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https://www.readbyqxmd.com/read/29120638/discovery-and-optimization-of-potent-cell-active-pyrazole-based-inhibitors-of-lactate-dehydrogenase-ldh
#1
Ganesha Rai, Kyle R Brimacombe, Bryan T Mott, Daniel J Urban, Xin Hu, Shyh-Ming Yang, Tobie D Lee, Dorian M Cheff, Jennifer Kouznetsova, Gloria A Benavides, Katie Pohida, Eric J Kuenstner, Diane K Luci, Christine M Lukacs, Douglas R Davies, David M Dranow, Hu Zhu, Gary Sulikowski, William J Moore, Gordon M Stott, Andrew J Flint, Matthew D Hall, Victor M Darley-Usmar, Leonard M Neckers, Chi V Dang, Alex G Waterson, Anton Simeonov, Ajit Jadhav, David J Maloney
We report the discovery and medicinal chemistry optimization of a novel series of pyrazole-based inhibitors of human lactate dehydrogenase (LDH). Utilization of a quantitative high-throughput screening paradigm facilitated hit identification, while structure-based design and multiparameter optimization enabled the development of compounds with potent enzymatic and cell-based inhibition of LDH enzymatic activity. Lead compounds such as 63 exhibit low nM inhibition of both LDHA and LDHB, submicromolar inhibition of lactate production, and inhibition of glycolysis in MiaPaCa2 pancreatic cancer and A673 sarcoma cells...
November 9, 2017: Journal of Medicinal Chemistry
https://www.readbyqxmd.com/read/29048945/cell-density-affects-the-detection-of-chk1-target-engagement-by-the-selective-inhibitor-v158411
#2
Clara C Geneste, Andrew J Massey
Understanding drug target engagement and the relationship to downstream pharmacology is critical for drug discovery. Here we have evaluated target engagement of Chk1 by the small-molecule inhibitor V158411 using two different target engagement methods (autophosphorylation and cellular thermal shift assay [CETSA]). Target engagement measured by these methods was subsequently related to Chk1 inhibitor-dependent pharmacology. Inhibition of autophosphorylation was a robust method for measuring V158411 Chk1 target engagement...
October 1, 2017: SLAS Discovery
https://www.readbyqxmd.com/read/29026104/cetsa-quantitatively-verifies-in-vivo-target-engagement-of-novel-ripk1-inhibitors-in-various-biospecimens
#3
Tsuyoshi Ishii, Takuro Okai, Misa Iwatani-Yoshihara, Manabu Mochizuki, Satoko Unno, Masako Kuno, Masato Yoshikawa, Sachio Shibata, Masanori Nakakariya, Takatoshi Yogo, Tomohiro Kawamoto
The proof of target engagement (TE) is a key element for evaluating potential investment in drug development. The cellular thermal shift assay (CETSA) is expected to facilitate direct measurement of intracellular TE at all stages of drug development. However, there have been no reports of applying this technology to comprehensive animal and clinical studies. This report demonstrates that CETSA can not only quantitatively evaluate the drug-TE in mouse peripheral blood, but also confirm TE in animal tissues exemplified by using the receptor interacting protein 1 kinase (RIPK1) lead compound we have developed...
October 12, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28957646/a-high-throughput-dose-response-cellular-thermal-shift-assay-for-rapid-screening-of-drug-target-engagement-in-living-cells-exemplified-using-smyd3-and-ido1
#4
Dean E McNulty, William G Bonnette, Hongwei Qi, Liping Wang, Thau F Ho, Anna Waszkiewicz, Lorena A Kallal, Raman P Nagarajan, Melissa Stern, Amy M Quinn, Caretha L Creasy, Dai-Shi Su, Alan P Graves, Roland S Annan, Sharon M Sweitzer, Marc A Holbert
A persistent problem in early small-molecule drug discovery is the frequent lack of rank-order correlation between biochemical potencies derived from initial screens using purified proteins and the diminished potency and efficacy observed in subsequent disease-relevant cellular phenotypic assays. The introduction of the cellular thermal shift assay (CETSA) has bridged this gap by enabling assessment of drug target engagement directly in live cells based on ligand-induced changes in protein thermal stability...
September 1, 2017: SLAS Discovery
https://www.readbyqxmd.com/read/28890935/ski-178-a-multitargeted-inhibitor-of-sphingosine-kinase-and-microtubule-dynamics-demonstrating-therapeutic-efficacy-in-acute-myeloid-leukemia-models
#5
Jeremy A Hengst, Taryn E Dick, Arati Sharma, Kenichiro Doi, Shailaja Hegde, Su-Fern Tan, Laura M Geffert, Todd E Fox, Arun K Sharma, Dhimant Desai, Shantu Amin, Mark Kester, Thomas P Loughran, Robert F Paulson, David F Claxton, Hong-Gang Wang, Jong K Yun
AIM: To further characterize the selectivity, mechanism-of-action and therapeutic efficacy of the novel small molecule inhibitor, SKI-178. METHODS: Using the state-of-the-art Cellular Thermal Shift Assay (CETSA) technique to detect "direct target engagement" of proteins intact cells, in vitro and in vivo assays, pharmacological assays and multiple mouse models of acute myeloid leukemia (AML). RESULTS: Herein, we demonstrate that SKI-178 directly target engages both Sphingosine Kinase 1 and 2...
July 2017: Cancer Translational Medicine
https://www.readbyqxmd.com/read/28666623/geranylnaringenin-cg902-inhibits-constitutive-and-inducible-stat3-activation-through-the-activation-of-shp-2-tyrosine-phosphatase
#6
Yena Jin, Yae Jin Yoon, Yoon Jung Jeon, Jiyeon Choi, Yu-Jin Lee, Joonku Lee, Sangho Choi, Oyekanmi Nash, Dong Cho Han, Byoung-Mog Kwon
The roles and significance of signal transducer and activator of transcription 3 (STAT3) in human cancers have been extensively studied and STAT3 is a promising therapeutic target for cancer drug discovery. During the screening of natural products to identify STAT3 inhibitors, we identified geranylnaringenin (CG902), which decreased luciferase activity in a dose-dependent manner. CG902 specifically inhibited STAT3 phosphorylation at Tyr-705 in DU145 prostate cancer cells and decreased the expression levels of STAT3 target genes, such as cyclin D1, cyclin A, and survivin...
October 15, 2017: Biochemical Pharmacology
https://www.readbyqxmd.com/read/28631436/characterization-of-a-novel-p110%C3%AE-specific-inhibitor-bl140-that-overcomes-mdv3100-resistance-in-castration-resistant-prostate-cancer-cells
#7
Chenchen He, Shaofeng Duan, Liang Dong, Yifen Wang, Qingting Hu, Chunjing Liu, Marcus L Forrest, Jeffrey M Holzbeierlein, Suxia Han, Benyi Li
BACKGROUND: Our previous studies demonstrated that the class IA PI3K/p110β is critical in castration-resistant progression of prostate cancer (CRPC) and that targeting prostate cancer with nanomicelle-loaded p110β-specific inhibitor TGX221 blocked xenograft tumor growth in nude mice, confirming the feasibility of p110β-targeted therapy for CRPCs. To improve TGX221's aqueous solubility, in this study, we characterized four recently synthesized TGX221 analogs. METHODS: TGX221 analog efficacy were examined in multiple prostate cancer cell lines with the SRB cell growth assay, Western blot assay for AKT phosphorylation and cell cycle protein levels...
August 2017: Prostate
https://www.readbyqxmd.com/read/28174211/overlapping-and-divergent-actions-of-structurally-distinct-histone-deacetylase-inhibitors-in-cardiac-fibroblasts
#8
Katherine B Schuetze, Matthew S Stratton, Weston W Blakeslee, Michael F Wempe, Florence F Wagner, Edward B Holson, Yin-Ming Kuo, Andrew J Andrews, Tonya M Gilbert, Jacob M Hooker, Timothy A McKinsey
Inhibitors of zinc-dependent histone deacetylases (HDACs) profoundly affect cellular function by altering gene expression via changes in nucleosomal histone tail acetylation. Historically, investigators have employed pan-HDAC inhibitors, such as the hydroxamate trichostatin A (TSA), which simultaneously targets members of each of the three zinc-dependent HDAC classes (classes I, II, and IV). More recently, class- and isoform-selective HDAC inhibitors have been developed, providing invaluable chemical biology probes for dissecting the roles of distinct HDACs in the control of various physiologic and pathophysiological processes...
April 2017: Journal of Pharmacology and Experimental Therapeutics
https://www.readbyqxmd.com/read/27641368/the-use-of-cellular-thermal-shift-assay-cetsa-to-study-crizotinib-resistance-in-alk-expressing-human-cancers
#9
Abdulraheem Alshareef, Hai-Feng Zhang, Yung-Hsing Huang, Chengsheng Wu, Jing Dong Zhang, Peng Wang, Ahmed El-Sehemy, Mohamed Fares, Raymond Lai
Various forms of oncogenic ALK proteins have been identified in various types of human cancers. While Crizotinib, an ALK inhibitor, has been found to be therapeutically useful against a subset of ALK(+) tumours, clinical resistance to this drug has been well recognized and the mechanism of this phenomenon is incompletely understood. Using the cellular thermal shift assay (CETSA), we measured the Crizotinib-ALK binding in a panel of ALK(+) cell lines, and correlated the findings with the ALK structure and its interactions with specific binding proteins...
2016: Scientific Reports
https://www.readbyqxmd.com/read/27401582/early-perspective-microplate-applications-of-the-cellular-thermal-shift-assay-cetsa
#10
Brinton Seashore-Ludlow, Thomas Lundbäck
The cellular thermal shift assay (CETSA) was introduced in 2013 as a means to assess drug binding in complex environments such as cell lysates, live cells, and even tissues. The assay principle relies on the well-proven biophysical concept of ligand-induced thermal stabilization of proteins, which in CETSA applications is measured as a persistent presence of soluble protein at elevated temperatures. Given its recent development, we have just started to learn about the benefits and pitfalls of the method as it is applied to a growing number of protein target classes, the majority of which are intracellular soluble proteins...
July 11, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27384680/compounds-from-the-marine-sponge-cribrochalina-vasculum-offer-a-way-to-target-igf-1r-mediated-signaling-in-tumor-cells
#11
Ana Zovko, Metka Novak, Petra Hååg, Dimitry Kovalerchick, Teresa Holmlund, Katarina Färnegårdh, Micha Ilan, Shmuel Carmeli, Rolf Lewensohn, Kristina Viktorsson
In this work two acetylene alcohols, compound 1 and compound 2, which were isolated and identified from the sponge Cribrochalina vasculum, and which showed anti-tumor effects were further studied with respect to targets and action mechanisms. Gene expression analyses suggested insulin like growth factor receptor (IGF-1R) signaling to be instrumental in controlling anti-tumor efficacy of these compounds in non-small cell lung cancer (NSCLC). Indeed compounds 1 and 2 inhibited phosphorylation of IGF-1Rβ as well as reduced its target signaling molecules IRS-1 and PDK1 allowing inhibition of pro-survival signaling...
August 2, 2016: Oncotarget
https://www.readbyqxmd.com/read/27266997/design-and-synthesis-of-n-5-chloro-2-4-dihydroxybenzoyl-r-1-2-3-4-tetrahydroisoquinoline-3-carboxamides-as-novel-hsp90-inhibitors
#12
Chuanpeng Liang, Huilin Hao, Xingkang Wu, Zhenyu Li, Jing Zhu, Chunhua Lu, Yuemao Shen
Heat shock protein 90 (Hsp90) is an attractive chemotherapeutic target for antitumor drug development. Herein, we reported the design and synthesis of two series of novel N-(5-chloro-2,4-dihydroxybenzoyl)-1,2,3,4-tetrahydroisoquinoline-3- carboxamides as Hsp90 inhibitors using (S)-Tic (1,2,3,4-tetrahydroisoquinoline-3- carboxylic acid) (A1-13) and (R)-Tic (B1-13) as scaffold, respectively. Cellular thermal shift assay (CETSA) screening showed that compounds B1-13 with (R)-Tic scaffold exhibited potent ability to stabilize Hsp90α...
October 4, 2016: European Journal of Medicinal Chemistry
https://www.readbyqxmd.com/read/27216142/cellular-thermal-shift-and-clickable-chemical-probe-assays-for-the-determination-of-drug-target-engagement-in-live-cells
#13
Hua Xu, Ariamala Gopalsamy, Erik C Hett, Shores Salter, Ann Aulabaugh, Robert E Kyne, Betsy Pierce, Lyn H Jones
Proof of drug-target engagement in physiologically-relevant contexts is a key pillar of successful therapeutic target validation. We developed two orthogonal technologies, the cellular thermal shift assay (CETSA) and a covalent chemical probe reporter approach (harnessing sulfonyl fluoride tyrosine labeling and subsequent click chemistry) to measure the occupancy of the mRNA-decapping scavenger enzyme DcpS by a small molecule inhibitor in live cells. Enzyme affinity determined using isothermal dose response fingerprinting (ITDRFCETSA) and the concentration required to occupy 50% of the enzyme (OC50) using the chemical probe reporter assay were very similar...
July 14, 2016: Organic & Biomolecular Chemistry
https://www.readbyqxmd.com/read/27115831/in-situ-proteome-profiling-and-bioimaging-applications-of-small-molecule-affinity-based-probes-derived-from-dot1l-inhibitors
#14
Biwei Zhu, Hailong Zhang, Sijun Pan, Chenyu Wang, Jingyan Ge, Jun-Seok Lee, Shao Q Yao
DOT1L is the sole protein methyltransferase that methylates histone H3 on lysine 79 (H3K79), and is a promising drug target against cancers. Small-molecule inhibitors of DOT1L such as FED1 are potential anti-cancer agents and useful tools to investigate the biological roles of DOT1L in human diseases. FED1 showed excellent in vitro inhibitory activity against DOT1L, but its cellular effect was relatively poor. In this study, we designed and synthesized photo-reactive and "clickable" affinity-based probes (AfBPs), P1 and P2, which were cell-permeable and structural mimics of FED1...
June 1, 2016: Chemistry: a European Journal
https://www.readbyqxmd.com/read/27010513/cetsa-screening-identifies-known-and-novel-thymidylate-synthase-inhibitors-and-slow-intracellular-activation-of-5-fluorouracil
#15
Helena Almqvist, Hanna Axelsson, Rozbeh Jafari, Chen Dan, André Mateus, Martin Haraldsson, Andreas Larsson, Daniel Martinez Molina, Per Artursson, Thomas Lundbäck, Pär Nordlund
Target engagement is a critical factor for therapeutic efficacy. Assessment of compound binding to native target proteins in live cells is therefore highly desirable in all stages of drug discovery. We report here the first compound library screen based on biophysical measurements of intracellular target binding, exemplified by human thymidylate synthase (TS). The screen selected accurately for all the tested known drugs acting on TS. We also identified TS inhibitors with novel chemistry and marketed drugs that were not previously known to target TS, including the DNA methyltransferase inhibitor decitabine...
2016: Nature Communications
https://www.readbyqxmd.com/read/26964663/advances-in-identification-and-validation-of-protein-targets-of-natural-products-without-chemical-modification
#16
REVIEW
J Chang, Y Kim, H J Kwon
Covering: up to February 2016Identification of the target proteins of natural products is pivotal to understanding the mechanisms of action to develop natural products for use as molecular probes and potential therapeutic drugs. Affinity chromatography of immobilized natural products has been conventionally used to identify target proteins, and has yielded good results. However, this method has limitations, in that labeling or tagging for immobilization and affinity purification often result in reduced or altered activity of the natural product...
May 4, 2016: Natural Product Reports
https://www.readbyqxmd.com/read/26566155/the-cellular-thermal-shift-assay-a-novel-biophysical-assay-for-in-situ-drug-target-engagement-and-mechanistic-biomarker-studies
#17
REVIEW
Daniel Martinez Molina, Pär Nordlund
A drug must engage its intended target to achieve its therapeutic effect. However, conclusively measuring target engagement (TE) in situ is challenging. This complicates preclinical development and is considered a key factor in the high rate of attrition in clinical trials. Here, we discuss a recently developed, label-free, biophysical assay, the cellular thermal shift assay (CETSA), which facilitates the direct assessment of TE in cells and tissues at various stages of drug development. CETSA also reveals biochemical events downstream of drug binding and therefore provides a promising means of establishing mechanistic biomarkers...
2016: Annual Review of Pharmacology and Toxicology
https://www.readbyqxmd.com/read/26159518/assessing-the-efficacy-of-mdm2-mdm4-inhibiting-stapled-peptides-using-cellular-thermal-shift-assays
#18
Ban Xiong Tan, Christopher J Brown, Fernando J Ferrer, Tsz Ying Yuen, Soo Tng Quah, Boon Hong Chan, Anna E Jansson, Hsiang Ling Teo, Pär Nordlund, David P Lane
Previous publications on stapled peptide inhibitors against Mdm2/Mdm4-p53 interactions have established that this new class of drugs have the potential to be easily optimised to attain high binding affinity and specificity, but the mechanisms controlling their cellular uptake and target engagement remain elusive and controversial. To aid in understanding the rules of peptide and staple design, and to enable rapid optimisation, we employed the newly-developed cellular thermal shift assay (CETSA). CETSA was able to validate stapled peptide binding to Mdm2 and Mdm4, and the method was also used to determine the extent of cellular uptake, cellular availability, and intracellular binding of the endogenous target proteins in its native environment...
July 10, 2015: Scientific Reports
https://www.readbyqxmd.com/read/26062395/cetsa-a-target-engagement-assay-with-potential-to-transform-drug-discovery
#19
EDITORIAL
Annika Jenmalm Jensen, Daniel Martinez Molina, Thomas Lundbäck
No abstract text is available yet for this article.
2015: Future Medicinal Chemistry
https://www.readbyqxmd.com/read/25101824/the-cellular-thermal-shift-assay-for-evaluating-drug-target-interactions-in-cells
#20
Rozbeh Jafari, Helena Almqvist, Hanna Axelsson, Marina Ignatushchenko, Thomas Lundbäck, Pär Nordlund, Daniel Martinez Molina
Thermal shift assays are used to study thermal stabilization of proteins upon ligand binding. Such assays have been used extensively on purified proteins in the drug discovery industry and in academia to detect interactions. Recently, we published a proof-of-principle study describing the implementation of thermal shift assays in a cellular format, which we call the cellular thermal shift assay (CETSA). The method allows studies of target engagement of drug candidates in a cellular context, herein exemplified with experimental data on the human kinases p38α and ERK1/2...
September 2014: Nature Protocols
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