3-(1-methylpyrrolidin-2-yl)pyridine (R,S)-nicotine (RS)-nicotine nicotine Nicotin US9284322, Nicotine Nikotin BDBM50004108 US11667638, Example Nicotine Nicotine-(+) US9993465, Nicotine (+-)-nicotine CHEMBL440464 US8609708, 54 Nicotine
BDBM82070 US9303017, Nicotine Nicotine-D salicylate US10667515, Compound (S)-nicotine CAS_29790-52-1 NICOTINE-L (BASE)
Nicotine-L salicylate Nicotine-D salicylate CHEMBL9732 BDBM82068
3-(2-(N-methylpyrrolidinyl))pyridine (S)-3-(N-methylpyrrolidin-2-yl)pyridine 3-(N-methylpyrollidino)pyridine (S)-(-)-nicotine (S)-nicotine BDBM50035403 3-[(2S)-1-methylpyrrolidin-2-yl]pyridine (-)-nicotine NICOTINE-L (BASE) L(-)-nicotine
(R)-3-(1-methyl-2-pyrrolidinyl)pyridine (+)-nicotine 3-[(2R)-1-methylpyrrolidin-2-yl]pyridine d-nicotine CHEMBL9732 BDBM50035407 (R)-nicotine pseudonicotine
S(-)nicotine-B BDBM50170270 CHEMBL182530
US8609708, 92 Nicotine N-1'-Oxide BDBM109779 US8609708, 92 Nicotine N-1′-Oxide
BDBM444834 US10667515, Compound 6-methyl-nicotine
BDBM50099560 CHEMBL282473 boron-containing nicotine analogue
BDBM50099563 CHEMBL21553 boron-containing nicotine analogue
CHEMBL277416 BDBM50099561 boron-containing nicotine analogue
BDBM109780 US8609708, 93 Nicotine Δ1'-5'-iminium ion US8609708, 93 Nicotine Delta 1'-5'-iminium ion
BDBM444833 US10667515, Compound (S)-5-bromo-nicotine
CHEMBL364003 BDBM50475399 US10667515, Compound 5-methyl-nicotine
US10667515, Compound (S)-nicotine-5-carboxaldehyde BDBM444838
nicotine 3-heteroaromatic analogue 25 BDBM12366 3-Phenylthiophene
(S)-1'-demethylnicotine 3-[(2S)-pyrrolidin-2-yl]pyridine CHEMBL6200 BDBM50035415 l-nor-nicotine 1'-demethyl nicotine S-(-)-nornicotine nornicotine US8609708, 90 Nornicotine
3-Methyl-4-phenylthiophene BDBM12365 nicotine 3-heteroaromatic analogue 24
(5-Phenylthiophen-2-yl)methanamine nicotine 3-heteroaromatic analogue 23 BDBM12364
nicotine 3-heteroaromatic analogue 22 BDBM12363 (5-Phenylfuran-2-yl)methanamine
nicotine 3-heteroaromatic analogue 30 BDBM12369 3-(Pyridin-3-yl)propan-1-amine
nicotine 3-heteroaromatic analogue 8 US8609708,11 CHEMBL361153 BDBM12355 3-(thiophen-3-yl)pyridine
nicotine 3-heteroaromatic analogue 6 3-(5-methyl-1H-imidazol-1-yl)pyridine BDBM12353
5-(1-Methyl-pyrrolidin-2-yl)-2-phenethyl-pyridine CHEMBL190295 (+/-)6-(2-Phenylethyl)nicotine BDBM50168255
US8609708, 4 CHEMBL179669 3-(3-methylthiophen-2-yl)pyridine nicotine 3-heteroaromatic analogue 4 BDBM12351
nicotine 3-heteroaromatic analogue 11 BDBM12358 CHEMBL179704 3-(4-methylthiophen-3-yl)pyridine US8609708, 17 US8609708,17
nicotine 3-heteroaromatic analogue 7 US8609708,10 BDBM12354 2-fluoro-5-(3-methylthiophen-2-yl)pyridine CHEMBL179621
nicotine 3-heteroaromatic analogue 9 US8609708,13 CHEMBL368883 BDBM12356 3-(2-methyl-1H-imidazol-1-yl)pyridine cid_16654686
3-(1H-pyrazol-5-yl)pyridine nicotine 3-heteroaromatic analogue 27 3-(1H-Pyrazol-3-yl)pyridine BDBM12368
3-(pyridin-3-yl)prop-2-yn-1-amine CHEMBL360541 US8609708, 3 BDBM12348 nicotine 3-heteroaromatic analogue 3a
US8609708, 5 3-(1-methyl-1H-imidazol-4-yl)pyridine BDBM12352 CHEMBL360999 nicotine 3-heteroaromatic analogue 5 cid_11344157
US8609708,16 CHEMBL178938 nicotine 3-heteroaromatic analogue 2b methyl({[5-(pyridin-3-yl)furan-2-yl]methyl})amine BDBM12346
CHEMBL369285 nicotine 3-heteroaromatic analogue 1b BDBM12343 methyl({[5-(pyridin-3-yl)thiophen-2-yl]methyl})amine US8609708, 9
US8609708, 2 nicotine 3-heteroaromatic analogue 2a BDBM12345 CHEMBL178090 US8609708, 47 [5-(pyridin-3-yl)furan-2-yl]methanamine
US8609708, 56 nicotine 3-heteroaromatic analogue 2c BDBM12347 dimethyl({[5-(pyridin-3-yl)furan-2-yl]methyl})amine CHEMBL360998
US8609708, 75 dimethyl({[5-(pyridin-3-yl)thiophen-2-yl]methyl})amine CHEMBL361969 BDBM12344 nicotine 3-heteroaromatic analogue 1c
US8609708,14 nicotine 3-heteroaromatic analogue 10 CHEMBL178516 BDBM12357 JMC514968 Compound 7 US8609708, 14 3-(1H-imidazol-4-yl)pyridine
4-methyl-2-(pyridin-3-yl)-1,3-thiazole BDBM12367 nicotine 3-heteroaromatic analogue 26 3-(4-Methylthiazol-2-yl)pyridine
BDBM12349 nicotine 3-heteroaromatic analogue 3b CHEMBL149808 US8609708, 26 methyl[3-(pyridin-3-yl)prop-2-yn-1-yl]amine
CHEMBL178681 US8609708, 31 [5-(pyridin-3-yl)thiophen-2-yl]methanamine US8609708, 1 nicotine 3-heteroaromatic analogue 1a BDBM12341 CHEMBL359657
US8609708, 76 nicotine 3-heteroaromatic analogue 3c CHEMBL180270 BDBM12350 dimethyl[3-(pyridin-3-yl)prop-2-yn-1-yl]amine
(5-Pyridin-3-yl-furan-2-yl)methanethiol BDBM12360 nicotine 3-heteroaromatic analogue 14 [5-(pyridin-3-yl)furan-2-yl]methanethiol
3-(5-((Methylthio)methyl)furan-2-yl)pyridine 3-{5-[(methylsulfanyl)methyl]furan-2-yl}pyridine nicotine 3-heteroaromatic analogue 15 BDBM12361
3-[5-(1,3-dithiolan-2-yl)furan-2-yl]pyridine 3-(5-[1,3]Dithiolan-2-yl-furan-2-yl)pyridine BDBM12362 nicotine 3-heteroaromatic analogue 17
1-({[5-(pyridin-3-yl)furan-2-yl]methyl}sulfanyl)ethan-1-one BDBM12359 S-(5-(Pyridin-3-yl)furan-2-yl)methyl ethanethioate nicotine 3-heteroaromatic analogue 13
[3-(3-pyridyl)isoxazol-5-yl]methylamine BDBM12371 (3-(Pyridin-3-yl)isoxazol-5-yl)methanamine Dihydrochloride nicotine 3-heteroaromatic analogue 35 [3-(pyridin-3-yl)-1,2-oxazol-5-yl]methanamine dihydrochloride
BDBM12370 [3-(pyridin-3-yl)-1H-pyrazol-5-yl]methanamine (3-(Pyridin-3-yl)-1H-pyrazol-5-yl)methanamine nicotine 3-heteroaromatic analogue 32 [3-(3-pyridyl)-1H-pyrazol-5-yl]methylamine;hydrochloride
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- ChEMBL_1504716 (CHEMBL3594852) Displacement of [3H]-nicotine from alpha4beta2 nicotine acetylcholine receptor (unknown origin)
- ChEMBL_144194 (CHEMBL750736) Inhibition of [3H]MLA binding in presence of (-)-nicotine
- ChEMBL_143082 (CHEMBL748732) inhibition of (-)-[3H]nicotine binding to torpedo electroplax membranes
- ChEMBL_144188 (CHEMBL750730) Inhibition of [125I]iodo-MLA binding in presence of (-)-nicotine
- ChEMBL_2023079 (CHEMBL4676892) Displacement of [3H]nicotine from NPY2R (unknown origin)
- ChEMBL_492116 (CHEMBL947458) Displacement of [3H]nicotine from chick alpha4beta2 nAChR
- ChEMBL_797629 (CHEMBL1943747) Displacement of [3H]nicotine from nAChR alpha4beta2 receptor
- ChEMBL_2023046 (CHEMBL4676859) Inhbition of [3H] nicotine binding to rat brain alpha4beta2 nAChR
- ChEMBL_2023078 (CHEMBL4676891) Displacement of [3H]nicotine from Thromboxane A2 (unknown origin)
- ChEMBL_947917 (CHEMBL2343660) Displacement of [3H]nicotine from alpha4beta2 nAChR (unknown origin)
- ChEMBL_1276107 (CHEMBL3089188) Antagonist activity at human alpha4beta2 nAChR assessed as inhibition of nicotine-induced [86Rb+] efflux preincubated for 10 mins before nicotine exposure by cell-based liquid scintillation counting
- ChEMBL_1276104 (CHEMBL3089185) Antagonist activity at rat alpha3beta4 nAChR expressed in HEK293 cells assessed as inhibition of nicotine-induced [86Rb+] efflux preincubated for 10 mins before nicotine exposure by liquid scintillation counting
- ChEMBL_1283691 (CHEMBL3108123) Positive allosteric modulation of human alpha4beta2alpha5 nACHR expressed in HEK-tsA201 cells assessed as potentiation of nicotine-induced current preincubated for 15 mins followed by nicotine-treatment by fluorometric analysis
- ChEMBL_142763 (CHEMBL750485) Binding affinity for nAChR with [3H]-nicotine in rat alpha4 beta2 membranes
- ChEMBL_143718 (CHEMBL753469) Inhibition of [3H]-nicotine binding at the nicotinic acetylcholine receptor alpha4-beta2
- ChEMBL_1928142 (CHEMBL4431214) Displacement of [3H]-nicotine from alpha4beta2 nAChR in rat cortical membranes
- ChEMBL_2023077 (CHEMBL4676890) Displacement of [3H]nicotine from dopamine D3 receptor (unknown origin)
- ChEMBL_2023082 (CHEMBL4676895) Displacement of [3H]nicotine from B2 bradykinin receptor (unknown origin)
- ChEMBL_2023083 (CHEMBL4676896) Displacement of [3H]nicotine from Histamine H3 receptor (unknown origin)
- ChEMBL_2023084 (CHEMBL4676897) Displacement of [3H]nicotine from muscarinic M3 receptor (unknown origin)
- ChEMBL_2023085 (CHEMBL4676898) Displacement of [3H]nicotine from muscarinic M2 receptor (unknown origin)
- ChEMBL_2023086 (CHEMBL4676899) Displacement of [3H]nicotine from muscarinic M1 receptor (unknown origin)
- ChEMBL_2023088 (CHEMBL4676901) Displacement of [3H]nicotine from Endothelin receptor type A (unknown origin)
- ChEMBL_2023090 (CHEMBL4676903) Displacement of [3H]nicotine from cholecystokinin receptor type A (unknown origin)
- ChEMBL_217461 (CHEMBL823510) Inhibition of [3H](-)-nicotine binding to recombinant rat alpha4-beta2 nAChR.
- ChEMBL_497243 (CHEMBL1006196) Displacement of [3H]nicotine from alpha4beta2 nAChR in rat brain membrane
- ChEMBL_509576 (CHEMBL1000462) Displacement of [3H]nicotine from alpha4beta2 nAChR in rat cortical membrane
- ChEMBL_509577 (CHEMBL1000463) Displacement of [3H]nicotine from alpha4beta2 nAChR in rat brain membrane
- ChEMBL_509580 (CHEMBL1000466) Displacement of [3H]nicotine from alpha4beta2 nAChR in rat cerebral membrane
- Inhibition Assay (Testosterone Hydroxylase) To gain insight into the selectivity of the synthetic compounds, nicotine, nicotine related alkaloids and nicotine metabolites for inhibition of other CYPs, we examined the major CYP present in human liver (i.e., CYP 3A4). That the CYP2A6 inhibitors showed low or no inhibitory activity against CYP3A4 suggests that the inhibitors examined selectively inhibited CYP2A6.
- ChEMBL_2023005 (CHEMBL4676818) Displacement of [3H]nicotine from rat alpha4beta2 nAChR by liquid scintillation counting
- ChEMBL_2023072 (CHEMBL4676885) Displacement of [3H]nicotine from 5-hydroxytryptamine receptor 3A (unknown origin)
- ChEMBL_2023073 (CHEMBL4676886) Displacement of [3H]nicotine from 5-hydroxytryptamine receptor 2A (unknown origin)
- ChEMBL_2023074 (CHEMBL4676887) Displacement of [3H]nicotine from 5-hydroxytryptamine receptor 1A (unknown origin)
- ChEMBL_2023087 (CHEMBL4676900) Displacement of [3H]nicotine from Endothelin receptor type B (unknown origin)
- ChEMBL_2023089 (CHEMBL4676902) Displacement of [3H]nicotine from cholecystokinin receptor type B (unknown origin)
- ChEMBL_303228 (CHEMBL827193) Inhibition of [3H]nicotine binding to rat alpha4-beta2 nicotinic acetylcholine receptor
- ChEMBL_877366 (CHEMBL2182411) Displacement of [3H]nicotine human alpha4beta2 nAChR in SH-EP1 cell membranes
- ChEMBL_946142 (CHEMBL2343544) Non-competitive antagonist activity at human alpha3beta4 nAChR expressed in CHO cells assessed as inhibition of nicotine-induced calcium influx incubated for 30 mins prior to nicotine-induction by calcium-4 based FLIPR analysis
- ChEMBL_143866 (CHEMBL751222) Inhibition of [3H]nicotine binding to Nicotinic acetylcholine receptor alpha4-beta2 in rat brain
- ChEMBL_143872 (CHEMBL751228) Inhibition of [3H]nicotine binding to alpha4-beta2 nACh receptor from rat membranes
- ChEMBL_1504717 (CHEMBL3594853) Displacement of [3H]-methyllycaconitine from alpha7 nicotine acetylcholine receptor in rat brain membranes
- ChEMBL_497108 (CHEMBL998586) Activation of human alpha3beta2 nAChR assessed as potentiation of submaximal response to nicotine induced current
- ChEMBL_497109 (CHEMBL998587) Activation of human alpha4beta2 nAChR assessed as potentiation of submaximal response to nicotine induced current
- ChEMBL_497110 (CHEMBL998588) Activation of human alpha3beta4 nAChR assessed as potentiation of submaximal response to nicotine induced current
- ChEMBL_497114 (CHEMBL998592) Activation of human alpha7 nAChR assessed as potentiation of submaximal response to nicotine induced current
- ChEMBL_143715 (CHEMBL753466) Binding affinity towards Nicotinic acetylcholine receptor alpha4-beta2 using [3H]nicotine as radioligand
- ChEMBL_143864 (CHEMBL750613) Inhibition of [3H]nicotine binding to Nicotinic acetylcholine receptor alpha4-beta2 from rat membranes
- ChEMBL_143867 (CHEMBL751223) Inhibition of [3H]nicotine binding to Nicotinic acetylcholine receptor alpha4-beta2 in rat brain membranes
- ChEMBL_2023033 (CHEMBL4676846) Inhbition of [3H] nicotine binding to rat brain alpha4beta2 nAChR by Cheng-Prusoff equation analysis
- ChEMBL_217462 (CHEMBL823511) Inhibition of [3H](-)-nicotine binding to recombinant rat alpha4-beta2 nAChR was determined; experiment 1
- ChEMBL_217463 (CHEMBL821650) Inhibition of [3H](-)-nicotine binding to recombinant rat alpha4-beta2 nAChR was determined; experiment 2
- ChEMBL_303459 (CHEMBL839719) Inhibition of [3H]nicotine binding to alpha4-beta2 nicotinic acetylcholine receptor of rat brain membrane
- ChEMBL_303479 (CHEMBL838853) Inhibition of [3H]nicotine binding to alpha4-beta2 nicotinic acetylcholine receptor of rat brain membrane
- ChEMBL_303639 (CHEMBL828849) Inhibition of [3H]nicotine binding to Nicotinic acetylcholine receptor alpha4-beta2 of rat cortical membranes
- ChEMBL_873560 (CHEMBL2187477) Displacement of [3H]nicotine from human alpha4beta2 nAChR expressed in human SH-EP1 cells
- ChEMBL_142765 (CHEMBL750486) Binding affinity towards Nicotinic acetylcholine receptor by the displacement of [3H]-nicotine from rat cortical membranes
- ChEMBL_143704 (CHEMBL756004) Inhibition of binding of [3H]nicotine to Nicotinic acetylcholine receptor alpha4-beta2 in rat cerebral cortical membranes
- ChEMBL_303690 (CHEMBL829735) Inhibition of [3H](-)-nicotine binding to rat brain (minus cerebellum) Nicotinic acetylcholine receptor alpha4-beta2
- ChEMBL_320944 (CHEMBL882463) Inhibition constant against [3H]nicotine binding to vesicular monoamine transporter-2 of rat brain membranes
- ChEMBL_320958 (CHEMBL885365) Binding affinity for rat brain Nicotinic acetylcholine receptor alpha4-beta2 using [3H]S-(-)-nicotine
- ChEMBL_582486 (CHEMBL1058163) Displacement of L-[3H]Nicotine from alpha4beta2 nAChR in rat brain homogenate by rapid filtration method
- ChEMBL_701134 (CHEMBL1648232) Antagonist activity at alpha6 nAChR in rat striatum assessed as inhibition of nicotine-induced [3H]dopamine release
- ChEMBL_143870 (CHEMBL751226) Inhibition of [3H]nicotine binding to alpha4-beta2 nACh receptor from rat membranes from ref 14
- ChEMBL_143871 (CHEMBL751227) Inhibition of [3H]nicotine binding to alpha4-beta2 nACh receptor from rat membranes from ref 16
- ChEMBL_144327 (CHEMBL751838) Binding affinity at homopentameric Nicotinic acetylcholine receptor alpha-7 subtype using [3H]S-(-)-nicotine as radioligand
- ChEMBL_1451534 (CHEMBL3365055) Displacement of [3H] nicotine from human alpha4beta2 nAChR transfected in SH-EP1 cells by liquid scintillation counting
- ChEMBL_1504713 (CHEMBL3594849) Binding affinity to alpha3beta4 nicotine acetylcholine receptor (unknown origin) after 90 mins by radioligand displacement assay
- ChEMBL_177560 (CHEMBL784318) Inhibitory activity against nAChR mediated nicotine-evoked [3H]DA overflow using rat striatal slices
- ChEMBL_223088 (CHEMBL841331) Inhibition of [3H]nicotine binding to alpha4-beta2 nACh receptor from rat membranes from ref 15
- ChEMBL_223089 (CHEMBL841332) Inhibition of [3H]nicotine binding to alpha4-beta2 nACh receptor from rat membranes from ref 18
- ChEMBL_223090 (CHEMBL841333) inhibition of [3H]nicotine binding to alpha4-beta2 nACh receptor from rat membranes from ref 18
- ChEMBL_851194 (CHEMBL2155622) Agonist activity at human alpha7 nAChR expressed in GH4C1 cells assessed as potentiation of nicotine-induced calcium signal
- ChEMBL_143415 (CHEMBL750057) Potency to displace [3H]nicotine binding to Nicotinic acetylcholine receptor alpha4-beta2 immuno-isolated from M10 cells
- ChEMBL_143865 (CHEMBL750614) Inhibition of [3H]nicotine binding to Nicotinic acetylcholine receptor alpha4-beta2 from rat membranes from ref 14
- ChEMBL_1438837 (CHEMBL3390305) Competitive inhibition of human CYP2A6 in baculovirus-infected insect cell system assessed as nicotine metabolism after 15 mins
- ChEMBL_1438839 (CHEMBL3390307) Irreversible inhibition of human CYP2A6 in baculovirus-infected insect cell system assessed as nicotine metabolism after 15 mins
- ChEMBL_223087 (CHEMBL841330) Inhibition of [3H]nicotine binding to Nicotinic acetylcholine receptor alpha4-beta2 from rat membranes from ref 15
- ChEMBL_303488 (CHEMBL838862) Binding affinity against nicotinic acetylcholine receptor alpha4-beta2 in human HEK293 cells using [3H]- nicotine as radioligand
- ChEMBL_320951 (CHEMBL882470) Binding affinity for human Nicotinic acetylcholine receptor alpha4-beta2 expressed in HEK 293 cells using [3H]nicotine
- ChEMBL_143705 (CHEMBL756005) Inhibition of binding of [3H]nicotine to Nicotinic acetylcholine receptor alpha4-beta2 in rat cerebral cortical membranes at 100 uM concentration
- ChEMBL_143707 (CHEMBL756007) The compound was evaluated for percentage inhibition of binding of [3H]- nicotine to Nicotinic acetylcholine receptor alpha4-beta2 in Rat Cerebral Cortical Membranes
- ChEMBL_143720 (CHEMBL753471) Non-specific binding in presence of 300 uM nicotine at nicotinic acetylcholine receptor alpha4-beta2 in rat cerebral cortex membranes
- ChEMBL_1438838 (CHEMBL3390306) Mixed-type inhibition of human CYP2A6 in baculovirus-infected insect cell system assessed as nicotine metabolism after 15 mins
- ChEMBL_527213 (CHEMBL972002) Activity at rat alpha3beta4 nicotinic receptor expressed in KXalpha-3-beta-4R2 cells assessed as nicotine-induced 86Rb+ efflux
- ChEMBL_872709 (CHEMBL2185122) Displacement of [3H]nicotine from alpha4beta2 nAChR in human SH-EP1 cells after 2 hrs by liquid scintillation assay
- ChEMBL_143553 (CHEMBL755326) Binding affinity to alpha4 beta-2 receptor subtype from mouse fibroblast M10 cells was measured using [3 H]nicotine
- ChEMBL_527212 (CHEMBL972001) Antagonist activity at rat alpha3beta4 nicotinic receptor expressed in KXalpha-3-beta-4R2 cells assessed as nicotine-induced calcium fluorescence
- ChEMBL_974586 (CHEMBL2410354) Displacement of [3H]Nicotine from alpha4beta2 nACHR in human SHEP1 cell membranes after 2 hrs by liquid scintillation counting analysis
- ChEMBL_1504714 (CHEMBL3594850) Displacement of [3H]-MLA from Sprague-Dawley rat brain alpha7 nicotine acetylcholine receptor after 90 mins by radioligand displacement assay
- ChEMBL_1504719 (CHEMBL3594924) Displacement of [3H]-MLA from alpha7 nicotine acetylcholine receptor in rat brain minus cerebellum after 60 mins by scintillation counting analysis
- ChEMBL_466430 (CHEMBL926873) Activation of human alpha-7 nAChR expressed in Xenopus oocytes assessed as modulation of nicotine-induced current by two-electrode voltage-clamp method
- ChEMBL_2355773 Antagonist activity at human alpha3/beta4 nAChR expressed in HEK293 cells assessed as nicotine-induced intracellular calcium release incubated for 15 mins by FLIPR analysis
- ChEMBL_2355774 Antagonist activity at human alpha4/beta2 nAChR expressed in HEK293 cells assessed as nicotine-induced intracellular calcium release incubated for 15 mins by FLIPR analysis
- ChEMBL_872698 (CHEMBL2184678) Apparent antagonist activity at alpha4beta2 nAChR in human SH-EP1 cells assessed as inhibition of nicotine-induced calcium flux by calcium4-based FLIPR assay
- ChEBML_1708057 Antagonist activity at rat alpha3beta4 nAChR expressed in HEK293 cells assessed as inhibition of (S)-nicotine-induced current response measured up to 90 secs by FMP assay
- ChEMBL_1335312 (CHEMBL3238809) Antagonist activity at mouse alpha4/beta2 nAChR expressed in HEK293 cells assessed as inhibition of (S)-nicotine-induced activity by FLIPR membrane potential blue assay
- ChEMBL_1335313 (CHEMBL3238810) Antagonist activity at rat alpha3/beta4 nAChR expressed in HEK293 cells assessed as inhibition of (S)-nicotine-induced activity by FLIPR membrane potential blue assay
- ChEMBL_1504725 (CHEMBL3594930) Displacement of [3H]-alpha-bungarotoxin from alpha7 nicotine acetylcholine receptor in Wistar rat cerebral cortex after 2 hrs by liquid scintillation counting analysis
- ChEMBL_915450 (CHEMBL3076134) Inhibition of [3H]nicotine binding to alpha4 nicotinic acetylcholine receptor in Mus musculus (mouse) M10 cells co-expressing beta2 nicotinic acetylcholine receptor subunits
- ChEMBL_986916 (CHEMBL2438882) Displacement of [3H]nicotine from human alpha4beta2 nAChR expressed in human SH-EP1 cell membranes after 2 hrs by liquid scintillation counting analysis
- ChEMBL_1280131 (CHEMBL3097702) Displacement of [3H]nicotine from alpha4beta2 nAChR expressed in human SH-EP1 clonal cell membranes after 2 to 3 hrs by liquid scintillation counting analysis
- ChEMBL_1708057 (CHEMBL4059290) Antagonist activity at rat alpha3beta4 nAChR expressed in HEK293 cells assessed as inhibition of (S)-nicotine-induced current response measured up to 90 secs by FMP assay
- ChEMBL_1708058 (CHEMBL4059291) Antagonist activity at mouse alpha4beta2 nAChR expressed in HEK293 cells assessed as inhibition of (S)-nicotine-induced current response measured up to 90 secs by FMP assay
- ChEMBL_2145372 (CHEMBL5029652) Uncompetitive type inhibition of human AOX assessed as inhibition constant using nicotine-1(S)-iminium ion as substrate incubated for 2 mins by HPLC-MS analysis
- ChEMBL_915451 (CHEMBL3076135) Inhibition of [3H]nicotine binding to alpha3 nicotinic acetylcholine receptor in Homo sapiens (human) SH-SY5Y cells co-expressing beta2beta4alpha5 Nicotinic acetylcholine receptor subunits
- ChEMBL_992027 (CHEMBL2447428) Type-1 positive allosteric modulation of human alpha7 nAChR expressed in Xenopus oocytes assessed as maximum modulation of nicotine EC5 response by two-electrode voltage clamp method
- ChEMBL_466431 (CHEMBL926874) Activation of human GABAA alpha-2-beta-3-gamma-2L receptor expressed in Xenopus oocytes assessed as modulation of nicotine-induced current by two-electrode voltage-clamp method
- ChEMBL_581328 (CHEMBL1050978) Antagonist activity at nAChR-alpha-6-beta-2 in Sprague-Dawley rat striatal slices assessed as inhibition of nicotine-evoked [3H]dopamine uptake by liquid beta-scintillation spectrometry
- ChEMBL_909609 (CHEMBL3057495) Displacement of [3H]nicotine from Homo sapiens (human) alpha4beta2 nicotinic acetylcholine receptor expressed in Homo sapiens (human) SH-EP1 cells after 2.5 min by scintillation counting
- ChEMBL_2315606 Antagonist activity at nAChR alpha3beta4 in human SH-SY5Y cells assessed as increase in intracellular calcium level preincubated for 10 mins followed by nicotine addition measured after 30 mins by FLIPR assay
- ChEMBL_1508833 (CHEMBL3603305) Antagonist activity at rat alpha-7 nicotinic acetylcholine receptor expressed in Xenopus oocytes assessed as inhibition of acetylcholine-induced currents preincubated for 10 mins followed by nicotine induction by two-electrode voltage-clamp electrophysiology assay
- Functional Assay IC50(10′): The functional properties of the ligands were determined by 86Rb+ efflux assays in cells expressing α3β4 and α4β2 nAChR subtypes. The functional activity of each ligand was measured for its agonism, antagonism and desensitization ability. Agonist activity for each of the ligands was tested at eight different concentrations. The responses were compared to that stimulated by 100 μM (−)-nicotine, a near maximally effective concentration. The potency (IC50(0′)) of each ligand as an antagonist was derived from the full concentration-effect curves. We determined the desensitization potency of each ligand by pre-treating cells with the test compound for 10 minutes before 100 μM (−)-nicotine was applied. The potency of a ligand to desensitize the receptor after a 10 minute exposure (IC50(10′) was obtained with full concentration-effect curves using at least 8 concentrations of the ligand.
- Inhibition Assay To gain insight into the selectivity of the synthetic compounds for inhibition of other CYPs, we examined the major CYPs present in human liver. Prior to these studies we showed that the nicotine analogs were quite metabolically stable in the presence of mouse or rat or human liver microsomes (i.e., T1/2>60 mins). That the nicotine analogs showed low or no inhibitory activity against other CYPs suggests that the inhibitors examined selectively inhibited CYP2A6. A typical incubation mixture (final volume 0.25 mL) contained 50 mM Tris or KPhos buffer (pH 7.5), 0.5 mM NADP+, 2.0 mM G6P, 1 U of G6P dehydrogenase and 0.6 mg DETAPAC and the inhibitor was added last to minimize interaction with the protein. After mixing on ice, the reaction was initiated by the addition of substrate and incubated at 37 ° C. with shaking in air. Organic extracts were analyzed by fluorescence (for fluorometric substrates) or injected onto a Hitachi L-7100 system equipped with a Hitachi L-7400 UV detector.
- Functional Assay The functional properties of the ligands were determined by 86Rb+ efflux assays in cells expressing α3β4 and α4β2 nAChR subtypes. The functional activity of each ligand was measured for its agonism, antagonism and desensitization ability. Agonist activity for each of the ligands was tested at eight different concentrations. The responses were compared to that stimulated by 100 μM (−)-nicotine, a near maximally effective concentration. The full concentration-effect curves generated potency (EC50) and efficacy (Emax) of each ligand.
- Steady-State Kinetics Initial rates of amine oxidation were routinely measured by monitoring oxygen consumption in 0.1 M Hepes (pH 8.0) and 0.1 M NaCl at 25 °C using an oxygen electrode (model 5300A, Yellow Springs Instruments, Yellow Springs, OH). With (R,S)-6-hydroxynicotine and (S)-6-hydroxynornicotine, the concentration of enzyme was typically 0.1 μM; this was increased to 4 μM for (S)-nicotine and (R,S)-4-(1-methyl pyrrolidine-2-yl)phenol. The concentration of oxygen was varied by bubbling the desired concentration of oxygen (62 μM to 1.25 mM) into the oxygen electrode cell for ∼10 min.
- Binding Assay The [3H]-methyllycaconitine binding assay is a modification of the method described by Davies et al. (Neuropharmacol. 1999, 38, 679-690). The P2 pellet is washed twice with binding buffer (50 mM Tris-HCl, 1 mM MgCl2, 120 mM NaCl, 5 mM KCl, 2 mM CaCl2, pH 7.4), and centrifuged (15 000×g, 4° C., 30 min). The P2 membranes are resuspended in binding buffer and incubated in a volume of 250 μl (amount of membrane protein 0.1-0.5 mg) in the presence of 1-5 nM [3H]-methyllycaconitine, 0.1% (w/v) BSA (bovine serum albumin) and various concentrations of the test substance at 21° C. for 2.5 h. The non-specific binding is determined by incubation in the presence of 1 μM □-bungarotoxin or 100 μM nicotine or 10 μM MLA (methyllycaconitine).
- [3H]-Epibatidine Radioligand Binding Assay Briefly, cultured cells at >80% confluence were removed from their flasks (80 cm2) with a disposable cell scraper and placed in 10 mL of 50 mM Tris.HCl buffer (pH 7.4, 4° C.). The cell suspension was centrifuged at 10,000×g for 5 min and the pellet was collected. The cell pellet was then homogenized in 10 mL buffer with a polytron homogenizer and centrifuged at 36,000 g for 10 min at 4° C. The membrane pellet was resuspended in fresh buffer, and aliquots of the membrane preparation were used for binding assays. The concentration of [3H]-epibatidine used was ˜500 pM for competition binding assays. Nonspecific binding was assessed in parallel incubations in the presence of 300 μM nicotine. Bound and free ligands were separated by vacuum filtration through Whatman GF/C filters treated with 0.5% polyethylenimine. The filter-retained radioactivity was measured by liquid scintillation counting. Specific binding was defined as the difference between total binding and nonspecific binding. Data from competition binding assays were analyzed using Prism 5 (GraphPad Software, San Diego, Calif.).
- [3H]-Epibatidine Radioligand Binding Assay Briefly, cultured cells at >80% confluence were removed from their flasks (80 cm^2) with a disposable cell scraper and placed in 10 mL of 50 mM Tris.HCl buffer (pH 7.4, 4 °C.). The cell suspension was centrifuged at 10,000×g for 5 min and the pellet was collected. The cell pellet was then homogenized in 10 mL buffer with a polytron homogenizer and centrifuged at 36,000 g for 10 min at 4 °C. The membrane pellet was resuspended in fresh buffer, and aliquots of the membrane preparation were used for binding assays. The concentration of [3H]-epibatidine used was ~500 pM for competition binding assays. Nonspecific binding was assessed in parallel incubations in the presence of 300 μM nicotine. Bound and free ligands were separated by vacuum filtration through Whatman GF/C filters treated with 0.5% polyethylenimine. The filter-retained radioactivity was measured by liquid scintillation counting. Specific binding was defined as the difference between total binding and nonspecific binding. Data from competition binding assays were analyzed using Prism 5 (GraphPad Software, San Diego, Calif.). The Kd values for [3H]-epibatidine used for calculating Ki values of nAChR subtypes were 0.02 nM for α2β2, 0.08 nM for α2β4, 0.03 nM for α3β2, 0.3 nM for α3β4, 0.04 nM for α4β2, 0.09 nM for α4β4, 1.8 nM for α7 and 0.05 for rat forebrain.
- The [3H]-methyllycaconitine binding assay Rat brain tissue (hippocampus or whole brain) is homogenized in homogenization buffer (10% w/v, 0.32 M sucrose, 1 mM EDTA, 0.1 mM phenylmethylsulphonyl fluoride (PMSF), 0.01% (w/v) NaN3, pH 7.4, 4° C.) at 600 rpm in a glass homogenizer. The homogenate is centrifuged (1000×g, 4° C., 10 min) and the supernatant is removed. The pellet is resuspended (20% w/v) and the suspension is centrifuged (1000×g, 4° C., 10 min). The two supernatants are combined and centrifuged (15 000×g, 4° C., 30 min). The pellet obtained in this way is referred to as the P2 fraction.The P2 pellet is suspended in, binding buffer (50 mM Tris-HCl, 1 mM MgCl2, 120 mM NaCl, 5 mM KCl, 2 mM CaCl2, pH 7.4), and the suspension is centrifuged (15 000×g, 4° C., 30 min), twice.The residue is resuspended in binding buffer and incubated in a volume of 250 (amount of membrane protein 0.1-0.5 mg) in the presence of 1-5 nM [3H]-methyllycaconitine 0.1% (w/v). BSA (bovine serum albumin) and various concentrations of the test substance at 21° C. for 2.5 h. Incubation is then carried out in the presence of 1 μM α-bungarotoxin or 100 μM nicotine or 10 μM MLA (methyllycaconitine).The incubation is stopped by adding 4 ml PBS (20 mM Na2HPO4, 5 mM KH2PO4, 150 mM NaCl, pH 7.4, 4° C.) and filtering through type A/E glass fibre filters (Gelman Sciences) which have previously been placed in 0.3% (v/v) polyethyleneimine (PEI) for 3 h. The filters are washed twice with 4 ml of PBS (4° C.), and the bound radioactivity is determined by scintillation measurement. All the assays are carried out in triplicate. The dissociation constant Ki of the test substance was determined from the IC50 of the compounds (concentration of the test substance at which 50% of the ligand bound to the receptor is displaced), the dissociation constant KD and the concentration L of [3H]-methyllycaconitine using the equation Ki=IC50/(1+L/KD).
- Alpha 7 UAChR Inhibition Assay The [3H]-methyllycaconitine binding assay is a modification of the method described by Davies et al. in Neuropharmacol, 1999, 38, 679-690.Rat brain tissue (hippocampus or whole brain) is homogenized in aqueous homogenization buffer (10% w/v, 0.32 M sucrose, 1 mM EDTA, 0.1 mM phenylmethylsulfonyl fluoride (PMSF), 0.01% (w/v) NaN3, pH 7.4, 4° C.) at 600 rpm in a glass homogenizer. The homogenate is centrifuged (1000×g, 4° C., 10 min) and the supernatant is removed. The pellet is resuspended (20% w/v) and the suspension is centrifuged (1000×g, 4° C., 10 min). The two supernatants are combined and centrifuged (15 000×g, 4° C., 30 min). The pellet obtained in this way is referred to as the P2 fraction.The P2 pellet is suspended in binding buffer (50 mM Tris-HCl, 1 mM MgCl2, 120 mM NaCl, 5 mM KCl, 2 mM CaCl2, pH 7.4), and the suspension is centrifuged (15 000×g, 4° C., 30 min), twice.The residue is resuspended in binding buffer to a concentration of 4 mg/ml and incubated in a volume of 250 μl (amount of membrane protein 0.4 mg) in the presence of 2 nM [3H]-methyllycaconitine, 0.1% (w/v) BSA (bovine serum albumin) and various concentrations of the test substance at 21° C. for 60 min.Incubation is then carried out in the presence of 1 μM α-bungarotoxin or 100 μM nicotine or 10 μM MLA (methyllycaconitine). The incubation is stopped by adding 4 ml of PBS (20 mM Na2HPO4, 5 mM KH2PO4, 150 mM NaCl, pH 7.4, 4° C.) and filtering through type A/E glass fiber filters (Gelman Sciences) which have previously been placed in 0.3% (v/v) polyethyleneirnine (PEI) for 3 h. The filters are washed twice with 4 ml of PBS (4° C.), and the bound radioactivity is determined by scintillation measurement. All the assays are carried out in triplicate. The dissociation constant Ki of the test substance was determined from the IC50 of the compounds (concentration of the test substance at which 50% of the ligand bound to the receptor is displaced), the dissociation constant KD and the concentration L of [3H]-methyllycaconitine using the equation K1=IC50/(1+L/KD).In place of [3H]-methyllycaconitine it is also possible to employ other α7 nAChR-selective radioligands such as, for example, [125I]-α-bungarotoxin or nonselective nAChR radioligands together with inhibitors of other nAChRs.The in vitro data for the effects of the compounds of the invention show a Ki of <300 nM.