US2020063110A1PendingUtilityA1
Thienopyrrole compounds and uses thereof
Est. expiryJun 25, 2035(~8.9 yrs left)· nominal 20-yr term from priority
C12N 9/0069C07D 491/048C12Y 113/12007C07D 493/04G01N 33/542C07D 207/34C07D 495/04C12Q 1/66G01N 33/581C07D 209/42C07K 5/0222C07K 5/0819
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Claims
Abstract
Thienopyrrole compounds that may inhibit Oplophorus-derived luciferases are disclosed, as well as compositions and kits comprising the thienopyrrole compounds, and methods of using the thienopyrrole compounds.
Claims
exact text as granted — not AI-modified1 . A compound of formula (I), or a salt thereof:
wherein:
the dashed line represents the presence or absence of a bond;
n is 0, 1, 2, 3, 4 or 5;
X is CH, N, O, or S;
wherein, when the dashed line represents the presence of a bond, X is CH or N, and when the dashed line represents the absence of a bond, X is O or S;
A is an optionally substituted phenyl ring, or an optionally substituted 5- or 6-membered heteroaryl ring;
R 1 and R 2 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted arylalkyl, optionally substituted alkoxyalkyl and optionally substituted alkoxyalkoxyalkyl; and
R 3 and R 4 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted C 3 -C 8 -cycloalkyl, optionally substituted C 3 -C 8 -cycloalkylalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, optionally substituted heterocyclyl, and optionally substituted heterocyclylalkyl; or R 3 and R 4 , together with the nitrogen atom to which they are attached, together form an optionally substituted ring.
2 . The compound of claim 1 , wherein n is 1.
3 . The compound of any of claims 1 - 2 , wherein the dashed line represents the presence of a bond, and X is CH.
4 . The compound of any of claims 1 - 3 , wherein A is a 5-membered heteroaryl ring.
5 . The compound of any of claims 1 - 4 , wherein A is a thienyl ring or a furanyl ring.
6 . The compound of any of claims 1 - 3 , wherein A is a phenyl ring.
7 . The compound of any of claims 1 - 6 , wherein R 1 is selected from the group consisting of hydrogen, C 1 -C 8 alkyl, halo-C 1 -C 8 -alkyl, alkoxyalkoxyalkyl and arylalkyl.
8 . The compound of any of claims 1 - 7 , wherein R 1 is selected from the group consisting of hydrogen, ethyl, n-hexyl, 2-(2-methoxyethoxy)ethyl and benzyl.
9 . The compound of any of claims 1 - 8 , wherein R 1 is ethyl.
10 . The compound of any of claims 1 - 9 , wherein R 2 is optionally substituted aryl.
11 . The compound of claim 10 , wherein R 2 is substituted phenyl.
12 . The compound of claim 11 , wherein R 2 is phenyl substituted with one substituent selected from the group consisting of C 1 -C 4 alkyl, cyano, amido, C 1 -C 4 alkoxy, and hydroxyalkyl.
13 . The compound of claim 12 , wherein R 2 is phenyl substituted with one methyl group.
14 . The compound of any of claims 1 - 13 , wherein R 3 and R 4 , together with the nitrogen atom to which they are attached, together form an optionally substituted ring.
15 . The compound of claim 14 , wherein R 3 and R 4 , together with the nitrogen atom to which they are attached, together form an optionally substituted monocyclic heterocycle.
16 . The compound of claim 15 , wherein the optionally substituted monocyclic heterocycle is selected from the group consisting of optionally substituted pyrrolidine, piperidine and piperazine.
17 . The compound of claim 15 , wherein the optionally substituted monocyclic heterocycle is selected from the group consisting of unsubstituted pyrrolidine, unsubstituted piperidine, piperidine substituted with one substituent, or piperazine substituted with one substituent.
18 . The compound of any of claims 1 - 17 , wherein R 3 is hydrogen.
19 . The compound of any of claims 1 - 18 , wherein R 4 is selected from the group consisting of unsubstituted C 1 -C 8 alkyl, halo-C 1 -C 8 -alkyl, carboxy-C 1 -C 8 -alkyl, C 1 -C 4 -alkoxycarbonyl-C 1 -C 8 -alkyl, optionally substituted phenyl, optionally substituted C 5 -C 6 cycloalkyl, optionally substituted C 5 -C 6 -cycloalkylalkyl, optionally substituted heterocyclyl, optionally substituted heteroarylalkyl, and optionally substituted heterocyclylalkyl.
20 . The compound of claim 19 , wherein R 4 is phenyl that is unsubstituted or substituted with one substituent.
21 . The compound of claim 20 , wherein the substituent is selected from the group consisting of C 1 -C 4 -alkoxycarbonyl and C 1 -C 4 -alkoxycarbonyl-C 1 -C 4 -alkyl.
22 . The compound of any of claims 1 - 18 , wherein R 4 is cyclohexyl substituted with one substituent selected from the group consisting of carboxy, C 1 -C 4 -alkoxycarbonyl, C 1 -C 8 -alkylamido, hydroxy-C 1 -C 8 -alkylamido, amido, optionally substituted amino-C 1 -C 8 -alkylamido, C 1 -C 4 -dialkylamino-C 1 -C 8 -alkylamido, carboxy-C 1 -C 8 -alkylamido, sulfonic acid-C 1 -C 8 -alkylamido, sulfonate-C 1 -C 8 -alkylamido, C 1 -C 4 -alkylcarbonyl-C 1 -C 8 -alkylamido, optionally substituted C 3 -C 6 -cycloalkylamido, and optionally substituted heterocyclylamido.
23 . The compound of claim 1 , wherein the compound has formula (Ia):
24 . The compound of claim 1 , wherein the compound has formula (Ib):
wherein:
Y is selected from the group consisting of —NR a R b and —OR c ;
R a and R b are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted C 3 -C 8 -cycloalkyl, and optionally substituted heterocyclyl; or R a and R b , together with the nitrogen atom to which they are attached, together form an optionally substituted ring; and
R c is selected from the group consisting of hydrogen and optionally substituted C 1 -C 4 alkyl.
25 . The compound of claim 24 , wherein Y is —OR c .
26 . The compound of claim 25 , wherein R c is selected from the group consisting of hydrogen and methyl.
27 . The compound of claim 24 , wherein Y is —NR a R b .
28 . The compound of claim 27 , wherein R a and R b , together with the nitrogen atom to which they are attached, together form an optionally substituted ring.
29 . The compound of claim 28 , wherein R a and R b , together with the nitrogen atom to which they are attached, together form an optionally substituted monocyclic heterocycle.
30 . The compound of claim 29 , wherein the optionally substituted monocyclic heterocycle is optionally substituted piperidine.
31 . The compound of claim 30 , wherein the optionally substituted monocyclic heterocycle is selected from the group consisting of unsubstituted piperidine and piperidine substituted with one substituent.
32 . The compound of claim 27 , wherein R a is hydrogen.
33 . The compound of claim 32 , wherein R b is selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted C 3 -C 8 -cycloalkyl, and optionally substituted heterocyclyl.
34 . The compound of claim 32 , wherein R b is selected from the group consisting of hydrogen, C 1 -C 6 alkyl, hydroxyalkyl, optionally substituted aminoalkyl, carboxyalkyl, sulfonic acid-alkyl, sulfonate-alkyl, alkylcarbonylalkyl, optionally substituted C 3 -C 6 -cycloalkyl, and optionally substituted six-membered heterocyclyl.
35 . The compound of any of claims 24 - 34 , wherein the compound has the following formula (Ib′)
36 . The compound of claim 1 , wherein the compound is selected from the group consisting of:
N-cyclohexyl-4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; N-ethyl-2-(5-(pyrrolidine-1-carbonyl)-4H-thieno[3,2-b]pyrrol-4-yl)-N-(m-tolyl)acetamide; N-ethyl-2-(5-(piperidine-1-carbonyl)-4H-thieno[3,2-b]pyrrol-4-yl)-N-(m-tolyl)acetamide; ethyl 1-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carbonyl)piperidine-4-carboxylate; 4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-N-phenyl-4H-thieno[3,2-b]pyrrole-5-carboxamide; ethyl 2-(4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)phenyl)acetate; methyl 3-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)benzoate; methyl-cis-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; 8-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)octanoic acid; 6-(4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)piperidin-1-yl)-6-oxohexanoic acid; trans-methyl-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylic acid; N-(trans-4-(butylcarbamoyl)cyclohexyl)-4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; 4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-N-(trans-4-((2-hydroxyethyl)carbamoyl)cyclohexyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; N-(trans-4-((2-(dimethylamino)ethyl)carbamoyl)cyclohexyl)-4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; 4-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)butanoic acid; N-(trans-4-carbamoylcyclohexyl)-4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; 4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-N-(trans-4-(hexylcarbamoyl)cyclohexyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; ethyl 1-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carbonyl)piperidine-4-carboxylate; methyl 6-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)hexanoate; 6-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)hexanoic acid; 1-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carbonyl)piperidine-4-carboxylic acid; 8-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)octanoic acid; N-(trans-4-(cyclohexylcarbamoyl)cyclohexyl)-4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; 4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-N-(trans-4-((1-methylpiperidin-4-yl)carbamoyl)cyclohexyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; tert-butyl 4-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)piperidine-1-carboxylate; 4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-N-(trans-4-(piperidin-4-ylcarbamoyl)cyclohexyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; N-(trans-4-((1-acetylpiperidin-4-yl)carbamoyl)cyclohexyl)-4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; tert-butyl (6-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)hexyl)carbamate; N-(trans-4-((6-(3′,6′-dihydroxy-3-oxo-3H-spiro[isobenzofuran-1,9′-xanthene]-5(6)-carboxamido)hexyl)carbamoyl)cyclohexyl)-4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; N-(trans-4-((6-aminohexyl)carbamoyl)cyclohexyl)-4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide hydrochloride; 4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-N-(trans-4-((6-hydroxyhexyl)carbamoyl)cyclohexyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; methyl-trans-4-(trans-4-(4-(2-(ethcyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)cyclohexane-1-carboxylate; trans-4-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)cyclohexane-1-carboxylic acid; (11S,14S,17S)-17-acetamido-11,14-bis(carboxymethyl)-1-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexyl)-1,10,13,16-tetraoxo-2,9,12,15-tetraazanonadecan-19-oic acid; 4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-N-methyl-4H-thieno[3,2-b]pyrrole-5-carboxamide; N-cyclopentyl-4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; 4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-N-(pyridin-4-ylmethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; 4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-N-(3-morpholinopropyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; N-ethyl-2-(5-(4-methylpiperazine-1-carbonyl)-4H-thieno[3,2-b]pyrrol-4-yl)-N-(m-tolyl)acetamide; methyl 4-(2-oxo-2-(m-tolylamino)ethyl)-4H-thieno[3,2-b]pyrrole-5-carboxylate; N-cyclohexyl-4-(2-oxo-2-(m-tolylamino)ethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; N-cyclohexyl-4-(24(2-(2-methoxyethoxy)ethyl)(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; methyl-trans-4-(4-(2-((2-(2-methoxyethoxy)ethyl)(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl 4-(2-(hexyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxylate; N-cyclohexyl-4-(2-(hexyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; methyl-trans-4-(4-(2-(hexyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl 4-(2-(benzyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxylate; 6-(cis-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)hexanoic acid; methyl 6-(trans-4-((4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)methyl)cyclohexane-1-carboxamido)hexanoate; 6-(trans-4-((4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)methyl)cyclohexane-1-carboxamido)hexanoic acid; sodium 6-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)hexane-1-sulfonate; potassium 6-(trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)hexane-1-sulfonate; trans-4-(4-(2-(hexyl(m-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylic acid; methyl trans-4-(4-(2-(ethyl(phenyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-((3-cyanophenyl)(ethyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-((3-carbamoylphenyl)(ethyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-(ethyl(3-ethylphenyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-(ethyl(3-methoxyphenyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-(ethyl(o-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-(6-methyl-3,4-dihydroquinolin-1(2H)-yl)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-(ethyl(p-tolyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-(ethyl(4-(hydroxymethyl)phenyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(1-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-1H-indole-2-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(1-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-6-methoxy-1H-indole-2-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-4H-furo[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; 4-(2-(ethyl(3-ethylphenyl)amino)-2-oxoethyl)-N-(trans-4-((6-hydroxyhexyl)carbamoyl)cyclohexyl)-4H-thieno[3,2-b]pyrrole-5-carboxamide; sodium 6-(trans-4-(4-(2-(ethyl(3-ethylphenyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxamido)hexane-1-sulfonate; methyl trans-4-(4-(2-(ethyl(3-isopropylphenyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-(ethyl(3-(hydroxymethyl)phenyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-((3-(bromomethyl)phenyl)(ethyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-((3-(dimethylamino)phenyl)(ethyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(4-(2-(ethyl(3-isobutylphenyl)amino)-2-oxoethyl)-4H-thieno[3,2-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate; methyl trans-4-(1-(2-(ethyl(3-ethylphenyl)amino)-2-oxoethyl)-1H-indole-2-carboxamido)cyclohexane-1-carboxylate; 1-(2-(ethyl(3-ethylphenyl)amino)-2-oxoethyl)-N-(trans-4-((6-hydroxyhexyl)carbamoyl)cyclohexyl)-1H-indole-2-carboxamide; sodium 6-(trans-4-(1-(2-(ethyl(3-ethylphenyl)amino)-2-oxoethyl)-1H-indole-2-carboxamido)cyclohexane-1-carboxamido)hexane-1-sulfonate; methyl trans-4-(1-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-1H-pyrrole-2-carboxamido)cyclohexane-1-carboxylate; and methyl trans-4-(6-(2-(ethyl(m-tolyl)amino)-2-oxoethyl)-6H-thieno[2,3-b]pyrrole-5-carboxamido)cyclohexane-1-carboxylate.
37 . A method of inhibiting an Oplophorus -derived luciferase the method comprising contacting the Oplophorus -derived luciferase with a compound of any of claims 1 - 36 .
38 . The method of claim 37 , wherein the Oplophorus -derived luciferase comprises a polypeptide sequence of SEQ ID NO: 2.
39 . A method of inhibiting an Oplophorus -derived luciferase, the method comprising contacting the Oplophorus -derived luciferase with a compound of formula (II):
wherein:
the dashed line represents the presence or absence of a bond;
n is 0, 1, 2, 3, 4 or 5;
X is CH, N, O, or S;
wherein, when the dashed line represents the presence of a bond, X is CH or N, and when the dashed line represents the absence of a bond, X is O or S;
A is an optionally substituted phenyl ring, or an optionally substituted 5- or 6-membered heteroaryl ring;
R 1 and R 2 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted arylalkyl, optionally substituted alkoxyalkyl and optionally substituted alkoxyalkoxyalkyl;
Z is selected from the group consisting of —NR 3 R 4 and —OR 5 ; and
R 3 , R 4 and R 5 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted C 3 -C 8 -cycloalkyl, optionally substituted C 3 -C 8 -cycloalkylalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, optionally substituted heterocyclyl, and optionally substituted heterocyclylalkyl; or R 3 and R 4 , together with the nitrogen atom to which they are attached, together form an optionally substituted ring.
40 . The method of claim 39 , wherein the Oplophorus -derived luciferase comprises a polypeptide sequence of SEQ ID NO: 2.
41 . A method for modulating luminescence of an Oplophorus -derived luciferase in a sample, the method comprising,
(a) contacting the sample with a coelenterazine substrate and the compound of any of claims 1 - 36 ; and (b) detecting luminescence in the sample, wherein the compound of any of claims 1 - 36 causes a decrease in the luminescence from the Oplophorus -derived luciferase.
42 . A method to detect an interaction between a first protein and a second protein in a sample, the method comprising:
(a) contacting a sample with a coelenterazine substrate and the compound of any one of claims 1 - 36 , wherein the sample comprises:
(xvii) a first polynucleotide encoding a first fusion protein, wherein the first fusion protein comprises a first fragment of an Oplophorus -derived luciferase and a first protein; and
(xviii) a second polynucleotide encoding a second fusion protein, wherein the second fusion protein comprises a second fragment of the Oplophorus -derived luciferase and a second protein; and
(b) detecting luminescence in the sample,
wherein the detection of luminescence indicates an interaction between the first protein and the second protein.
43 . The method of any of claims 41 - 42 , comprising contacting the sample with the coelenterazine substrate prior to contacting the sample with the compound of any one of claims 1 - 36 .
44 . The method of claim 42 , wherein when the first protein and second protein interact, the first fragment of the Oplophorus -derived luciferase and the second fragment of the Oplophorus -derived luciferase reconstitute a full-length enzyme capable of stably binding the coelenterazine substrate.
45 . A method to detect an interaction between a first protein and a second protein in a sample, the method comprising:
(a) contacting a sample with a coelenterazine substrate and the compound of any one of claims 1 - 36 , wherein the sample comprises:
(xix) a first polynucleotide encoding a first fusion protein, wherein the first fusion protein comprises an Oplophorus -derived luciferase and a first protein, wherein the Oplophorus -derived luciferase is a bioluminescent donor; and
(xx) a second polynucleotide encoding a second fusion protein, wherein the second fusion protein comprises a fluorescent acceptor molecule and a second protein;
(b) detecting bioluminescence resonance energy transfer (BRET) in the sample, indicating an interaction or close proximity of the bioluminescent donor and the fluorescence acceptor.
46 . The method of any one of claims 41 - 45 , wherein the sample comprises a cell.
47 . The method of claim 46 , wherein the cell comprises the Oplophorus -derived luciferase.
48 . The method of claim 46 , wherein the cell expresses the Oplophorus -derived luciferase.
49 . The method of any one of claims 41 - 48 , wherein the coelenterazine substrate is a coelenterazine, coelenterazine derivatives, coelenterazine analogs, pro-coelenterazine, or quinone-masked coelenterazine.
50 . A bioluminescence resonance energy transfer (BRET) system comprising: a first fusion protein including a first target protein and a bioluminescence donor molecule, wherein the bioluminescence donor molecule is an Oplophorus -derived luciferase ; a second fusion protein including a second target protein and a fluorescent acceptor molecule; a coelenterazine substrate, and the compound of any one of claims 1 - 36 .
51 . A kit comprising:
(a) a compound of any one of claims 1 - 36 ; and (b) an Oplophorus -derived luciferase.
52 . The kit of claim 51 , wherein the Oplophorus -derived luciferase comprises a polypeptide sequence of SEQ ID NO: 2.
53 . The kit of any of claims 51 - 52 , further comprising a coelenterazine substrate.
54 . The kit of any of claims 51 - 53 , further comprising instructions for carrying out a luminescent assay.
55 . A method for modulating luminescence of an Oplophorus -derived luciferase in a sample, the method comprising:
(a) contacting the sample with a coelenterazine substrate and a compound of formula (II):
wherein:
the dashed line represents the presence or absence of a bond;
n is 0, 1, 2, 3, 4 or 5;
X is CH, N, O, or S;
wherein, when the dashed line represents the presence of a bond, X is CH or N, and when the dashed line represents the absence of a bond, X is O or S;
A is an optionally substituted phenyl ring, or an optionally substituted 5- or 6-membered heteroaryl ring;
R 1 and R 2 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted arylalkyl, optionally substituted alkoxyalkyl and optionally substituted alkoxyalkoxyalkyl;
Z is selected from the group consisting of —NR 3 R 4 and —OR 5 ; and
R 3 , R 4 and R 5 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted C 3 -C 8 -cycloalkyl, optionally substituted C 3 -C 8 -cycloalkylalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, optionally substituted heterocyclyl, and optionally substituted heterocyclylalkyl; or R 3 and R 4 , together with the nitrogen atom to which they are attached, together form an optionally substituted ring; and
(b) detecting luminescence in the sample,
wherein the compound of formula (II) causes a decrease in the luminescence from the Oplophorus -derived luciferase.
56 . A method to detect an interaction between a first protein and a second protein in a sample, the method comprising:
(a) contacting a sample with a coelenterazine substrate and a compound of formula (II):
wherein:
the dashed line represents the presence or absence of a bond;
n is 0, 1, 2, 3, 4 or 5;
X is CH, N, O, or S;
wherein, when the dashed line represents the presence of a bond, X is CH or N, and when the dashed line represents the absence of a bond, X is O or S;
A is an optionally substituted phenyl ring, or an optionally substituted 5- or 6-membered heteroaryl ring;
R 1 and R 2 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted arylalkyl, optionally substituted alkoxyalkyl and optionally substituted alkoxyalkoxyalkyl;
Z is selected from the group consisting of —NR 3 R 4 and —OR 5 ; and
R 3 , R 4 and R 5 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted C 3 -C 8 -cycloalkyl, optionally substituted C 3 -C 8 -cycloalkylalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, optionally substituted heterocyclyl, and optionally substituted heterocyclylalkyl; or R 3 and R 4 , together with the nitrogen atom to which they are attached, together form an optionally substituted ring,
wherein the sample comprises:
(xxi) a first polynucleotide encoding a first fusion protein, wherein the first fusion protein comprises a first fragment of an Oplophorus -derived luciferase and a first protein; and
(xxii) a second polynucleotide encoding a second fusion protein, wherein the second fusion protein comprises a second fragment of the Oplophorus -derived luciferase and a second protein; and
(b) detecting luminescence in the sample,
wherein the detection of luminescence indicates an interaction between the first protein and the second protein.
57 . The method of any of claims 55 - 56 , comprising contacting the sample with the coelenterazine substrate prior to contacting the sample with the compound of formula (II).
58 . The method of claim 57 , wherein when the first protein and second protein interact, the first fragment of the Oplophorus -derived luciferase and the second fragment of the Oplophorus -derived luciferase reconstitute a full-length enzyme capable of stably binding the coelenterazine substrate.
59 . A method to detect an interaction between a first protein and a second protein in a sample, the method comprising:
(a) contacting a sample with a coelenterazine substrate and a compound of formula (II):
wherein:
the dashed line represents the presence or absence of a bond;
n is 0, 1, 2, 3, 4 or 5;
X is CH, N, O, or S;
wherein, when the dashed line represents the presence of a bond, X is CH or N, and when the dashed line represents the absence of a bond, X is O or S;
A is an optionally substituted phenyl ring, or an optionally substituted 5- or 6-membered heteroaryl ring;
R 1 and R 2 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted arylalkyl, optionally substituted alkoxyalkyl and optionally substituted alkoxyalkoxyalkyl;
Z is selected from the group consisting of —NR 3 R 4 and —OR 5 ; and
R 3 , R 4 and R 5 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted C 3 -C 8 -cycloalkyl, optionally substituted C 3 -C 8 -cycloalkylalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, optionally substituted heterocyclyl, and optionally substituted heterocyclylalkyl; or R 3 and R 4 , together with the nitrogen atom to which they are attached, together form an optionally substituted ring,
wherein the sample comprises:
(xxiii) a first polynucleotide encoding a first fusion protein, wherein the first fusion protein comprises an Oplophorus -derived luciferase and a first protein, wherein the Oplophorus -derived luciferase is a bioluminescent donor; and
(xxiv) a second polynucleotide encoding a second fusion protein, wherein the second fusion protein comprises a fluorescent acceptor molecule and a second protein;
(b) detecting bioluminescence resonance energy transfer (BRET) in the sample, indicating an interaction or close proximity of the bioluminescent donor and the fluorescence acceptor.
60 . The method of any one of claims 55 - 59 , wherein the sample comprises a cell.
61 . The method of claim 60 , wherein the cell comprises the Oplophorus -derived luciferase.
62 . The method of claim 60 , wherein the cell expresses the Oplophorus -derived luciferase.
63 . The method of any one of claims 55 - 62 , wherein the coelenterazine substrate is coelenterazine substrate is a coelenterazine, coelenterazine derivatives, coelenterazine analogs, pro-coelenterazine, or quinone-masked coelenterazine.
64 . A bioluminescence resonance energy transfer (BRET) system comprising: a first fusion protein including a first target protein and a bioluminescence donor molecule, wherein the bioluminescence donor molecule is an Oplophorus -derived luciferase; a second fusion protein including a second target protein and a fluorescent acceptor molecule; a coelenterazine substrate; and a compound formula (II):
wherein:
the dashed line represents the presence or absence of a bond;
n is 0, 1, 2, 3, 4 or 5;
X is CH, N, O, or S;
wherein, when the dashed line represents the presence of a bond, X is CH or N, and when the dashed line represents the absence of a bond, X is O or S;
A is an optionally substituted phenyl ring, or an optionally substituted 5- or 6-membered heteroaryl ring;
R 1 and R 2 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted arylalkyl, optionally substituted alkoxyalkyl and optionally substituted alkoxyalkoxyalkyl;
Z is selected from the group consisting of —NR 3 R 4 and —OR 5 ; and
R 3 , R 4 and R 5 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted C 3 -C 8 -cycloalkyl, optionally substituted C 3 -C 8 -cy cl oal kyl al kyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, optionally substituted heterocyclyl, and optionally substituted heterocyclylalkyl; or R 3 and R 4 , together with the nitrogen atom to which they are attached, together form an optionally substituted ring.
65 . A kit comprising:
(a) a compound of formula (II):
wherein:
the dashed line represents the presence or absence of a bond;
n is 0, 1, 2, 3, 4 or 5;
X is CH, N, O, or S;
wherein, when the dashed line represents the presence of a bond, X is CH or N, and when the dashed line represents the absence of a bond, X is O or S;
A is an optionally substituted phenyl ring, or an optionally substituted 5- or 6-membered heteroaryl ring;
R 1 and R 2 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted arylalkyl, optionally substituted alkoxyalkyl and optionally substituted alkoxyalkoxyalkyl;
Z is selected from the group consisting of —NR 3 R 4 and —OR 5 ; and
R 3 , R 4 and R 5 are each independently selected from the group consisting of hydrogen, optionally substituted C 1 -C 8 alkyl, optionally substituted C 3 -C 8 -cycloalkyl, optionally substituted C 3 -C 8 -cycloalkylalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, optionally substituted heterocyclyl, and optionally substituted heterocyclylalkyl; or R 3 and R 4 , together with the nitrogen atom to which they are attached, together form an optionally substituted ring; and
(b) an Oplophorus -derived luciferase.
66 . The kit of claim 65 , wherein the Oplophorus -derived luciferase comprises a polypeptide sequence of SEQ ID NO: 2.
67 . The kit of any of claims 65 - 66 , further comprising a coelenterazine substrate.
68 . The kit of any of claims 65 - 67 , further comprising instructions for carrying out a luminescent assay.Join the waitlist — get patent alerts
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