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1.
J Med Chem ; 65(19): 12802-12824, 2022 10 13.
Article in English | MEDLINE | ID: mdl-36153998

ABSTRACT

Ligand-targeting drug delivery systems have made significant strides for disease treatments with numerous clinical approvals in this era of precision medicine. Herein, we report a class of small molecule-based immune checkpoint-targeting maytansinoid conjugates. From the ligand targeting ability, pharmacokinetics profiling, in vivo anti-pancreatic cancer, triple-negative breast cancer, and sorafenib-resistant liver cancer efficacies with quantitative mRNA analysis of treated-tumor tissues, we demonstrated that conjugate 40a not only induced lasting regression of tumor growth, but it also rejuvenated the once immunosuppressive tumor microenvironment to an "inflamed hot tumor" with significant elevation of gene expressions that were not accessible in the vehicle-treated tumor. In turn, the immune checkpoint-targeting small molecule drug conjugate from this work represents a new pharmacodelivery strategy that can be expanded with combination therapy with existing immune-oncology treatment options.


Subject(s)
Phosphatidylserines , Triple Negative Breast Neoplasms , Humans , Ligands , RNA, Messenger , Sorafenib/pharmacology , Sorafenib/therapeutic use , Tumor Microenvironment
2.
Pharmaceutics ; 14(7)2022 Jun 21.
Article in English | MEDLINE | ID: mdl-35890212

ABSTRACT

Ligand-targeting drug conjugates are a class of clinically validated biopharmaceutical drugs constructed by conjugating cytotoxic drugs with specific disease antigen targeting ligands through appropriate linkers. The integrated linker-drug motif embedded within such a system can prevent the premature release during systemic circulation, thereby allowing the targeting ligand to engage with the disease antigen and selective accumulation. We have designed and synthesized new thioester-linked maytansinoid conjugates. By performing in vitro cytotoxicity, targeting ligand binding assay, and in vivo pharmacokinetic studies, we investigated the utility of this new linker-drug moiety in the small molecule drug conjugate (SMDC) system. In particular, we conjugated the thioester-linked maytansinoids to the phosphatidylserine-targeting small molecule zinc dipicolylamine and showed that Zn8_DM1 induced tumor regression in the HCC1806 triple-negative breast cancer xenograft model. Moreover, in a spontaneous sorafenib-resistant liver cancer model, Zn8_DM1 exhibited potent antitumor growth efficacy. From quantitative mRNA analysis of Zn8_DM1 treated-tumor tissues, we observed the elevation of gene expressions associated with a "hot inflamed tumor" state. With the identification and validation of a plethora of cancer-associated antigens in the "omics" era, this work provided the insight that antibody- or small molecule-based targeting ligands can be conjugated similarly to generate new ligand-targeting drug conjugates.

3.
Redox Biol ; 46: 102126, 2021 10.
Article in English | MEDLINE | ID: mdl-34509914

ABSTRACT

Nitro-fatty acids are a class of endogenous electrophilic lipid mediators with anti-inflammatory and cytoprotective effects in a wide range of inflammatory and fibrotic disease models. While these beneficial biological effects of nitro-fatty acids are mainly attributed to their ability to form covalent adducts with proteins, only a small number of proteins are known to be nitro-alkylated and the scope of protein nitro-alkylation remains undetermined. Here we describe the synthesis and application of a clickable nitro-fatty acid probe for the detection and first global identification of mammalian proteins that are susceptible to nitro-alkylation. 184 high confidence nitro-alkylated proteins were identified in THP1 macrophages, majority of which are novel targets of nitro-fatty acids, including extended synaptotagmin 2 (ESYT2), signal transducer and activator of transcription 3 (STAT3), toll-like receptor 2 (TLR2), retinoid X receptor alpha (RXRα) and glucocorticoid receptor (NR3C1). In particular, we showed that 9-nitro-oleate covalently modified and inhibited dexamethasone binding to NR3C1. Bioinformatic analyses revealed that nitro-alkylated proteins are highly enriched in endoplasmic reticulum and transmembrane proteins, and are overrepresented in lipid metabolism and transport pathways. This study significantly expands the scope of protein substrates targeted by nitro-fatty acids in living cells and provides a useful resource towards understanding the pleiotropic biological roles of nitro-fatty acids as signaling molecules or as multi-target therapeutic agents.


Subject(s)
Fatty Acids , Nitro Compounds , Alkylation , Animals , Protein Binding , Signal Transduction
4.
J Med Chem ; 62(13): 6047-6062, 2019 07 11.
Article in English | MEDLINE | ID: mdl-31181158

ABSTRACT

We report that compound 13, a novel phosphatidylserine-targeting zinc(II) dipicolylamine drug conjugate, readily triggers a positive feedback therapeutic loop through the in situ generation of phosphatidylserine in the tumor microenvironment. Linker modifications, pharmacokinetics profiling, in vivo antitumor studies, and micro-Western array of treated-tumor tissues were employed to show that this class of conjugates induced regeneration of apoptotic signals, which facilitated subsequent recruitment of the circulating conjugates through the zinc(II) dipicolylamine-phosphatidylserine association and resulted in compounding antitumor efficacy. Compared to the marketed compound 17, compound 13 not only induced regressions in colorectal and pancreatic tumor models, it also exhibited at least 5-fold enhancement in antitumor efficacy with only 40% of the drug employed during treatment, culminating in a >12.5-fold increase in therapeutic potential. Our study discloses a chemically distinct apoptosis-targeting theranostic, with built-in complementary functional moieties between the targeting module and the drug mechanism to expand the arsenal of antitumor therapy.


Subject(s)
Antineoplastic Agents/therapeutic use , Coordination Complexes/therapeutic use , Indolizines/therapeutic use , Neoplasms/drug therapy , Phosphatidylserines/metabolism , Picolines/therapeutic use , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Apoptosis/drug effects , Cell Line, Tumor , Coordination Complexes/chemical synthesis , Coordination Complexes/chemistry , Drug Design , Humans , Indolizines/chemical synthesis , Indolizines/chemistry , Male , Mice, Inbred ICR , Mice, Nude , Molecular Structure , Picolines/chemical synthesis , Picolines/chemistry , Structure-Activity Relationship , Topoisomerase I Inhibitors/chemical synthesis , Topoisomerase I Inhibitors/chemistry , Topoisomerase I Inhibitors/therapeutic use , Xenograft Model Antitumor Assays , Zinc/chemistry
5.
Sci Rep ; 7(1): 12336, 2017 09 26.
Article in English | MEDLINE | ID: mdl-28951584

ABSTRACT

Influenza is an annual seasonal epidemic that has continually drawn public attentions, due to the potential death toll and drug resistance. Neuraminidase, which is essential for the spread of influenza virus, has been regarded as a valid target for the treatment of influenza infection. Although neuraminidase drugs have been developed, they are susceptible to drug-resistant mutations in the sialic-binding site. In this study, we established computational models (site-moiety maps) of H1N1 and H5N1 to determine properties of the 150-cavity, which is adjacent to the drug-binding site. The models reveal that hydrogen-bonding interactions with residues R118, D151, and R156 and van der Waals interactions with residues Q136, D151, and T439 are important for identifying 150-cavitiy inhibitors. Based on the models, we discovered three new inhibitors with IC50 values <10 µM that occupies both the 150-cavity and sialic sites. The experimental results identified inhibitors with similar activities against both wild-type and dual H274Y/I222R mutant neuraminidases and showed little cytotoxic effects. Furthermore, we identified three new inhibitors situated at the sialic-binding site with inhibitory effects for normal neuraminidase, but lowered effects for mutant strains. The results suggest that the new inhibitors can be used as a starting point to combat drug-resistant strains.


Subject(s)
Antiviral Agents/pharmacology , Drug Discovery/methods , Influenza, Human/drug therapy , Molecular Dynamics Simulation , Neuraminidase/antagonists & inhibitors , Viral Proteins/antagonists & inhibitors , Antiviral Agents/therapeutic use , Binding Sites/genetics , Computer Simulation , Drug Resistance, Viral/drug effects , Drug Resistance, Viral/genetics , Humans , Influenza A Virus, H1N1 Subtype/drug effects , Influenza A Virus, H1N1 Subtype/genetics , Influenza A Virus, H1N1 Subtype/metabolism , Influenza A Virus, H5N1 Subtype/drug effects , Influenza A Virus, H5N1 Subtype/genetics , Influenza A Virus, H5N1 Subtype/metabolism , Influenza, Human/virology , Inhibitory Concentration 50 , Mutation , Neuraminidase/chemistry , Neuraminidase/genetics , Protein Structure, Tertiary , Viral Proteins/chemistry , Viral Proteins/genetics
6.
Bioconjug Chem ; 28(7): 1878-1892, 2017 07 19.
Article in English | MEDLINE | ID: mdl-28581724

ABSTRACT

A series of zinc(II) dipicolylamine (ZnDPA)-based drug conjugates have been synthesized to probe the potential of phosphatidylserine (PS) as a new antigen for small molecule drug conjugate (SMDC) development. Using in vitro cytotoxicity and plasma stability studies, PS-binding assay, in vivo pharmacokinetic studies, and maximum tolerated dose profiles, we provided a roadmap and the key parameters required for the development of the ZnDPA based drug conjugate. In particular, conjugate 24 induced tumor regression in the COLO 205 xenograft model and exhibited a more potent antitumor effect with a 70% reduction of cytotoxic payload compared to that of the marketed irinotecan when dosed at the same regimen. In addition to the validation of PS as an effective pharmacodelivery target for SMDC, our work also provided the foundation that, if applicable, a variety of therapeutic agents could be conjugated in the same manner to treat other PS-associated diseases.


Subject(s)
Antineoplastic Agents/pharmacokinetics , Immunoconjugates/therapeutic use , Molecular Targeted Therapy/methods , Organometallic Compounds/immunology , Phosphatidylserines/immunology , Picolinic Acids/immunology , Animals , Antineoplastic Agents/chemical synthesis , Cell Line, Tumor , Humans , Immunoconjugates/chemistry , Immunoconjugates/pharmacokinetics , Organometallic Compounds/chemical synthesis , Organometallic Compounds/pharmacology , Phosphatidylserines/metabolism , Picolinic Acids/chemical synthesis , Picolinic Acids/pharmacology , Structure-Activity Relationship , Xenograft Model Antitumor Assays
7.
Journal of Preventive Medicine ; (12): 770-775, 2016.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-792529

ABSTRACT

Objective To evaluate the association of cervical cancer and the G/C single nucleotide polymorphism at codon 72 in p53 gene exon -4 in Chinese population.Methods Literatures were searched in CNKI,VIP,CBM, PubMed,EMBase,the Cochrane Library (Issue 1,2015),WanFang Data Database (1980—2015)with computer. Results Thirteen studies involving 1 780 cervical cancer patients and 1 5 17 healthy volunteers were included.According to the situation of the HPV infection in research object,thirteen studies were divided into two subgroups.The results of meta-analyses in the subgroup of non-HPV infection showed that the Chinese women having the genotype G/G presented no increased relative risk compared to that with the genotype C/C(OR=0.980,95%CI:0.558 -1.723).The allele G showed no increased relative risk compared to the allele C (OR=1.070,95%CI:0.789-1.449)The results of meta-analyses in the subgroup of HPV infection showed that the Chinese women having the genotype G/G presented no increased relative risk compared to that with the genotype C/C(OR=0.891,95%CI:0.213 -3.722).The allele G showed no increased relative risk compared to the allele C (OR =0.981,95%CI:0.493 -1.951 ).Conclusion This meta -analysis demonstrates that G/C single nucleotide polymorphism at codon 72 in p53 gene exon-4 were not associated with cervical cancer in Chinese women.

8.
Journal of Preventive Medicine ; (12): 995-998, 2015.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-792453

ABSTRACT

Objective To investigate the metastatic risk factors of pelvic lymph nodes in patients with cervical carcinoma of stage Ia2 and IIa2.Methods The clinic opathologicparameters in 337 patients with stage Ia2 -IIa2 cervical carcinoma were retrospectively analyzed,and were studied the related factors of pelvic lymph node metastasis.Results The lymph nodes metastasis rate was 11.87%,with the obturator and internal iliaclymph nodes mostly involved.Multivariate analysis showed that the myometrial invasion,size of tumor,lymph -vascular space involvement and utero -tissue infiltration were major risk factors.The OR value and 95%CI were 3.464(1.502 -7.985)、4.316 (1.164 -7.833)、6.167 (2.592 -14.674)和 8.507(1.966 -36.808)respectively.Conclusion The myometrial invasion≥ 2 /3,size of tumor≥ 4 cm, lymph -vascular space involvement and utero -tissue infiltration positive lymph node metastasis of cervical cancer are easy to occur.

9.
PLoS One ; 9(11): e111331, 2014.
Article in English | MEDLINE | ID: mdl-25412347

ABSTRACT

Enterovirus 71 (EV-A71) is a neurotropic virus that can cause severe complications involving the central nervous system. No effective antiviral therapeutics are available for treating EV-A71 infection and drug discovery efforts are rarely focused to target this disease. Thus, the main goal of this study was to discover existing drugs with novel indications that may effectively inhibit EV-A71 replication and the inflammatory cytokines elevation. In this study, we showed that LiCl, a GSK3ß inhibitor, effectively suppressed EV-A71 replication, apoptosis and inflammatory cytokines production (Interleukin 6, Interleukin-1ß) in infected cells. Furthermore, LiCl and an immunomodular agent were shown to strongly synergize with each other in suppressing EV-A71 replication. The results highlighted potential new treatment regimens in suppressing sequelae caused by EV-A71 replication.


Subject(s)
Aniline Compounds/administration & dosage , Antiviral Agents/administration & dosage , Enterovirus A, Human/drug effects , Enterovirus Infections/drug therapy , Hydroxybutyrates/administration & dosage , Lithium Chloride/administration & dosage , Aniline Compounds/pharmacology , Animals , Antiviral Agents/pharmacology , Apoptosis/drug effects , Cell Line , Chlorocebus aethiops , Crotonates , Cytokines/genetics , Cytokines/metabolism , Drug Repositioning , Drug Synergism , Drug Therapy, Combination , Enterovirus A, Human/physiology , Enterovirus Infections/immunology , Humans , Hydroxybutyrates/pharmacology , Lithium Chloride/pharmacology , Nitriles , Toluidines , Vero Cells , Virus Replication/drug effects
11.
PLoS One ; 8(2): e56704, 2013.
Article in English | MEDLINE | ID: mdl-23437217

ABSTRACT

Infection with influenza virus is a major public health problem, causing serious illness and death each year. Emergence of drug-resistant influenza virus strains limits the effectiveness of drug treatment. Importantly, a dual H275Y/I223R mutation detected in the pandemic influenza A 2009 virus strain results in multidrug resistance to current neuraminidase (NA) drugs. Therefore, discovery of new agents for treating multiple drug-resistant (MDR) influenza virus infections is important. Here, we propose a parallel screening strategy that simultaneously screens wild-type (WT) and MDR NAs, and identifies inhibitors matching the subsite characteristics of both NA-binding sites. These may maintain their potency when drug-resistant mutations arise. Initially, we analyzed the subsite of the dual H275Y/I223R NA mutant. Analysis of the site-moiety maps of NA protein structures show that the mutant subsite has a relatively small volume and is highly polar compared with the WT subsite. Moreover, the mutant subsite has a high preference for forming hydrogen-bonding interactions with polar moieties. These changes may drive multidrug resistance. Using this strategy, we identified a new inhibitor, Remazol Brilliant Blue R (RB19, an anthraquinone dye), which inhibited WT NA and MDR NA with IC(50) values of 3.4 and 4.5 µM, respectively. RB19 comprises a rigid core scaffold and a flexible chain with a large polar moiety. The former interacts with highly conserved residues, decreasing the probability of resistance. The latter forms van der Waals contacts with the WT subsite and yields hydrogen bonds with the mutant subsite by switching the orientation of its flexible side chain. Both scaffolds of RB19 are good starting points for lead optimization. The results reveal a parallel screening strategy for identifying resistance mechanisms and discovering anti-resistance neuraminidase inhibitors. We believe that this strategy may be applied to other diseases with high mutation rates, such as cancer and human immunodeficiency virus type 1.


Subject(s)
Drug Resistance, Viral/genetics , Influenza A Virus, H1N1 Subtype/genetics , Influenza, Human/drug therapy , Influenza, Human/genetics , Neuraminidase/antagonists & inhibitors , Anthraquinones/administration & dosage , Anthraquinones/chemistry , Antiviral Agents , Drug Resistance, Multiple/genetics , Genotype , Humans , Influenza A Virus, H1N1 Subtype/pathogenicity , Influenza, Human/virology , Mutation , Neuraminidase/genetics , Neuraminidase/metabolism , Pandemics
12.
ChemMedChem ; 7(9): 1546-50, 2012 Sep.
Article in English | MEDLINE | ID: mdl-22821876

ABSTRACT

From a high-throughput screening (HTS) hit with inhibitory activity against virus-induced cytophathic in the low micromolar range, we have developed a potent anti-influenza lead through careful optimization without compromising the drug-like properties of the compound. An orally bioavailable compound was identified as a lead agent with nanomolar activity against influenza, representing a 140-fold improvement over the initial hit.


Subject(s)
Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Influenza A Virus, H1N1 Subtype/drug effects , Quinolines/chemistry , Quinolines/pharmacology , Animals , Antiviral Agents/pharmacokinetics , Cell Line , Drug Discovery , Humans , Influenza, Human/drug therapy , Male , Orthomyxoviridae Infections/drug therapy , Quinolines/pharmacokinetics , Rats , Rats, Sprague-Dawley , Structure-Activity Relationship
13.
Anal Chem ; 84(15): 6391-9, 2012 Aug 07.
Article in English | MEDLINE | ID: mdl-22712523

ABSTRACT

Recent studies have shown that NP (nucleoprotein), which possesses multiple functions in the viral life cycle, is a new potential anti-influenza drug target. NP inhibitors reliably induce conformational changes in NPs, and these changes may confer inhibition of the influenza virus. The six conserved tryptophan residues in NP can be used as an intrinsic probe to monitor the change in fluorescence of the tryptophan residues in the protein upon binding to an NP inhibitor. In the present study, we found that the fluorescence of recombinant NP proteins was quenched following the binding of available NP inhibitors (such as nucleozin) in a concentration- and time-dependent manner, which suggests that the inhibitor induced conformational changes in the NPs. The minimal fluorescence-quenching effect and weak binding constant of nucleozin to the swine-origin influenza virus H1N1pdm09 (SOIV) NP revealed that the SOIV is resistant to nucleozin. We have used the fluorescence-quenching property of tryptophans in NPs that were bound to ligands in a 96-well-plate-based drug screen to assess the ability of promising small molecules to interact with NPs and have identified one new anti-influenza drug, CSV0C001018, with a high SI value. This convenient method for drug screening may facilitate the development of antiviral drugs that target viruses other than the influenza virus, such as HIV and HBV.


Subject(s)
Influenza A Virus, H1N1 Subtype/metabolism , Nucleoproteins/antagonists & inhibitors , Spectrometry, Fluorescence , Tryptophan/chemistry , Amino Acid Sequence , Animals , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Dogs , Drug Evaluation, Preclinical , Humans , Influenza A Virus, H1N1 Subtype/drug effects , Madin Darby Canine Kidney Cells , Molecular Sequence Data , Nucleoproteins/genetics , Nucleoproteins/metabolism , Recombinant Proteins/antagonists & inhibitors , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Swine
14.
Anticancer Res ; 32(1): 147-51, 2012 Jan.
Article in English | MEDLINE | ID: mdl-22213300

ABSTRACT

A high-throughput 32D(L858R/T790M) cell-based assay to identify inhibitors of the L858R/T790M mutant epidermal growth factor receptor (EGFR) pathway was established. After screening, ten hits from among 60,000 compounds in our in-house compound library were initially identified. In the secondary assays, one hit, 1-[2-(decyloxy)-2-oxoethyl]-3-methyl-2-[(4-methylphenoxy) methyl]-1H-benzimidazol-3-ium, was confirmed to directly inhibit the kinase activity of recombinant L858R/T790M EGFR and the phosphorylation of EGFR-L858R/T790M in gefitinib-resistant H1975 cells. Thus, this high-throughput assay system may be useful for identifying novel inhibitors which suppress mutant EGFR-T790M signalling and for overcoming T790M-mediated acquired resistance for future anticancer drug discovery.


Subject(s)
Carcinoma, Non-Small-Cell Lung/drug therapy , Epidermal Growth Factor/pharmacology , ErbB Receptors/antagonists & inhibitors , ErbB Receptors/genetics , Lung Neoplasms/drug therapy , Mutation/drug effects , Quinazolines/pharmacology , Carcinoma, Non-Small-Cell Lung/genetics , Cell Line, Tumor , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/genetics , Gefitinib , High-Throughput Screening Assays , Humans , Lung Neoplasms/genetics , Phosphorylation/drug effects , Protein Kinase Inhibitors/pharmacology , Signal Transduction/drug effects
15.
Antimicrob Agents Chemother ; 56(2): 647-57, 2012 Feb.
Article in English | MEDLINE | ID: mdl-21930871

ABSTRACT

The aim of this study was to identify the antiviral mechanism of a novel compound, BPR3P0128. From a large-scale screening of a library of small compounds, BPR3P compounds were found to be potent inhibitors of influenza viral replication in Madin-Darby canine kidney (MDCK) cells. BPR3P0128 exhibited inhibitory activity against both influenza A and B viruses. The 50% inhibitory concentrations were in the range of 51 to 190 nM in MDCK cells, as measured by inhibition-of-cytopathic-effect assays. BPR3P0128 appeared to target the viral replication cycle but had no effect on viral adsorption. The inhibition of cap-dependent mRNA transcription by BPR3P0128 was more prominent with a concurrent increase in cap-independent cRNA replication in a primer extension assay, suggesting a role of BPR3P0128 in switching transcription to replication. This reduction in mRNA expression resulted from the BPR3P-mediated inhibition of the cap-dependent endoribonuclease (cap-snatching) activities of nuclear extracts containing the influenza virus polymerase complex. No inhibition of binding of 5' viral RNA to the viral polymerase complex by this compound was detected. BPR3P0128 also effectively inhibited other RNA viruses, such as enterovirus 71 and human rhinovirus, but not DNA viruses, suggesting that BPR3P0128 targets a cellular factor(s) associated with viral PB2 cap-snatching activity. The identification of this factor(s) could help redefine the regulation of viral transcription and replication and thereby provide a potential target for antiviral chemotherapeutics.


Subject(s)
Antiviral Agents/pharmacology , Endonucleases/antagonists & inhibitors , Orthomyxoviridae/drug effects , Pyrazoles/pharmacology , RNA Caps/drug effects , Animals , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , Cell Line , Cytopathogenic Effect, Viral/drug effects , Dogs , Endonucleases/metabolism , HEK293 Cells , Humans , Influenza A Virus, H1N1 Subtype/drug effects , Influenza A Virus, H1N1 Subtype/metabolism , Influenza A Virus, H3N2 Subtype/drug effects , Influenza A Virus, H3N2 Subtype/metabolism , Influenza B virus/drug effects , Influenza B virus/metabolism , Orthomyxoviridae/metabolism , Orthomyxoviridae/physiology , Pyrazoles/chemical synthesis , Pyrazoles/chemistry , Quinolines/chemical synthesis , Quinolines/chemistry , Quinolines/pharmacology , RNA Caps/metabolism , RNA, Viral/biosynthesis , Transcription, Genetic/drug effects , Viral Proteins/antagonists & inhibitors , Viral Proteins/metabolism , Virus Replication/drug effects
16.
J Med Chem ; 53(20): 7316-26, 2010 Oct 28.
Article in English | MEDLINE | ID: mdl-20961149

ABSTRACT

HTS hit 7 was modified through hybrid design strategy to introduce a chiral side chain followed by introduction of Michael acceptor group to obtain potent EGFR kinase inhibitors 11 and 19. Both 11 and 19 showed over 3 orders of magnitude enhanced HCC827 antiproliferative activity compared to HTS hit 7 and also inhibited gefitinib-resistant double mutant (DM, T790M/L858R) EGFR kinase at nanomolar concentration. Moreover, treatment with 19 shrinked tumor in nude mice xenograft model.


Subject(s)
Antineoplastic Agents/chemical synthesis , ErbB Receptors/antagonists & inhibitors , Heterocyclic Compounds, 3-Ring/chemical synthesis , Pyrimidines/chemical synthesis , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Carcinoma, Non-Small-Cell Lung , Cell Line, Tumor , Drug Design , Drug Resistance, Neoplasm , Drug Screening Assays, Antitumor , ErbB Receptors/genetics , Gefitinib , Heterocyclic Compounds, 3-Ring/chemistry , Heterocyclic Compounds, 3-Ring/pharmacology , Humans , Male , Mice , Mice, Nude , Mutation , Neoplasm Transplantation , Pyrimidines/chemistry , Pyrimidines/pharmacology , Quinazolines/pharmacology , Rats , Rats, Sprague-Dawley , Stereoisomerism , Structure-Activity Relationship , Transplantation, Heterologous
17.
J Med Chem ; 53(13): 4980-8, 2010 Jul 08.
Article in English | MEDLINE | ID: mdl-20550212

ABSTRACT

A focused library of furanopyrimidine (350 compounds) was rapidly synthesized in parallel reactors and in situ screened for Aurora and epidermal growth factor receptor (EGFR) kinase activity, leading to the identification of some interesting hits. On the basis of structural biology observations, the hit 1a was modified to better fit the back pocket, producing the potent Aurora inhibitor 3 with submicromolar antiproliferative activity in HCT-116 colon cancer cell line. On the basis of docking studies with EGFR hit 1s, introduction of acrylamide Michael acceptor group led to 8, which inhibited both the wild and mutant EGFR kinase and also showed antiproliferative activity in HCC827 lung cancer cell line. Furthermore, the X-ray cocrystal study of 3 and 8 in complex with Aurora and EGFR, respectively, confirmed their hypothesized binding modes. Library construction, in situ screening, and structure-based drug design (SBDD) strategy described here could be applied for the lead identification of other kinases.


Subject(s)
Carcinoma, Non-Small-Cell Lung/enzymology , ErbB Receptors/antagonists & inhibitors , Furans/chemistry , Lung Neoplasms/enzymology , Protein Kinase Inhibitors/chemistry , Protein Serine-Threonine Kinases/antagonists & inhibitors , Pyrimidines/chemistry , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Aurora Kinases , Blotting, Western , Carcinoma, Non-Small-Cell Lung/drug therapy , Cell Line, Tumor , Cell Survival/drug effects , Crystallography, X-Ray , ErbB Receptors/metabolism , Furans/chemical synthesis , Furans/pharmacology , Humans , Inhibitory Concentration 50 , Lung Neoplasms/drug therapy , Magnetic Resonance Spectroscopy , Models, Molecular , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/metabolism , Pyrimidines/chemical synthesis , Pyrimidines/pharmacology , Spectrometry, Mass, Fast Atom Bombardment
18.
J Antimicrob Chemother ; 65(4): 676-83, 2010 Apr.
Article in English | MEDLINE | ID: mdl-20089540

ABSTRACT

OBJECTIVES: Enterovirus 71 (EV71) causes serious diseases in humans. The aim of this study was to examine the effects of aurintricarboxylic acid (ATA) on EV71 replication and to explore the underlying mechanism. METHODS: To measure the activity of ATA in inhibiting the cytopathic effect (CPE) of EV71, a cell-based neutralization (inhibition of virus-induced CPE) assay was performed. The effect of ATA was further confirmed using plaque reduction and viral yield reduction assays. A time of addition assay was performed to identify the mechanisms of ATA's anti-EV71 activity. We examined the effects of ATA on the following key steps involved in virus replication: (i) translation of the internal ribosomal entry site (IRES)-mediated viral polyprotein; (ii) the proteolytic activity of viral proteases 2A and/or 3C; and (iii) the viral 3D RNA-dependent RNA polymerase (RdRp) activity. RESULTS: In this study, ATA was found to be a potent inhibitor of the replication of EV71. In the antiviral neutralization assay, ATA exhibited inhibitory activity against EV71 (TW/4643/98) and EV71 (TW/2231/98). Plaque assay further demonstrated that ATA inhibited EV71 replication with an EC(50) (effective concentration at which 50% of plaques were removed) of 2.9 microM. Studies on the mechanism of action revealed that ATA targets the early stage of the viral life cycle after viral entry. ATA was able to inhibit the RdRp activity of EV71, while neither the IRES-mediated translation of viral polyprotein nor the viral 3C protease activity was affected. CONCLUSIONS: Overall, the findings in this study suggest that ATA is able to effectively inhibit EV71 replication through interfering with the viral 3D polymerase.


Subject(s)
Antiviral Agents/pharmacology , Aurintricarboxylic Acid/pharmacology , Enterovirus A, Human/drug effects , Enzyme Inhibitors/pharmacology , RNA-Dependent RNA Polymerase/antagonists & inhibitors , Viral Proteins/antagonists & inhibitors , Virus Replication/drug effects , Animals , COS Cells , Cell Line, Tumor , Chlorocebus aethiops , Cytopathogenic Effect, Viral , Humans , Vero Cells , Viral Plaque Assay
19.
ChemMedChem ; 5(2): 255-67, 2010 Feb 01.
Article in English | MEDLINE | ID: mdl-20039358

ABSTRACT

Herein we reveal a simple method for the identification of novel Aurora kinase A inhibitors through substructure searching of an in-house compound library to select compounds for testing. A hydrazone fragment conferring Aurora kinase activity and heterocyclic rings most frequently reported in kinase inhibitors were used as substructure queries to filter the in-house compound library collection prior to testing. Five new series of Aurora kinase inhibitors were identified through this strategy, with IC(50) values ranging from approximately 300 nM to approximately 15 microM, by testing only 133 compounds from a database of approximately 125,000 compounds. Structure-activity relationship studies and X-ray co-crystallographic analysis of the most potent compound, a furanopyrimidine derivative with an IC(50) value of 309 nM toward Aurora kinase A, were carried out. The knowledge gained through these studies could help in the future design of potent Aurora kinase inhibitors.


Subject(s)
Furans/chemistry , Protein Kinase Inhibitors/chemistry , Protein Serine-Threonine Kinases/antagonists & inhibitors , Pyrimidines/chemistry , Aurora Kinases , Binding Sites , Crystallography, X-Ray , High-Throughput Screening Assays , Protein Binding , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/metabolism , Pyrimidines/chemical synthesis , Pyrimidines/pharmacology , Structure-Activity Relationship
20.
Anal Biochem ; 377(1): 89-94, 2008 Jun 01.
Article in English | MEDLINE | ID: mdl-18358823

ABSTRACT

Epidermal growth factor receptor (EGFR) is a valid drug target for development of target-based therapeutics against non-small-cell lung cancer. In this study, we established a high-throughput cell-based assay to screen for compounds that may inhibit EGFR activation and/or EGFR-mediated downstream signaling pathway. This drug screening platform is based on the characterization of an EGFR-transfected 32D cell line (32D-EGFR). The expression of EGFR in 32D cells allowed cell proliferation in the presence of either epidermal growth factor (EGF) or interleukin 3 (IL-3) and provided a system for both screening and counterscreening of EGFR pathway-inhibitory compounds. After the completion of primary and secondary screenings in which 32D-EGFR cells were grown under the stimulation of either EGF or IL-3, 9 of 20,000 compounds were found to selectively inhibit the EGF-dependent proliferation, but not the IL-3-dependent proliferation, of 32D-EGFR cells. Subsequent analysis showed that 3 compounds of the 9 initial hits directly inhibited the kinase activity of recombinant EGFR in vitro and the phosphorylation of EGFR in H1299 cells transfected with EGFR. Thus, this 32D-EGFR assay system provides a promising approach for identifying novel EGFR and EGFR signaling pathway inhibitors with potential antitumor activity.


Subject(s)
Drug Evaluation, Preclinical/methods , ErbB Receptors/antagonists & inhibitors , ErbB Receptors/metabolism , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Signal Transduction/drug effects , Animals , Cell Line , Cell Survival/drug effects , Enzyme Activation/drug effects , Humans , Mice , Phosphorylation/drug effects , Reproducibility of Results
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