23, 2017 Sheet 26 of 155 US 2017 /0335401 A1
5'GAGCGTATACGAGAAAGTTTTTACG(SEQIDNO:128) 5'CCACGGACGGCGTTCGCGTTT/3C6(SEQIDNO:130) 5'AACGCCTTCCCGTCGAA(SEQIDNO:129)
FIG.1(contd)
Bisulfite-convertedTargetDNA:(SEQIDNO127) FLJ34208TargetDNA:(SEQIDNO126) CGGGAAGGCGCCTCTTCCGACCGTCCTGGATG3' CGGGAAGGCGTTTTTTTCGATCGTTTTGGATG3' PCRandFlapAssayOligonucleotides: FLJ34208Flapoligo.i 5'GCGCCCCGGCCGCAGGCGGAGGACAGGGAGGAGCGCACACGAGAAAGCTCCCACGCGCCCGCGCCTCGCCTCCGA 5'GCGTTTCGGTCGTAGGCGGAGGATAGGGAGGAGCGTATACGAGAAAGTTTTTACGCGTTCGCGTTTCGTTTTCGA FLJ34208ForwardPrimer: FLJ34208ReversePrimer: Patent Application Publication Nov . 23 , 2017 Sheet 27 of 155 US 2017 /0335401 A1
5'CGCCGAGGCCGTCACCTCCA/3C6(SEQIDNO:135) 5'GGGCGCGAGTATAGTCG3(SEQIDNO:133) 5'CAACGCGACTAATCCGC3(SEQIDNO:134)
FIG.1(cont'd)
3
1 Bisulfite-convertedTargetDNA:(SEQIDNO132)
I 131)SEQNO:IDDNA(TargetFLJ45983 GCCGTGCTCAACGGGCAGCACCCGGACACGCAC3' GTCGTGTTTAACGGGTAGTATTCGGATACGTAT3' 5'CGAGAGGGCGCGAGCACAGCCGAGGCCATGGAGGTGACGGCGGACCAGCCGCGCTGGGTGAGCCACCACCACCCC 5'CGAGAGGGCGCGAGTATAGTCGAGGTTATGGAGGTGACGGCGGATTAGTCGCGTTGGGTGAGTTATTATTATTTC PCRandFlapAssayOligonucleotides: FLJ45983ForwardPrimer: FLJ45983ReversePrimer: FLJ45983Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 28 of 155 US 2017 /0335401 A1
3
I IIII. 5'TCGATTATGTCGTTTTAGACGTTATCG3(SEQIDNO:138) 5'TCTACATCGACATTCTAAAACGACTAAC3(SEQIDNO:139) 5'CCACGGACGCGCATACCATCG/3C6(SEQIDNO:140) IT: II
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO137) TAGAGGTCGTAGCTGAAGCCGATGGTATGCGCCAGCCGCTTCAGAATGTCGATGCAGAAACCCTTG3' TAGAGGICGTAGTTGAAGTCGATGGTATGCGTTAGTCGTITTAGAATGICGATGTAGAAATTTTTG3'Ar17 GRIN2DTargetDNA:(SEQIDNO136) PCRandFlapAssayoligonucleotides: 5'CGCCCCCTCACCTCCCCGATCATGCCGTTCCAGACGCCATCGATCTTCTTTCCGTGCTTGCCATTGGTGACCAGG 5'CGTTTTTTTATTTTTTCGATTATGTCGTTTTAGACGTTATCGATTTTTTTTTCGTGTTTGTTATTGGTGATTAGG GRIN2DForwardPrimer: GRIN2DReversePrimer: GRIN2DFlapoligo.i Patent Application Publication Nov . 23, 2017 Sheet 29 of 155 US 2017 /0335401 A1
5'TGGCGTATTATAATAAGCGTTCG3(SEQIDNO:143) 5'CCACGGACGCGTCTAAATCTC/306)(SEQIDNO:145 5'AACAACAAACGCACGACC3(SEQIDNO:144)
FIG.1(contd)
4 Bisulfite-convertedTargetDNA:(SEQIDNO142) HISTIH2BETargetDNA:(SEQIDNO141) CCTGCTGCTTCCCGGGGA3' TTTGTTGTTTTTCGGGGA3' PCRandFlapAssayoligonucleotides: HIST1H2BEForwardPrimer: HIST1H2BEReversePrimer: HIST1H2BEFlapoligo.:
I mond 1 5'CGGCGAGGCTTCCCGCCTGGCGCATTACAACAAGCGCTCGACCATCACCTCCAGGGAGATCCAGACGGCCGTGCG 5'CGGCGAGGTTTTTCGTTTGGCGTATTATAATAAGCGTTCGATTATTATTTTTAGGGAGATTTAGACGGTCGTGCG TT Patent Application Publication Nov . 23 , 2017 Sheet 30 of 155 US 2017 /0335401 A1
17
TIIIMGIIIII 777
71711721£££A{ 5'TTGGGTAATTATTACGTGGATTCG3(SEQIDNO:148) 5'CCACGGACGCGACGCCCAACA/3C6(SEQIDNO:150) 5'ACTCATCCGCGACGTC3(SEQIDNO:149)
77
7
4
1
FIG.1(cont'd) 17/ IVGIIIIII Bisulfite-convertedTargetDNA:(SEQIDNO147) mnm777
HOXA9TargetDNA:(SEQIDNO146) TGGGCGTCGACGTCGCGGATGAGTTG3' PCRandFlapAssayoligonucleotides: 5'GGGCGGGCCAGGCGCTGGGCACGGTGATGGCCACCACTGGGGCCCTGGGCAACTACTACGTGGACTCGTTCCTGC TGGGCGCCGACGCCGCGGATGAGCTG3 5'GGGCGGGTTAGGCGTTGGGTACGGTGATGGTTATTATTGGGGTTTTGGGTAATTATTACGTGGATTCGTTTTTGT HOXA9ForwardPrimer: HOXA9ReversePrimer: HOXA9Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 31 of 155 US 2017 /0335401 A1
YOU TIMMT TITI 5'GTTAGAAGACGTTTTTTCGGGG3(SEQIDNO:153) 5'AAAACAAAAATCGACCGCGA3(SEQIDNO:154) 5'CGCCGAGGGCGTTAGGATTT/306)(SEQIDNO:155 III
2
5 paat mnmtim FIG.1(cont'd)
7
1
1 Bisulfite-convertedTargetDNA:(SEQIDNO152) La HOXB2TargetDNA:(SEQIDNO151) PCRandFlapAssayOligonucleotides: GACTCCTGTCTCCAGCTATC3' GATTTTTGTTTTTAGTTATT3' HOXB2ForwardPrimer: HOXB2ReversePrimer: HOXB2Flapoligo.:
u 5'GGGCCATTGCCAGAAGACGTCTTCTCGGGGCGCCAGGATTCACCTTTCCTTCCCGACCTCAACTTCTTCGCGGCC 5'GGGTTATTGTTAGAAGACGTTTTTTCGGGGCGTTAGGATTTATTTTTTTTTTTCGATTTTAATTTTTTCGCGGTC13 Patent Application Publication Nov . 23 , 2017 Sheet 32 of 155 US 2017 /0335401 A1
5'AGTTTTCGGGTTTTGGAGTTCGTTA3(SEQIDNO:158) 5'CGCCGAGGACGGCGGTAGTT/3C6(SEQIDNO:160) 5'CCAAATCCAACCGCCGC3(SEQIDNO:159) TTTT? VII
FIG.1(contd) KLHDC7B.chr2250987185-50987290TargetDNA:(SEQIDNO156) 157)SEQIDNO:(TargetDNAconvertedBisulfite-
GTTGCCTGGACGTGCTGGCCTTTGCCCAGCA3' GTTGTTTGGACGTGTTGGTTTTTGTTTAGTA3' PCRandFlapAssayOligonucleotides: 5'GGCCCCGGAAGCCCAGCTCCCGGGCCCTGGAGCCCGCCACGGCGGCAGCCCTGCGGCGGCGGCTGGACCTGGGCA 5'GGTTTCGGAAGTTTAGTTTTCGGGTTTTGGAGTTCGTTACGGCGGTAGTTTTGCGGCGGCGGTTGGATTTGGGTA KLHDC7BForwardPrimer: KLHDC7BReversePrimer:KLHDC7B Flapoligo.: Patent Application Publication Nov . 23, 2017 Sheet 33 of 155 US 2017 /0335401 A1
parut LOC100129726ForwardPrimer:5'TTGATTGCGGTAGTAGGCG3(SEQIDNO163) LOC100129726ReversePrimer:5'AACCCACAACTTTACCGCC3(SEQIDNO164) LOC100129726Flapoligo.i5'CGCCGAGGCGTAACCGCCGC/3C6(SEQIDNO:165) FIG.1(cont'd)
162)IDNO:DNA(SEQTargetBisulfiteconverted- LO LOC100129726TargetDNA:(SEQIDNO161) 5'GGCGGCGCCGGCGGCTGCGCGGGGGGCGCCAGGCCCTGCTGCTGCTGCTGCTGCTGACTGCGGTAGTAGGCGGCG GCGGCCACGGCGGCAAAGTTGTGGGTCTGGA 5'GGCGGCGTCGGCGGTTGCGCGGGGGGCGTTAGGTTTTGTTGTTGTTGTTGTTGTTGATTGCGGTAGTAGGCGGCG GCGGTTACGGCGGTAAAGTTGTGGGTTTGGA PCRandFlapAssayoligonucleotides: Patent Application Publication Nov . 23 , 2017 Sheet 34 of 155 US 2017 /0335401 A1
LI
5'GTTTCGGGGTTTTTTTACGTTTTTTCG3(SEQIDNO:168) 5'CGCCGAGGGTCGACGTGTTT/3C6(SEQIDNO:170) 5'AAACGCGACTCGAAAACGC3(SEQIDNO:169)
FIG.1(cont'd)
£
£
£ Bisulfite-convertedTargetDNA:(SEQIDNO167)
U MATKTargetDNA:(SEQIDNO166) YOTE PCRandFlapAssayoligonucleotides: MATKForwardPrimer: MATKReversePrimer: MATKFlapoligo.i 5'GGTTTccccCCAccccGGCCTcGGGGTCTICTCCACGTCTcccccccGACGTGCTCACCTGCTCAGGGGGCGcccc CGAGCCGCGCCCCGCGCCCGCCCCCAGGAGGGCCTCCGCGAGCCGGCTGCACACCCCGAGGCGGTCCCGGCTGCACA AC3) 5'GGTTTTTTTTTATTTCGGTTTCGGGGTTTTTTTACGTTTTTTCGTCGACGTGTTTATTTGTTTAGGGGGCGTTTT CGAGTCGCGTTTCGCGTTCGTTTTTAGGAGGGTTTTCGCGAGTCGGTTGTATATTTCGAGGCGGTTTCGGTTGTATA 'AT3 Patent Application Publication Nov . 23, 2017 Sheet 35 of 155 US 2017 /0335401 A1
ITTT1
7 TITI IT 1 3
Timur??
17 17 TIF FIG.1(cont'd) MAXChr10.225ForwardPrimer:5'CGGTTTTTAGCGATATTAGGCG3(SEQIDNO173) MAXChr10.225Flapoligo:5'CGCCGAGGGCGGTTAGTGTT/3C6(SEQIDNO175) MAX_Chr10.225ReversePrimer:CCCAAAACGTAACCCCGA35'(SEQIDNO174)
MAX.chr1022541891-22541946TargetDNA:(SEQIDNO171) Bisulfite-convertedTargetDNA:(SEQIDNO172) IT 17"LE
7M PCRandFlapAssayoligonucleotides:
I 5'CTCCGGTTTTCGCGGTTCTCAGCGATATTAGGCGCGGCCAGTGTCTGAAAGCTCCTCGGGGTTACGTCCTGGGGCim GACTGGAGGCGGCTCACGAC3' 5'TTTCGGTTTTCGCGGTTTTTAGCGATATTAGGCGCGGTTAGTGTTTGAAAGTTTTTCGGGGTTACGTTTTGGGGC GATTGGAGGCGGTTTACGAT3' Patent Application Publication Nov . 23 , 2017 Sheet 36 of 155 US 2017 /0335401 A1
??
1 VI
I
FIG.1(cont'd) TY MAXChr10.226ForwardPrimer:5'GGGAAATTTGTATTTCGTAAAATCG3(SEQIDNO178) MAX_Chr10.226ReversePrimer:5'ACAACTAACTTATCTACGTAACATCGT3(SEQIDNO179) MAXChr10.226Flapoligo:5'CCACGGACGGCGGTTAAGAAA/3C6(SEQIDNO180) MAX.chr1022624430-22624544TargetDNA:(SEQIDNO176) GTAGACAAGCCAGTTGTGACGTTCAGCACAACGTGCTACTGAACTACCGAGATCCGCCACCAAATGGC3' Bisulfite-convertedTargetDNA:(SEQIDNO177) GTAGATAAGTTAGTTGTGACGTTTAGTATAACGTGTTATTGAATTATCGAGATTCGTTATTAAATGGT3' 1m(217IIIAULALA 5'CGACGGCCGCGGAGGAGGAAGGCCAGGGGGAAATTTGCATTTCGTAAAACCGCGGTTAAGAAATGACGATGCCAC 5'CGACGGTCGCGGAGGAGGAAGGTTAGGGGGAAATTTGTATTTCGTAAAATCGCGGTTAAGAAATGACGATGTTAC PCRandFlapAssayOligonucleotides: Patent Application Publication Nov . 23, 2017 Sheet 37 of 155 US 2017 /0335401 A1
8
1
FIG.1(cont'd) MAX.chr1252ForwardPrimer:5'TCGTTCGTTTTTGTCGTTATCG3(SEQIDNO183) 5'AACCGAAATACAACTAAAAACGC3(SEQIDNO:184)MAX.chr1252ReversePrimer MAX.chr1252Flapoligoi5'CCACGGACGCGAACCCCGCAA/3C6(SEQIDNO:185) MAX.chr1252652268-52652362TargetDNA:(SEQIDNO181) CGGGGTCCGCGCCTTCAGCTGCATCTCGGCCTGCGGGCCCC3' Bisulfite-convertedTargetDNA:(SEOIDNO182) CGGGGTTCGCGTTTTTAGTTGTATTTCGGTTTGCGGGTTTT3' 5'GGCTIGGGGTCCAGcccccccccccTGcccccAccGCACCATGTccTGcc??????ccccccTCAGcccccccTG 5'GGTTTGGGGTTTAGTCGTTCGTTTTTGTCGTTATCGTATTATGTTTTGTTTTTATTTTCGTTTTAGCGTTTTTTG PCRandFlapAssayOligonucleotides: Patent Application Publication Nov . 23 , 2017 Sheet 38 of 155 US 2017 /0335401 A1 niya 77777
IV AA17T
N
FIG.1(cont'd) ATT71 JL MAX.chr1650ForwardPrimerv2:5'TTCGAGTTTAAGGAAGTCG3(SEQIDNO188):189)TCTAAAAACGTAAATTTCGAACT3(SEQIDNO 'Primer5Reversev2:chr16.50MAX 5'CCACGGACGGCGATAATGTAG/3C6(SEQIDNO:190)MAX.chr1650Flapoligoi MAX.chr1650875223-50875241TargetDNA:(SEQIDNO186) Bisulfite-convertedTargetDNA:(SEQIDNO187) cCAcccccccAGACCITATCT'Gccccccccccc3 TTACGTTTTTAGATTTTATTTGCGCGTTTCGTT3' PCRandFlapAssayOligonucleotides:
5'GGAAGGCTGCAGCGAGAGATTTACATATTCATCCGAGCTTAAGGAAGCCGCGATAATGCAGGTACAGCCCGAAAC 5'GGAAGGTTGTAGCGAGAGATTTATATATTTATTCGAGTTTAAGGAAGTCGCGATAATGTAGGTATAGTTCGAAAT 4 Patent Application Publication Nov . 23, 2017 Sheet 39 of 155 US 2017 /0335401 A1
fe
FIG.1(cont'd) MAX.chr1916ForPrimer:5'TTTAATTATCGTTTTTTAGAATATTACGGA3(SEQIDNO193) MAX.chr1916ReversePrimer:5'ACTATTATTCTCAACAATCGCAAAAC3(SEOIDNO194) MAX.chr1916Flapoligo;5'CCACGGACGCCTCGTATTAAC/3C6(SEQIDNO:195) MAX.chr1916394489-16394575TargetDNA:(SEQIDNO191) CTGTGCTTCCTGCTAACACGAGGCCCTGCGATTGCTGAGAACAACAGCCCCGAGACTGCGCG3' Bisulfite-convertedTargetDNA:(SEOIDNO192) TTGTGTTTTTTGTTAATACGAGGTTTTGCGATTGTTGAGAATAATAGTTTCGAGATTGCGCG3' 5'GGAGTTATTTTTAACCATCGCCTC?CAGAACATTACGGAGCTICCTCTCTCCAACACGCAGGAAAC?CTACTIGG 5'GGAGTTATTTTTAATTATCGTTTTTTAGA?TATTACGGAGTTTTTTTTTTTTAATACGTAGGAAA????????GG PCRandFlapAssayOligonucleotides: Patent Application Publication Nov . 23 , 2017 Sheet 40 of 155 US 2017 /0335401 A1
5'AGTTTAAGATTGCGGAATTATATTCGT3(SEQIDNO:198) 5'CGCCGAGGAACGCTAAATCT/3C6(SEQIDNO:200)
5'TTCCGAAATCCCCGCAA3(SEQIDNO:199) V
FIG.1(cont’d)
MAII.I MAX.chr1937288426-37288480TargetDNA:(SEQIDNO196) CCAGCGCCTGCGGGGATTCCGGAAACGGGAGAGCGGGCTTCC3'. Bisulfite-convertedTargetDNA:(SEQIDNO197)
TTAGCGTTTGCGGGGATTTCGGAAACGGGAGAGCGGGTTTTT3' i PCRandFlapAssayoligonucleotides: 5'GGCGGGCGCTTGGCCAAACAGCCCAAGACTGCGGAATCACACTCGCCACTGTGTACCTGGACGCCATCTGCAGAC 5'GGCGGGCGTTTGGTTAAATAGTTTAAGATTGCGGAATTATATTCGTTATTGTGTATTTGGACGTTATTTGTAGAT MAX.chr1937ForPrimerv2: MAX.chr1937ReversePrimer: MAX.chr1937Flapoligo: Patent Application Publication Nov . 23 , 2017 Sheet 41 of 155 US 2017 /0335401 A1
117
777IAI SITIITIA
- FIG.1(cont'd) 170CT? LL MAXChr8.124ForwardPrimer:5'GGTTGAGGTTTTCGGGTTTTTAG3(SEQIDNO203) MAXChr8.124Flapoligo,5'CGCCGAGGGCGGGTTTTCGT/3C61(SEQIDNO:205) MAX_Chr8.124ReversePrimerv2:5'ACGAAAATAATCAAAATCGCCTCC3(SEQIDNO207) MAXChr8.124Flapoligo,v2:5'CGCCGAGGCCCACGAAATCG)3061(SEQIDNO208 MAXChr8.124ReversePrimer:5'CCTCCCCACGAAATCGC3(SEQIDNO204) MAXChr8.124ForwardPrimerv2:5'AGGAGTTTTTTTGCGCGG3(SEQIDNO206) ILA ITUTUT17017IL MAX.chr8124173236-124173370TargetDNA:(SEQIDNO201) Bisulfite-convertedTargetDNA:(SEQIDNO202) IL rir?riIII
I
I TUITIIII riri PCRandFlapAssayOligonucleotides:
I
I 5'CGCAGGCTGAGGCCCTCGGGTCCCCAGCGGGTCCTCGCCATCAGTCACTCTCTACGGGCCAGGCCTGGGGGTCAC GGCCTGCAGGAGCCTCCCTGCGCGGCCCCACTCCCTCATCTGCGACCCCGTGGGGAGGCGACCCTGACCACCCTCGT TCCG3? 5'CGTAGGTTGAGGTTTTCGGGTTTTTAGCGGGTTTTCGTTATTAGTTATTTTTTACGGGTTAGGTTTGGGGGTTAC GGTTTGTAGGAGTTTTTTTGCGCGGTTTTATTTTTTTATTTGCGATITCGTGGGGAGGCGATTTTGATTATTTTCGTT: TTCG3' Patent Application Publication Nov . 23 , 2017 Sheet 42 of 155 US 2017 /0335401 A1
TTTTLIII AMMI JI
17 T
TN771???
FIG.1(cont'd) my MAXChr8.145ForwardPrimer:5'GCGGTATTAGTTAGAGTTTTAGTCG3(SEQIDNO211) MAXChr8.145ReversePrimer:5'ACAACCCTAAACCCTAAATATCGT3(SEOIDNO212) MAX_Chr8.145Flapoligoi5'CCACGGACGGACGGCGTTTTT/3061(SEQIDNO:213) MAX.chr8145105646-145105653TargetDNA(SEQIDNO:209) CCTAAGCACGATACCCAGGGCCCAGGGCTGCTCTTGGCG3' Bisulfite-convertedTargetDNA:(SEQIDNO210) TTTAAGTACGATATTTAGGGTTTAGGGTTGTTTTTGGCG3' 5'CGGGGGAGGGCGGCATCAGCCAGAGCCTCAGCCGACGGCGCTCCCCAGGTCCACTTCCCGCTCCGATACCCTCCC 5'CGGGGGAGGGCGGTATTAGTTAGAGTTTTAGTCGACGGCGTTTTTTAGGTTTATTTTTCGTTTCGATATTTTTTT Oligonucleotides:AssayandFlapPCR Patent Application Publication Nov . 23, 2017 Sheet 43 of 155 US 2017 /0335401 A1
7777ITI7IIII TIIM 17TITIAII
A
ITI7 TI IULAZA T7 IIb IIGI T 5'AACCTAAATAATTAATAACAACCCGC3(SEQIDNO:217)MAXChr1.110ReversePrimer FIG.1(contd) MAXChr1.110(chr1110627198-110627213)TargetDNA:SEQIDNO214 YA 777T7 MAXChr1.110ForwardPrimer:5'TTTCGTAGGTTTGGTCGCG3(SEQIDNO216) MAXChr1.110Flapoligo:5'CCACGGACGGCGACGGGTATT/3C6(SEQIDNO218) Bisulfite-convertedTargetDNA:(SEQIDNO215) Tim
11 GGGCACTGCATCCATTTC?CTCGCGTGCGC?C3" MIT GGGTATTGTATTTATTTTTTTCGCGTGCGTTT3'I Oligonucleotides:AssayandFlapPCR
I
ITIFT ?? 1
? LV
I - 5'CTCCGCTCCCCGCAGGCCTGGCCGCGCGACGGGCACCCAGCGGGTTGTTATCAATTATTCAGGCCCCAAGTTCAC 5'TTTCGTTTTTCGTAGGTTTGGTCGCGCGACGGGTATTTAGCGGGTTGTTATTAATTATTTAGGTTTTAAGTTTAC Patent Application Publication Nov . 23 , 2017 Sheet 44 of 155 US 2017 /0335401 A1
III
in 117.71OLIA
7717 ITI 222)SEQIDNO:TCTAACCCTATTTAACCAACCGA3'(5' 5'CGCCGAGGGCGGTTAAAGTG/3C6(SEQIDNO:223) TIN JLI 221)SEQNO:IDTGGTTCGGGCGTGACGCG3'(5'
?
?
?
?
FIG.1(cont'd) ??? T 7??
.i AyonF71 Bisulfite-convertedTargetDNA:(SEQIDNO220) III
TI1191 JU 219)NO:SEQIDDNA(TargetNFIX TinUI PCRandFlapAssayOligonucleotides: GCGAAGCAGAAGG3'. GCGAAGTAGAAGG3' NFIXHMForwardPrimer: NFIXHMReversePrimer: NFIX_HMFlapoligo.: 5'GTGGGCCGGGCGTGACGCGCGGTCAAAGTGCAATGATTTTTCAGTTCGGTTGGCTAAACAGGGTCAGAGCTGAGA I Patent Application Publication Nov . 23, 2017 Sheet 45 of 155 US 2017 /0335401 A1
5'GATTTTTTCGGTTTCGTTTTATTCG3(SEQIDNO:226) 5'CAATCGCAAAATATCCTTAACCGA3(SEQIDNO:227) 5'CCACGGACGGTTTTCGGGATG/306!(SEQIDNO:228)
FIG.1(contd)
2 2 Bisulfite-convertedTargetDNA:(SEQIDNO225)
NKX2-6TargetDNA:(SEQIDNO224) PCRandFlapAssayOligonucleotides: 5'GG????ccTCGGccccccccCATccGcc??cGGGATGCTGCTGAGccccGTCACCTCCAccccc????cGGTCAA GGACATCCTGCGACTGGAG3' 5'GGATTTTTTCGGTTTCGTTTTATTCGTTTTCGGGATGTTGTTGAGTTTCGTTATTTTTATTTTTTTTTCGGTTAA GGATATTTTGCGATTGGAG3' NKX2-6ForwardPrimer: NKX2-6ReversePrimer: NKX2-6Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 46 of 155 US 2017 /0335401 A1
aTITUIT r17171TIlirn I.Lebruit
m 5'CGTCGCGTTTTTCGGTTATACG3(SEQIDNO:231) 5'CGCGAAAACTAAAAAACCGCG3(SEQIDNO:232) 5'CCACGGACGGCACCGTAAAAC/3C6(SEQIDNO:233)
17617 ir 3
FIG.1(cont'd) 7 de
ITI7 who Hn
T
230)SEQIDNO:DNA(TargetBisulfiteconverted- 7TIITIITI UI 229)NO:SEQIDDNA(TargetOPLAH ITI PCRandFlapAssayOligonucleotides:
? I OPLAHForwardPrimer: OPLAHReversePrimer: OPLAHFlapoligo.:
CCTCCCCGCCCCCG3' 1 I TUTI mnm 5'CTGTCAGTGCTGACCGAGCGCCGCGCCTTCCGGCCATACGGGCTCCACGGTGCGCGGTTCCCCAGCCCTCGCGGC 5'TTGTTAGTGTTGATCGAGCGTCGCGTTTTTCGGTTATACGGGTTTTACGGTGCGCGGTTTTTTAGTTTTCGCGGTI Patent Application Publication Nov . 23, 2017 Sheet 47 of 155 US 2017 /0335401 A1
5'CGTAACGTAAAATCTCTACGCCC3(SEQIDNO:237) 5'CCACGGACGCGCTCGAACTAC/3C61(SEQIDNO:238) 5'GGTTGAGTTTGGGGTTCG3(SEQIDNO:236)
FIG.1(cont'd)
235)IDNO:(SEQTargetDNABisulfiteconverted- PARP15TargetDNA:(SEQIDNO234) AGCAGCCCGAGCGGGCGCAGAGACCCCACGCCACGCACA3' AGTAGTTCGAGCGGGCGTAGAGATTTTACGTTACGTATA3' PCRandFlapAssayOligonucleotides: 5'CGGAGTATGGTGAGGAGCGCGGGGGACGGGTGCGGGAAGGGGACAGCAGGGCTGAGCCTGGGGCCCGCAAGACCC 5'CGGAGTATGGTGAGGAGCGCGGGGGACGGGTGCGGGAAGGGGATAGTAGGGTTGAGTTTGGGGTTCGTAAGATTT PARP15ForwardPrimer: PARP15ReversePrimer: PARP15Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 48 of 155 US 2017 /0335401 A1
Tim
onT ima LLLLL PRDM14ReversePrimerv2:5'CCACGATAACCAAACTATTTCTATAATATCC3(SEQIDNO242) 11 PRDM14ForwardPrimer:5'GAGTAGTTCGTAGAATTTGGTCG3(SEQIDNO241) PRDM14Flapoligo.:5'CCACGGACGGCGTATTATACG/3C6(SEQIDNO243) PRDM14Rev.Primerv3:5CTATTTCTATAATATCCATAAAACGAAAAAAACGT3'(SEQIDNO:245) FIG.1(cont'd) ????? PRDM14ForwardPrimerv3:5'GGAGAGTAGTTCGTAGAATTTGG3(SEQIDNO244) 246)SEQIDNO:CCACGGACGGTCGCGTATTAT/3C6(oligov35'.PRDM14Flap
240)IDNO:(SEQTargetDNABisulfiteconverted-
PRDM14TargetDNA:(SEQIDNO239) 5'GGAGAGCAGCCCGCAGAACCTGGCCGCGTACTACACGCCTTTCCCGTCCTATGGACACTACAGAAACAGCCTGGC CACCGTGGAGGAAGACTTCC3'. 5'GGAGAGTAGTTCGTAGAATTTGGTCGCGTATTATACGTTTTTTTCGTTTTATGGATATTATAGAAATAGTTTGGT TATCGTGGAGGAAGATTTTT3' PCRandFlapAssayOligonucleotides: Patent Application Publication Nov . 23, 2017 Sheet 49 of 155 US 2017 /0335401 A1
5'CTTCTACAACCACTACACCGA3(SEQIDNO:250) 5'CCACGGACGGCGCGCGTTTAT/306)(SEQIDNO:251 5'GGAAGGTGTTTTGCGCG3(SEQIDNO:249)
Y
FIG.1(cont'd)
M Bisulfite-convertedTargetDNA:(SEQIDNO248)
PRKCB28TargetDNA:(SEQIDNO247) PCRandFlapAssayoligonucleotides: 5'GGGAAGGTGCCCTGCGCGCGCGCGCTCACCAGATGAAGTCGGTGCAGTGGCTGCAGAAGGTGGGCTGCTTGAAGA AGCGGGCGGTGAATTTG3' 5'GGGAAGGTGTTTTGCGCGCGCGCGTTTATTAGATGAAGTCGGTGTAGTGGTTGTAGAAGGTGGGTTGTTTGAAGA AGCGGGCGGTGAATTTG3' PRKCB28ForwardPrimer: PRKCB28ReversePrimer: PRKCB28Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 50 of 155 US 2017 /0335401 A1
WAIT 5'GGGTTCGGGGATTTATAATTACGG3(SEQIDNO:254) 5'CCTCCTACTACTAACGCTAATAACC3(SEQIDNO:255) 5'CCACGGACGCGTACTCTTCAC/306(SEQIDNO:256)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO253)
PTGDRTargetDNA:(SEQIDNO252) GGAGGTGATCCAGCTCCTCCAGGGGCTGAGGG3' GGAGGTGATTTAGTTTTTTTAGGGGTTGAGGG3' oligonucleotides:AssayandFlapPCR 5'GCCTCGGGGCCCGGGGACTCACAATTACGGGCAGAGAACACATAGTGAAGAGCACGGTCATCAGCGCCAGCAGCA 5'GTTTCGGGGTTCGGGGATTTATAATTACGGGTAGAGAATATATAGTGAAGAGTACGGTTATTAGCGTTAGTAGTA PTGDRForwardPrimer: PTGDRReversePrimer: PTGDRFlapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 51 of 155 US 2017 /0335401 A1
YAULLI
5'CGCCGAGGCGCGACTCGACA/3C6(SEQIDNO:261) 5'GTTGTAGCGGTATTCGCG3(SEQIDNO:259) 5'CTTCTCTCCCGTCCGCGC3!(SEQIDNO:260)
FIG.1(cont'd) 47mmJUIII
Bisulfite-convertedTargetDNA:(SEQIDNO258) mmmmIUI 257)NO:SEQIDDNA(TargetPTGDR9_ mi JU PCRandFlapAssayOligonucleotides: PTGDR9ForwardPrimer: PTGDR9ReversePrimer: PTGDR9Flapoligo.:
promene 5'GGCGGCTGCAGCGGCACCCGCGCTCCTGCACCAGGGACTGTGCCGAGCCGCGCGCGGACGGGAGGGAAGCGTCCC CTCAG 5'GGCGGTTGTAGCGGTATTCGCGTTTTTGTATTAGGGATTGTGTCGAGTCGCGCGCGGACGGGAGGGAAGCGTTTT TTTAG Patent Application Publication Nov . 23, 2017 Sheet 52 of 155 US 2017 /0335401 A1
I
HumanRASSF1ForwardPrimerv2:5'AGAAATACGGGTATTTTCGCG3(SEQIDNO264) HumanRASSF1ReversePrimerv2:5'CCACAACGACGACGACC3(SEQIDNO265):266) SEQIDNOCCACGGACGCGCAAAACACCA/3C6(oligo.V2:5'HumanRASSF1Flap FIG.1(contd) GGGGACGAAGGAGGGAAGGAAGGGCAAGGCGGGGGGGGCTCTGCGAGAGCGCGCCCAGCCCCGCCTTC3' Bisulfite-convertedTargetDNA:(SEQIDNO263) GGGGACGAAGGAGGGAAGGAAGGGTAAGGCGGGGGGGGTTTTGCGAGAGCGCGTTTAGTTTCGTTTTT3' HumanRASSF1TargetDNA:(SEQIDNO262) PCRandFlapAssayOligonucleotides: 5'TCCAGAAACACGGGTATCTCCGCGTGGTGCTTTGCGGTCGCCGTCGTTGTGGCCGTCCGGGGTGGGGTGTGAGGA 5'TTTAGAAATACGGGTATTTTCGCGTGGTGTTTTGCGGTCGTCGTCGTTGTGGTCGTTCGGGGTGGGGTGTGAGGA F Patent Application Publication Nov . 23 , 2017 Sheet 53 of 155 US 2017 /0335401 A1
TI
ITI7 III
19 QUIIIII 11TIITIT 5'CCACGGACGATCCGCAAACGC/3061(SEQIDNO:271) 5'GTTCGAGTTTAGGGGTAGCG3'(SEQIDNO:269) 5'CCGCACAAAAAACCGCA3(SEQIDNO:270) mynin17IT17
QUMLIIIALGIN 077 FIG.1(cont'd) TipyTryitim IVOIII IL Bisulfite-convertedTargetDNA:(SEQIDNO268)
pornynr7 SHOX2TargetDNA:(SEQIDNO267) AGCGAGGGGCGTCTGCGGATGCGGCTCCCTGTGCGGCACAACACC3' AGCGAGGGGCGTTTGCGGATGCGGTTTTTTGTGCGGTATAATATT3' PCRandFlapAssayOligonucleotides: 5'CGGTCGGGCAGGCGGGACGGAGATTACCTGGCTGTCCAGGGGACCTTATGCAGGGTTTGGCCCGAGCCCAGGGGC SHOX2ForwardPrimer: SHOX2ReversePrimer: SHOX2Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 54 of 155 US 2017 /0335401 A1
A
5'CGCCGAGGGCGTTTTGAAGT/3C6(SEQIDNO:276)
£ ? 5'GGAGTATTCGTCGTTGCG3(SEQIDNO:274) J 5'CGAAAACCGCCTACCGC3(SEQIDNO:275)
-Tin,3
FIG.1(cont'd)
I JLI 1917 273)IDNO:(SEQTargetDNABisulfiteconverted- ?
Tin ?
? ? i SHROOM1TargetDNA:(SEQIDNO272) *
* PCRandFlapAssayOligonucleotides: 5'ccGGAGCACTcGccGcTGCGCGC?CTGAAGCCGCTGGCGGTAGGCGGC?CTCGAG3 * KUIVUILQ SHROOM1_HMForwardPrimer: SHROOMIHMReversePrimer: SHROOM1_HMFlapoligo.: Tim GI
TI
5 Patent Application Publication Nov . 23 , 2017 Sheet 55 of 155 US 2017 /0335401 A1
when I
L
L.LA 5'ACGGTTTTTTCGTTATTTTTACGGG3(SEQIDNO:279) 5'CAACGCCTAAAAACACGACTC3(SEQIDNO:280)5CGCCGAGGGGCGGTTGTTGG'/3C61( SEQIDNO:281)
FIG.1(cont'd)
i71
Bisulfite-convertedTargetDNA:(SEQIDNO278) I YAT II SKITargetDNA:(SEQIDNO277) JU TH CGTTGGTTTTTATGGAGGTGG3' PCRandFlapAssayOligonucleotides: CGTTGGCTCTCATGGAGGTGG3' SKIFlapoligo.: 5'CCCGGGCCTACGGTCCTCCCGCCACCTCCACGGGGCGGCTGTTGGGGCCCCACCAGGCAGAGCCGTGTTCTCAGG 5'TTCGGGTTTACGGTTTTTTCGTTATTTTTACGGGGCGGTTGTTGGGGTTTTATTAGGTAGAGTCGTGTTTTTAGGYTM SKIForwardPrimer: SKIReversePrimer: Patent Application Publication Nov . 23, 2017 Sheet 56 of 155 US 2017 /0335401 A1
YAT S1PR4HMReversePrimer:5'CTTACGTATAAATAATACAACCACCGAATA3(SEQIDNO285) FIG.1(cont'd) S1PR4HMFlapoligo.:5'CCACGGACGACGTACCAAACA/3C6(SEQIDNO286) I S1PR4HMForwardPrimer:5'TTATATAGGCGAGGTTGCGT3(SEQIDNO284)
TTTGGTACGTATTCGGTGGTTGTATTATTTATACGTAAGATTGTAATTTTAGATGTTTCGTACGTTGGAGATG3' L ccTGGCACGCAcccGGTGGCTGCACCATCCACACGCAAGACTGCAACTICAGATCCTccGCACGCTGGAGATG3' TITI7 283)SEQIDNO:(TargetDNABisulfiteconverted-
S1PR4TargetDNA:(SEQIDNO282) PCRandFlapAssayOligonucleotides: 5'GGGCCTGTCCCGTTCCCTGCTCCCCATACAGGCGAGGCTGCGTGCACACAGCTTCCTGTACCCCAGGAGGGCCTG 5'GGGTTTGTTTCGTTTTTTGTTTTTTATATAGGCGAGGTTGCGTGTATATAGTTTTTTGTATTTTAGGAGGGTTTG N Patent Application Publication Nov . 23, 2017 Sheet 57 of 155 US 2017 /0335401 A1
5'CCACGGACGGGAGGGCGTAGG/3C6(SEQIDNO:291) 5'TTAGGAGGGTGGGGTTCG3(SEQIDNO:289) 5'CTTTCCTCGCAAAACCGC3(SEQIDNO:290)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO288)
SLC12A8TargetDNA:(SEQIDNO287) PCRandFlapAssayoligonucleotides: TGGGAAGGGACCC3' TGGGAAGGGATTT3' SLC12A8ForwardPrimer: SLC12A8ReversePrimer: SLC12A8Flapoligo.: 5'CGGAGCTAGGAGGGTGGGGCTCGGAGGGCGCAGGAAGAGCGGCTCTGCGAGGAAAGGGAAAGGAGAGGCCGCTTC 5'CGGAGTTAGGAGGGTGGGGTTCGGAGGGCGTAGGAAGAGCGGTTTTGCGAGGAAAGGGAAAGGAGAGGTCGTTTT ed Patent Application Publication Nov . 23 , 2017 Sheet 58 of 155 US 2017 /0335401 A1
5'CGTACCGTTCACGATAACGT3(SEQIDNO:295) 5'CGCCGAGGGGCGGTCGCGGT/306)(SEQIDNO:296 5'CGCCGAGGTTACAAACCGCG/3061(SEQIDNO:297) AN JT 5'TTTCGGCGGGTTTCGAG3(SEDIDNO:294)
.
T
FIG.1(cont'd) I
Bisulfite-convertedTargetDNA:(SEQIDNO293) I
292)NO:SEQIDDNA(TargetSOBP
- 5'cccccGGCGGGccccGAGGCGGccGcGGCCTGCAACGTCATCGTGAACGGCACGCGCGG3" 5'GTTTCGGCGGGTTTCGAGGCGGTCGCGGTTTGTAACGTTATCGTGAACGGTACGCGCGG3T PCRandFlapAssayOligonucleotides: SOBPHMForwardPrimer: SOBP_HMReversePrimer: SOBP_HMFlapoligo.i SOBP_HMFlapoligo.V2: Patent Application Publication Nov . 23 , 2017 Sheet 59 of 155 US 2017 /0335401 A1
TTT
7
5'CCACGGACGGTGTTCGTACGG/3C6(SEQIDNO:302) 5'CGAGATGGTGGAAGGCG3(SEOIDNO:300) TITET.LII 5'GCGCCCTTCTACACGAA3(SEQIDNO:301)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO299) MOTO SPOCK2TargetDNA:(SEQIDNO298) PCRandFlapAssayOligonucleotides: AGGCTGGTTTTCCC3' AGGTTGGTTTTTTT3' SPOCK2ForwardPrimer: SPOCK2ReversePrimer: SPOCK2Flapoligo.: 5'CTAGGCGAGATGGTGGAAGGCGTGTCCGTACGGGGGTGGGCTGGGGTCCCCGTGCAGAAGGGCGCGCGAGGACCC 5'TTAGGCGAGATGGTGGAAGGCGTGTTCGTACGGGGGTGGGTTGGGGTTTTCGTGTAGAAGGGCGCGCGAGGATTTTT Patent Application Publication Nov . 23 , 2017 Sheet 60 of 155 US 2017 /0335401 A1
5'CGCCGAGGCGACGAAAAACG/3C6(SEQIDNO:307) 5'AAATATGTGGTTCGTGGCGTT3(SEQIDNO:305) 5'ACGCAACAACGCGAAAAAC3(SEQIDNO:306)
FIG.1(cont'd)
VV TCCCCTTGTCCCCTCCGAGCGATGCTCCTGCGCCCTICGCCGCCTCCCGCGCTGCTGCGCCGCCAGGC???? Bisulfite-convertedTargetDNA:(SEQIDNO304) O STESIA1TargetDNA:(SEQIDNO303) TGTTTTTGCGTTTTTCGTCGTTTTTCGCGTTGTTGCGTCGTTAGGTAA3' PCRandFlapAssayoligonucleotides: 5'GCGCTGCTGCGCCGCCAGGCAAGGCGAGGGTCCGGGAGAAGGCTCGGCTCCCTCCTAAACATGTGGCCCGTGGCG 5'GGCGAGGGTTCGGGAGAAGGTTCGGTTTTTTTTTAAATATGTGGTTCGTGGCGTTTTTTTGTTTTTTTCGAGCGA ST8SIA1ForwardPrimer: STOSIA1ReversePrimer: ST8SIA1Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 61 of 155 US 2017 /0335401 A1
YnmJULI
5'CCACGGACGGGCGTGGAGGAG/306)(SEQIDNO:312 IT 5'GGGGTGTTGTAATAGGGTACG3(SEQIDNO:310) 5'CTAAACGCTCTAACCGCGA3(SEQIDNO:311) TMT FIG.1(cont'd) TCAGGTACTccTGCC??cGCGGC????ccccTCTAGCGTCC??????ccccGACTGCAGAGG3" 309)IDNO:(SEQTargetDNAconvertedBisulfite- TTAGGTATTTTTGTTTTCGCGGTTTTTTTTTTTTAGCGTTTTTTTTTTTTCGAGTGTAGAGG3' STESIA1_22TargetDNA:(SEQIDNO308) 5'ACGAGAAAGAGATCGTGCAGGGGGTGCTGCAACAGGGCACGGCGTGGAGGAGGAACCAGACCGCGGCCAGAGCGT 5'ACGAGAAAGAGATCGTGTAGGGGGTGTTGTAATAGGGTACGGCGTGGAGGAGGAATTAGATCGCGGTTAGAGCGT PCRandFlapAssayoligonucleotides; ST8SIA122ForwardPrimer: STOSIAL22ReversePrimer: ST8SIA122Flapoligo,i Patent Application Publication Nov . 23 , 2017 Sheet 62 of 155 US 2017 /0335401 A1
1
TO
5'TAGCGTCGAATGGAAGTTCGA3(SEQIDNO:315) MAIGII IITLGATTIGIATTAGAIILGI 5'GCGCGTAAACATAACGCACC3(SEQIDNO:317) 5'GCGCGTAAACATAACGCACC3(SEQIDNO:317) '5CCACGGACGCCGTACGAATCC/306(SEQIDNO:318) 4 3 TTTTTTT7 SP9ForwardPrimerUniversal:5'GGTCGTTAGCGTCGAATG3(SEQIDNO316) FIG.1(contd) SP9Flapoligo.i5'CCACGGACGCCGTACGAATCC/3061(SEQIDNO:318) YAMA GGIU Bisulfite-convertedTargetDNA:(SEQIDNO314) JULVIVI
SP9TargetDNA:(SEQIDNO313) CATGCCCACGCGCGGGTACAGCCCGTCGGCTGCCGTCGTGTG3' TUVU LOVIJUIIAIIIWIVULJIVI man PCRandFlapAssayOligonucleotides: SP9ForwardPrimer: SP9ReversePrimer: SP9Flapoligo.: SP9ReversePrimer: 7 UUU ? YTTmammmmmmmm ? ??3 wana ?? 5'CGCGCCGTTGGTCACCTCGCCGGCCGCCAGCGTCGAATGGAAGCCCGACTTGTACCAGGACTCGTACGGGTGCGC 2 Patent Application Publication Nov . 23 , 2017 Sheet 63 of 155 US 2017 /0335401 A1
4 iAIO 2 JIHALLI TL11 VA
?????1 LILILAIVGILIWIIGIIIII07/07dadanII SUCLG2HMForwardPrimer:5'TCGTGGGTTTTTAATCGTTTCG3(SEQIDNO321) It FIG.1(cont'd) norryAnn SUCLG2HMReversePrimer:5/TCACGCCATCTTTACCGC3'(SEQIDNO322) SUCLG2HMFor.PrimerUniv:5'GGTTTTTTTTCGTGGGTTTTTAATCG3(SEQIDNO324) SUCLG2HMFlapoligo.Univ:5'CCACGGACGGTTTCGTTGATT/3C6(SEQIDNO326) IIIIWW SUCLG2HMFlapoligo.:5'CCACGGACGCGAAAATCTACA/3C6(SEQIDNO323) SUCLG2HMRev.PrimerUniv:5'CTAATCACGCCATCTTTACCG3(SEQIDNO325) Bisulfite-convertedTargetDNA:(SEQIDNO320) SUCLG2TargetDNA:(SEQIDNO319) ?nimannman777777 - - -
T 3 PCRandFlapAssayoligonucleotides: VU £ 1
£ F
£
I GTGACCAGAA3' T GTGATTAGAA3' Patent Application Publication Nov . 23 , 2017 Sheet 64 of 155 US 2017 /0335401 A1
) 5'TCTAAACAAAACTAACTTTAATTAACGCCC3(SEQIDNO:330) 5'CGTAGGTCGGATTGATTTTTCGT3(SEQIDNO:329) 5'CCACGGACGCGAACACCCGCA/306)(SEQIDNO:331
Tim
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO328) TBX15TargetDNARegion1:(SEQIDNO327) CGGGCGCCAACTAAAGCCAGCTCTGTCCAGACGCGGAAAG3' CGGGCGTTAATTAAAGTTAGTTTTGTTTAGACGCGGAAAG3' oligonucleotides:AssayandFlapPCR TBX15ForwärdPrimer: TBX15ReversePrimer: TBX15Flapoligo.: 5'GGAGTGAGTGCCTACAACGCGCAGGCCGGA?TGATCCCCCGTTGCTGCAGGTTGGTGCCCCAAGCTGCGGGTGCT 5'GGAGTGAGTGTTTATAACGCGTAGGTCGGATTGATTTTTCGTTGTTGTAGGTTGGTGTTTTAAGTTGCGGGTGTT U Patent Application Publication Nov . 23 , 2017 Sheet 65 of 155 US 2017 /0335401 A1
V
L
FIG.1(cont'd) TBX15Reg.2ForwardPrimerUniv:5'GGAAGGAAATTGCGGGTTTTC3(SEQIDNO333) TBX15Reg.2ReversePrimer:5'CCAAAAATCGTCGCTAAAAATCAAC3(SEQIDNO334) TBX15Reg.2Flapoligo:5'CCACGGACGCGCGCATTCACT/3061(SEQIDNO335) I :332)SEQIDNOAGGAAATTGCGGGTTTTCG3'(Primer5Forward:TBX15Reg.2 (SEQIDNO:403)
404)IDNO:(SEQTargetDNAconvertedBisulfite- TITIE I
TBX15TargetDNARegion2: GACTTTCAGCGACGACTCCTGGAAGCAACGCCA3' I GATTTTTAGCGACGATTTTTGGAAGTAACGTTA3' PCRandFlapAssayOligonucleotides: 5'GGAAGGAAATTGCGGGTTCCCGTCTGCCTTGTCTCCAGCTTCTCTGCTGAAGCCCGGTAGCAGTGAATGCGCGCT 5'GGAAGGAAATTGCGGGTTTTCGTTTGTTTTGTTTTTAGTTTTTTTGTTGAAGTTCGGTAGTAGTGAATGCGCGTT 1 Patent Application Publication Nov . 23 , 2017 Sheet 66 of 155 US 2017 /0335401 A1
MATITITA'
5'TTTCGTTGATTTTATTCGAGTCG3(SEQIDNO:338) 777 5'TCTTCAAATAAACCGCCGC3(SEQIDNO:339)
TIT 5'CGCCGAGGGTCGTTTGGCGT/3C6(SEQIDNO:340)
FIG.1(cont'd) Bisulfite-convertedTargetDNAwithprimerandFlapoligo.sites TITETITIITIATI TRHTargetDNA:(SEQIDNO336) GAGGGCTCGGCAGGCGCCCGGGGTC3' GAGGGTTCGGTAGGCGTTCGGGGTT3' PCRandFlapAssayoligonucleotides: TRHForwardPrimer: TRHReversePrimer: TRHFlapoligo.: 5'GGCGGCCGCGACCCCTCCCCGCTGACCTCACTCGAGCCGCCGCCTGGCGCAGATATAAGCGGCGGCCCATCTGAA (SEQIDNO:337) 5'GGCGGTCGCGATTTTTTTTCGTTGATTTTATTCGAGTCGTCGTTTGGCGTAGATATAAGCGGCGGTTTATTTGAA Patent Application Publication Nov . 23, 2017 Sheet 67 of 155 US 2017 /0335401 A1
7 min MT III A 5'GTTTGGGAGAGTTTTATCGTCG3(SEQIDNO:343) 5'CGCCGAGGGCGGTCGTTGGA/306;(SEQIDNO:345) 5'CCTCCAAAAATCCGCCGA3(SEQIDNO:344)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO342) TSC22D4TargetDNA:(SEQIDNO341) CGAGACCTGGAGCCCCACAGCTTCGGCGGACTCCTGGAGGGAA3' CGAGATTTGGAGTTTTATAGTTTCGGCGGATTTTTGGAGGGAA3' 5'CGGGTGGTGAAGCTGCCCCACGGCCTGGGAGAGCCTTATCGCCGCGGTCGCTGGACGTGTGTGGATGTTTATGAG 5'CGGGTGGTGAAGTTGTTTTACGGTTTGGGAGAGTTTTATCGTCGCGGTCGTTGGACGTGTGTGGATGTTTATGAG PCRandFlapAssayOligonucleotides: TSC22D4ForwardPrimer: TSC22D4ReversePrimer: TSC22D4Flapoligo.i Patent Application Publication Nov . 23 , 2017 Sheet 68 of 155 US 2017 /0335401 A1
5'CGCCGAGGGACGAACGCACG/3C6(SEQIDNO:350) 5'GTCGGGGTCGATAGTTTACG3(SEQIDNO:348) 5'ACTCGAACTCACGAAAACG3(SEQIDNO:349)
FIG.1(cont'd) ZDHHC1-zincfinger,DHHCtypecontaining1TargetDNA:(SEQIDNO346) 5'GGGGCGGGGGCCGACAGCCCACGCTGGCGCGGCAGGCGCGTGCGCCCGCCGTTTTCGTGAGCCCGAGCAG3 Bisulfite-convertedTargetDNA:(SEQIDNO347) 5'GGGGTCGGGGTCGATAGTTTACGTTGGCGCGGTAGGCGCGTGCGTTCGTCGTTTTCGTGAGTTCGAGTAG3 PCRandFlapAssayoligonucleotides: ZDHHC1ReversePrimerv3: ZDHHC1ForwardPrimer: ZDHHC1Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 69 of 155 US 2017 /0335401 A1
5'CGCCGAGGGTTCGTAGGGTT/3061(SEQIDNO:355) 5'GTAGGGTGGGTGGTTACG3'(SEQIDNO:353) 5'AACTTCCCACGACCCGC3(SEQIDNO:354)
FIG.1(cont'd)
Bisulfite-convertedTargetDNA:(SEQIDNO352)
ZMIZ1TargetDNA:(SEQIDNO351) PCRandFlapAssayoligonucleotides: GCGGGCCGTGGGAAGTTTCTC3' GCGGGTCGTGGGAAGTTTTTT3' ZMIZ1ForwardPrimer: ZMIZ1ReversePrimer: ZMIZ1Flapoligo.i 5'GGAGCCCCCAGCCCCACGCGGGCACACGCAGGGTGGGTGGTCACGCCCGCAGGGTCCGCGAGCGCGGCGCAGAGC 5'GGAGTTTTTAGTTTTACGCGGGTATACGTAGGGTGGGTGGTTACGTTCGTAGGGTTCGCGAGCGCGGCGTAGAGC 1 Patent Application Publication Nov . 23 , 2017 Sheet 70 of 155 US 2017 /0335401 A1
I
?rrrrr
ZNF132ReversePrimer:5'ACACTATAATACGTATAAAAACGCGATA3(SEQIDNO359) FIG.1(cont'd) ZNF132ForwardPrimerv2:5'TGGAGTTATATTAGGGTACGGGA3(SEQIDNO358) ZNF1325'CCACGGACGAACGATCCCTAC/3061(SEQIDNO:360)Flapoligo.:
my II Bisulfite-convertedTargetDNA:(SEQIDNO357) CGGGAGTGGGTGCAGGGACCGTCACCGCGCCTTCACACGCACCATAGTGCCC3' IMITA CGGGAGTGGGTGTAGGGATCGTTATCGCGTTTTTATACGTATTATAGTGTTT3' ZNF132TargetDNA:(SEQIDNO356) MIT PCRandFlapAssayoligonucleotides: 5.GGCGCCGCCATTGCGGTCCTCATTTTGCTGCTGGTGGGTTGGGCTACAGCAGGCCTCTGGAGCCACACCAGGGCA 5'GGCGTCGTTATTGCGGTTTTTATTTTGTTGTTGGTGGGTTGGGTTATAGTAGGTTTTTGGAGTTATATTAGGGTA ??? ) ?? ) ? Patent Application Publication Nov . 23 , 2017 Sheet 71 of 155 US 2017 /0335401 A1
5'CCACGGACGGGTTCGTTAGGG/306)(SEQIDNO:365 5'GGTGGTGGGTATGTCGG3(SEQIDNO:363) 5'CCAATCGCCACGAAACG3(SEQIDNO:364)
FIG.1(cont'd)
GCCAGGGGCGTAGTTCTCCTGGTGGGTGGGGACGCTCCGTGGCGATTGGGGTCACTCCTCTGAGG3' Bisulfite-convertedTargetDNA:(SEQIDNO362) GTTAGGGGCGTAGTTTTTTTGGTGGGTGGGGACGTTTCGTGGCGATTGGGGTTATTTTTTTGAGG3' ZNF329TargetDNA:(SEQIDNO361) PCRandFlapAssayoligonucleotides: 5'GGCGGCGAGGGGCGCGTCCGCGGGTGGGTTTCACCTGGGTGGTGGGCATGTCGGGCCCGCTAGGGCGAGGGTCTG 5'GGCGGCGAGGGGCGCGTTCGCGGGTGGGTTTTATTTGGGTGGTGGGTATGTCGGGTTCGTTAGGGCGAGGGTTTG ZNF329ForwardPrimer: ZNF329ReversePrimer: ZNF329Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 72 of 155 US 2017 /0335401 A1
I.
YTTYM.I III. TnTM 5'CCAATATCCCGAAACGCGTCT3(SEQIDNO:369) 5'CCACGGACGGCGTTTCGATCG/3C61(SEQIDNO:370) 5'GTTGTCGGGAGCGGTAGG3(SEQIDNO:368)
FIG.1(cont'd) JULI1 mommy NYTT '! I Bisulfite-convertedTargetDNA:(SEQIDNO367) RE TI VILLA ZNF671TargetDNA:(SEQIDNO366) TCAGACGCGTCTCGGGACACTGGGGACAACATCICCTCCGCG3' TTAGACGCGTTTCGGGATATTGGGGATAATATTTTTTTCGCG3' PCRandFlapAssayoligonucleotides: ZNF671ForwardPrimer: ZNF671ReversePrimer: ZNF671Flapoligo. 5'CCGTGGGCGCGGACAGCTGCCGGGAGCGGCAGGCGTCTCGATCGGGGACGCAGGCACTTCCGTCCCTGCAGAGCA 5.TCGTGGGCGCGGATAGTTGTCGGGAGCGGTAGGCGTTTCGATCGGGGACGTAGGTATTTTCGTTTTTGTAGAGTAGGGG M Patent Application Publication Nov . 23 , 2017 Sheet 73 of 155 US 2017 /0335401 A1
117I OTTITIT VIII 177TIeen TIIT7
In ? IIIII. Tin71
5'CGTTTTTTTGTTTTTCGAGTGCG3'(SEQIDNO:373) 5'TCAATAACTAAACTCACCGCGTC3(SEQIDNO:374) 5'CCACGGACGGCGGATTTATCG/3C6(SEQIDNO:375) ninHII
FIG.1(cont'd) ZNF781:Chr1938183137-38183018(GRCh37/hg19)TargetDNASEQIDNO371
I I 793ITI TIT TTATGACGCGGTGAGTCCAGCCACTGATTCCTAACGGTTTAGAGT3'. Bisulfite-convertedTargetDNA:(SEQIDNO372) TTATGACGCGGTGAGTTTAGTTATTGATTTTTAACGGTTTAGAGT3' PCRandFlapAssayOligonucleotides: 5'AAGCTGCGCCCGGAGACGTGGGAGCGTTCTCTTGTTTTCCGAGTGCGCGGACTCATCGGGTCACAGTTTATGCTT 5'AAGTTGCGTTCGGAGACGTGGGAGCGTTTTTTTGTTTTTCGAGTGCGCGGATTTATCGGGTTATAGTTTATGTTT ZNF781ForwardPrimer: ZNF781ReversePrimer: ZNF781Flapoligo.: Patent Application Publication Nov . 23 , 2017 Sheet 74 of 155 US 2017 /0335401 A1
64.67 64.51 12.88 195.13 35.41 95.52 36.73 10.87 65.19 172.75 270.65 45.26 5.83 97.00 29.84 15.74 15.35 29.35 31.95 22.59 22.96 BC / UND
2.58 4.44 3.55 BC / SQ 28.13 22.18 27.10 11.70 62.77 10.27 68.54 91.45 18.61 10.45 26.04 28.54 13.54 30.2112354 20.181771 19.25 49.28 68.55 .80 26.82 22.47 28.73 43.76 31.02 24.44 28.68 3.80 82.74 170.64 9.23 5.68
BC/ SC 5 33.329437371507 42.51309545311574 26.10388954763096 BC/ LC 68.62 36.73 15.89 71.50 42.10 132.84 108.02 31.27 84.15 14.00 57.98 35.38 14.15 42.49 25.41 16.71 27.463421 27.625051 60.90 24.05 20.38 59.66 44.26 91.85 87.76 60.14 29.73 26.57 40.31 40.71 36.37 61.89 29.77 24. AdBC
1.31 3.25 1.55 4.69 3.10 1.51 1.59 2.42 4.89 1.74 4.72 5.40 3.56 1.55 2.04 4.05 2.07 4.29 2.69 3.15 1.79 BC / Norm
island .Lung cancer 32% 59%%8 8% 56% 49% 35% 19% 13% 26% 40% 44% 14% 2% 39% 16% 5% 9% 14% 5% 23% 13% undefined mean
island. Lung %8 28% 9% 7% 31% 10% 1% 14% 25% %1 2% 21% 1% 37% 13% 6% 12% 5% 4% 29% 7% Squamous mean
sland .Lung 22% 50% 5% 5% 68% 25% 3% 32% 9% 7% 7% 10% %7 16% 7% 2% 40% 28% 9% 3% FIG.2 cell Small mean 19%| island. Lung 15% 36% 6% 20% 47% 18% 19% 29% 10% 21% 34% 9% 34% 7% 20% 22% 7% 7% 26% 9% cell Large mean
island. Lung 21% 24% 21% 19% 38% 10% 12% 24% 24% 10% 15% 26% 16% 12% 14% 14% 25% 18% 10% 30% 13% Adenocarcinoma mean
island %1 3% 1% 4% 1% 1% 3% 2% 0% 1% 2% 1% 1% 1% 1% 1% 2% 0% 3% 1% .normal lug. mean
0% 1% 1% 0% 1% 0% 1% 1% 0% 0% %0 0% 0% 0% 1% 0% 1% 0% 0% %1 1% island . BC mean 25056162| position Stop 3785923 74726617 52401020 27135679 27135823 830382 97193639 65731507 968584 27467467 88137628 7314772073147790 3743553037435681 156863554 145106439 145106827 124860665 101033620 145105675 position Start 8100292681002992 3785837 74726554 25056015 52400959 156863477 145106353 27135634 27135772 145106.742 830323 124860573 97193509 65731423 968477 101033514 27467359 145105570 88137543 Chromosome 12 Om- 1 17 # . ref region marker noghot Adenocarcinoma AC1 AC2 AC3 AC4 AC5 AC6 ACT AC8 AC9 AC10 AC11 AC12 AC13 AC14 AC15 AC16 AC17 AC18 AC19 AC20 AC21 Patent Application Publication Nov . 23 , 2017 Sheet 75 of 155 US 2017 /0335401 A1
title Gene phosphoinositides1)-associated (potassiumchloridetransporters), associatedhomolog(rat) zincfinger,MIZ-typecontaining1 homeobox1spiraclesempty |proteinphosphatase1,regulatory megakaryocyte-associatedtyrosine branchedchainamino-acid transaminase1,cytosolic GRP1(generalreceptorfor 5-oxoprolinase(ATPhydrolysing) 5-oxoprolinase(ATPhydrolysing) CD151molecule(Raphblood Dnaj(Hsp40)homolog,subfamily brain-enrichedguanylatekinase. (inhibitor)subunit16B ladybirdhomeobox2 plateletendothelialaggregation solutecarrierfamily12 proteinpha scaffaldprotein receptor1 homeoboxA1 homeoboxA1 member8 C,member6 myosinXVIIIA kinase group) agrin
ID Entrez 2016 26051 4145 85474 586 26873 3198 3198 26873 977 84561 9829 57596 399687 - - 571781 1126051 160622 375033 9829Dnali| 375790
1 Island CpG In 11 O O 0
Distance Tss +1-26282620; 01183; 10-147 0893;2617 1040048:40048 -45 2628 0174135 |03117 1212 06967071036;-| O994 @12975 Exon In -03889 0 -08842 |-19231 10?? ? ? ? + + + + + II FIG.2(cont'd) Strand + + : : |- 01178094;NM001178091001|693:46378;-6 NM_002378;1393551393-015973578: NM001178092;_005504NMO-04637846378
Transcript NM004357;_1390291390 NM_001172735;015568 INM001195483_;NM024628 NM001159531:_020836 NM005522;_153620 NM_005522;153620 NM078471203318 NM020338 NM004097 NM_181711 NM_001080471 NM017570 NM017570 30;NM001039490 014787NM NM198576 54 NM001009812 178093
Gene MAX.chr297193509-97193639 MAX.chr8,145105570-145105675 MAX.chr988137543-88137628 hoede581975 ZMIZ1 EMX1 PPP1R16B MATK GRASP PEAR1 OPLAH HOXA1 HOXA1 OPLAH CD151 SLC12A8 ONAJC6 AGRN BEGAIN MYO18A Chromosome 10 2 20 2AC5LBX2 AC612BCAT1 12 1 8 7 8 1 1 14 17 # .ref region marker AC10 14AC heten Adenocarcinoma ACi AC2 AC3 AC4 AC7 ACB AC9 AC117 AC12 AC13 AC15 AC16 AC17 AC18 AC19 AC20 AC219 Patent Application Publication Nov . 23 , 2017 Sheet 76 of 155 US 2017 /0335401 A1
1.28 8.10 56.57| BC/ UND 50.29 59.05 67.24 144.38 105.40 7.42 24.93 14.59 19.04 209.78 34.94 40.51 41.48 18,37 63.96 12.623440 38.31101 BC/ SQ 22.88 81.37 20.22 12.26 37.48 30.71 11.44 5.88 9.22 10.13 14.20 13.87 64.67 24.73 1.96 6.89 19.39
12.58| 116.74| BC/ SC 5.10 44.00 232.04 55.00 5.77 45.37 13.18 148.49 66. 34.88 2.55 77.49 4.971 5.84543693771200194213515 #DIV/0!OIVIO 25.58293,90123797119965410229801 76.85 36.5322371750 42.2025795543 27.00| 39.95 39.16 56.94 30,28 32.21 24.19136616503 BC/ LC 11.75 63.35 361.21800144431 119.15 11.93 25.97 189.49 26.41 41.95 84.98899440660
BC /Ad 29.45 176.53 45.20 110.62 27.79 22.36 22.43 31.36 119.07 45,55 75.23 25.92 47.29 25.89 56.31
1.86 6.00 1.37 2.71 1.71 5.69 10.25 2.75 2.26 2.78 2.00 12.77 3.87 4,80 2.72 1.60 2.31 5.21 2.51 6.77 BC/ Norm
island .Lung cancer 0% %4 16% 4% 21% 10% 12% 4% 19% T14% 17% 26% 16% 18% 19% 15% 34% 30% 13% 7% 17% 27% 16% undefined mean
1% 1% island. Lung 8% 7% 6% 7% 12% 2% 3% 7% 10% 12% 2% 15% 21% 2% 2% 5% 18% 3% 5% 10% 6% Squamous mean
jsland. Lung 2% 4%4% 64% 9% 18% 32% 6% 7% 4% 44% 11% 18% 54% 21% 6% 10% 3% 26% 1% 31% 1% 52% 62% FIG.2(cont'd) cell Small mean island .Lung 15% 6% 17% 12% 8% 6% 14% 6% 20% 25% 35% 23% 18% 18% 11% 12% 24% 22% 10% 11% 11% 29% 14% cel Large mean
island . Lung 11% 15% 12% 20% 13% 26% 13% 15% 17% 22% 27% 15% 21% 24% 10% 15% 14% 19% 14% 10% 15% 19% 13% Adenocarcinoma mean island 1% 1%14 0% 1% 0% 1% 2% 3% 2% 2% 2% 2% 1% 1% 1% 2% 0% 1% 1% 2% 1% .normal . lung mean
%0 0%0% 0% 1% 0% 0% 0% 1% 1% 1% 1% 0% 0% 0% 0% 0% 0% 1% 0% 0% 0% 1% 0% island . BC mean position Stop 36909447 96991212 2166058 26699117 58662026 50297912 32237880 65731666 23847925 23848020 3786032 37288615 103046982 25439264 61638536 32237998 50378540 5866175758661861 26699039) 2384758623847684 4403159944031658 25439185| 61638472) position Start 36909350 96991058 2165937 9719316697193253 1145013661145013775 58661880 50297879 32237695 65731622 100075307100075425 23847871 23847938 3785977 37288523 103046898 32237893 50378496 Chromosome 2 17 16 13 C # .ref region marker Adenocarcinoma AC22 AC23 AC24 AC25 AC26 AG27 AC28 AC29 AC30 AC31 AC32 AC33 AC34 AC35 AC36 AC37 AC38 AC39 AC40 AC412 AC42 AC43 AC44 Patent Application Publication Nov . 23 , 2017 Sheet 77 of 155 US 2017 /0335401 A1
Rasassociation(RalGOS/AF-6 title Gene interactingprotein-like1 Fasapoptoticinhibitorymolecule2 TSC22domainfamily,mernber4 megakaryocyte-associatedtyrosine inositol1,45-triphosphatereceptor sterilealphaandTIRmotif Onaj(Hsp40)homolog,subfamily proteinkinaseC,beta proteintyrosinephosphatase, proteinkinaseC,beta proteinkinaseC,beta fibroblastgrowthfactor14 basichelix-loopfamily, domainfamilymember1 Jacyl-CoAthioesterase4 v-skisarcomaviraloncogene homolog(avian) zincfingerprotein329 containing1 zincfingerprotein329 receptortype,F pseudogene plectin C,member6 kinase membere23 10 Entrez 644189 150771 6497 79673 5339 23098 79673 23017 9829 5579 81628 5792 557955795579 5579 4145 22592259 128408 11186 Island CpG In 1 11
Distance Tss 36,11383;974449144 15244;303137252340 01-129:28323601 0-876;886310 15833;438: 1287; 035053,35053 572; 0639, 0-3088 0391 153 0268 0-159 01193 11595 05804 07226 -085 O O O Exon In O- -107226 -O + - FIG.2(cont'd) Strand + + + - - - + - - - + + - + + + -
Transcript NM_178495;0011635240 45;NM_20137820138020 NM_002378;1393551393 NM170714;007182:1707 NM201381;2013830004 201379201382NM_1384; 01008949;NM001163523 NM002738212535 |NM_002840;130440 NM002738;212535 212535002738;NM NM_170714;NM_007 NR033748 NM_003036 024620NM_ NM_015077 NM_012306 NM030935 NM175929 12:NM170713 NM024620 NM014787 54 NM080606
Gene MAX.chr297193166-97193253 MAX.chr132237695-32237880 MAX.chr1937288523-37288615 MAX.chr225439185-25439264 MAXCET1.32237893-32297938MAX.chr132237893-32237998
LOC644189 ITPRIPL7 SKI IZNF329 PLEC SARM1 ZNF329 FAM2 ONAJC6 PRKCB TSC22D4 PTPRE PRKCB PRKCB MARK FGF14 BHLHE23 TV RASSF1 Chromosome 2 1 19 V . 19 12 76 7 1 16 19 19 13 2 20 1
# . ref region marker Adenocarcinoma AC22 AC23 AC24 AC25 AC26 AG27 AC28 AC30 AC31 AC32 AC33 AC34 AC35 AC36 AC37 AC38 AC39 AC40 AC41 AC42 AC43 AC44 Patent Application Publication Nov . 23 , 2017 Sheet 78 of 155 US 2017 /0335401 A1
14.12| BC/ UND 57.44 49.816901 11.96687 114.941041 1.90 SQIBC 23.63
BC/ SC 8.77 78.85 35.67 22.13 0.57
55.94| BC/ LC 40.63 17.97 98.18 8.72
BC/ Ad 31.01 39.22 29.13 51.82 21.31
1.44 1.79 3.211 5.61 0.77 BC / Norm
island . Lung cancer 26% 27% 6% 2% 11% undefined mean island .Lung 1% 19% 10% 25% 19% Squamous mean
island .Lung 4% 30% 29% 5% 0% FIG.2(cont'd) cell Small mean island .Lung 18% 22% 15% 21% 7% cell Large mean
island .Lung 14% 15% 24% 11% 17% Adenocarcinoma mean
island 1% 1% 3% %1 701% .normal . lung mean
0% 0% 1% 0% %1 island . BC mean position Stop 99439192 37288510 96715360 65116440 17627534 position Start 99439151 37288426 96715209 65116396 17627469 |
Chromosome 8 6 15 17 # . ref region marker Adenocarcinoma AC45 AC46 AC47 AC48 AC49 Patent Application Publication Nov . 23, 2017 Sheet 79 of 155 US 2017 /0335401 A1
potassiumvoltage-gatedchannel, title Gene delayed-rectifier,subfamilyS BARXhomeobox1 PIF15-03'ONAhelicasehomolog Tretinoicacidinduced1 member2 (S.cerevisiae)
ID Entrez 3788 56033 80119 10743
Island CpG In 1
Distance Tss
14421 +10-98 142683 Exon In 10- -12399 FIG.2(cont'd) Strand +
Transcript
NM020697 NM021570 NM_025049 NM030665
Gene MAX.chr1937288426-37288510 KCNS2 BARX1 PIFT RAI1
Chromosome B 19 9 15 17
# .ref region marker Adenocarcinoma AC45 AC46 AC47 AC48 AC49 Patent Application Publication Nov . 23 , 2017 Sheet 80 of 155 US 2017 /0335401 A1
209.78 362.13 390.901 159.79 254.67 152.65 1.25 72.91 105.40 96.24 58.29 45.26 37.60 80.58 68.97 206,97 135.15 72.85 UNDBC
#DIV/0! 55.3133916 156.4333425 9.2620341 41.0229801 132.8824758 13.87 198.95 203.93 6.18 158.96 39.55 35.25 15.20 11.44 17.90 37.61 68.54 2.231 48.17 70.91 19.42 18.74 BC/ SQ 117.62
277.57| 288.47 72.25 199.65 0.58 148.49 325,38| 33.00 245.53 259.29 71.80 43.30 55.00 46.66 35.52 31.02 17.41 50.15 266.28 167.27 12.00 84.05 BC/ SC #DIV/0! 1054,27
110.72 109.94 108.02 107.93 99.18 99.05 95.09 93.77 92.41 BC/ LC 29.5315586| 82.1314162| 119.07)18949 77.5018542 94.6317566 23.9213794 17.8712928 36.9912111 110.62|11915 29.141796225275| 29.902554729559 5.2136,3523874 25.582939023797 17.37847320232 64,06 43.34 81.76 17.71 32.43 18.97 71.79 54.36 78.53 AdIBC #DIV/0!|
BC/ Norm 12.77 24.55 8.091 2.85 18.40 3.49 3.89 6.92 10.25 8.85 4.38 5.401 1.28 8.25 8.70 1.27 5.84 764
island. Lung cancer 19% %13 50% 16% 15% 26% 26%CUISINE 55% 33% 12% 62% %31 0%SUR 9% 12% 9% 11% 14% 5% 19% 20% 32% 34% 8% undefed ean
island .Lung 2% 2% 23% 1% 2% 14% 2% 30% 17% 0% 39% 8% 6% 2% 1% 7% 21% 0% 11% 21% 18% 5% 2% Squamous mean
island .Lung 6% 40% 43% 6% 10% 0% 18% 50% 24% 2% 60% 52% 12% 5% 6% 4% 6% 10% 2% 12% 78% 26% 3% 10% cell Small mean FIG.3 island. Lung 11% 11% 38% 19% 12% 21% 23% 28% 15% 11% 35% 28% 22% 15% 14% 10% 20% 34% 15% 23% 29% 15% 24% 11% cell Large mean
island -Lung 10% 10% 27% 15% 9% 15% 12% 8% 2% 20% 5% 3% 5% 13% 6% 8% 26% 2% 8% 11% 14% 9% Adenocarcinoma mean
island 1% 1% 4% 0% 1% 2% 2% 4% 1% 0% 5% 1% 1% 1% 1% 1% 2% 0% 3% 0% 1% 1% .normal . lung mean
0% 0% 0% 0% 0% 0% 0% 0% 0% 0% 0% %0 0% 0% 0% 0% 0% 0% 0% %0 0% 0% 0%0% 0% island . BC mean position Stop 23847925 169064446 42992393 37488105 23848020 17346549 23847684 42995393 58238942 37488163 42995528 110627325 58011488 49484184 26699117 47443961 46675237 145106827 75447708 102711959 14926955 42287988 3786032 182322403
. position Start 23847871 169064343 42992328 37488072 23847938 17346461 23847586 42995328 58238816 37488118 42995477 110627264 58011421 49484143 26699039 47443870 46675164 145106742 75447560 102711879 14926886 42287927 3785977 182322274
|
Chromosome 5 1 20 8 17 11 17 2 16LCLCE19 LC919 # .ref region |LC416 LC1319 marker cell Large LC2 LC3 LC4 LCO LC716 LC85 LC10 |LC11 LC12 LC1412 LC15 LC16 LC17 LC18 LC19 LC20 LC21 LC22 LC23 LC24 Patent Application Publication Nov . 23 , 2017 Sheet 81 of 155 US 2017 /0335401 A1
nuclearreceptorsubfamily2,group phosphatidylinositol-3,45 title Gene engulfmentandcellmotility1 lengulfmentandcellmotility1 trisphosphate-dependentRac 5-oxoprolinase(ATPhydrolysing) megakaryocyte-associatedtyrosine integrin,alpha4(antigenCD490. alpha4subunitofVLA-receptor) proteinkinaseC,beta dedicatorofcytokinesis2 proteinkinaseC,beta proteinkinaseC,beta zincfingerprotein671 zincfingerprotein773 sterilealphaandTIRmotif exchangefactor1 B-cellscaffoldproteinwithankyrin upstreambindingtranscription factor,RNApolymerase| containing1 homeoboxB5 repeats1 F,member6 deserthedgehog septin9 kinase ID Entrez 5579 1794 9844 5579 2063 5579 79891 9844 374928 50846 23098 57580 3215 26873 10801 55024 7343 4145 3676 Island CpG In 1 -1 -1 - - - 1 -- 114145
Distance Tss O8997;103237737 015833;438 +01572:572 0639: 9690 1287; 8842 0439 1179 0393 0- 1113 114459 5500 11-4061 1656 Exon in +193 O 1531]+ O O - |-O - - - ?? - - - ? ? FIG.3(cont'd) + - - Strand 1 + 1 + 1 + | | | | | | + 1 + ] ' 1 + * NM_006640;001113494O+01319647539694816351 01113496;NM_001113492001|88:78289170069 001127507T+1[116;017935NM_|
Transcript NM_001076684;001076683N NM_002378;1393551393 NM002738;212535 NM002738;212535 NM_002738;212535 (113493;NM001113491 NM004946 NM014800 NM005234 NM024833 NM014800 NM198542 NM021044 NM_015077 NM020820 NM002147 NM_017570 M014233 NM_000885 - 54
Gene LC115MAX.chr542995477-42995528 LC121MAX.chr1110627264-110627325 |11MAX.chr1114926886-14926955 LC35MAX.chr542992328-42992393 LC85MAX.chr542995328-42995393 BANK1 ITGA4 17ELMO 19ZNF773 12DHH 17SARMI PREX120 17HOXB5 8OPLAM 17se 17UBTF 19MATK Chromosome LC116PRKCB LC25DOCK2 |16PRKCBLC5 LC619NR2F6 LC716PRKCB LCO19ZNF671 LC107ELMO1 F Farm # .ref region marker cell Large LC4 LC13 LC14 LG15 LC16 LC17 LC18 LC19 LC20 LC21 LC22 LC23 LC24 Patent Application Publication Nov . 23 , 2017 Sheet 82 of 155 US 2017 /0335401 A1
70.13 78.56 97.00 70.81 140.87 117.27 65.19 59.41 11.52 195.13 99.50 77.02 120.57 15.74 123.00 UNDIBC 38.31101 91.526427 78.491666378.4916663 1.976409 54,5099.42 52.166990 59.052022 BC/ SQ 7.67 34.47 91.45 6.84 44.17 55.43 62.77 25.10 19.33 26.13 57.13 9.07 14.64 17.71 12.03
24.55| 19.25 53.16 276,90276.90 37.84 13,37 28.60 103.02 125.02 22.47 43.06 30.54 32.54 93.83 14.174 232.04 158.81 142.53 32.31 20.18 28.83 BC/ SC
84.98899440660 84.15 BC/ LC 89.03 86.8986.89 85.84 84.33 82.00 77.51 74.49 71.50 70.90 69.46 68.62 66.89 64.23 63.36 63.35 57.98 57,90 26.296320 62.474184 15.226067 45.71 63.7963.79 36.03 25.74 29.73 22.77 36.00 38.31 59.66 22.12 17.04 66.901 17.32 33.74 36.56 45.20 40.31 34.83 Ad?BC
BC/ Norm 6.77 5.53 7.507.50 2.47 5.69 1.55 2.22 4.32 6.97 4.89 2.90 3.23 4.691 3.09 7.31 5.71 1.371 2.13 3.69 2:36 4.05
island .Lung cancer 76% 13% 29%29% 10% 14% 39% 13% 40% 41% 26% 29% 3% 56% 11% 61% 24% 16% 28% 14% 25% 5% 23% undefined mean
island .Lung 6% 2% 14%14% 6% 37% 1% 12% 19% 25% 12% 4% 7% 0% 34% 18% 6% 16% 2% 3% 6% 29 Squamous mean islandpue?s! . Lung Bunge 62% 11% SK49% 6% 2% 12% 4% 29% 43% 9% 21% 7% 5% 6% 58% 49% 64% 45% 26% 7% 5% FIG.3(cont'd) cell Small mean island. Lung 14% 18% 15% 13% 15% 34% 15% 22% 26% 29% 34% 16% 20% 12% 40% 22% 17% 18% 12% 13% 20% 11% cell Large mean
island. Lung p 13% 9% %11 5% 5% 12% 4% 10% 13% 24% 11% 4% 19% 3% 21% 13% 12% 7% 8% 3% 14% 7% Adenocarcinomapuepsibun ???? e
island 1% 1% 0% %1 0% 1% 2% 2% 1% 1% 1% 1% 5% 2% 0% 1% 1% %0 1% .normal .fung mean
0% 0% 0% 0% 0% 0% 0% 0% %0 0% 0% 0% 0% 1% 0% 0% 0% 0% 0% 0% 0% island. BC mean
position Stop 50378540 157098381 124173386 42153601 17346450 124860665 37488303 25055997 194118919 145106439 46620622 173099817 3785923 37487893 6004338 22541996 2166058 223936997 1295314 128564773 65731507 142494904
position Start 50378496 157098339 124173236 42153564 17346401 124860573 37488225 25055873 194118747 145106353 46620564 173099757 3785837 37487776 6004298 22541891 2165937 223936868 1295194 128564667 65731423 142494769
Chromosome NO 19 7 10 LC3212 | # .ref region LC3117 marker cell Large LC25 LC26 LC27 LC28 LC29 LC30 LC33 LC34 LC35 LC36 LC37LC37 LC38 LC39 |LC40 LC41 L042 LC43 LC44 LC45 LC46 Patent Application Publication Nov . 23 , 2017 Sheet 83 of 155 US 2017 /0335401 A1
Inuclearreceptorsubfamily2,group title Gene 11186Rasassociation(RalGDS/AF-6) (potassium/chloridetransporters), engulfmentandcellmotility1 branchedchainamino-acid glycoproteinV(platelet) 5-oxoprolinase(ATPhydrolysing) megakaryocyte-associatedtyrosine Jengulfmentandcellmotility1 domainfamilymember1 100190949chromosome5openreadingframe BENdomaincontaining4 transaminase1,cytosolic calpain2,(m/II)largesubunit |Friendleukemiavirusintegration1 Onaj(Hsp40)homolog,subfamily solutecarrierfamily12 JV-skisarcomaviraloncogene homolog(avian) telomerasereversetranscriptase F,member6 member8 homeoboxB2 Ineuritin1 C,member6 52 kinase ID Entrez 389206 2063 84561 9844 586 2814 26873 3212 4145 9844 51299 6497 824 7015 2313 98299829
Island CpG In 1 - - - 1 1 T17015
Distance Tss 0-129:;28323601 046520; 551;46520136 015973;578 01331; 1047574;36750 0-221 11829 1735 -032; +0855;2279 994 97501 1.069670;71036 0286 |11248 19231 05804 Exon in 0 Holo -0735bololo -03335 toto O FIG.3(cont'd) - - - - . - - - + Strand + +
01178094;NM_001178091001 Transcript NM_170714;0071821707 NM_001178092;0055040 NM_002378;1393551393 NM_001195483;024628 |NM_001159547;207406 NM001146068;001748 NM001193376;198253 NM_002017;001167681 12:NM170713NM_001145132 NM_005234 NM_014800 NM004488 NM017570 NM002145 NM014800 NM016588 NM_003036 NM_014787 178093 54
Gene MAX.chr8124173236-124173386 MAX.chr7142494769-142494904 2MAX.chr2173099757-173099817LC36 10MAX.chr1022541891-22541996 C5orf52 SLC12AB SKI DNAJC6 3RASSF1 4BENDA 19NR2F6 19MATK LC4411F2!1 5 8 3 |7ELMO1 |6NRN1 Chromosome ELMO1LC317 LC3212BCAT1 LC333GP5 LC348OPLAH LC3517HOXB2 T1CAPNZLC42 |5TERTLC43 # . ref region marker cell Large LC25 LC26 LC27 LC28 LC29 LC30 LC37 LC39 |LC40 LC41 LC45 LC46 Patent Application Publication Nov . 23 , 2017 Sheet 84 of 155 US 2017 /0335401 A1
2.04 33.86| BC /UND 46.87 56.57 143.28 41.94 61.88 83.04 51.80 95.52 46.42 34.97 93.33 35.85 27.46 71,09 17.15 83.14 47.14
32.84 23.82 64.67 258.74790811584 57.49 49.84 12.07 27.48 10.61 84.00 27.10 26.78 5.33 38.92 5.10 4.51 5.35 7.65 28.60 2.82 BC/ SQ
1.19 100.17 66. 15139) 13.90 18.35 61.39 9.59 20.78 68.55 59,12 4.01 43.26 17.50 10.38 93.41 24.63 51.11 17.94 BC/ SC
50.24) BC/ LC 57.68 57. 56.94 56.55 54.78 54.77 54.63 51.32 50.51 49.38 47.57 47. 46.30 46.14 45.94 44.87 44.54 17.135363 22.915255 15.83 53.16 75.23 24.19 39.97 26.23 10.87 23.37 27.62 25.97 12.28 15.10 6.49 13.39 29.12 22.67| 23.18 51.11 BC/ Ad
3.04) 1.99| 2.20 7.651 4.80 5.65 5.87 3.80 5.10 2.24 1.98 1.51 3.67 3.60 2.121 2.24 3.71 3.02 5.65 2.26 BC/ Norm
island .Lung cancer 0% 28% 18% 28% 61% 10% 21% 19% 28% 22% 35% 44% 17% 40% %8 9% 22% 6% 50% 42% undefined mean
island . Lung 6% 14% 21% 19% 25% 12% 7% 8% 4% 35% 10% 25% 3% 17% 1% 1% 2% 3% 17% 3% Squamous mean
island .Lung 0% 60% 21% 62% 65% 3% 11% 19% 3% 25% 56% 2% 19% 4% 3% 30% 9% 30% 16% FIG.3(cont'd) cell Small mean island . Lung 11% 35% 18% 13% 23% 13%13% 34% 16% 18% 21% 18% 47% 24% 20% 11% 14% 15% 17% 27% 40% cellpuesrbung Large mean
island . Lung 3% 32% 24% 6% 17% 6% 7% 5% 0% 10% 10% 24% 6% 6% 1% 4% 970 8% 14% 46% Adenocarcinoma mean FER
2% 1% 1% 1% island 0% 5% 2% 1% 3% 1% 2% 1% 1% 1% 3% 2% 0% 1% 3% 2% .normal . lung mean TEEK
0% 1% 0% 0% 0% 0% 1% 0% 0% 0% 1% 0% 0% 0% 0% 0% 0% 1% 1% island . BC mean
position Stop 161275801 32237654 44031658 61560749 122296828 145104342 140201277 15780191 111217682 124860798 25056162 48042606 107713254 37464219 65218413 94834171 169064314 4274003 25055804 13115073
position Start 161275664 32237619 44031599 61560692 122296709 145104291 140201231 15780145 111217635 124860704 25056015 48042562 107713157 37464151 65218381 94834101 169064211 4273887 25055634 13115009
Chromosome * 3 12 LC5912 12LC61 10LC62 LC6512 # .ref region LC6315 marker cell Large LC471 LC48 LC49 LC50 LC51 LC52 LC53 |LC544LC54 LC55 LC56LC56 LC57 LC5817 LC60 LC64 LC66 Patent Application Publication Nov . 23 , 2017 Sheet 85 of 155 US 2017 /0335401 A1
Ipotassiumvoltage-gatedchannel, title Gene chromosome4openreadingframe shaker-relatedsubfamily,member3 (potassium/chloridetransporters), TBC1domainfamily,member30 cytochromeP450,family26 proteintyrosinephosphatase, Ideathinducer-obliterator1 branchedchainamino-acid transaminase1,cytosolic BT8(POZ)domaincontaining11 subfamilyA,polypeptide1 dedicatorofcytokinesis2 branchedchainamino-acid transaminase1,cytosolic IQmotifandSec7domain1 myelinproteinzero receptortype,F poly(ADP-ribose)polymerase solutecarrierfamily12 distal-lesshomeobox4 family,member15 CD38molecule member8 49 D : Entrez 4359 5792 11083 165631 84709 952 3738 84561 586 1748 121551 23329 1592 1794 586 9922
? 0 1 my 1 0 Island CpG In - - |1 ??????
Distance Tss 08612;-2789 693;46378-6 27892789;- 046759;,37546759;312 035053; 069539;70905 -1046378;46378 1455,870 14098 0261 2611 1215 120 0-3999 1961 130 0-39 0-392 Expn In 0 0 O O
- FIG.3(cont'd) Strand - * - + - + + + - + + + - -
|-NM_001178092;005504O Transcript NM033081;001193369O NM_001178092;005504O01178094;NM_001178091001 01170094;NM_001178091001 22105;NM080797001193370 NM001195483;024628 NM_002840;130440 NM000783057157 NM000530 NM080796 NM_001113523 NM_032623 NM001775 NM002232 NM138281 001018072NM NM_015279 NM004946 NM001134382 178093 - 178093
Gene MAX.chr132237619-32237654 MAX.chr8145104291-145104342 LC6019MAX.chr1937464151-37464219 LC6412MAX.chr124273887-4274003 1MPZ PTPRF PARP15 C4orf49 CD38 KCNA3 SLC12A8 12BCAT1 IQSEC1 Chromosome LC5020DIDO1 4 3 17DLX4LC58 128TBD11LC59 TBC1030LC6112 10CYP26A1LC62 5DOCK2LC63 BCAT112LC65| # .ref region marker cell Large LC47 LC48 LC49 LC51 LC52 LC53 LC54 LC55 LC56 LC57 33333333 LC663 Patent Application Publication Nov . 23 , 2017 Sheet 86 of 155 US 2017 /0335401 A1
BC/ UND 43.59 38.77 36.89 66.72 50.89 38.23 19.04 62.64 34.94 0.42 64.51 62.03 75. 35.54 63.62 25.40 38.73 34.4035.13 46.41 11.01
SQBC 3.70 37.98 35.86 32.63 6.32 12.77 14.20 18.15 32.72 46.29 30.96 15.06 14.11 10.93 29.21 8.97 2.58 12.62 30.77 1618. 2.12
2.57 0.23 53.71 74,59 3.78 61.10 13.18 29.28 21.72 54.76 3.39 82.32 20.42 58.88 1.01 5.80 65.0360.30 35.88 5.94 BC/ SC 116,741
BC/ LC 44.50 44.09 42.45 41.91 40.85 40.13 39.95 39.63 39.16 39.05 38.89 38.77 38.62 38.51 38.15 36.80 36.73 36.61 36.52 36.29 36.12
31.25 7.67 18.40 30.29 20.85 21.82 31.36 18.47 45.55 1.07 26.10 12.32 9.42 10.47 19.49 16.56 24.05 24.19 20.79 9.54 16.83 BC/ Ad
2.23 0.72 4.14 5.02 2.59 3.72 2.00 2.91 3.871 1.261 3.25 2.48 2.800 2.98 5.37 2.55 1.59 2.515.20 3.88 3.03 BC/ Norm
island. Lung cancer 10% 10% 29% 58% 18% 21% 17% 51% 16% %0 59% 16% 58% 17% 80% 13% 19% 27% 48% 18% %5 undefined mean
island. Lung 1% 9% 28% 28% 2% 7% 12% 15% 15% 17% 28% 4% 11% 5% 37% 5% 1% 10% 42% 7% 1% Squamous mean
%1 1% 1% 1% island . Lung 0% 42% 65% %34 11% 24% 54% 8% 50% 63% 10% 74% 3% 52% 83% 14% 2% FIG.3(cont'd) cell Small mean island .Lung 10% 11% 33% 36% 15% 22% 35% 32% 18% 15% 36% 10% 30% 18% 48% 19% 19% 29% 50% 14% 15% cell Large mean
island -Lung 7% 2% 14% 26% 7% 12% 27% 15% 21% 0% 24% 3% 7% %5 24% 9% 12% 19% 29% 4% 7% Adenocarcinoma mean
% island 0% 3% 4% %1 2% 2% 2% 0% 3% 1% 2% %1 7% 1% 2% 7% 1% 1% .normal . lung mean
0% 0% 1% %1 %0 1% 1% 1% 0% 0% 1% 0% 1% 0% 1% %1 1% %1 1% 0% 0% island . BC mean
position Stop 37434793 100273857 118981945 70982028 61638352 77268718 65731666 78511827 100075425 73073810 73147790 103352138 22624553 98290161 66808728 84748909 52401020 32237998 27205081 8859363 47444682
position Start 37434670 100273764 118981859 70981960 61638221 77268624 65731622 78511734 100075307 73073700 73147720 103352075 22624410 98290125 66808687 84748786 52400959 32237893 27205013 8859253 47444641
Chromosome 17 # .ref region marker cell Large LC67 LC6B LC69 LC70 LC71 LC72 LC73 LC74 LC75 LC76 LC77 LC78 LC79 LC80 LC81 LC82 LC8312LC83 LC84 LC85 LC86 LC87 Patent Application Publication Nov . 23 , 2017 Sheet 87 of 155 US 2017 /0335401 A1
Iphosphatidylinositol-3,45 title Gene TSC22domainfamily,member4 lelongationfactorTuGTP-binding phosphoinositides1)-associated proteinphosphatase1,regulatory Iguaninenucleotidebindingprotein basichelix-loopfamily, achaete-scutecomplexhomolog1 trisphosphate-dependentRac (Gprotein),betapolypeptide2 Dnaj(Hsp40)homolog,subfamily Temptyspiracleshomeobox1 domain-containingprotein1 GRP1(generalreceptorfor (inhibitor)subunit16B PRdomaincontaining14 GIPCPDZdomaincontaining exchangefactor1 (Drosophila) TSPY-like5 homeoboxA9 membere23 C,member6 family,member2 pseudogene scaffoldprotein ID Entrez 26051 2783 63978 128408 9829 54810 81628 2016 429 85453 160622 3205 57580
Island CpG In - 1 1 1 1
Distance Tss
0323, 212 - 136 02402 11602 1166 11930 1146 11595 03117 1624 151 0-152 1212 10-221 Exon In | - - 0 - - 0 - 0 - bol0 - bolo|1
FIG.3(cont'd) + + + + # - - + Strand + + P1
-
Transcript |NM_001172735;015568 NM_005273 NM024504 NM_080606 014787NM NM017655 NM_030935 NM004097 NM004316 NM033512 NR036652 NM_181711 NM152739 NM020820
Gene MAX.chr2118981859-118981945 15MAX.chr577268624-77268718LC72 LC7617MAX.chrt773073700-73073810 LC7910MAX.chr1022624410-22624553 LC81|2MAX.chr266808687-66808728 LC841MAX.chrt32237893-32237998 LC864MAX.chr48859253-8859363 20PPP1R168 7GNB2 |GIPC2 Chromosome LC708PROM14 LC7120BHLHE23 1DNAJC6LC73 |1 LC757TSC2204 LC772EMX1 LC7812ASCL1 LC8215LOC648809 LC8312GRASP LC857ROXAS LC8720PREX1 # .ref region marker cell Large LC67 LC68 LC692 55 56 LC74 9555 555LC80BTSPYL5 3885 Patent Application Publication Nov . 23 , 2017 Sheet 88 of 155 US 2017 /0335401 A1
57.48 56.07 37.29 62.78 15.35 58.67 45.24 8.78 44.43 35.41 15.29 56.82 57.40 59.56 49.05 36.14 64.67 21.41 63.00 47.74 45.33 12:27 41.54 BC { UND 64.49
60.97| 5.84 8.28 6.57 7,43 14.87 6.231 22.18 12.22 9.28 2.98 7.48 4.02 15.74 3.28 BC/ SQ 20.22 18.61 3181. 17.49 31.77 11.55 24.66 16.07 16.83
16.74 160.15 42.10 16.30 4.29 3.80 3.79 78.81 9,04 43.50 49.28 3.28 65.55 5.06 12.05 5.47 43.87 45.31 119.53 10.9B 47.26 57.24 12.44 45.85 BC/ SC
35.68| BC/ LC 36.01 36.01 35.89 35.70 35.38 35.26 34.94 34.87 34.38 33.99 33.63 33.62 33.53 33.24 32.37 30.95 30.66 30.46 30.04 30.02 29.85 29.64
18.45 8.36 24.3214.171 5.87 40.71 22.18 29.20 12.26 29.10 27.46 19.47 8.06 16.32 16.75 20.44 3710. 42.51 10,57 11.98 23.46 14.93 21.52 17.95 BC / Ad
2.95 2.39 3.723.811 2.42 2.07 3.36 3.97 1.83 10.3 3.08 3.43 2.19 3.42 3.10 3.50 1.31 1.78 2.93 3.31 2.84 3.39 BC/ Norm 4.03 3.41
island. Lung cancer 34% 18% 56%21% 25% 9% 23% 24% 3% 26% 49% 7% 35% 19% %24 23% 23% 32% 11% 38% 63% 59% 8% 46% undefined mean
%3 3% island .Lung 3% 18% 18% 5% 3% 12% 17% 5% 4% 31% 5% 6% 1% 2% 11% 8% 16% 7% 33% 21% 2% 18% Squamous mean
island. Lung 10% 47% 37% 9% 2% 2% 1% 41% 3% 26% 68% 1% 40% 2% %5 3% 28% 22% 62% 7% 63% 74% 8% 50% FIG.3(cont'd) cell Small mean island . Lung 21% 11% 31% 20% 14% 22% 14% 18% 11% 20% 47% 15% 21% 11% 13% 16% 21% 15% 16% 18% 40% 39% 20% 33% cell Large mean
island. Lung 11% 2% 21% 8% 2% 25% 9% 15% 4% 17% 38% 9% 5% 5% 7% 10% 7% 21% 5% 7% 31% 19% 14% 20% Adenocarcinoma mean
2% 1% 3% 1% 1% 1% 1% 2% 1% 1% 1% 1% %1 2% 2% 4% island 2% 4% 2% 4% 4% . normal. lung mean
1% 0% 1% 1% 0% 1% 0% 0% 1% 1% 0% %1 0% 0% 0% 1% 0% 1% 1% 1% 1% 1% %1 island. BC mean
position Stop 54486134 26407876 27196154 158937476 103352268 968684 157483425 178957695 46675464 51831160 74726617 70654616 134879709 107715084 52734592 101004860 13524378 81002992 14926716 176945135 248020722 31820418 26699456 214158969 968477 position Start 54486055 26407721 27196035 158937376 103352229 157483370 178957576 46675383 51831114 74726554 70654555 134879621 107715041 52734489 101004818 13524253 81002926 14926627 176945102 248020671 31820365 26699373 214158912
Chromosome 19 M 127 5 17 19 2 14 5 12 14 1 10 11 77 17 # .ref region 1999T14 markercell Large LC88 LC89 LC90 LC91 LC92 LC93 LC94 LC95 LC96 LC97 LC98 LC99 LC100 LC101 LC102 |LC103 LC104 LC105 LC106 LC307 LC108 LC109 LC110 LC111 Patent Application Publication Nov . 23 , 2017 Sheet 89 of 155 US 2017 /0335401 A1
familywithsequencesimilarity59. prostaglandinD2receptor(DP) title Gene dependent,gammasubunit8 achaete-scutecomplexhomolog1 proteintyrosinephosphatase,Treceptortype,Npolypeptide2 zincfinger,MIZ-typecontaining1 calciumchannel,voltage vasoactiveintestinalpeptide IgLONfamilymember5 solutecarrierfamily8 (sodiurn/calciumexchanger), BTB(POZ)domaincontaining11 Gprotein-coupledreceptor88 even-skippedhomeobox2 Fripartitemotif-containing58 sterilealphaandTIRmotif prosperohomeobox1 memberB homeoboxA7 receptor2 (Drosophila) homeobox85 member3 pairedbox6 containing1 agrin ladybirdhomeobox2 ID Entrez 59283 150946 3204 7434 429 375790 5799 3215 402665 85474 6547 121551 5729 54112 57178 344191 25893 5080 23098 5629
Island CpG in 1 TE
Distance Tss -0897112;897112,8971121 01232; 012514;19144 14137;11762 119766 1261 2730 1778 1-4280 116013 159 11091 0174135 O 13588 1171 3871387 0-2947 Exon In - - +012975 03889 02845 000 - 0 FIG.3(cont'd) Strand + ++ 1 - ++ 1 + - + + + + ++ ++ +
Transcript NM130842;_0028471308 NM183002;058240_1829 NM_000280;0016040011- NM_001191033;001168241 NM_031895 NM006896 NM003382 NM_004316 NM_198576 NM_002147 32;NM_033262 NM000953 NM022049 015431NM NM015077_015077 NM002763 43 NM001101372 NM001009812 NM001018072 NM020338 NM001080458 27612
Gene MAX.chr5178957576-178957695 MAX.chr5134879621-134879709 LC1044MAX.chr413524253-13524378 11MAX.chr1114928627-14926716LC106 19CACNGS FAM59B2 7HOXA7 7VIPR2 12ASCL1 AGRN PTPRN2 SARM117 1PROX1 Chromosome 5 LC9617HOXBS LC9719GLONS LC982LBX2 14SLC8A3LC99 5 |12BTBD11LC101 14PTGDRLC102 LC1031GPR88 LC10510ZMIZ1 LC1072EVX2 1TRIM58LC108 11PAX6LC109 # .ref region markercell Large LCB8 LC89 LC90 L291 LC92 LC93 LC94 LC95 LC100 LC110LC110 LC111 Patent Application Publication Nov . 23 , 2017 Sheet 90 of 155 US 2017 /0335401 A1
36.31 39,68 31.74 61.82 14.73 1.601 25.68 61.08 20.21 47.09 24.93 74.66 18:37 14.59 20.46 22.59 19.80 48.45 45.23 BC/ UND
BC / SQ 12.51 10.94 14. 4.24 4.89 1.82 14.91 4.201 21,36 9.22 8.92 6.89 10.131 6.68 28.54 2.89 16.93 26.30
7.65 80.54 19.00 17.82 3.39 45.90 7.161 36.65 13.50 39.72 5.77 17.16 77.49 45.37 10.49 9.23 2.04 26.39 66.32 BC/ SC
29.63 29.3829.38 28.82 28.53 28.44 28.42 28.37 27.91 27.33 27.03 27.00 26.97 26.41 25.97 25.58 25.41 25.37 25.20 25.16 BC/ LC
19.29) BC/ Ad 16.30 10.5610,56 12.04 15.29 9.00 3.90 14.60 18.06 13.60 22.36 14.58 25.891 22.43 16115, 29.77 19.11 17.21 19.21
2.31 3.40 3.15 2.85 3.09 2.01 1.58 1.61 2.76 1.800 4.03 2.26 2.15 2.78 3.07 3.15 1.36 3.68 2.54 BC/ Norm
island . Lung cancer 31% 17% 34% 63% 5% 1% 23% 50% 14% 64% 19% 62% 7% 14% 33% 23% 12% 69% 56% undefined mean
island. Lung 11% 5% 15% 15% 2% 2% 2% 12% 3% 29% 7% 7% 3% 10% 11% 29% 2% 24% 32% Squamous mean islandpuejs?funz. Lung 6% 35% 21% 18% 1% 21% 6% 30% 10% 54% 14% 31% 44% 17% 9% 1% 38% 81% FIG.3(cont'd) cell Small mean island .Lung 25% 13% 31% 29% 10% 13% 25% 23% 19% 37% 20% 22% 11% 25% 42% 26% 16% 36% 31% puestburcell Large mean
island .Lung 14% 5% 13% 15% 3% 2% 17% 12% 13% 19% 17% 12% 10% 22% 25% 30% 12% 25% 24% AdenocarcinomaBunz mean
30% 1% 3% 3% 1% 1% 1% 2% 1% 5% 2% 2% 1% 3% 3% 1% 5% 3% island 5% .normal . lung mean
1% 0% 1% 1% 0% 0% 1% 1% 1% 1% 1% 0% 1% 2% 1%1 1% 1% 1% island . BC mean
position Stop 13984972 134879483 8860023 52735485 12254366 2903705 72756053 23564136 30017896 25056246 50297912 52735373 25439264 32237880 32713669 145105675 52401138 27195829 99439356
position Start 13984886 134879362 8859990 52735425 12254306 2903614 72755971 23564059 30017795 25056183 50297879 52735223 25439185 32237695 32713586 145105570 52401041 27195748 99439270
Chromosome 4 14 12 14 12 # .ref region marker cell Large LC112 LC113 LC114 LC115 LC116 LC117 LC118 LC119 LC120 LC121 LC122 LC123 LC124 LC125 LC126 LC127 LC128 LC129 LC130 Patent Application Publication Nov . 23 , 2017 Sheet 91 of 155 US 2017 /0335401 A1
prostaglandinD2receptor(OP) (atrionatriureticpeptidereceptorC) title Gene prostaglandinD2receptor(DP)5729 serpinpeptidaseinhibitor,cladeB5272 Fasapoptoticinhibitorymolecule2 Iphosphoinositides1)-associated chromosome2openreadingframe celldeath-inducingDFFAlike1149 transaminase1,cytosolic GRP1(generalreceptorfor spondin1,extracellularmatrix (ovalbumin),member9 137814NK2transcriptionfactorrelated, locus6(Drosophila) 586branchedchainamino-acid natriureticpeptidereceptor C/guanylatecyclase effectora 342865V-setandtransmembranedomain containing28 scaffoldprotein homeoboxA7 protein 100132891hypotheticalLOC100132891 55 ID Entrez 10418 23017 5729 4883 160622 3204 343990
Island CpG In 1
Distance Tss 17446210;-861 1-379;614 -046210;46210 19 |+10-11;53 69-0 137-0 0-159 1793 19220 1294 0548 1113414 ExonExon nIn } • ? +0995 ? ? 0ololo lo |-O +0305 FIG.3(cont'd) + - + - + - + StrandStrand + + + +
Transcript NM_001178092;005504001178094;NM_001178091001 NM001279NR036468 NR033651;033652 NM_006108 NM000953 004155NM_ |NM_001136271 NM_001146339 178093 NM012306 NM000953 NM_000908 NM_181711 NM006896 |NM_207362
Gene MAX.chr5134879362-134879483 4MAX.chr48859990-8860023 2MAX.chr225439185-25439264 1MAX.chr132237695-32237880 MAX.chr8145105570-145105675 NKX26- 11SPON1 14PTGDR 18CIDEA SERPINB9 LOC100132891 19VSTM2B BCAT112 12FAIM2 14PTGDR 5NPR3 12GRASP 7HOXA7 2C2orf55 Chromosome 5 8 8 8 # .ref region marker cell Large LC112 LC113 LC114 LC115 LC116 LC117 LC118 LC119 LC120 LC121 LC122 LC123 LC124 LC125 LC126 LC127 LC128 LC129 LC130 Patent Application Publication Nov . 23, 2017 Sheet 92 of 155 US 2017 /0335401 A1
BC/ UND 13.03 31.02 2.90 3.79 23. 0.45 39.93 57.60 21.94 46.04 48.13 30.81 29.27 22.04 1721. 9.25 26.04 19.36 26.96 1.26 32.65 25.28
.77 4.31 9.72 14.12 12.84 18.68 4.40 13.73 3.82 10.21 5.11 6.54 5.54 23.48 4.33 3.68 3.65 13.18 26.30 6.18 5.76 SQIBC 19.55
20.868 21.20 1.32 10.80 45.83 13.65 50.79 43.13 3.72 18.91 12.57 36.30 9.72 10.90 82.48 23.61 3.21 11.77 32.73 13.62 8.65 2.19 8C / SC
24.47| 23.92| meLCIBC 24.87 24.80 24.72 24.70 24.28 23.37 22.92 22.80 22.74 22.73 22.64 22.63 22.19 22.08 21.93 21.79 21.67 21.62 2161 21.57 5.42 11.74 7.61 10.77 19.071 2.43 13.94 3.80 9.63 16.40 15.78 6.55 15.63 6.84 8.50 12.66 11.60 10.12 12.74 8.67 2.73 BC/ Ad 13.80
1.19 1.13 2.91 88.1 2.36 2.01 2.37 1.45 1.14 2.36 1.13 BC/ Norm 2.57 3.29 1.54 2.73 1.40 2.49 3.07 57.2 2.01 1.50 1.58
2% 2% 0% 10% 1% island. Lung cancer 16% 55% 29% 61% 55% 18% 42% 53% 33% 16% 29% 1.% 11% 26% 38% 21% 31% undefined mean
island .Lung 11% 17% 12% 6% 23% 3% 30% 13% 3% 9% 6% 7% 2% 7% 12% 5% 4% 2% 19% 18% 4% 7% Squamous mean
3%. island . Lung 26% 37% 1% 5% 56% %9 78% 41% 3% 17% 14% 39% 5% 14% 41% 28% 6% 46% 9% 6% 3% FIG.3(cont'd) }} ce Small mean island .Lung 31% 44% 20% 11% 30% 16% 37% 22% 18% 21% 25% 25% 12% 30% 11% 26% 22% 12% 31% 15% 14% 26% fa5abje7 ueau
island .Lung 7% 21% 6% 5% 23% %2 21% 4% 8% 15% 17% %7 9% 4% 15% 12% 5% 19% 9% 0% 3% Adenocarcinoma mean *
island 3% 6% 1% 1% 7 4% 2% 1% 2% 4% 1% 3% 1% 2% 1% 2% %1 .normal . lung mean
%1 2% 1% 0% 1% 1% 2% 1% 1% 1% 1% 1% %1 1% %1 %1 10% 1% 1% 1% 1% 1% island . BC mean
position Stop 124910715 35293764 161275649 44896855 108440760 2903104 57618831 36042110 233352699 52536240 105036777 110627221 143592285 158938133 43339345 119312997 77179841 155412375 70947084 6511655B 52535870 103352323
position Start 124910562 35293717 161275580 44896701 108440646 2903051 57618791 36042030 233352635 52536106 105036730 110627121 143592229 158938047 43339264 119312919 77179798 155412291 70947017 65116474 52535777 103352291
17 12 2 17 10 17 14 Chromosome 21 |12 | | 4LC148 # .ref region 114LC140 18LC149 marker cell Large LC131 LC132 LC133 LC134 LC135 LC136 LC137 LC138 LC139 LC141 LC142 LC143 LC144 LC145 LC146 LC147 LC150 LC151 LC152 Patent Application Publication Nov . 23 , 2017 Sheet 93 of 155 US 2017 /0335401 A1
endothelinconvertingenzyme-like1 benzimidazoles3homolog(yeast) cladeinhibitor8,serpinpeptidase title Gene wingless-typeMMTVintegrationsite chlorideintracellularchannel6 chromosome17openreadingframe RNAbindingprotein,fox-thomolog achaete-scutecomplexhomolog1 nidogen2(osteonidogen) filamentprotein,alpha brain-specificangiogenesisinhibitor dachsous2(Drosophila) PIF15-103'DNAhelicasehomolog Inidogen2(osteonidogen) buddinguninhibitedby myelinproteinzeru family,member3 (ovalbumin),merberg neurexophilin4 internexinneuronalintermediate vasoactiveintestinalpeptide receptor2 (C.elegans3 (S.cerevisiae) (Drosophila) T-box20 46 ID Entrez 9184 57057 4359 7473 5272 11247 54102 9427 22795 9118 575 7434 124783 146713 80119 22795 429
Island CpG in to - o ------1
Distance Tss
0-3197; 0-6; 619-0 0-103 0-160 0-189 398-0 -1586,639 O494 0- 1169 |14182 046853 |11364 |1840 ExonExon InIn - +18214- +1343 O 0215- 0298765- 10. FIG.3(cont'd) + - - - - + - + - - Strand + + + +
Transcript NM001007793004725 INM001166220;NM001077653 NM001142552;001142553 NM000530 NM030753 NM_004155 NM007224 NM053277 NM004826 NM007361 NM032727 NM_001702 NM_003382 152343NM_ NM001082575 NM_025049 NM007361 NM004316
Gene MAX.chr6108440646-108440760 MAX.chr1110627121-110627221 119312997119312919-MAX.chr1010 MAX.chr870947017-70947084 BAI1 10BUB3 MPZ SERPINDY 12NXPH4 21CLIC6ECELT 2 NID214 JINA10 VIPR2 C170r4617 17RBFOX3 DCHS2 PIF1 Chromosome TBX20LC1327 WNT3LC13417| 4 8 15 - | LC151|14NID2 LC152|12ASCL1 # .ref region marker cel Largeen LC131 LC133 LC135 LC136 LC137 LC138 LC139 LC140 LC141 LC142 LC143 LC1441 LC145 LC146 LC147 LC148 LC149 LC150 Patent Application Publication Nov . 23, 2017 Sheet 94 of 155 US 2017 /0335401 A1
BC/ UND 14.91 28.67 11.06 32.61 16.97 15.52 30.00 24.97 36.27 18.84 15.82 25.10 4.191831 14.8911 9.423478 56.502957 7.602348 11.71754 2.74| 4.36 49.01 4.01 5.17 4.08 10.20 302. 7.05 19.46 7.96 10.27 BC/ SQ
9.66 9.30 7.07 4.83 12.33 2.63 3.27 2.45 SCIBC 112.60 26.15 18.12 45.68 12.20 13.46 29.02 19.04 11.47 13.43
20.66 20.35 2720. 20.26 20.12 BC/ LC 21.51 21,33 21.26 21.25 21.20 21.05 20.82 20.71 20.70 20.67 20.56 20.19 20.04
9.78 9.12 4.64 7.35 57.8 9.26 7.48 5.07 .967 9.08 5.44 5.94 BC/ Ad 11.00 16.72 11.42 10.14 3.68 12.72
1.46 1.73 2.19 2.16 2.06 2.001 1.74 1.64 2.12 1.59 2.00 1.42 1.91 1.93 2.78 1.27 2.36 1.75 BC/ Norm
island. Lung cancer 10% 16% 18% 20% 34% 16% 45% 30% 23% 59% 23% 22% 4% 39% 62% 22% 16% 58% undefined mean
island .Lung 3% 27% 7% 3% 8% 10% 10% 24% 6% 31% 2% 7% 7% 11% 33% 9% 3% 24% Squamous mean
island .Lung 6% %62 15% 4% 10% 12% 64% 5% 12% 48% 9% 13% 2% 4% 49% 22% 11% 31% FIG.3(cont'd) cell Small mean island. Lung 14% 12% 35% 13% %42 21% 51% 42% 14% 22% 16% 19% 12% 32% 34% 24% 20% 46% cell Large mean
island .Lung 7% %5 15% 10% 23% 5% 18% 21% 6% 10% %6 5% 2% 13% 15% 6% 6% 29% Adenocarcinoma mean
island %1 %1 4% 1% 4% 2% 4% 3% 1% %2 2% 1% %1 3% 5% 1% 2% 4% .normal . lung mean
1% %1 2% 1% 2% 1% 2% 1% 1% 1% %1 2% 2% 1% 1% %2 island . BC mean
position Stop 114141 26407639 80329526 946438 87088997 78512047 46832494 1876340 8859184 8277571 10563607 74726257 129377714 106979932 170137439 74726331 50343395 687729 114075 946378 position Start 26407567 80329497 87088958 78511892 46832435 1876308 8859049 8277482 105635691 74726082 129377645 106979840 170137371 74726270 50343331 687683
Chromosome 2 VN 8 # .ref region marker cell Large LC153 LC154 LC155 LC156 LC1572 LC158 LC159 LC160 LC1614 LC162 LC1635 LC164 LC165 LC166 LC167 LC168 LC169 LC170 Patent Application Publication Nov. 23, 2017 Sheet 95 of 155 US 2017 / 0335401 A1
familywithsequencesimilarity59, IKAROSfamilyzincfinger1(Ikaros) title Gene LIMhomeoboxtranscriptionfactor regulatoryfactorX,4(influences urotensin2receptor syntrophin,gamma2 GIPCPDZdomaincontaining lankyrinrepeatdomain33B ladybirdhomeobox2 HLAclassIlexpression) ladybirdhomeobox2 memberB CD8bmolecule family,member2 claudin11 1,beta ? 1 D Entrez 150946 2837 54221 926 54810 651746 85474 4010 5992 5010 85474 10320
Island CpG in 1 1 - - 1 1
Distance Tss
13983;11608 O924; +0-1656;719 0-865 175-0+ 3041 O4361 O2808 +0-1046 Exon in O +10-2703 to odolaO O 417310-
FIG.3(cont'd) - + + - + - + 1-10 Strand 89:;-11(172102NM0011004931 + |+
Transcript NM002316;001174146O NM_001191033;001168241 78100;NM_172213172101 |NM_001185056;005602 NM018949 NM018968 NM_017655 NM001164440 NM001009812 NM213594 001009812NM NM_006060 ? ? 1 01174147
Gene MAX.chr9114075-114141 LC15917MAX.chr1746832435-4683249418763401876308 -LC1605MAX.chr5 LC161MAX.Chr48859049-8859184 MAX.chr18277482-8277571LC162 MAX.chr8687683-687729 FAM59B |GIPC2 RFX412 CLON11 18X2 IKZF1 Chromosome 9 2 17UTS2RLC155 LC1572CD8B 1 ANKRD33BLC163N03 LBX2 LC1642 LC1659LMX1B 3 # .ref region SNTG2LC1562| marker cell Large LC153 LC154 LC158 LC166 LC167 LC1682LC1697 LC1708 Patent Application Publication Nov . 23 , 2017 Sheet 96 of 155 US 2017 /0335401 A1
627.38 68.97 33.11 254.67 246.29 31.95 33.15 87,14 74.38 143.28 69.90 91.67 34.94 68.46 29.71 75, 66.72 79.20 45.23 45.05 29.92 35.13 46.42 45.33 46.07 17.68 32.83 13.65 35.41 UNDBC
BC/ SQ 158.96 15.55 57.49 52.16 39.04 32.72 21.45 20.71 14.11 32.63 34.74 26.30 13.58 22.68 30.77 26.78 16.07 13.59 6.30 10.38 15.00 22.18 459.6933517 266.287091 249.9018619 45.0021746 170.642604 168.753565 66.1516484 BC/ SC 154.96 151.39 83.04 82.32 74.59 68.59 64.55 62.06 60.30 59.12 57.24 55.76 51.66 50.49 49.97 49.28 141,74163.36 42.5711967 39.1611674 41.0110181 25.166632 BC/ LC 220.17 99.05 84.20 141.62 37.71 42.49 42.26 54.98 14.66 54.78 24.03 38.62 41.91 39.97 19.50 28.80 36.52 50.24 30.02 21.37 19.00 23.92 8.26 34.21
3.73) 9.57 9.42 11 20.79| 8.36 9.00 BC / Ad 88.00 18.97 27.57 82.13 27,54 61.89 16.01 19.63 39.97 36.56 33.54 45.55 32.52 11.67 30.29 21.20 19.21 22.10 14.48 25.97 14.93 12.56 27.46
8.04) 37. 8.701 6.70 18.40 15.58 2.69 10.65 14.98 8.35 5.87 5.71 3.87 5.38 2.86 2.80 5.02 6.02 2.54 483. 4.191 5.20 3.67 3.41 4.02 2.14 3.65 2.44 3.10 BC/ Norm
island. Lung cancer 59% 20% 10% 62% 55% 5% 13% 25% 22% 61% 24% 50% 16% 22% 28% 58% 58% 58% 56% 49% 31% 48% 44% 59% 51% 15% 42% 16% 49% undefined mean 8
island . Lung 32% 21% 27% 39% 10% 4% 14% 19% 5% 25% 18% 21% 15% 7% 19% 11% 28% 25% 32% 15% 24% 42% 25% 21% 15% 5% 13% 18% 31% Squamous mean
island. Lüng 43% 78% 78% 60% 49% 28% 66% 48% 46% 65% 49% 65% 54% 32% 77% 63% 65% 50% 81% 71% 65% 83% 56% 74% 62% 42% 64% 59% 68% cell Small mean FIG.4 pue?si' bun ? 21% 29% 26% 35% 8% 7% 16% 16% 4% 23% 22% 23% 18% 13% 22% 30% 36% 29% 31% 21% 30% 50% 47% 39% 24% 16% 31% 10% 47% cell Large mean
island Lung 8% 6% 9% 20% 6% 10% 6% 6% 3% 17% 13% 18% 21% 10% 11% 7% 26% 15% 24% 24% 15% 29% 24% 19% 14% 7% 11% 4% 38% Adenocarcinoma mean
island 4% 3% 2% 5% 3% 0% 4% 4% 2% %3 2% 4% 2% 2% 3% 2% 4% 4% 3% 4% 4% 7% 3% 4% 4% 2% 5% 3% 4% .normal . tung mean
0% 0% 0% 0% 0% 0% 0% %0 0% 0% 0% 1% 0% 0% 1% 1% 1% 1% 1% 1% 1% 1% 1% 1% 1% 1% 1% 1% 1% island . BC mean
position Stop 42995171 14926955 14926853 42995528 124132861 27467467 14927132 149803586 17403270 122296828 22541996 72754648 100075425 144328649 96721564 22624553 70982028 138658705 99439356 33402304 223936773 27205081 48042606 31820418 71116269 239140351 55095201 89914776 74726617 position Start 42995102 14926886 14926795 42995477 124132797 27467359 14926995 149803478 17403238 122296709 22541891 72754556 100075307 144328573 96721507 22624410 70981960 138658597 99439270 33402226 223936705 27205013 48042562 31820365 71116233 239140226 55095158 89914730 74726554
Chromosome 19 10 14 15 10SC16 # .ref region SC1519 marker Cell Small SC15 SC2 SC3 SC4 SC5 SC6 SC7 SC8 SC9 SC10 SC11 SC12 SC13 SC14 SC17 SC18 SC19 SC20 SC21 SC22 SC23 SC24 SC25 SC26 SC27 SC28 SC29 Patent Application Publication Nov . 23, 2017 Sheet 97 of 155 US 2017 /0335401 A1
title Gene abhydrolasedomaincontaining8 chromosome2openreadingframe zincfingerCCCH-typecontaining 81628ITSC22domainfamily,member4 Ipoly(ADP-ribose)polymerase PRdomaincontaining14 calpain2,(m/fl)largesubunit distal-lesshombobox4 ladybirdhomeobox2 family,member15 286128zincfingerprotein41homolog pairedbox6 hypotheticalLOC151174 myosinXVIIA (mouse) 56033BARXhomeobox1 homeoboxA9 stomatin 120 100132891hypotheticalLOC100132891 55 ID Entrez 17 . 1 2040 399687 340152 79575 165631 63978 343990 824 3205 1748 5080 151174 85474
Island CpG In O O 7 puejsisdo un ------1- - -
Distance Tss
40048; -1794;801 0-252 92;1 1595 -535 0-3899 113414 110441 O261 11602 1136 +0-3999 0 O 0 1 1 0 0 O Exon in ??????? 1-0 02670- -03889 + + 1-01 FIG.4(cont'd) Strand - 111i11 - + 1 1 1 12 TOP |+047411;36587NM001146068001748 TO NM000280;001604:001101251419144
Transcript NM198194;004099 NM078471;203318 NR_033651;033652 NR026926:026925 NM_207360 NM024527NM001113523 NM030935 NM173832 NM_021570 NM024504 NM207362 NM152739 NM138281 NM001009812 - - 27612
Gene MAX.chr542995102-42995171 2254199622541891-MAX.chr10 MAX.chr1022624410-22624553 MAX.chr3138658597-138658705 14MAX.chr1433402226-33402304 MAX.chr271116233-71116269 MAX.chr1855095158-55095201 MAX.chr1589914730-89914776 SC211MAX.chr1114926886-14926955 SC311MAX.chr1114926795-14926853 SC45MAX.chr542995477-42995528 SC7|11MAX.chr1114926995-14927132 PARP15 LOC100132891 TSC22D4 ZFP41 BARX1 PRDM14 C2orf55 1CAPN2 7HOXA9 17OLX4 11PAX6 LOC151174 L8X2 Chromosome STOMSCS9SC6|17MYO18A 6ZC3H12DSC8 10 8 7 8 9 10 8 3 - - = - ABHD8SC919------18SC2722 SC2815 # .ref region markerCell Small SC1 SC10 SC11 SC12 SC13 SC14 SC15 SC16 SC17 SC18 SC19 SC20 SC21 SC22 SC23 SC24 SC25SC262 SC292 Patent Application Publication Nov . 23 , 2017 Sheet 98 of 155 US 2017 /0335401 A1
25.84| UNDBC 43.20 50.06 15.47 20.83 32,46 27.92 27.11 21.30 35.78 43.54 32.15 17.08 17.01 34.09 23.32 27.57 61.08 36.91 24.53 34.56 21.87
18.39| 11:24 17.75 9.64 8.35 7.09 9.17 3.55 12.92 7.83 6.15 15.47 5.12 11.04 15.88 .59 14.91 5.35 15.71 10.85 6.17 SQBC 48.262358 5
47.17 42.16 1042. 42.04 41.49 40.62 40.60 40.60 39.04 38.741 38.09 37.74 37.51 37.26 36.66 36.65 36.50 36.28 35.93 35.861 35.74 BC/ SC
24.67 18.41 11.30 24.13 15.80 17.57 9.211 12.14 20.02 13.85 15.05 10.15 9.61 19.16 16.95 5.141 27.91 12.56 18.83 16.83 9.67 18.371 BC/ LC
22.69 5.10 5.48 12.47 8.89 9.82 6.40 7.95 8.471 10.92 9.27 13.16 13.80 10.30 7.42 5.53 14.60 12.78 10.01 10.55 6.44 17.27 AdBC
2.44) 1.911 81.1 1.93 2.23 1.810 1.52 2913. 3.76 2.15 1.72 2.54 2.18 3.36 2.76 2.39 2.35 3.19 BC /Nory 4.21 4.65 1.82 2.83
island. Lung cancer 49% 56% 26% 34% 58% 54% 54% 42% 49% 51% 60% 23% 15% 66% 48% 38% 50% 55% 56% 60% 44% 39% undefined mean
island .Lung 26% 13% 30% 16% 15% 14% 18% 7% 18% 9% 11% 21% 4% 21% 33% 8% 12% 8% 36% 19% 13% 28% Squamouspuejsbunz mean
island. Lung 54% 53% 72% 68% 76% 80% 81% 79% 56% 46% 72% 52% 32% 73% 77% 50% 30% 54% 83% 62% 73% 54% FIG.4(cont'd) cellpurys Small :sunt mean pumperbungisland. Lung 28% 21% 19% 39% 28% 34% 18% 24% 28% 16% 28% 14% 8% 37% 35% 7% %23 19% 43% 29% 20% 28% cell Large mean
island .Lung 25% 6% 9% 20% 16% 19% 13% 16% 12% 13% 17% 18% 12% 20% 15% 8% 12% 19% 23% 18% 13% 26% Adenocarcinoma mean
island 5% 2% 3% 8% 3% 4% 4% 3% 5% 4% 4% 2% 2% 5% 5% 5% 2% 4% 5% 5% 5% 5% .normal . lung mean
1% 1% 2% 2% 2% 2% 2% 2% %1 1% 2% 1% 1% 2% 2% 1% 1% 1% 2% 2% 2% 2% island. BC mean
position Stop 133481521 22624375 61560535 151811523 1295496 8482670 96722762 13617235 31825955 47696674 1295587 16394575 101033775 71116131 157821378 39597893 23564136 25061911 171678966 165323624 2765768 38724946
13617-166 position Start 133481464 22624260 61560462 151811410 1295444 8482598 96722680 31825851 47696594 1295519 16394457 101033663 71116047 157821297 39597822 23664059 25061836 171678927 165323561 2765684 38724873
Chromosome 12 10 20 7 5 1914 NS Alan 14 # .ref region markerSome Cel Small SC30 SC31 SC32 SC33 SC34 SC35 SC369 SC37 SC3811SC38 SC39 SC40 SC41SOLIS SC42 SC43 SC44 SC45 SC46 SC47 SC48 SC49 SC50 SC51 Patent Application Publication Nov . 23 , 2017 Sheet 99 of 155 US 2017 /0335401 A1
glutamatedecarboxylase1(brain, title Gene dependent,P/Qtypealpha1A associatedhomolog(rat) INK2transcriptionfactorrelated, deathinducer-obliterator1 C2calcium-dependentdomain telomerasereversetranscriptase calciumchannel,voltage T-cellacutelymphocyticleukemia1 telomerasereversetranscriptase brain-enrichedguanylatekinase shortstaturehomeobox2 visualsystemhomeobox1 LIMhomeoboxtranscriptionfactor C-typelectindomainfamily14, neurexophilin1 pairedbox6 locUS6(Drosophila) containing40 67kDa) memberA subunit 1.alpha D { Entrea 11083 100191040 7015 30010 773 5080 6886 7015 . 57596 6474 137814 30813 2571 4009 161198 sland( CpG In 1 1 1 1 - - - 1 F - o - - o
Distance Tss 8842;-2559: 108; 7028;13658 2655; 2559;- -282; -357; 5728: 23911917; 0744;2468 1623 9014 -1151 . 137-10 -0931:931 11701 1 1 uoxg u 0O 0 O 0 -01 -0 FIG.4(cont'd) - +- - - - 1 1 1 U Strand ! 1- -1
NM_001127222;0230350 Transcript NM033081;001193369O 22105;NM080797001193370 01127221;NM_000068001174 NM_000280;0016040011 NM001163678;006884O NM001193376:198253 |NM001193376;198253 NM_001159531;020836 NM014588;199425 NM_000817;013445 NM_001174069;177398 NM080796 NM001136003 NM152745 NM003189 NM001136271 175060NM_ 080 27612 - 03030
Gene 12MAX.chr12133481464-133481521 9672276296722680-SC369MAX.chr9 MAX.chr1916394457-1639457519 MAX.chr271116047-71116131 |MAX.chr2039597822-39597893 MAX.chr42765684-2765768 SC31|10MAX.chr1022624260-22624375 NKX2-6 TAL1 lGA- C2CD40 5TERT 14BEGAIN SHOX2 20VSX1 GAD1 LMXIA CLEC14A Chromosome 20DIDO1SC32 TERT5SC34 SC3517NXPKA 19CACNA1ASC37 SC3811PAXG 1123 Tralis2 pela20 & 2 4 14 # .ref region marker Cell Small SC30 SC33 SC39 SC40 SC41 SC42 SC43 SC44 SC45 SC46 SC47 SC48 SC49 SC50 SC51