Granted Patent
B2
US 12,467,082 · App. 19/028,618 · Granted Nov 11, 2025
Selective oxidation of 5-methylcytosine by tet-family proteins
Inventors:
Anjana Rao (La Jolla, CA); Mamta Tahiliani (New York, NY); Kian Peng Koh (Jamaica Plain, MA); Suneet Agarwal (Belmont, MA); Aravind Iyer (Bethesda, MD)
C12Q1/6827C12N9/1007C12Q1/26C12Q1/6806C12Q1/6869G01N33/5308C12N2501/70C12N2501/71C12N2501/724C12Q2521/531C12Q2522/10C12Q2537/164C12Q2600/154
View Patent ↗
Loading inventors, assignments & file history…
Abstract
The present invention provides for novel methods for regulating and detecting the cytosine methylation status of DNA. The invention is based upon identification of a novel and surprising catalytic activity for the family of TET proteins, namely TET1, TET2, TET3, and CXXC4. The novel activity is related to the enzymes being capable of converting the cytosine nucleotide 5-methylcytosine into 5-hydroxymethylcytosine by hydroxylation.
Claims (27)
1 . A method comprising:
(a) modifying a hydroxyl group of a hydroxymethylated cytosine residue in a mammalian deoxyribonucleic acid to generate a modified hydroxymethylated cytosine residue, wherein said modifying comprises glycosylating said hydroxyl group to generate a glycosylated hydroxymethylated cytosine residue; and
(b) sequencing said mammalian deoxyribonucleic acid comprising said glycosylated hydroxymethylated cytosine residue.
2 . The method of claim 1 , wherein said sequencing comprises high-throughput sequencing.
3 . The method of claim 1 , wherein said sequencing comprises whole genome sequencing.
4 . The method of claim 1 , wherein said sequencing comprises array hybridization.
5 . The method of claim 1 , wherein said isolated mammalian deoxyribonucleic acid is from a somatic cell.
6 . The method of claim 1 , wherein said mammalian deoxyribonucleic acid is obtained from a cell sample or a tissue sample of a subject.
7 . The method of claim 1 , wherein said mammalian deoxyribonucleic acid is obtained from an extracellular fluid sample.
8 . The method of claim 1 , wherein said mammalian deoxyribonucleic acid is obtained from a subject having cancer or suspected of having cancer.
9 . The method of claim 1 , further comprising detecting a methylation status of said mammalian deoxyribonucleic acid based on a presence or absence of said glycosylated hydroxymethylated cytosine residue.
10 . The method of claim 1 , wherein said glycosylating comprises glycosylating said hydroxymethylated cytosine residue with a glucose residue or a modified glucose residue.
11 . The method of claim 1 , wherein said glycosylating comprises contacting said mammalian deoxyribonucleic acid with a glycosyltransferase to glycosylate said hydroxyl group.
12 . The method of claim 1 , further comprising, before (a), contacting said mammalian deoxyribonucleic acid with: (i) a TET polypeptide; (ii) a catalytically active fragment of said TET polypeptide; (iii) a variant of said TET polypeptide; or any combination thereof.
13 . The method of claim 12 , wherein said TET polypeptide comprises: (i) a TET1 polypeptide; (ii) a TET2 polypeptide, or (iii) a CXXC4 polypeptide, and comprises a polypeptide having at least 90% sequence identity to any one of SEQ ID NOs: 27, 28, or 29.
14 . The method of claim 12 , wherein said TET polypeptide further comprises SEQ ID NO: 1 or a variant thereof.
15 . The method of claim 12 , wherein said TET polypeptide comprises a TET2 polypeptide.
16 . The method of claim 12 , wherein said TET polypeptide comprises a catalytically active fragment of a TET2 polypeptide.
17 . The method of claim 12 , wherein said TET polypeptide comprises a polypeptide having at least 90% sequence identity to SEQ ID NO: 28.
18 . The method of claim 12 , wherein said catalytically active fragment of said TET polypeptide comprises a polypeptide having at least 90% sequence identity to a catalytically active fragment of SEQ ID NO: 28.
19 . The method of claim 17 , wherein said TET polypeptide comprising said polypeptide having at least 90% sequence identity to SEQ ID NO: 28 comprises SEQ ID NO: 1 or a variant thereof.
20 . The method of claim 18 , wherein said catalytically active fragment of said TET polypeptide comprising said polypeptide having at least 90% sequence identity to said catalytically active fragment of SEQ ID NO: 28 comprises SEQ ID NO: 1 or a variant thereof.
21 . The method of claim 17 , wherein said TET polypeptide comprising said polypeptide having at least 90% sequence identity to SEQ ID NO: 28 comprises a conservatively-substituted variant of SEQ ID NO: 28.
22 . The method of claim 18 , wherein said catalytically active fragment of said TET polypeptide comprising said polypeptide having at least 90% sequence identity to said catalytically active fragment of SEQ ID NO: 28 comprises a conservatively-substituted catalytically active fragment of SEQ ID NO: 28.
23 . The method of claim 15 , wherein said TET2 polypeptide comprises SEQ ID NO: 28.
24 . The method of claim 16 , wherein said catalytically active fragment of said TET2 polypeptide comprises a catalytically active fragment of SEQ ID NO: 28.
25 . The method of claim 1 , wherein said modified hydroxymethylated cytosine residue comprising said hydroxymethylation comprises a hydroxymethylcytosine ether.
Assignments (5)
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded Feb 10, 2025
From: RAO, ANJANA; TAHILIANI, MAMTA; KOH, KIAN PENG
To: IMMUNE DISEASE INSTITUTE, INC.
Reel/Frame 070157/0817 →
MERGER
Recorded Feb 10, 2025
From: IMMUNE DISEASE INSTITUTE, INC.
To: CHILDREN'S HOSPITAL CORPORATION, THE
Reel/Frame 070157/0820 →
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded Feb 10, 2025
From: IYER, ARAVIND
To: THE UNITED STATES OF AMERICA, AS REPRESENTED BY THE SECRETARY, DEPARTMENT OF HEALTH AND HUMAN SERVICES
Reel/Frame 070157/0839 →
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded Feb 10, 2025
From: AGARWAL, SUNEET
To: CHILDREN'S MEDICAL CENTER CORPORATION
Reel/Frame 070157/0845 →
CHANGE OF NAME
Recorded Feb 10, 2025
From: CHILDREN'S HOSPITAL CORPORATION, THE
To: CHILDREN'S MEDICAL CENTER CORPORATION
Reel/Frame 070157/0855 →
Continuity (13)
Continuation
17350181
· Jun 17, 2021
Continuation
13120861
Provisional Application
61121844
· Dec 11, 2008
Provisional Application
61100995
· Sep 29, 2008
Provisional Application
61100503
· Sep 26, 2008
References Cited (400)
US 4115538A
· Satoh et al.
· 1978
[cited by applicant]
US 4889798A
· Rabbani
· 1989
[cited by applicant]
US 5241060A
· Engelhardt et al.
· 1993
[cited by applicant]
US 5260433A
· Engelhardt et al.
· 1993
[cited by applicant]
US 5405760A
· Raleigh et al.
· 1995
[cited by applicant]
US 7220854B1
· Engelhardt et al.
· 2007
[cited by applicant]
US 7399614B2
· Zon
· 2008
[cited by applicant]
US 8257950B2
· Berlin et al.
· 2012
[cited by applicant]
US 8653007B2
· Zheng et al.
· 2014
[cited by applicant]
US 8679745B2
· Ballhause et al.
· 2014
[cited by applicant]
US 8741567B2
· He et al.
· 2014
[cited by applicant]
US 8822146B2
· Klimasauskas et al.
· 2014
[cited by applicant]
US 8889352B2
· Klimasauskas et al.
· 2014
[cited by applicant]
US 8951736B2
· Schmidt
· 2015
[cited by applicant]
US 8962246B2
· Ballhause et al.
· 2015
[cited by applicant]
US 8969061B2
· Zhu et al.
· 2015
[cited by applicant]
US 9029087B2
· Zheng et al.
· 2015
[cited by applicant]
US 9034597B2
· Bitinaite et al.
· 2015
[cited by applicant]
US 9040239B1
· Zheng et al.
· 2015
[cited by applicant]
US 9115386B2
· Rao et al.
· 2015
[cited by applicant]
US 9121061B2
· Vaisvila et al.
· 2015
[cited by applicant]
US 9145580B2
· Feehery et al.
· 2015
[cited by applicant]
US 9150918B2
· Turner et al.
· 2015
[cited by applicant]
US 9175338B2
· Flusberg et al.
· 2015
[cited by applicant]
US 9175341B2
· Flusberg et al.
· 2015
[cited by applicant]
US 9175348B2
· Korlach et al.
· 2015
[cited by applicant]
US 9200260B2
· Correa, Jr. et al.
· 2015
[cited by applicant]
US 9200316B2
· Zheng et al.
· 2015
[cited by applicant]
US 9238836B2
· Korlach et al.
· 2016
[cited by applicant]
US 9243233B2
· Rimseliene et al.
· 2016
[cited by applicant]
US 9267117B2
· Guan et al.
· 2016
[cited by applicant]
US 9290807B2
· Booth et al.
· 2016
[cited by applicant]
US 9297806B2
· Yegnasubramanian et al.
· 2016
[cited by applicant]
US 9347093B2
· Klimasauskas et al.
· 2016
[cited by applicant]
US 9447452B2
· Rao et al.
· 2016
[cited by applicant]
US 9464277B2
· Zheng et al.
· 2016
[cited by applicant]
US 9505797B2
· Klimasauskas et al.
· 2016
[cited by applicant]
US 9546400B2
· Turner et al.
· 2017
[cited by applicant]
US 9567633B2
· Gao et al.
· 2017
[cited by applicant]
US 9611510B2
· He et al.
· 2017
[cited by applicant]
US 9650675B2
· Rimseliene et al.
· 2017
[cited by applicant]
US 9677128B2
· Robertson et al.
· 2017
[cited by applicant]
US 9816986B2
· Rao et al.
· 2017
[cited by applicant]
US 9822394B2
· Ost et al.
· 2017
[cited by applicant]
US 9879315B2
· Summerer et al.
· 2018
[cited by applicant]
US 9915655B2
· Bensimon et al.
· 2018
[cited by applicant]
US 9988673B2
· Klimasauskas et al.
· 2018
[cited by applicant]
US 20030017454A1
· Sukumar et al.
· 2003
[cited by applicant]
US 20030211522A1
· Landes et al.
· 2003
[cited by applicant]
US 20040048279A1
· Olek et al.
· 2004
[cited by applicant]
US 20040132026A1
· Olek
· 2004
[cited by applicant]
US 20050153296A1
· Berlin
· 2005
[cited by applicant]
US 20050245737A1
· Cummings
· 2005
[cited by applicant]
US 20060183128A1
· Berlin et al.
· 2006
[cited by applicant]
US 20070026393A1
· Berlin et al.
· 2007
[cited by applicant]
US 20070243161A1
· Olek
· 2007
[cited by applicant]
US 20070269824A1
· Albrecht et al.
· 2007
[cited by applicant]
US 20100167942A1
· Zheng et al.
· 2010
[cited by applicant]
US 20100197510A1
· Spain et al.
· 2010
[cited by applicant]
US 20120064521A1
· Yen et al.
· 2012
[cited by applicant]
US 20130230856A1
· Schneider et al.
· 2013
[cited by applicant]
US 20140030727A1
· Pfeifer et al.
· 2014
[cited by applicant]
US 20140178873A1
· Brachmann et al.
· 2014
[cited by applicant]
US 20140179564A1
· Korlach et al.
· 2014
[cited by applicant]
US 20140272970A1
· Zegzouti et al.
· 2014
[cited by applicant]
US 20150056616A1
· He et al.
· 2015
[cited by applicant]
US 20150240310A1
· Bitinaite et al.
· 2015
[cited by applicant]
US 20150285807A1
· Shi et al.
· 2015
[cited by applicant]
US 20150307542A1
· Roy et al.
· 2015
[cited by applicant]
US 20160046981A1
· Correa, Jr. et al.
· 2016
[cited by applicant]
US 20160186207A1
· Reik et al.
· 2016
[cited by applicant]
US 20160194696A1
· Guan et al.
· 2016
[cited by applicant]
US 20160222448A1
· Horvath
· 2016
[cited by applicant]
US 20160304552A1
· Roy et al.
· 2016
[cited by applicant]
US 20170051354A1
· Davis et al.
· 2017
[cited by applicant]
US 20170067093A1
· Klimasauskas et al.
· 2017
[cited by applicant]
US 20170175085A1
· Rao et al.
· 2017
[cited by applicant]
US 20170175129A1
· Roy et al.
· 2017
[cited by applicant]
US 20170176420A1
· Rao et al.
· 2017
[cited by applicant]
US 20170219589A1
· Rao et al.
· 2017
[cited by applicant]
US 20170283863A1
· Robertson et al.
· 2017
[cited by applicant]
US 20170283870A1
· Ost et al.
· 2017
[cited by applicant]
US 20170298422A1
· Song et al.
· 2017
[cited by applicant]
US 20180044632A1
· Rao et al.
· 2018
[cited by applicant]
US 20180044633A1
· Rao et al.
· 2018
[cited by applicant]
US 20180119225A1
· Rao et al.
· 2018
[cited by applicant]
US 20180179587A1
· Rao et al.
· 2018
[cited by applicant]
US 20180223332A1
· Ost et al.
· 2018
[cited by applicant]
US 20180224434A9
· Rao et al.
· 2018
[cited by applicant]
US 20180237839A1
· Rao et al.
· 2018
[cited by applicant]
US 20180251815A1
· Okamoto et al.
· 2018
[cited by applicant]
US 20180258149A1
· Motz et al.
· 2018
[cited by applicant]
US 20180327855A1
· Ebenstein et al.
· 2018
[cited by applicant]
US 20190048407A1
· Rao et al.
· 2019
[cited by applicant]
US 20190055593A1
· Rao et al.
· 2019
[cited by applicant]
US 20200040381A1
· Rao et al.
· 2020
[cited by applicant]
US 20200087715A1
· Rao et al.
· 2020
[cited by applicant]
US 20200087716A1
· Rao et al.
· 2020
[cited by applicant]
US 20210230679A1
· Rao et al.
· 2021
[cited by applicant]
US 20220090176A1
· Rao et al.
· 2022
[cited by applicant]
US 20240191284A1
· Rao et al.
· 2024
[cited by applicant]
US 20250019747A1
· Rao et al.
· 2025
[cited by applicant]
CN 105648537A
· 2016
[cited by applicant]
EP 1394173A1
· 2004
[cited by applicant]
EP 1568786A2
· 2005
[cited by applicant]
EP 1614149A4
· 2006
[cited by applicant]
EP 1394173B9
· 2008
[cited by applicant]
EP 2292797A1
· 2011
[cited by applicant]
EP 2292797B1
· 2013
[cited by applicant]
EP 1614149B1
· 2013
[cited by applicant]
EP 2414528B1
· 2014
[cited by applicant]
EP 2414527B1
· 2015
[cited by applicant]
EP 2470675B1
· 2016
[cited by applicant]
EP 3053585A1
· 2016
[cited by applicant]
EP 3061764A1
· 2016
[cited by applicant]
EP 2737085B1
· 2016
[cited by applicant]
EP 2825645B1
· 2016
[cited by applicant]
EP 2376632B1
· 2016
[cited by applicant]
EP 3124605A1
· 2017
[cited by applicant]
EP 2776575B1
· 2017
[cited by applicant]
EP 3013979B1
· 2017
[cited by applicant]
EP 3214183A1
· 2017
[cited by applicant]
EP 2630257B1
· 2017
[cited by applicant]
EP 2694686B1
· 2017
[cited by applicant]
EP 2948774B1
· 2018
[cited by applicant]
GB 1603778A
· 1981
[cited by applicant]
WO 2006109300A1
· 2006
[cited by applicant]
WO 2008150853A1
· 2008
[cited by applicant]
WO 2009092035A2
· 2009
[cited by applicant]
WO WO2010037001A2
· 2010
[cited by examiner]
WO 2013017853A2
· 2013
[cited by applicant]
WO 2014083118A1
· 2014
[cited by applicant]
WO 2015124955A1
· 2015
[cited by applicant]
WO 2015145133A1
· 2015
[cited by applicant]
WO 2016016639A1
· 2016
[cited by applicant]
WO 2016034908A1
· 2016
[cited by applicant]
WO 2016063034A1
· 2016
[cited by applicant]
WO 2016063059A1
· 2016
[cited by applicant]
WO 2016079509A1
· 2016
[cited by applicant]
WO 2016170319A1
· 2016
[cited by applicant]
WO 2016189288A1
· 2016
[cited by applicant]
WO 2017190215A1
· 2017
[cited by applicant]
WO 2018129120A1
· 2018
[cited by applicant]
WO 2018165459A1
· 2018
[cited by applicant]
Hayatsu et al., “Reaction of bisulfite with the 5-hydroxymethyl group in pyrimidines and in phage DNAs”, Biochemistry 18(4) 632-637 (1979).
[cited by applicant]
Hayatsu et al., “Reaction of sodium bisulfite with uracil, cytosine, and their derivatives”, Biochemistry 9(14) 2858-2865 (1970).
[cited by applicant]
He et al., “Tet-mediated formation of 5-carboxylcytosine and its excision by TDG in mammalian DNA”, Science 333 (6047) 1303-1307 (2011).
[cited by applicant]
Hegde et al., “Early steps in the DNA base excision/single-strand interruption repair pathway in mammalian cells”, Cell Res 18(1) 27-47 (2008).
[cited by applicant]
Hochedlinger et al., “Nuclear reprogramming and pluripotency”, Nature 441(7097) 1061-1067 (2006).
[cited by applicant]
International Search Report with Written Opinion for PCT/US2009/058562, dated May 20, 2010.
[cited by applicant]
Ito et al., “Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine”, Science 333(6047) 1300-1303 (2011).
[cited by applicant]
Jabbari et al., “Evolutionary changes in CpG and methylation levels in the genome of vertebrates”, Gene 205(1-2) 109-118 (1997).
[cited by applicant]
Jiricny et al., “DNA Cytosine demethylation: are we getting close?”, Cell 135(7) 1167-1169 (2008).
[cited by applicant]
Johnson et al., “5-Hydroxymethylcytosine localizes to enhancer elements and is associated with survival in glioblastoma patients”, Nat Commun 7: 13177 (2016).
[cited by applicant]
Kangaspeska et al., “Transient cyclical methylation of promoter DNA”, Nature 452(7183) 112-115 (2008).
[cited by applicant]
Kawasaki et al., “A Novel method for the simultaneous identification of methylcytosine and hydroxymethylcytosine at a single base resolution”, Nucleic Acids Res 45(4) e24 (2017).
[cited by applicant]
Kim, et al., “CREB/ATF-dependent T cell receptor-induced FoxP3 gene expression: a role for DNA methylation”, The Journal of Experimental Medicine 204(7):1543-1551 (2007).
[cited by applicant]
Kothari et al., “5-Methylcytosine content in the vertebrate deoxyribonucleic acids: species specificity”, J Mol Evol 7 (4) 625-629 (1976).
[cited by applicant]
Kriaucionis et al., “The nuclear DNA base 5-hydroxymethylcytosine is present in Purkinje neurons and the brain”, Science 324(5929) 929-930 (2009).
[cited by applicant]
Kriukiene et al., “5-Hydroxymethylcytosine—the elusive epigenetic mark in mammalian DNA”, Chem Soc Rev 41(21) 6916-6930 (2012).
[cited by applicant]
Lariviere et al., “Crystal structures of the T4 phage beta-glucosyltransferase and the D100A mutant in complex with UDP-glucose: glucose binding and identification of the catalytic base for a direct displacement mechani…
[cited by applicant]
Lariviere et al., “Structural evidence of a passive base-flipping mechanism for AGT, an unusual GT-B glycosyltransferase”, J Mol Biol 352(1) 139-150 (2005).
[cited by applicant]
Lariviere et al., “Structural evidence of a passive base-flipping mechanism for beta-glucosyltransferase”, J Biol Chem 279(33) 34715-34720 (2004).
[cited by applicant]
Lee et al., “Th2 lineage commitment and efficient IL-4 production involves extended demethylation of the IL-4 gene”, Immunity 16(5) 649-660 (2002).
[cited by applicant]
Leonhardt et al., “A targeting sequence directs DNA methyltransferase to sites of DNA replication in mammalian nuclei”, Cell 71(5) 865-873 (1992).
[cited by applicant]
Li et al., “Mechanisms and functions of DNA mismatch repair”, Cell Res 18(1) 85-98 (2008).
[cited by applicant]
Loenarz et al., “Expanding chemical biology of 2-oxoglutarate oxygenases”, Nat Chem Biol 4(3) 152-156 (2008).
[cited by applicant]
Lorsbach et al., “TET1, a member of a novel protein family, is fused to MLL in acute myeloid leukemia containing the t(10;11)(q22;q23)”, Leukemia 17(3) 637-641 (2003).
[cited by applicant]
Lu et al., “TET family proteins: oxidation activity, interacting molecules, and functions in diseases.”, Chem Rev 115(6) 2225-2239 (2015).
[cited by applicant]
Luscombe et al., “An overview of the structures of protein-DNA complexes”, Genome Biol 1(1) REVIEWS001 (2000).
[cited by applicant]
Mayer et al., “Demethylation of the zygotic paternal genome”, Nature 403(6769) 501-502 (2000).
[cited by applicant]
Mellen et al., “MeCP2 binds to 5hmC enriched within active genes and accessible chromatin in the nervous system”, Cell 151(7) 1417-1430 (2012).
[cited by applicant]
Metivier et al., “Cyclical DNA methylation of a transcriptionally active promoter”, Nature 452(7183) 45-50 (2008).
[cited by applicant]
Morera et al., “T4 phage beta-glucosyltransferase: substrate binding and proposed catalytic mechanism”, J Mol Biol 292(3) 717-730 (1999).
[cited by applicant]
Morris, “Liquid”, Academic Press Dictionary of Science and Technology (4th Ed.), Elsevier Science & Technology (1992).
[cited by applicant]
Netto et al., “The iron-catalyzed oxidation of dithiothreitol is a biphasic process: hydrogen peroxide is involved in the initiation of a free radical chain of reactions”, Arch Biochem Biophys 333(1) 233-242 (1996).
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 13/120,861, dated Apr. 22, 2015.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 13/795,739, dated Apr. 22, 2016.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 15/440,284, dated Apr. 4, 2018.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 15/440,408, dated Sep. 18, 2017.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 15/440,408, dated Aug. 23, 2017.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 15/440,826, dated Jun. 1, 2018.
[cited by applicant]
Notice of Allowance issued during the Prosecution of U.S. Appl. No. 15/193,796, dated Oct. 16, 2018.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 15/440,826, dated Nov. 22, 2017.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 13/120,861, dated Mar. 13, 2013.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 13/795,739, dated Mar. 24, 2014.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 14/235,707, dated Aug. 4, 2015.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 14/363,442, dated Nov. 10, 2016.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 14/363,442, dated Jul. 11, 2017.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 14/363,442, dated Mar. 7, 2016.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 14/648,527, dated Jan. 18, 2018.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/054,227, dated Oct. 3, 2016.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/193,796, dated Jun. 20, 2018.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/341,344, dated Apr. 16, 2018.
[cited by applicant]
Alegria et al., “Hydroxymethylation of pyrimidine mononucleotides with formaldehyde”, Biochim Biophys Acta 149(2) 317-324 (1967).
[cited by applicant]
Allen et al., “Solution structure of the nonmethyl-CpG-binding CXXC domain of the leukaemia-associated MLL histone methyltransferase”, EMBO J 25(19) 4503-4512 (2006).
[cited by applicant]
Aller et al., “A structural rationale for stalling of a replicative DNA polymerase at the most common oxidative thymine lesion, thymine glycol”, Proc Natl Acad Sci USA 104(3) 814-818 (2007).
[cited by applicant]
An et al., “TET family dioxygenases and DNA demethylation in stem cells and cancers”, Exp Mol Med 49(4) e323 (2017).
[cited by applicant]
Aravind et al., “The DNA-repair protein AlkB, EGL-9, and leprecan define new families of 2-oxoglutarate- and iron-dependent dioxygenases”, Genome Biol 2(3): 1-8 (2001).
[cited by applicant]
Arita et al., “Recognition of hemi-methylated DNA by the SRA protein UHRF1 by a base-flipping mechanism”, Nature 455(7214) 818-822 (2008).
[cited by applicant]
Avvakumov et al., “Structural basis for recognition of hemi-methylated DNA by the SRA domain of human UHRF1”, Nature 455(7214) 822-826 (2008).
[cited by applicant]
Bird et al., “DNA methylation patterns and epigenetic memory”, Genes Dev 16(1) 6-21 (2002).
[cited by applicant]
Blelloch et al., “Reprogramming efficiency following somatic cell nuclear transfer is influenced by the differentiation and methylation state of the donor nucleus”, Stem Cells 24(9) 2007-2013 (2006).
[cited by applicant]
Booth et al., “Oxidative bisulfite sequencing of 5-methylcytosine and 5-hydroxymethylcytosine”, Nat Protoc 8(1) 1841-1851 (2013).
[cited by applicant]
Booth et al., “Quantitative sequencing of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution”, Science 336(6083) 934-937 (2012).
[cited by applicant]
Borst et al., “Base J: discovery, biosynthesis, and possible functions”, Annu Rev Microbiol 62: 235-251 (2008).
[cited by applicant]
Breton et al., “Structures and mechanisms of glycosyltransferases”, Glycobiology 16(2) 29R-37R (2006).
[cited by applicant]
Bullard et al., “Identification of the glucosyltransferase that converts hydroxymethyluracil to base J in the trypanosomatid genome”, J Biol Chem 289(29) 20273-20282 (2014).
[cited by applicant]
Cannon et al., “5-Hydroxymethylcytosine DNA glycosylase activity in mammalian tissue”, Biochem Biophys Res Commun 151(3) 1173-1179 (1988).
[cited by applicant]
Castro et al. “5-Methylcytosine attack by hydroxyl free radicas and during carbon tetrachloride promoted liver microsomal lipid peroxidation.” Chemico-Biological Interactions 99(1-3): 289-299 (1996).
[cited by applicant]
Cedar et al., “Gene expression. The amazing demethylase”, Nature 397(6720) 568-569 (1999).
[cited by applicant]
Chuang et al., “Human DNA-(cytosine-5) methyltransferase-PCNA complex as a target for p21WAF1”, Science 277 (5334) 1996-2000 (1997).
[cited by applicant]
Clark et al., “Enhanced 5-methylcytosine detection in single-molecule, real-time sequencing via Tet1 oxidation”, BMC Biol 11:4 (2013).
[cited by applicant]
Cliffe et al., “JBP1 and JBP2 are two distinct thymidine hydroxylases involved in J biosynthesis in genomic DNA of African trypanosomes”, Nucleic Acids Res 37(5) 1452-1462 (2009).
[cited by applicant]
Coulter et al., “Hydroquinone increases 5-hydroxymethylcytosine formation through ten eleven translocation 1 (TET1) 5-methylcytosine dioxygenase”, The Journal of Biological Chemistry 288(40):28792-28800 (2013).
[cited by applicant]
Dai et al., “Evaluation of UDP-GlcN derivatives for selective labeling of 5-(hydroxymethyl)cytosine”, Chembiochem 14(16) 2144-2152 (2013).
[cited by applicant]
De Kort et al., “Chemical and Enzymatic Synthesis of DNA Fragments Containing 5-(β-D-Glucopyranosyloxymethyl)-2′-deoxycytidine—a Modified Nucleoside in T4 Phage DNA”, European Journal of Organic Chemistry 2075-2082 (200…
[cited by applicant]
De Waard et al., “On the specificity of bacteriophage-induced hydroxymethylcytosine glucosyltransferases. II. Specificities of hydroxymethylcytosine alphaand beta-glucosyltransferases induced by bacteriophage T4”, Eur J…
[cited by applicant]
Delhommeau et al., “TET2 Is a Novel Tumor Suppressor Gene Inactivated in Myeloproliferative Neoplasms: Identification of a Pre-JAK2 V617F Event” Blood 112(11) 1ba-3 (2008). Paper presented at the Americna Society of Hem…
[cited by applicant]
Ehrlich et al., “5-Methylcytosine in eukaryotic DNA”, Science 212(4501) 1350-1357 (1981).
[cited by applicant]
Esteller et al., “Cancer epigenomics: DNA methylomes and histone-modification maps”, Nat Rev Genet 8(4) 286-298 (2007).
[cited by applicant]
Esteller et al., “Epigenetics in cancer”, N Engl J Med 358(11) 1148-1159 (2008).
[cited by applicant]
Farthing et al., “Global mapping of DNA methylation in mouse promoters reveals epigenetic reprogramming of pluripotency genes”, PLoS Genet 4(6) e1000116 1-8 (2008).
[cited by applicant]
Ficz et al., “Reprogramming by cell fusion: boosted by Tets”, Mol Cell 49(6) 1017-1018 (2013).
[cited by applicant]
Flaks et al., “Virus-induced acquisition of metabolic function. I. Enzymatic formation of 5-hydroxymethyldeoxycytidylate”, J Biol Chem 234(6) 1501-1506 (1959).
[cited by applicant]
Flusberg et al., “Direct detection of DNA methylation during single-molecule, real-time sequencing”, Nat Methods 7(6) 461-465 (2010).
[cited by applicant]
Fraga et al., “DNA methylation: a profile of methods and applications”, Biotechniques 33(3) 632, 634, 636-649 (2002).
[cited by applicant]
Franco et al., “Oxidative stress, DNA methylation and carcinogenesis”, Cancer Lett 266(1) 6-11 (2008).
[cited by applicant]
Frommer et al., “A genomic sequencing protocol that yields a positive display of 5-methylcytosine residues in individual DNA strands”, Proc Natl Acad Sci USA 89(5) 1827-1831 (1992).
[cited by applicant]
Fu et al., “Nucleic acid modifications with epigenetic significance”, Curr Opin Chem Biol 16(5-6) 516-524 (2012).
[cited by applicant]
Fukushige et al., “Methyl-CpG targeted transcriptional activation allows re-expression of tumor suppressor genes in human cancer cells”, Biochem Biophys Res Commun 377(2) 600-605 (2008).
[cited by applicant]
Gal-Yam et al., “Cancer epigenetics: modifications, screening, and therapy”, Annu Rev Med 59: 267-280 (2008).
[cited by applicant]
Geijsen et al., “Derivation of embryonic germ cells and male gametes from embryonic stem cells”, Nature 427(6970) 148-154 (2004).
[cited by applicant]
Globisch et al., “Tissue distribution of 5-hydroxymethylcytosine and search for active demethylation intermediates”, PLoS One 5(12) e15367 (2010).
[cited by applicant]
Goll et al., “Eukaryotic cytosine methyltransferases”, Annu Rev Biochem 74: 481-514 (2005).
[cited by applicant]
Goode et al. “Identification of Promiscuous Small Molecule Activators in High-Throughput Enzyme Activation Screens.” Journal of Medicinal Chemistry 51(8): 2346-2349 (2008).
[cited by applicant]
Green, “General PCR Protocol”, http://labs.mcbd.lsa.umich.edu/labs/maddock/protocols/PCR/general_pcr_protocol.html, 2 pages, 1996.
[cited by applicant]
Gros et al., “The major human AP endonuclease (Ape1) is involved in the nucleotide incision repair pathway”, Nucleic Acids Res 32(1) 73-81 (2004).
[cited by applicant]
Gruenbaum et al., “Methylation of CpG sequences in eukaryotic DNA”, FEBS Lett 124(1) 67-71 (1981).
[cited by applicant]
Gruenewald et al., “The role of antigenic determinants in the control of IgM and IgG antibody responses to denatured DNA”, J Immunol 111(1) 106-113 (1973).
[cited by applicant]
Guerrero et al., “Determination of 5-methyl-cytosine and cytosine in tumor DNA of cancer patients”, Electrophoresis 26(6) 1057-1062 (2005).
[cited by applicant]
Gurdon et al., “From nuclear transfer to nuclear reprogramming: the reversal of cell differentiation”, Annu Rev Cell Dev Biol 22: 1-22 (2006).
[cited by applicant]
Hajkova et al., “Chromatin dynamics during epigenetic reprogramming in the mouse germ line”, Nature 452(7189) 877-881 (2008).
[cited by applicant]
Hannum et al., “Genome-wide methylation profiles reveal quantitative views of human aging rates”, Mol Cell 49(2) 259-267 (2013).
[cited by applicant]
Vrielink et al., “Crystal structure of the DNA modifying enzyme beta-glucosyltransferase in the presence and absence of the substrate uridine diphosphoglucose”, EMBO J 13(15) 3413-3422 (1994).
[cited by applicant]
Wang et al. “Antibody Structure, Instability, and Formulation.” Journal of Pharmaceutical Sciences 96(1): 1-26 (2007).
[cited by applicant]
Wang et al., “Chemoenzymatic synthesis and antibody detection of DNA glycoconjugates”, Bioconjug Chem 14(6) 1314-1322 (2003).
[cited by applicant]
Wang et al., “Comparison of bisulfite modification of 5-methyldeoxycytidine and deoxycytidine residues”, Nucleic Acids Res 8(20) 4777-4790 (1980).
[cited by applicant]
Wu et al., “Mechanisms and functions of Tet protein-mediated 5-methylcytosine oxidation”, Gene Dev 25(23) 2436-2452 (2011).
[cited by applicant]
Wyatt et al., “The bases of the nucleic acids of some bacterial and animal viruses: the occurrence of 5-hydroxymethylcytosine”, Biochem 55(5) 774-782 (1953).
[cited by applicant]
Xia et al., “Bisulfite-free, base-resolution analysis of 5-formylcytosine at the genome scale”, Nat Methods 12(11) 1047-1050 (2015).
[cited by applicant]
Xiong et al., “Cooperative Action between SALL4A and TET Proteins in Stepwise Oxidation of 5-Methylcytosine”, Mol Cell 64(5) 913-925 (2016).
[cited by applicant]
Yu et al., “Base-resolution analysis of 5-hydroxymethylcytosine in the mammalian genome”, Cell 149(6) 1368-1380 (2012).
[cited by applicant]
Yu et al., “Induced Pluripotent Stem Cell Lines Derived from Human Somatic Cells”, Science 318(5858) 1917-1920 (2007).
[cited by applicant]
Yu et al., “Tet-assisted bisulfite sequencing of 5-hydroxymethylcytosine”, Nat Protoc 7(12) 2159-2170 (2012).
[cited by applicant]
Yu et al., “The protein that binds to DNA base J in trypanosomatids has features of a thymidine hydroxylase”, Nucleic Acids Res 35(7) 2107-2115 (2007).
[cited by applicant]
Zhu et al., “5-methylcytosine-DNA glycosylase activity is present in a cloned G/T mismatch DNA glycosylase associated with the chicken embryo DNA demethylation complex”, Proc Natl Acad Sci USA 97(10) 5135-5139 (2000).
[cited by applicant]
Bair et al., “Exclusion of Glucosyl-Hydroxymethylcytosine DNA Containing Bacteriophages”, J. Mol. Biol. 366(3) 779-789 (2007).
[cited by applicant]
Comeau et al., “Modular architecture of the T4 phage superfamily: A conserved core genome and a plastic periphery”, Virology 362 384-396 (2007).
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 16/012,280 dated Aug. 27, 2018.
[cited by applicant]
Petrov et al., “Plasticity of the Gene Functions for DNA Replication in the T4-like Phages”, J. Mol. Biol. 361 46-68 (2006).
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/440,319, dated Oct. 26, 2018.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/952,352, dated Nov. 14, 2018.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/341,344, dated Dec. 17, 2018.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/725,917, dated, Dec. 27, 2018.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 15/440,815; Dated Jan. 10, 2019.
[cited by applicant]
Li et al., “CD4 CD25+ regulatory T-Cell lines from human cord blood have functional and molecular properties of T-Cell anergy”, Blood 106(9) 3068-3073 (2005).
[cited by applicant]
Office Action issued duing the prosecution of U.S. Appl. No. 15/483,282; Dated Feb. 13, 2019.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 16/012,280; Dated Feb. 13, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 16/169,801; Dated Feb. 15, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 16/172,369; Dated Feb. 19, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 16/175,353; Dated Mar. 6, 2019.
[cited by applicant]
Hori et al., “Identification of high excision capacity for 5-hydroxymethyluracil mispaired with guanine in DNA of
[cited by applicant]
GenBank “
[cited by applicant]
GenBank “PREDICTED:
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/890,034; Dated Mar. 7, 2019.
[cited by applicant]
Notice of Allowance during the prosecution of U.S. Appl. No. 16/012,510; Dated Mar. 13, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 16/180,450; Dated Mar. 19, 2019.
[cited by applicant]
Notice of Allowance during the prosecution of U.S. Appl. No. 15/193,796; Dated Apr. 16, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/722,183; Dated Apr. 24, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/440,424; dated May 1, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/722,202; dated May 2, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/725,917; dated May 14, 2019.
[cited by applicant]
Notice of Corrected Allowability during the prosecution of U.S. Appl. No. 15/440,815; dated May 15, 2019.
[cited by applicant]
Chun-Xiao Song et al., “The hunt for 5-hydroxymethylcytosine: the sixth base”, Epigenomics, vo. 3, No. 5, 521-523, Oct. 1, 2011.
[cited by applicant]
European Search Report dated Oct. 10, 2018 for EP Application No. 18174572.
[cited by applicant]
Saori Takahashi et al., “A novel method to analyze 5-hydroxymethylcytosine in CpG sequences using maintenance DNA methyltransferase, DNMT1”, FEBS Open Bio, vol. 5, No. 1, 741-747, Jan. 1, 2015.
[cited by applicant]
Notice of Allowance during the prosecution of U.S. Appl. No. 16/012,510; Dated Jul. 15, 2019.
[cited by applicant]
Notice of Allowance during the prosecution of U.S. Appl. No. 16/169, 801; Dated Jul. 25, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/952,352; dated Aug. 8, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 16/180,450; dated Aug. 6, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 16/172,369; dated Aug. 6, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 16/175,353; dated Aug. 15, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 16/411,998; dated Aug. 19, 2019.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 15/193,796; Dated Aug. 29, 2019.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 16/169,801; Dated Sep. 16, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/483,282; dated Oct. 7, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/483,297; dated Oct. 10, 2019.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 16/012,510; Dated Oct. 22, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/725,917; dated Oct. 24, 2019.
[cited by applicant]
Office Action issued during the prosecution of U.S. Appl. No. 15/722,202; dated Nov. 12, 2019.
[cited by applicant]
Notice of Allowance issued during the prosecution of U.S. Appl. No. 15/341,344; Dated Nov. 19, 2019.
[cited by applicant]
Alves et al. “Differential methylation of human LINE-1 retrotransposons in malignant cells.” Gene 176(1-2): 39-44 (1996).
[cited by applicant]
Bastian et al. “Prognostic value of preoperative serum cell-free circulating DNA in men with prostate cancer undergoing radical prostatectomy.” Clinical Cancer Research 13(18): 5361-5367 (2007).
[cited by applicant]
Bayer et al. “[55] The avidin-biotin complex in affinity cytochemistry.” Methods in enzymology. Academic Press Bol. 62: 308-315 (1979).
[cited by applicant]
Bogani et al. “Hypermethylation of CXCR4 promoter in CD34+ cells from patients with primary myelofibrosis.” Stem Cells 26(8): 1920-1930 (2008).
[cited by applicant]
Ciccarone et al. “DNA hydroxymethylation levels are altered in blood cells from Down syndrome persons enrolled in the MARK-AGE project.” The Journals of Gerontology: Series A 73(6): 737-744 (2018).
[cited by applicant]
Cliffe et al. “JBP1 and JBP2 proteins are Fe2+/2-oxoglutarate-dependent dioxygenases regulating hydroxylation of thymidine residues in trypanosome DNA.” Journal of Biological Chemistry 287(24): 19886-19895 (2012).
[cited by applicant]
Cross et al. “J-binding protein increases the level and retention of the unusual base J in trypanosome DNA.” Molecular microbiology 46(1): 37-47 (2002).
[cited by applicant]
Deligezer et al. “Frequent copresence of methylated DNA and fragmented nucleosomal DNA in plasma of lymphoma patients.” Clinica Chimica Acta 335(1-2): 89-94 (2003).
[cited by applicant]
Gao et al. “Integrated detection of both 5-mC and 5-hmC by high-throughput tag sequencing technology highlights methylation reprogramming of bivalent genes during cellular differentiation.” Epigenetics 8(4): 421-430 (20…
[cited by applicant]
Gilat et al. “Single-molecule quantification of 5-hydroxymethylcytosine for diagnosis of blood and colon cancers.” Clinical Epigenetics 9(70): 1-8 (2017).
[cited by applicant]
Goldstein et al. “Carbohydrate binding properties of banana (
[cited by applicant]
Gommers-Ampt et al. “A novel DNA nucleotide in Trypanosoma brucei only present in the mammalian phase of the life-cycle.” Nucleic acids research 19(8): 1745-1751 (1991).
[cited by applicant]
Gommers-Ampt et al. “β-D-glucosyl-hydroxymethyluracil: a novel modified base present in the DNA of the parasitic protozoan T. brucei.” Cell 75(6): 1129-1136 (1993).
[cited by applicant]
Gommers-Ampt et al. “The identification of hydroxymethyluracil in DNA of Trypanosoma brucei.” Nucleic acids research 21(9): 2039-2043 (1993).
[cited by applicant]
Hatfull et al. “Bacteriophages and their genomes.” Current opinion in virology 1(4): 298-303 (2011).
[cited by applicant]
Jesaitis. “Differences in the chemical composition of the phage nucleic acids.” Nature 178(4534): 637-637 (1956).
[cited by applicant]
Josse et al. “Glucosylation of Deoxyribonucleic Acid III. α-AND β-Glucosyl Transferases From T4-Infected
[cited by applicant]
Kinney et al. “Tissue-specific distribution and dynamic changes of 5-hydroxymethylcytosine in mammalian genomes.” Journal of Biological Chemistry 286(28): 24685-24693 (2011).
[cited by applicant]
Koressaar et al. “Enhancements and modifications of primer design program Primer3.” Bioinformatics 23(10): 1289-1291 (2007).
[cited by applicant]
Kornberg et al. “Glucosylation of deoxyribonucleic acid by enzymes from bacteriophage-infected
[cited by applicant]
Kuno et al. “Gentiobiose, a constituent of deoxyribonucleic acid from coliphage T6.” J Biol Chem 237: 1266-1270 (1962).
[cited by applicant]
Lehman et al. “On the structure of the glucosylated hydroxymethylcytosine nucleotides of coliphages T2, T4, and T6.” Journal of Biological Chemistry 235(11): 3254-3259 (1960).
[cited by applicant]
Leone et al. “Inhibitors of DNA methylation in the treatment of hematological malignancies and MDS.” Clinical Immunology 109(1): 89-102 (2003).
[cited by applicant]
Lisowska et al. “Preparation of biotinylated lectins and application in microtiter plate assays and Western blotting.” BioMethods 7: 115-129 (1996).
[cited by applicant]
Monod et al. “The genome of the pseudo T-even bacteriophages, a diverse group that resembles T4.” Journal of Molecular Biology 267(2): 237-249 (1997).
[cited by applicant]
Panigrahi et al. “Four related proteins of the Trypanosoma brucei RNA editing complex.” Molecular and cellular biology 21(20): 6833-6840 (2001).
[cited by applicant]
Cited By (1)