Evaluation of a Covalent Library of Diverse Warheads (CovLib) Binding to JNK3, USP7, or p53

被引：2

作者：

Klett, Theresa ^{[1
]}

Schwer, Martin ^{[1
]}

Ernst, Larissa N. ^{[1
]}

Engelhardt, Marc U. ^{[1
]}

Jaag, Simon J. ^{[2
]}

Masberg, Benedikt ^{[2
]}

Knappe, Cornelius ^{[2
]}

Laemmerhofer, Michael ^{[2
]}

Gehringer, Matthias ^{[3
,4
]}

Boeckler, Frank M. ^{[1
,5
]}

机构：

[1] Eberhard Karls Univ Tubingen, Inst Pharmaceut Sci, Dept Pharm & Biochem, Lab Mol Design & Pharmaceut Biophys, Morgenstelle 8 Haus B, D-72076 Tubingen, Germany

[2] Eberhard Karls Univ Tubingen, Inst Pharmaceut Sci, Dept Pharm & Biochem, Pharmaceut Bio Anal, D-72076 Tubingen, Germany

[3] Eberhard Karls Univ Tubingen, Inst Pharmaceut Sci, Dept Pharm & Biochem, Pharmaceut Chem, D-72076 Tubingen, Germany

[4] Eberhard Karls Univ Tubingen, Inst Biomed Engn, Med Chem, D-72076 Tubingen, Germany

[5] Eberhard Karls Univ Tubingen, Interfac Inst Biomed Informat IBMI, D-72076 Tubingen, Germany

来源：

DRUG DESIGN DEVELOPMENT AND THERAPY | 2024年 / 18卷

关键词：

covalent fragment-based drug discovery; differential scanning fluorimetry; 5; 5'-dithiobis-(2-nitrobenzoic acid); intact protein mass spectrometry; glutathione; ENRICHED FRAGMENT LIBRARIES; ACTIVATED PROTEIN-KINASES; MUTANT P53; CRYSTAL-STRUCTURE; NITROGEN-HETEROCYCLES; STRUCTURAL BASIS; DRUG DISCOVERY; CORE DOMAIN; INHIBITORS; CANCER;

D O I：

10.2147/DDDT.S466829

中图分类号：

R914 [药物化学];

学科分类号：

100701 ;

摘要：

Purpose: Over the last few years, covalent fragment-based drug discovery has gained significant importance. Thus, striving for more warhead diversity, we conceived a library consisting of 20 covalently reacting compounds. Our covalent fragment library (CovLib) contains four different warhead classes, including five alpha-cyanoacacrylamides/acrylates (CA), three epoxides (EO), four vinyl sulfones (VS), and eight electron-deficient heteroarenes with a leaving group (SNAr/SN). Methods: After predicting the theoretical solubility of the fragments by LogP and LogS during the selection process, we determined their experimental solubility using a turbidimetric solubility assay. The reactivities of the different compounds were measured in a high-throughput 5,5'-dithiobis-(2-nitrobenzoic acid) DTNB assay, followed by a (glutathione) GSH stability assay. We employed the CovLib in a (differential scanning fluorimetry) DSF-based screening against different targets: c-Jun N-terminal kinase 3 (JNI(3), ubiquitin-specific protease 7 (USP7), and the tumor suppressor p53. Finally, the covalent binding was confirmed by intact protein mass Results: In general, the purchased fragments turned out to be sufficiently soluble. Additionally, they covered a broad spectrum of reactivity. All investigated alpha-cyanoacrylamides/acrylates and all structurally confirmed epoxides turned out to be less reactive compounds, possibly due to steric hindrance and reversibility (for alpha-cyanoacrylamides/acrylates). The SNAr and vinyl sulfone fragments are either highly reactive or stable. DSF measurements with the different targets JNI(3, USP7, and p53 identified reactive fragment hits causing a shift in the melting temperatures of the proteins. MS confirmed the covalent binding mode of all these fragments to USP7 and p53, while additionally identifying the SNAr-type electrophile SN002 as a mildly reactive covalent hit for p53. Conclusion: The screening and target evaluation of the CovLib revealed first interesting hits. The highly cysteine-reactive fragments VS004, SN001, SN006, and SN007 covalently modify several target proteins and showed distinct shifts in the melting temperatures up to +5.1 degrees C and -9.1 degrees C.

引用

页码：2653 / 2679

页数：27