US2017166603A1PendingUtilityA1
Large stokes shift dyes
Est. expiryFeb 11, 2029(~2.6 yrs left)· nominal 20-yr term from priority
C09K 2211/1007C09K 2211/1074C07H 19/207C09K 11/06C09K 2211/1022C09B 23/164C09K 2211/1014C09K 2211/1059C09K 2211/1044C07D 209/26C07D 401/14G01N 33/52C09B 23/0066C07D 401/10C09B 23/0041C09B 23/086C07D 209/14
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Claims
Abstract
Provided herein are heptamethine cyanine dyes having a large Stokes shift, and the salts and conjugates thereof. Also provided are methods of using and making such large Stokes shift dyes as fluorescence resonance energy transfer (FRET) acceptors or donors.
Claims
exact text as granted — not AI-modified1 . A compound of Formula I or II:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein
m and n are independently integers from 1 to 6;
R 5 and R 6 , when taken alone, are independently H, C 1-6 carbonyl or optionally substituted C 1-6 alkyl;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl;
X and Y are independently CR 9 2 , Ne, O or S; and
each R 9 and R 10 is independently C 1-4 alkyl.
2 . The compound of claim 1 , wherein each of X and Y is CR 9 2 .
3 . The compound of claim 1 , wherein each of R 7 and R 8 is H.
4 . The compound of claim 1 , wherein at least one of R 5 and R 6 is C 1-6 alkyl substituted with a substituent selected from the group consisting of SO 3 H, COOH, OH, NH 2 , maleimide, thiol, isocyanate, isothiocyanate, disulfide, alkynyl, azidoyl and trialkoxysilane.
5 . The compound of claim 1 , wherein at least one of R 3 and R 4 is C 1-6 alkyl substituted with a substituent selected from the group consisting of halo, hydroxyl, C 1-6 alkyl, or amino
6 . The compound of claim 1 , wherein each of R 1 and R 2 is SO 3 H.
7 . The compound of claim 1 , wherein each of R 3 and R 4 is (CH 2 ) m SO 3 H.
8 . The compound of claim 1 which has a Stokes shift of about 30 nm to about 250 nm.
9 . The compound of claim 1 which has a Stokes shift of about 50 nm to about 225 nm.
10 . The compound of claim 1 which has a Stokes shift of about 70 nm to about 200 nm.
11 . A compound of Formula I or II:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein
m and n are independently integers from 1 to 6;
R 5 and R 6 are taken together with the nitrogen atom to which they are attached to form an optionally substituted 5- or 6-membered azacyclic ring substituted, optionally containing an additional heteroatom selected from N, O and S as a ring member;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl;
X and Y are independently CR 9 2 , NR 10 , O or S; and
each R 9 and R 10 is independently C 1 -4 alkyl.
12 . The compound of claim 11 , wherein said azacyclic ring is substituted with a substituent (CH 2 ) p Z, where p is an integer from 0 to 6, and Z is SO 3 H, COOH, OH, and NH 2 .
13 . The compound of claim 11 , wherein each of X and Y is CR 9 2 .
14 . The compound of claim 11 , wherein each of R 7 and R 8 is H.
15 . The compound of claim 11 , wherein each of R 1 and R 2 is SO 3 H.
16 . The compound of claim 11 , wherein at least one of R 3 and R 4 is C 1-6 alkyl substituted with a substituent selected from the group consisting of halo, hydroxyl, C 1-6 alkyl, or amino
17 . The compound of claim 11 which has a Stokes shift of about 30 nm to about 250 nm.
18 . The compound of claim 11 which has a Stokes shift of about 50 nm to about 225 nm.
19 . The compound of claim 11 which has a Stokes shift of about 70 nm to about 200 nm.
20 . A compound of Formula III:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein
m and n are independently integers from 1 to 6;
R 5 and R 6 , when taken alone, are independently H, C 1-6 carbonyl or optionally substituted C 1-6 alkyl;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl; and
each R 9 is independently C 1-4 alkyl.
21 . The compound of claim 20 , wherein each R 9 is methyl.
22 . The compound of claim 20 , wherein each of R 7 and R 8 is H.
23 . The compound of claim 20 , wherein at least one of R 5 and R 6 is C 1-6 alkyl substituted with a substituent selected from the group consisting of SO 3 H, COOH, OH, and NH 2 , maleimide, thiol, isocyanate, isothiocyanate, disulfide, alkynyl, azidoyl and trialkoxysilane.
24 . The compound of claim 20 , wherein each of R 1 and R 2 is SO 3 H.
25 . The compound of claim 20 , wherein at least one of R 3 and R 4 is C 1-6 alkyl substituted with a substituent selected from the group consisting of halo, hydroxyl, C 1-6 alkyl, or amino
26 . The compound of claim 20 , wherein each of R 3 and R 4 is (CH 2 ) m SO 3 H.
27 . The compound of claim 20 which has a Stokes shift of about 30 nm to about 250 nm.
28 . The compound of claim 20 which has a Stokes shift of about 50 nm to about 225 nm.
29 . The compound of claim 20 which has a Stokes shift of about 70 nm to about 200 nm.
30 . A compound of Formula III:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein
m and n are independently integers from 1 to 6;
R 5 and R 6 are taken together with the nitrogen atom to which they are attached to form an optionally substituted 5- or 6-membered azacyclic ring substituted, optionally containing an additional heteroatom selected from N, O and S as a ring member;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl; and
each R 9 is independently C 1-4 alkyl.
31 . The compound of claim 30 , wherein said azacyclic ring is substituted with a substituent (CH 2 ) p Z, where p is an integer from 0 to 6, and Z is SO 3 H, COOH, OH, and NH 2 .
32 . The compound of claim 30 , wherein each R 9 is methyl.
33 . The compound of claim 30 , wherein each of R 7 and R 8 is H.
34 . The compound of claim 30 , wherein each of R 1 and R 2 is SO 3 H.
35 . The compound of claim 30 , wherein each of R 3 and R 4 is C 1-6 alkyl substituted with a substituent selected from the group consisting of halo, hydroxyl, C 1-6 alkyl, or amino
36 . The compound of claim 30 which has a Stokes shift of about 30 nm to about 250 nm.
37 . The compound of claim 30 which has a Stokes shift of about 50 nm to about 225 nm.
38 . The compound of claim 30 which has a Stokes shift of about 70 nm to about 200 nm.
39 . A compound selected from:
or a salt or conjugate thereof.
40 . The compound of any one of claim 1 , 11 , 20 , 30 , or 39 which is conjugated to a nucleotide, nucleoside, nucleic acid, oligonucleotide, deoxyoligonucleotide, DNA fragment, RNA fragment, or a derivatized variant of one of these; or an antibody, amino acid, peptide, or protein.
41 . The compound of any one of claim 1 , 11 , 20 , 30 , or 39 , wherein the compound has an excitation wavelength between 400 and 900 nm.
42 . The compound of claim 41 , wherein the compound has an emission wavelength that is at least 70 nm greater than the excitation wavelength.
43 . A fluorescing molecular complex comprising:
a donor dye capable of absorbing light at a first wavelength and emitting excitation energy in response; and an acceptor dye capable of absorbing the excitation energy emitted by the donor dye and fluorescing at a second wavelength in response; wherein either said donor dye or said acceptor dye has the structure of Formula III:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein
m and n are independently integers from 1 to 6;
R 5 and R 6 , when taken alone, are independently H, C 1-6 carbonyl or optionally substituted C 1-6 alkyl;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl; and
each R 9 is independently C 1-4 alkyl.
44 . The donor dye or acceptor dye of claim 43 which has a Stokes shift of about 30 nm to about 250 nm.
45 . The donor dye or acceptor dye of claim 43 which has a Stokes shift of about 50 nm to about 225 nm.
46 . The donor dye or acceptor dye of claim 43 which has a Stokes shift of about 70 nm to about 200 nm.
47 . The donor dye or acceptor dye of claim 43 , wherein the compound has an excitation wavelength between 400 and 900 nm.
48 . The donor dye or acceptor dye of claim 47 , wherein the compound has an emission wavelength that is at least 70 nm greater than the excitation wavelength.
49 . A fluorescing molecular complex comprising:
a donor dye capable of absorbing light at a first wavelength and emitting excitation energy in response; and an acceptor dye capable of absorbing the excitation energy emitted by the donor dye and fluorescing at a second wavelength in response; wherein either said donor dye or said acceptor dye has the structure of Formula III:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein m and n are independently integers from 1 to 6;
R 5 and R 6 are taken together with the nitrogen atom to which they are attached to form an optionally substituted 5- or 6-membered azacyclic ring substituted, optionally containing an additional heteroatom selected from N, O and S as a ring member;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl; and
each R 9 is independently C 1-4 alkyl.
50 . The donor dye or acceptor dye of claim 49 , wherein said azacyclic ring is substituted with a substituent (CH 2 ) p Z, where p is an integer from 0 to 6, and Z is SO 3 H, COOH, OH, and NH 2 .
51 . The donor dye or acceptor dye of claim 49 which has a Stokes shift of about 30 nm to about 250 nm.
52 . The donor dye or acceptor dye of claim 49 which has a Stokes shift of about 50 nm to about 225 nm.
53 . The donor dye or acceptor dye of claim 49 which has a Stokes shift of about 70 nm to about 200 nm.
54 . The donor dye or acceptor dye of claim 49 , wherein the acceptor dye has an excitation wavelength between 400 and 900 nm.
55 . The donor dye or acceptor dye of claim 54 , wherein the acceptor dye has an emission wavelength that is at least 70 nm greater than the excitation wavelength.
56 . A substantially pure composition of Formula I or II:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein
m and n are independently integers from 1 to 6;
R 5 and R 6 , when taken alone, are independently H, C 1-6 carbonyl or optionally substituted C 1-6 alkyl;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl;
X and Y are independently CR 9 2 , NR 10 , O or S; and
each R 9 and R 19 is independently C 1-4 alkyl.
57 . The substantially pure composition of claim 56 , wherein each of X and Y is CR 9 2 .
58 . The substantially pure composition of claim 56 , wherein each of R 7 and R 8 is H.
59 . The substantially pure composition of claim 56 , wherein at least one of R 5 and R 6 is C 1-6 alkyl substituted with a substituent selected from the group consisting of SO 3 H, COOH, OH, NH 2 , maleimide, thiol, isocyanate, isothiocyanate, disulfide, alkynyl, azidoyl and trialkoxysilane.
60 . The substantially pure composition of claim 56 , wherein at least one of R 3 and R 4 is C 1-6 alkyl substituted with a substituent selected from the group consisting of halo, hydroxyl, C 1-6 alkyl, or amino.
61 . The substantially pure composition of claim 56 , wherein each of R 1 and R 2 is SO 3 H.
62 . The substantially pure composition of claim 56 , wherein each of R 3 and R 4 is (CH 2 ) m SO 3 H.
63 . The substantially pure composition of claim 56 which has a Stokes shift of about 30 nm to about 250 nm.
64 . The substantially pure composition of claim 56 which has a Stokes shift of about 50 nm to about 225 nm.
65 . The substantially pure composition of claim 56 which has a Stokes shift of about 70 nm to about 200 nm.
66 . A substantially pure composition of Formula I or II:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein
m and n are independently integers from 1 to 6;
R 5 and R 6 are taken together with the nitrogen atom to which they are attached to form an optionally substituted 5- or 6-membered azacyclic ring substituted, optionally containing an additional heteroatom selected from N, O and S as a ring member;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl;
X and Y are independently CR 9 2 , Ne, O or S; and
each R 9 and R 10 is independently C 1-4 alkyl.
67 . The substantially pure composition of claim 66 , wherein said azacyclic ring is substituted with a substituent (CH 2 ) p Z, where p is an integer from 0 to 6, and Z is SO 3 H, COOH, OH, and NH 2 .
68 . The substantially pure composition of claim 66 , wherein each of X and Y is CR 9 2
69 . The substantially pure composition of claim 66 , wherein each of R 7 and R 8 is H.
70 . The substantially pure composition of claim 66 , wherein each of R 1 and R 2 is SO 3 H.
71 . The substantially pure composition of claim 66 , wherein at least one of R 3 and R 4 is C 1-6 alkyl substituted with a substituent selected from the group consisting of halo, hydroxyl, C 1-6 alkyl, or amino.
72 . The substantially pure composition of claim 66 which has a Stokes shift of about 30 nm to about 250 nm.
73 . The substantially pure composition of claim 66 which has a Stokes shift of about 50 nm to about 225 nm.
74 . The substantially pure composition of claim 66 which has a Stokes shift of about 70 nm to about 200 nm.
75 . A substantially pure composition of Formula III:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein
m and n are independently integers from 1 to 6;
R 5 and R 6 , when taken alone, are independently H, C 1-6 carbonyl or optionally substituted C 1-6 alkyl;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl; and
each R 9 is independently C 1-4 alkyl.
76 . The substantially pure composition of claim 75 , wherein each R 9 is methyl.
77 . The substantially pure composition of claim 75 , wherein each of R 7 and R 8 is H.
78 . The substantially pure composition of claim 75 , wherein at least one of R 5 and R 6 is C 1-6 alkyl substituted with a substituent selected from the group consisting of SO 3 H, COOH, OH, and NH 2 , maleimide, thiol, isocyanate, isothiocyanate, disulfide, alkynyl, azidoyl and trialkoxysilane.
79 . The substantially pure composition of claim 75 , wherein each of R 1 and R 2 is SO 3 H.
80 . The substantially pure composition of claim 75 , wherein at least one of R 3 and R 4 is C 1-6 alkyl substituted with a substituent selected from the group consisting of halo, hydroxyl, C 1-6 alkyl, or amino.
81 . The substantially pure composition of claim 75 , wherein each of R 3 and R 4 is (CH 2 ) m SO 3 H.
82 . The substantially pure composition of claim 75 which has a Stokes shift of about 30 nm to about 250 nm.
83 . The substantially pure composition of claim 75 which has a Stokes shift of about 50 nm to about 225 nm.
84 . The substantially pure composition of claim 75 which has a Stokes shift of about 70 nm to about 200 nm.
85 . A substantially pure composition of Formula III:
or a salt or conjugate thereof;
wherein R 1 and R 2 , when taken alone, are independently H, halo, hydroxyl, amino, C 1-6 alkylamino, C 2-12 dialkylamino, C 1-6 perfluoroalkyl, C 1-6 alkyl, C 1-6 alkoxy, SO 3 H or COOH;
R 3 and R 4 , when taken alone, are independently (CH 2 ) m SO 3 H, (CH 2 ) n COOH or optionally substituted C 1-6 alkyl, wherein
m and n are independently integers from 1 to 6;
R 5 and R 6 are taken together with the nitrogen atom to which they are attached to form an optionally substituted 5- or 6-membered azacyclic ring substituted, optionally containing an additional heteroatom selected from N, O and S as a ring member;
R 7 and R 8 are independently H, C 1-4 alkyl, or phenyl; and
each R 9 is independently C 1-4 alkyl.
86 . The substantially pure composition of claim 85 , wherein said azacyclic ring is substituted with a substituent (CH 2 ) p Z, where p is an integer from 0 to 6, and Z is SO 3 H, COOH, OH, and NH 2 .
87 . The substantially pure composition of claim 85 , wherein each R 9 is methyl.
88 . The substantially pure composition of claim 85 , wherein each of R 7 and R 8 is H.
89 . The substantially pure composition of claim 85 , wherein each of R 1 and R 2 is SO 3 H.
90 . The substantially pure composition of claim 85 , wherein each of R 3 and R 4 is C 1-6 alkyl substituted with a substituent selected from the group consisting of halo, hydroxyl, C 1-6 alkyl, or amino
91 . The substantially pure composition of claim 85 which has a Stokes shift of about 30 nm to about 250 nm.
92 . The substantially pure composition of claim 85 which has a Stokes shift of about 50 nm to about 225 nm.
93 . The substantially pure composition of claim 85 which has a Stokes shift of about 70 nm to about 200 nm.
94 . A process for preparing a compound of the formula:
said process comprising:
a. dissolving a first compound having the formula
a second compound having the formula
and triethylamine into a mixture comprising dimethylformamide (DMF) and H 2 O; and
b. heating the mixture containing the first compound, the second compound, and triethylamine to between about 80° C. to about 85° C. for a period of about 2 hours.
95 . A process for preparing a compound of the formula:
said process comprising:
a. dissolving a first compound having the formula
β-alanine, and triethylamine into a mixture comprising dimethylformamide (DMF) and H 2 O; and
b. heating the mixture containing the first compound, β-alanine, and triethylamine to between about 80° C. to about 85° C. for a period of about 2 hours.Join the waitlist — get patent alerts
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