- Development of Covalent Ligands for G Protein-Coupled Receptors: A Case for the Human Adenosine A3 Receptor
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The development of covalent ligands for G protein-coupled receptors (GPCRs) is not a trivial process. Here, we report a streamlined workflow thereto from synthesis to validation, exemplified by the discovery of a covalent antagonist for the human adenosine A3 receptor (hA3AR). Based on the 1H,3H-pyrido[2,1-f]purine-2,4-dione scaffold, a series of ligands bearing a fluorosulfonyl warhead and a varying linker was synthesized. This series was subjected to an affinity screen, revealing compound 17b as the most potent antagonist. In addition, a nonreactive methylsulfonyl derivative 19 was developed as a reversible control compound. A series of assays, comprising time-dependent affinity determination, washout experiments, and [35S]GTPγS binding assays, then validated 17b as the covalent antagonist. A combined in silico hA3AR-homology model and site-directed mutagenesis study was performed to demonstrate that amino acid residue Y2657.36 was the unique anchor point of the covalent interaction. This workflow might be applied to other GPCRs to guide the discovery of covalent ligands.
- Yang, Xue,Van Veldhoven, Jacobus P. D.,Offringa, Jelle,Kuiper, Boaz J.,Lenselink, Eelke B.,Heitman, Laura H.,Van Der Es, Daan,Ijzerman, Adriaan P.
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p. 3539 - 3552
(2019/04/16)
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- Sulfur(VI) fluoride compounds and methods for the preparation thereof
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This application describes a compound represented by Formula (I): (I) wherein: Y is a biologically active organic core group comprising one or more of an aryl group, a heteroaryl aryl group, a nonaromatic hydrocarbyl group, and a nonaromatic heterocyclic group, to which Z is covalently bonded; n is 1, 2, 3, 4 or 5; m is 1 or 2; Z is O, NR, or N; X1 is a covalent bond or —CH2CH2—, X2 is O or NR; and R comprises H or a substituted or unsubstituted group selected from an aryl group, a heteroaryl aryl group, a nonaromatic hydrocarbyl group, and a nonaromatic heterocyclic group. Methods of preparing the compounds, methods of using the compounds, and pharmaceutical compositions comprising the compounds are described as well.
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Page/Page column 20; 45; 46
(2018/11/23)
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- A study of the reactivity of S(VI)-F containing warheads with nucleophilic amino-acid side chains under physiological conditions
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Sulfonyl fluorides (SFs) have recently emerged as a promising warhead for the targeted covalent modification of proteins. Despite numerous examples of the successful deployment of SFs as covalent probe compounds, a detailed exploration of the factors influencing the stability and reactivity of SFs has not yet appeared. In this work we present an extensive study on the influence of steric and electronic factors on the reactivity and stability of the SF and related SVI-F groups. While SFs react rapidly with N-acetylcysteine, the resulting adducts were found to be unstable, rendering SFs inappropriate for the durable covalent inhibition of cysteine residues. In contrast, SFs afforded stable adducts with both N-acetyltyrosine and N-acetyllysine; furthermore, we show that the reactivity of arylsulfonyl fluorides towards these nucleophilic amino acids can be predictably modulated by adjusting the electronic properties of the warhead. These trends were largely conserved when the covalent reaction occurred within a protein binding pocket. We have also obtained a crystal structure depicting covalent modification of the catalytic lysine of a tyrosine kinase (FGFR1) by the ATP analog 5′-O-3-((fluorosulfonyl)benzoyl)adenosine (m-FSBA). Highly reactive warheads were demonstrated to be unstable with respect to hydrolysis in buffered aqueous solutions, indicating that warhead reactivity must be carefully tuned to provide optimal rates of protein modification. Our results demonstrate that the reactivity of SFs complements that of more commonly studied acrylamides, and we hope that this work spurs the rational design of novel SF-containing covalent probe compounds and inhibitors, particularly in cases where a suitably positioned cysteine residue is not present.
- Mukherjee,Debreczeni,Breed,Tentarelli,Aquila,Dowling,Whitty,Grimster
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supporting information
p. 9685 - 9695
(2017/11/30)
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- Aromatic sulfonyl fluorides covalently kinetically stabilize transthyretin to prevent amyloidogenesis while affording a fluorescent conjugate
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Molecules that bind selectively to a given protein and then undergo a rapid chemoselective reaction to form a covalent conjugate have utility in drug development. Herein a library of 1,3,4-oxadiazoles substituted at the 2 position with an aryl sulfonyl fluoride and at the 5 position with a substituted aryl known to have high affinity for the inner thyroxine binding subsite of transthyretin (TTR) was conceived of by structure-based design principles and was chemically synthesized. When bound in the thyroxine binding site, most of the aryl sulfonyl fluorides react rapidly and chemoselectively with the pK a-perturbed K15 residue, kinetically stabilizing TTR and thus preventing amyloid fibril formation, known to cause polyneuropathy. Conjugation t50s range from 1 to 4 min, ～1400 times faster than the hydrolysis reaction outside the thyroxine binding site. X-ray crystallography confirms the anticipated binding orientation and sheds light on the sulfonyl fluoride activation leading to the sulfonamide linkage to TTR. A few of the aryl sulfonyl fluorides efficiently form conjugates with TTR in plasma. Eleven of the TTR covalent kinetic stabilizers synthesized exhibit fluorescence upon conjugation and therefore could have imaging applications as a consequence of the environment sensitive fluorescence of the chromophore.
- Grimster, Neil P.,Connelly, Stephen,Baranczak, Aleksandra,Dong, Jiajia,Krasnova, Larissa B.,Sharpless, K. Barry,Powers, Evan T.,Wilson, Ian A.,Kelly, Jeffery W.
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p. 5656 - 5668
(2013/06/04)
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