EP0375408A1
Expired / Lapsed
EP
⏱️ 0.0 years remaining
Method for making synthetic oligonucleotides which bind specifically to target sites on duplex DNA molecules, by forming a colinear triplex, the synthetic oligonucleotides and methods of use
Assignee: Baylor College of Medicine
•
Filing: 1989-12-20
•
Granted: —
•
Expires: 2009-12-20
Abstract
Abstract A method for making synthetic oligonucleotides which bind to target sequences in a duplex DNA forming colinear triplexes by binding to the major groove. The method includes scanning genomic duplex DNA and identifying nucleotide target sequences of greater than about 20 nucleotides having either about at least 65% purine bases or about at least 65% pyrimidine bases; and synthesizing synthetic oligonucleotides complementary to identified target sequences. The synthetic oligonucleotides have a G when the complementary location in the DNA duplex has a GC base pair and have a T when he complementary location in the DNA duplex has an AT base pair. The synthetic oligonucleotides are oriented 5′ to 3′ and bind parallel or 3′ to 5′ and bind anti-parallel to the about at least 65% purine strand. Also described are synthetic oligonucleotides made by the above methods. The oligonucleotides can be altered by modifying and/or changing the bases, adding linkers and modifying groups to the 5′ and/or 3′ termini, and changing the backbone. These synthetic oligonucleotides bind to duplex DNA to form triplexes. This process alters the functioning of the genes which are bound. This process can be used to inhibit cell growth, alter protein ratios, treat diseases including cancer and permanently alter the DNA.
Inventors
- 👤 Michael Edward Hogan
- 👤 Donald Joseph Kessler
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-29 20:17
CPC Classifications
C; C12; C12N; C12N15/00; C12N15/09; C12N15/11; C12N15/113; A; A61; A61P; A61P25/00; A61P25/28; A61P3/00; A61P31/00; A61P31/04; C07; C07H; C07H21/00; C07H21/04; C12N15/1137; C12N15/1138; C12Q; C12Q1/00; C12Q1/68; C12Q1/6813; C12Q1/6839; C12Q1/6876; C12Q1/6883; C12Q1/6886; C12Q1/70; C12Q1/701; C12Q1/702; C12Q1/703; C12Q1/705; C12Q1/708; C12Y; C12Y101/00; C12Y101/01; C12Y101/01034; C12Y205/00; C12Y205/01; C12Y205/01018; C12Y207/00; C12Y207/07; C12Y207/07007; A61K; A61K38/00; C12N2310/00; C12N2310/10; C12N2310/15; C12N2310/30; C12N2310/32; C12N2310/321
Claims Analysis
Total Claims: 49
Claim 1
Independent Claim
108 words
A method for making a synthetic oligonucleotide which binds to a target sequence in duplex DNA forming a colinear triplex by binding to the major groove, said method comprising the steps of: scanning genomic duplex DNA and identifying nucleotide target sequences of greater than about 20 nucleotides having either about at least 65% purine bases or about at least 65% pyrimidine bases; and synthesizing said synthetic oligonucleotide complementary to said identified target sequence, said synthetic oligonucleotide having a G when the complementary location in the DNA duplex has a GC base pair, having a T when the complementary location in the DNA duplex has an AT base pair.
Claim 2
Independent Claim
108 words
A method for making a synthetic oligonucleotide which binds to a target sequence in duplex DNA forming a colinear triplex by binding to the major groove, said method comprising the steps of: scanning genomic duplex DNA and identifying nucleotide target sequences of greater than about 20 nucleotides having either about at least 65% purine bases or about at least 65% pyrimidine bases; and synthesizing said synthetic oligonucleotide complementary to said identified target sequence, said synthetic oligonucleotide having a G when the complementary location in the DNA duplex has a GC base pair, having a T when the complementary location in the DNA duplex has an AT base pair.
Claim 3
Dependent Claim
51 words
The method of Claim 1, wherein said synthetic oligonucleotide is selected from the group consisting of an oligonucleotide oriented 3′ to 5′ and binding anti-parallel to be about at least 65% purine strand and an oligonucleotide oriented 5′ to 3′ and binding parallel to the about at least 65% purine strand.
Claim 4
Dependent Claim
142 words
A synthetic oligonucleotide for forming a colinear triplex with a target sequence in a duplex DNA when said target sequence is either about at least 65% purine bases or about at least 65% pyrimidine bases, comprising, a nucleotide sequence of at least about 20 nucleotides; said nucleotide sequence including G and T, wherein G is used when the complementary location in the duplex DNA is a GC base pair and T is used when the complementary location in the duplex DNA is an AT base pair; and said sequence selected from the group consisting of an oligonucleotide oriented 3′ to 5′ and binding anti-parallel to the about at least 65% purine strand in the duplex DNA target sequence and an oligonucleotide oriented 5′ to 3′ and binding parallel to the about at least 65% purine strand in the duplex DNA target sequence.
Claim 5
Dependent Claim
142 words
A synthetic oligonucleotide for forming a colinear triplex with a target sequence in a duplex DNA when said target sequence is either about at least 65% purine bases or about at least 65% pyrimidine bases, comprising, a nucleotide sequence of at least about 20 nucleotides; said nucleotide sequence including G and T, wherein G is used when the complementary location in the duplex DNA is a GC base pair and T is used when the complementary location in the duplex DNA is an AT base pair; and said sequence selected from the group consisting of an oligonucleotide oriented 3′ to 5′ and binding anti-parallel to the about at least 65% purine strand in the duplex DNA target sequence and an oligonucleotide oriented 5′ to 3′ and binding parallel to the about at least 65% purine strand in the duplex DNA target sequence.
Claim 6
Dependent Claim
33 words
The synthetic oligonucleotide of claim 3, wherein, at least one T is replaced with a compound selected from the group consisting of X, halogenated derivatives of X, I and halogenated derivatives of I.
Claim 7
Dependent Claim
19 words
The synthetic oligonucleotide of claim 3, wherein, at least one G is replaced with a halogenated derivative of G.
Claim 8
Dependent Claim
23 words
The synthetic oligonucleotide of claim 3, wherein, at least one base is substituted at the 2′ furanose position with a non-charged bulky group.
Claim 9
Dependent Claim
27 words
The synthetic oligonucleotide of claim 6, wherein, said non-charged bulky group is selected from the group consisting of a branched alkyl, a sugar and a branched sugar.
Claim 10
Dependent Claim
21 words
The synthetic oligonucleotide of claim 3, wherein, the backbone is a phosphodiester analogue which is not readily hydrolyzed by cellular nucleases.
Claim 11
Dependent Claim
31 words
The synthetic oligonucleotide of claim 8, wherein, said phosphodiester analogue is selected from the group consisting of phosphorothioate, phosphoroselenoate, methyl phosphate, phosphoramidite, phosphotriester and the alpha enantiomer of naturally occurring phosphodiester.
Claim 12
Dependent Claim
13 words
The synthetic oligonucleotide of claim 3, further including a linker at a terminus.
Claim 13
Dependent Claim
55 words
The synthetic oligonucleotide of claim 10, wherein, said linker is attached to the 3′ terminus and is selected from the group consisting of a base analogue with a primary amine affixed to the base plane through an alkyl linkage and a base analogue with a sulfhydryl affixed to the base plane through an alkyl linkage.
Claim 14
Dependent Claim
83 words
The synthetic oligonucleotide of claim 10, wherein said linker is attached to the 5′ terminus and is selected from the group consisting of a base analogue with a primary amine affixed to the base plane through an alkyl linkage, a base analogue with a sulfhydryl affixed to the base plane through an alkyl linkage, a long chain amine coupled directly to the 5′ hydroxyl group of the oligonucleotide and a long chain thiol coupled directly to the 5′ hydroxyl group of the oligonucleotide.
Claim 15
Dependent Claim
48 words
The synthetic oligonucleotide of claims 10, 11 or 12 further including a modifying group attached to said linker, wherein, said modifying group binds to duplex DNA and is selected from the group of molecules consisting of an intercalator, a groove-binding molecule, a cationic amine and a cationic polypeptide.
Claim 16
Dependent Claim
54 words
The synthetic oligonucleotide of claims 10, 11 or 12, further including a modifying group attached to said linker, wherein said modifying group damages DNA and is selected from the group of molecules consisting of a catalytic oxidant, nitrogen mustard, alkylator, photochemical crosslinker, photochemical sensitizer of singlet oxygen and reagent capable of direct photochemical damage.
Claim 17
Dependent Claim
23 words
The synthetic oligonucleotide of claim 14, wherein said photochemical sensitizer of singlet oxygen is eosin, methylene blue, acridine orange, or 9 amino acridine.
Claim 18
Dependent Claim
14 words
The synthetic oligonucleotide of claim 14, wherein said reagent is ethidium or pyrene derivatives.
Claim 19
Dependent Claim
47 words
A method of inhibiting the growth of cells, comprising the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular uptake and binding to the target sequence, wherein said target sequence is positioned within the DNA domain adjacent to the RNA transcription origin.
Claim 20
Dependent Claim
21 words
The method of claim 17, wherein the cells are cancerous cells and the synthetic oligonucleotide is specific to the C-myc gene.
Claim 21
Dependent Claim
28 words
The method of claim 18, wherein the synthetic oligonucleotide is selected from the group consisting of: 3′-TGGTGTGTGGGTTTTGTGGGGGGTGGGGGGGTTTTTTTTGGGTGGG-5′ 5′-GTGGTGGGGTGGTTGGGGTGGGTGGGGTGGGTGGGGT-3, 3′-TGTGGTGGGGTGGTTGGGGTGGGTGGGGTGGGTGGG-5′, 5′-TTTGGTGTGGGGGTGGGGGTTTTGTTTTTTGT-3′, 3′-GGTTGGGGTGGGTGGGGTGGGTGGGGT-5′, 5′-GGTTGGGGTGGGTGGGGTGGGTGGGGT-3′, 5′-TTTGGTGTGGGGGTGGGGGTTTTGTTTTTTGT-3′ and fragments and analogues thereof.
Claim 22
Dependent Claim
14 words
The method of claim 19, wherein, said oligonucleotide includes a linker and modifying group.
Claim 23
Dependent Claim
45 words
A method of inhibiting the growth of pathogens comprising the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular uptake and binding to the target sequence, wherein said sequence binds within the nucleic acid domain adjacent the RNA transcription origin.
Claim 24
Dependent Claim
21 words
The method of claim 21, wherein the pathogen is HIV-1 virus and the synthetic oligonucleotide is within the viral LTR region.
Claim 25
Dependent Claim
31 words
The method of claim 22, wherein the synthetic oligonucleotide is selected from the group consisting of: 5′-TTTTGTTTTGGGGGGTGTGGTTGGG-5′, 3′-TTTTGTTTTGGGGGGTGTGGTTGGG-5′, 5′-TGTGTTGGTTGTTGTGGGGTTTGTTGGTGTGTT-3′, 3′-TGTGTTGGTTGTTGTGGGGTTTGTTGGTGTGTT-5′, 5′-GGGTTGGTGGTGGGGGTGTTTGTTG-3′, 3′-GGGTTGGTGGTGGGGGTGTTTGTTG-5′, 3′-TGGGTGGGGTGGGGTGGGGGGGTGTGCCCTCTGGGG-5′, 5′-GTTTTTGGGTGTTGTGGGTGTGTGTGGTTTG-3′, 3′-GTTTTTGGGTGTTGTGGGTGTGTGTGGTT-5′, 5′-TGGGTGGGGTGGGGTGGGGGGGTGTGGGGTGTGGGGTG-3′ and fragments and analogues thereof.
Claim 26
Dependent Claim
14 words
The method of claim 23, wherein, said oligonucleotide includes a linker and modifying group.
Claim 27
Dependent Claim
34 words
A method of manipulating the structural protein content of epidermal tissue comprising the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular uptake and binding to the target sequence.
Claim 28
Dependent Claim
29 words
The method of claim 25 for inhibiting a collagen gene, wherein the synthetic oligonucleotide is selected from the group consisting of: 3′-TGGGTTGGGTGGTGGTGGGGGTGTGGTTTGGTTGTGGGTTTTT-5′, 3′-GGGTTGGGTGTGGTTTGGGGTGGGGTTTGG-5′, 3′-GTGGGTTGGGTGGTGGTGGGGGTGTGGTTTGG-5′ and fragments and analogues thereof.
Claim 29
Dependent Claim
28 words
The method of claim 25, for inhibiting a collagenase gene, wherein the synthetic oligonucleotide is selected from the group consisting of 5′GGTTGGGGTTGGTGTGTTTTTTTTGTGTGGGTG-3′, 5′-TTGTGGTTGTTTTTTTGGTTGTGTGTGT-3′ and fragments and analogues thereof.
Claim 30
Dependent Claim
30 words
A method of permanently inhibiting gene expression comprising the step of administering the synthetic oligonucleotide of claim 14 in sufficient amount for cellular uptake and binding to the target sequence.
Claim 31
Dependent Claim
26 words
The method of claim 28, wherein the synthetic oligonucleotide is selected from the group consisting of eosin isothiocyanate, psoralin derivatives, metal chelates, ethidium and pyrene derivatives.
Claim 32
Dependent Claim
35 words
The method of altering the characteristics of muscle proteins in food animals comprising the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular uptake and binding to the target sequence.
Claim 33
Dependent Claim
24 words
The method of claim 30, wherein the synthetic oligonocleotide is selected from the group consisting of: 5′ - GTTTTTTGGGTGGGGGGGGGGGGGGGG-3′ and fragments and analogues thereof.
Claim 34
Dependent Claim
33 words
A method of inhibiting the interluken 2 alpha chain receptor comprising the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular uptake and binding to the target sequence.
Claim 35
Dependent Claim
30 words
The method of claim 32 for inhibiting interluken 2 alpha chain receptor, wherein the synthetic oligonucleotide is selected from the group consisting of: 5′-TTGGGGTGGGGTTTGTGGGTGTGGTTTT-3′, 3′TTGGGGTGGGGTTTGTGGGTGTGGTTTT-5′ and fragments and analogues thereof.
Claim 36
Dependent Claim
32 words
A method of disbursing plaque formation and Altzheimer's Disease, comprising the step of administering the synthetic oligonucleotide of claim 3 in suffient amount for cellular uptake and binding to the target sequence.
Claim 37
Dependent Claim
44 words
The method of claim 34 for disbursing plaque formation in Alzheimer's Disease, wherein the synthetic oligonucleotide is selected from the group consisting of: 5′-TTTTTGTTTGTTTTTTTTTTCTTTCTTTCTTTT-3′, 3′-TTTTTGTTTGTTTTTTTTTTCTTTCTTTCTTTT-5′, 3′-TGGTGGGGGTTGGTGGTTTGGTTGGTTGT-5′, 5′-TGGTGGGGGTTGGTGGTTTGGTTGGTTGT-3′, 3′-TTGTGTTTGTGTTGGTGTTTGGGGTGGGGGTGGTGTGG-5′, 5′-TTGTGTTTGTGTTGGTGTTTGGGGTGGGGGTGGTGTGG-3′, 5′-GTGTGTTTTTTGGTTTTGGGGTTTTTTT-3′, 3′-GTGTGTTTTTTGGTTTTGGGGTTTTTTT-5′, 3′-GTGTGGTTTGGGTGTTGGTGGTGGGTGGGTGTGGT-5′, 5′-GTGTGGTTTGGGTGTTGGTGGTGGGTGGGTGTGGT-3′, 3′-GGGTGGGTGTGGTGGGGGGTGTGTGTGGGTGGG-5′, 5′-GGGTGGGTGTGGTGGGGGGTGTGTGTGGGTGGG-3′, 5′-GGGGTGGGGTGGGGGGGGGTGGGG-3′, 3′-GGGGTGGGGTGGGGGGGGGTGGGG-5′, 3′-GTGGGGTGGGTGTGTGGGGGGGGGGGGGGGTG-5′, 5′-GTGGGGTGGGTGTGTGGGGGGGGGGGGGGGTG-3′ and fragments and analogues thereof.
Claim 38
Dependent Claim
35 words
The method of repressing the expression of the epidermal growth factor gene comprising the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular uptake and binding to the target sequence.
Claim 39
Dependent Claim
38 words
The method of claim 36 for repressing expression of the epidermal growth factor gene, wherein the synthetic oligonucleotids is selected from the group consisting of: 3′-TGGTGGGGGTTGGTGGTTTGGTTGGTTGT-5′, 5′-TGGTGGGGGTTGGTGGTTTGGTTGGTTGT-3′, 3′-TGGGTGGTGGTGGGGGGGTGGGTGGG-5′, 5′-TGGGTGGTGGTGGGGGGGTGGGTGGG-3′, 3′-TTGTGGTGGTGGTGTGGTGGTGGGGTTGGGTGGTGG-5′, 5′-TTGTGGTGGTGGTGTGGTGGTGGGGTTGGGTGGTGG-3′, 3′-TTGTGGTGGGTGGTGGTGGGTGGGTGGTGGTGGTGT-5′, 5′-TTGTGGTGGGTGGTGGTGGGTGGGTGGTGGTGGTGT-3′ and fragments and analogues thereof.
Claim 40
Dependent Claim
30 words
A method of repressing the GSTpi gene comprising the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular uptake and binding to the target sequence.
Claim 41
Dependent Claim
32 words
The method of claim 38 for repressing the GSTpi gene, wherein the synthetic oligonucleotide is selected from the group consisting of: 5′-GTGTGTGGTGTGGGGGGGTGGGGGGGGGGT-3′, 3′-GTGTGTGGTGTGGGGGGGTGGGGGGGGGGT-5′, 5′-GGGGTGGTGGGTTTGTGGGTTTGG-3′, 3′-GGGGTGGTGGGTTTGTGGGTTTGG-5′, 5′-TTTTTTTTTTTTTTTTTTTTTTTTTTTTTT-3′, 3′-TTTTTTTTTTTTTTTTTTTTTTTTTTTTTT-5′ and fragments and analogues thereof.
Claim 42
Dependent Claim
40 words
A method of intervening into the program of cholesterol synthesis by modulating the transcription of HMGCoA comprised in the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular uptake and binding to the target sequence.
Claim 43
Dependent Claim
33 words
The method of claim 40 for modulating the transcription of HMGCOA, wherein the synthetic oligonucleotide is selected from the group consisting of: 3′-GGTGTGTGTTGGTGGGGTGGGGGTTGTGGGGGG-5′, 5′-GGTGTGTGTTGGTGGGGTGGGGGTTGTGGGGGG-3′ 5′-GGGTGGGTGGTGTGGGGGGTTGTTTTGGGGT-3′, 3′-GGGTGGGTGGTGTGGGGGGTTGTTTTGGGGT-5′, 3′-TGGGGTTGGGTGGTTGGTTTGTTTTTGGGGGGGGT-5′, 5′-TGGGGTTGGGTGGTTGGTTTGTTTTTGGGGGGGGT-3′ and fragments and analogues thereof.
Claim 44
Dependent Claim
35 words
A method of suppression of expression of the nerve growth factor receptor comprising the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular uptake and binding to the target sequence.
Claim 45
Dependent Claim
40 words
The method of claim 42 for suppressing the gene encoding nerve growth factor receptor, wherein the synthetic oligonucleotide is selected from a group consisting of: 5′-GGGTTGTGGGTTGGTGGGGGGGTTGGGTGTGTGG-3′, 3′-GGGTTGTGGGTTGGTGGGGGGGTTGGGTGTGTGG-5′, 5′-TGGGGGGGTTGGGTGTGTGGGTGTTTGGGTGTTGG-3′, 3′-TGGGGGGGTTGGGTGTGTGGGTGTTTGGGTGTTGG-5′, 5′-TTGGGTGTTGGGTGGGTGTTGGGGTGGGGTGGGGGTT-3′, 3′-TTGGGTGTTGGGTGGGTGTTGGGGTGGGGTGGGGGGTT-5′, 5′-GGGTGGGTTTGGGTGTGGTTGGGTGGGG-3′, 3′-GGGTGGGTTTGGGTGTGGTTGGGTGGGG-5′, 5′-GGGGGTGGGGGGGGTGGGGGGGGGGTGGGGGGGG-3′, 3′-GGGGGTGGGGGGGTGGGGGGGGGGTGGGGGGGGG-5′ and fragments and analogues thereof.
Claim 46
Dependent Claim
35 words
A method for arresting final replication of the Herpes Simplex Virus 1 comprising the step of administering the synthetic oligonucleotide of claim 3 in sufficient amount for cellular update and binding to the target sequence.
Claim 47
Dependent Claim
37 words
A method of claim 44 for arresting viral replication, wherein the synthetic oligonucleotide is selected from a group consisting of: 3′-TTTTTGTGTTGGGGGGTGGGGTGTGGGGGGTGTTT-5′, 5′-TTTTTGTGTTGGGGGGGTGGGGTGTGGGGGGTGTTT-3′, 3′-TTTTTGTGTTGGGGGGGTGGGGTGTTGGGGGGTGTTT-5′, 5--TTTTTGTGTTGGGGGGGTGGGGTGTTGGGGGGTGTTT-3′, 5′-TTTTTGGGGGGGGGGGGGGGGTGGGTGTTGGGGTGGG-3′, 3′-TTTTTGGGGGGGGGGGGGGGGTGGGTGTTGGGGTGGG-5′, 5′-TTTTTTTGTGTGTTGGGGTTGGGTTGGGTGTTTGTGGT-3′, 3′-TTTTTTTGTGTGTTGGGGTTGGGTTGGGTGTTTGTGGT-5′, 3′-TGGTGTTTGTGGGTTGGGTTGGGGTTGTGTGTTTTTTT-5′, 5′-TGGTGTTTGTGGGTTGGGTTGGGGTTGTGTGTTTTTTT-3′, 5′-TTGGGGGGGGGGGGGGGGGGTTTTTGTTGTGTGTT-3′, 3′-TTGGGGGGGGGGGGGGGGGGTTTTTGTTGTGTGTT-5′ and fragments and analogues thereof.
Claim 48
Dependent Claim
36 words
The method of suppressing beta globin genes synthesis in thallassemics and sickle cell anemia comprising the step of administering the synthetic oligonucleotide of claim 3 and sufficient amount for cellular update combining to the target sequence.
Claim 49
Dependent Claim
41 words
The method of claim 46 for suppressing the synthesis of beta globin gene, wherein the synthetic oligonucleotide is selected from the group consisting of: 3′-GGTTTTGGGGTGGTTGGGGTTGTTTGT-5′, 5′-GGTTTTGGGGTGGTTGGGGTTGTTTGT-3′, 5′-TGGTGGTGGGTGGGGTGGTGGGTGGGGTGGGGTTTTTTG-3′, 3′-TGGTGGTGGGTGGGGTGGTGGGTGGGGTGGGGTTTTTTG-5′, 5′-TGGGGTGGGGTTTTTTGTGTGGGGTGTG-3′, 3′-TGGGGTGGGGTTTTTTGTGTGGGGTGTG-5′, 3′-GGGTTGTTGTTTTGTTTGGGGTTGTTTTGT-5′, 5′-GGGTTGTTGTTTTGTTTGGGGTTGTTTTGT-3′, 3′-TTGTTGGTTTGTTTTTTTTTGTTGTGG-5′, 5′-TTGTTGGTTTGTTTTTTTTTGTTGTGG-3′, 3′-GTGGGTTGTTTTTTTTGTTTTTTTTTT-5′, 5′-GTGGGTTGTTTTTTTTGTTTTTTTTTT-3′ and fragments and analogues thereof.
References Cited (Backward Citations)
No citation records indexed.
USPTO Annuity & Maintenance Windows
Maintenance schedule is calculated for granted US utility patents.