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1.
J Med Chem ; 66(4): 2566-2588, 2023 02 23.
Article in English | MEDLINE | ID: mdl-36749735

ABSTRACT

The development of orally bioavailable, furanopyrimidine-based double-mutant (L858R/T790M) EGFR inhibitors is described. First, selectivity for mutant EGFR was accomplished by replacing the (S)-2-phenylglycinol moiety of 12 with either an ethanol or an alkyl substituent. Then, the cellular potency and physicochemical properties were optimized through insights from molecular modeling studies by implanting various solubilizing groups in phenyl rings A and B. Optimized lead 52 shows 8-fold selective inhibition of H1975 (EGFRL858R/T790M overexpressing) cancer cells over A431 (EGFRWT overexpressing) cancer cells; western blot analysis further confirmed EGFR mutant-selective target modulation inside the cancer cells by 52. Notably, 52 displayed in vivo antitumor effects in two different mouse xenograft models (BaF3 transfected with mutant EGFR and H1975 tumors) with TGI = 74.9 and 97.5% after oral administration (F = 27%), respectively. With an extraordinary kinome selectivity (S(10) score of 0.017), 52 undergoes detailed preclinical development.


Subject(s)
Antineoplastic Agents , Carcinoma, Non-Small-Cell Lung , ErbB Receptors , Lung Neoplasms , Protein Kinase Inhibitors , Pyrimidines , Animals , Humans , Mice , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/pharmacology , Carcinoma, Non-Small-Cell Lung/drug therapy , Cell Line, Tumor , Cell Proliferation , Drug Resistance, Neoplasm , ErbB Receptors/antagonists & inhibitors , ErbB Receptors/genetics , Lung Neoplasms/drug therapy , Mutation , Protein Kinase Inhibitors/administration & dosage , Protein Kinase Inhibitors/pharmacology , Administration, Oral , Pyrimidines/administration & dosage , Pyrimidines/pharmacology
2.
PLoS One ; 12(10): e0186447, 2017.
Article in English | MEDLINE | ID: mdl-29036218

ABSTRACT

Human farnesyl pyrophosphate synthase (hFPPS) catalyzes the production of the 15-carbon isoprenoid farnesyl pyrophosphate. The enzyme is a key regulator of the mevalonate pathway and a well-established drug target. Notably, it was elucidated as the molecular target of nitrogen-containing bisphosphonates, a class of drugs that have been widely successful against bone resorption disorders. More recently, research has focused on the anticancer effects of these inhibitors. In order to achieve increased non-skeletal tissue exposure, we created phenylaminopyridine bisphosphonates (PNP-BPs) that have bulky hydrophobic side chains through a structure-based approach. Some of these compounds have proven to be more potent than the current clinical drugs in a number of antiproliferation assays using multiple myeloma cell lines. In the present work, we characterized the binding of our most potent PNP-BPs to the target enzyme, hFPPS. Co-crystal structures demonstrate that the molecular interactions designed to elicit tighter binding are indeed established. We carried out thermodynamic studies as well; the newly introduced protein-ligand interactions are clearly reflected in the enthalpy of binding measured, which is more favorable for the new PNP-BPs than for the lead compound. These studies also indicate that the affinity of the PNP-BPs to hFPPS is comparable to that of the current drug risedronate. Risedronate forms additional polar interactions via its hydroxyl functional group and thus exhibits more favorable binding enthalpy; however, the entropy of binding is more favorable for the PNP-BPs, owing to the greater desolvation effects resulting from their large hydrophobic side chains. These results therefore confirm the overall validity of our drug design strategy. With a distinctly different molecular scaffold, the PNP-BPs described in this report represent an interesting new group of future drug candidates. Further investigation should follow to characterize the tissue distribution profile and assess the potential clinical benefits of these compounds.


Subject(s)
Diphosphonates/metabolism , Geranyltranstransferase/chemistry , Geranyltranstransferase/metabolism , Amino Acid Motifs , Amino Acid Sequence , Crystallography, X-Ray , Diphosphonates/chemistry , Humans , Protein Binding , Thermodynamics
3.
Eur J Med Chem ; 124: 186-199, 2016 Nov 29.
Article in English | MEDLINE | ID: mdl-27573544

ABSTRACT

Aurora kinases have emerged as important anticancer targets so that there are several inhibitors have advanced into clinical study. Herein, we identified novel indazole derivatives as potent Aurora kinases inhibitors by utilizing in silico fragment-based approach and knowledge-based drug design. After intensive hit-to-lead optimization, compounds 17 (dual Aurora A and B), 21 (Aurora B selective) and 30 (Aurora A selective) possessed indazole privileged scaffold with different substituents, which provide sub-type kinase selectivity. Computational modeling helps in understanding that the isoform selectivity could be targeted specific residue in the Aurora kinase binding pocket in particular targeting residues Arg220, Thr217 or Glu177.


Subject(s)
Aurora Kinase A/antagonists & inhibitors , Computer Simulation , Drug Design , Indazoles/chemistry , Indazoles/pharmacology , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Aurora Kinase A/chemistry , Cell Proliferation/drug effects , HCT116 Cells , Humans , Molecular Docking Simulation , Protein Conformation
4.
Nat Chem ; 8(4): 338-46, 2016 Apr.
Article in English | MEDLINE | ID: mdl-27001729

ABSTRACT

A new class of broadly neutralizing antibodies (bNAbs) from HIV donors has been reported to target the glycans on gp120--a glycoprotein found on the surface of the virus envelope--thus renewing hope of developing carbohydrate-based HIV vaccines. However, the version of gp120 used in previous studies was not from human T cells and so the glycosylation pattern could be somewhat different to that found in the native system. Moreover, some antibodies recognized two different glycans simultaneously and this cannot be detected with the commonly used glycan microarrays on glass slides. Here, we have developed a glycan microarray on an aluminium-oxide-coated glass slide containing a diverse set of glycans, including homo- and mixed N-glycans (high-mannose, hybrid and complex types) that were prepared by modular chemo-enzymatic methods to detect the presence of hetero-glycan binding behaviours. This new approach allows rapid screening and identification of optimal glycans recognized by neutralizing antibodies, and could speed up the development of HIV-1 vaccines targeting cell surface glycans.


Subject(s)
Antibodies, Neutralizing/immunology , HIV Antibodies/immunology , HIV-1/immunology , Polysaccharides/chemical synthesis , AIDS Vaccines/immunology , HIV Envelope Protein gp120/immunology , Humans , Ligands , Polysaccharides/chemistry , Polysaccharides/immunology
5.
Proc Natl Acad Sci U S A ; 112(34): 10611-6, 2015 Aug 25.
Article in English | MEDLINE | ID: mdl-26253764

ABSTRACT

Antibodies have been developed as therapeutic agents for the treatment of cancer, infection, and inflammation. In addition to binding activity toward the target, antibodies also exhibit effector-mediated activities through the interaction of the Fc glycan and the Fc receptors on immune cells. To identify the optimal glycan structures for individual antibodies with desired activity, we have developed an effective method to modify the Fc-glycan structures to a homogeneous glycoform. In this study, it was found that the biantennary N-glycan structure with two terminal alpha-2,6-linked sialic acids is a common and optimized structure for the enhancement of antibody-dependent cell-mediated cytotoxicity, complement-dependent cytotoxicity, and antiinflammatory activities.


Subject(s)
Immunoglobulin Fc Fragments/chemistry , Immunoglobulin G/chemistry , Polysaccharides/chemistry , Rituximab/chemistry , Acetylglucosamine/chemistry , Acetylglucosamine/immunology , Animals , Antibodies, Viral/chemistry , Antibodies, Viral/immunology , Antibodies, Viral/therapeutic use , Antibody-Dependent Cell Cytotoxicity , Bacterial Proteins/metabolism , Bacteroides fragilis/enzymology , Cell Line, Tumor , Female , HEK293 Cells , Humans , Immunoglobulin Fc Fragments/immunology , Immunoglobulin G/immunology , Lymphoma, B-Cell/pathology , Mice , Mice, Inbred BALB C , Neuraminidase/metabolism , Orthomyxoviridae Infections/prevention & control , Protein Engineering , Receptors, IgG/immunology , Rituximab/immunology , Sialic Acids/chemistry , Sialic Acids/immunology , Streptococcus pyogenes/enzymology , Structure-Activity Relationship , Trastuzumab/chemistry , Trastuzumab/immunology , alpha-L-Fucosidase/metabolism
6.
Bioorg Med Chem Lett ; 25(5): 1117-23, 2015 Mar 01.
Article in English | MEDLINE | ID: mdl-25630225

ABSTRACT

In order to explore the interactions of bisphosphonate ligands with the active site and an allosteric pocket of the human farnesyl pyrophosphate synthase (hFPPS), substituted indole and azabenzimidazole bisphosphonates were designed as chameleon ligands. NMR and crystallographic studies revealed that these compounds can occupy both sub-pockets of the active site cavity, as well as the allosteric pocket of hFPPS in the presence of the enzyme's Mg(2+) ion cofactor. These results are consistent with the previously proposed hypothesis that the allosteric pocket of hFPPS, located near the active site, plays a feed-back regulatory role for this enzyme.


Subject(s)
Diphosphonates/metabolism , Geranyltranstransferase/chemistry , Geranyltranstransferase/metabolism , Allosteric Site , Catalytic Domain , Diphosphonates/chemistry , Humans , Ligands , Magnesium/metabolism , Molecular Docking Simulation , Protein Binding
7.
J Med Chem ; 57(17): 7435-49, 2014 Sep 11.
Article in English | MEDLINE | ID: mdl-25144111

ABSTRACT

Bisphosphonates can mimic the pyrophosphate leaving group of the nucleotidyl transfer reaction and effectively inhibit RNA/DNA polymerases. In a search of HIV-1 reverse transcriptase (RT) inhibitors, a new chemotype of nonhydrolyzable purine diphosphate mimic was synthesized. A modular synthetic protocol was developed, utilizing 2-amino-6-(methylthio)-4-(trimethylsilyl)nicotinonitrile as the key synthon in the preparation of highly substituted 2-aminonicotinonitriles. These building blocks were subsequently elaborated to the pyrido[2,3-d]pyrimidine bisphosphonates (PYPY-BPs). Biochemical screening identified analogs of PYPY-BPs that inhibit HIV-1 RT-catalyzed DNA synthesis.


Subject(s)
Diphosphonates/pharmacology , HIV Reverse Transcriptase/antagonists & inhibitors , HIV-1/drug effects , Reverse Transcriptase Inhibitors/pharmacology , Diphosphonates/chemical synthesis , Diphosphonates/chemistry , Dose-Response Relationship, Drug , HIV Reverse Transcriptase/genetics , HIV Reverse Transcriptase/metabolism , HIV-1/enzymology , HIV-1/genetics , Models, Chemical , Molecular Structure , Mutation , Purines/chemistry , Reverse Transcriptase Inhibitors/chemical synthesis , Reverse Transcriptase Inhibitors/chemistry , Structure-Activity Relationship
8.
J Med Chem ; 57(13): 5764-76, 2014 Jul 10.
Article in English | MEDLINE | ID: mdl-24911527

ABSTRACT

Human farnesyl pyrophosphate synthase (hFPPS) is the gate-keeper of mammalian isoprenoids and the key target of bisphosphonate drugs. Bisphosphonates suffer from poor "drug-like" properties and are mainly effective in treating skeletal diseases. Recent investigations have implicated hFPPS in various nonskeletal diseases, including Alzheimer's disease (AD). Analysis of single nucleotide polymorphisms in the hFPPS gene and mRNA levels in autopsy-confirmed AD subjects was undertaken, and a genetic link between hFPPS and phosphorylated tau (P-Tau) levels in the human brain was identified. Elevated P-Tau levels are strongly implicated in AD progression. The development of nonbisphosphonate inhibitors can provide molecular tools for validating hFPPS as a therapeutic target for tauopathy-associated neurodegeneration. A multistage screening protocol led to the identification of a new monophosphonate chemotype that bind in an allosteric pocket of hFPPS. Optimization of these compounds could lead to human therapeutics that block tau metabolism and arrest the progression of neurodegeneration.


Subject(s)
Enzyme Inhibitors/pharmacology , Geranyltranstransferase/antagonists & inhibitors , Organophosphonates/pharmacology , Allosteric Site/drug effects , Alzheimer Disease/genetics , Alzheimer Disease/physiopathology , Catalytic Domain , Crystallography, X-Ray , Diphosphonates/pharmacology , Drug Evaluation, Preclinical , Geranyltranstransferase/genetics , Geranyltranstransferase/metabolism , Humans , Ligands , Neurodegenerative Diseases/drug therapy , Organophosphonates/chemical synthesis , Phosphorylation , Polymorphism, Single Nucleotide , Pyrimidines/pharmacology , tau Proteins/metabolism
9.
Acta Crystallogr F Struct Biol Commun ; 70(Pt 3): 299-304, 2014 Mar.
Article in English | MEDLINE | ID: mdl-24598914

ABSTRACT

Human farnesyl pyrophosphate synthase (hFPPS) produces farnesyl pyrophosphate, an isoprenoid essential for a variety of cellular processes. The enzyme has been well established as the molecular target of the nitrogen-containing bisphosphonates (N-BPs), which are best known for their antiresorptive effects in bone but are also known for their anticancer properties. Crystal structures of hFPPS in ternary complexes with a novel bisphosphonate, YS0470, and the secondary ligands inorganic phosphate (Pi), inorganic pyrophosphate (PPi) and isopentenyl pyrophosphate (IPP) have recently been reported. Only the co-binding of the bisphosphonate with either PPi or IPP resulted in the full closure of the C-terminal tail of the enzyme, a conformational change that is required for catalysis and that is also responsible for the potent in vivo efficacy of N-BPs. In the present communication, a co-crystal structure of hFPPS in complex with YS0470 and two molecules of Pi is reported. The unusually close proximity between these ligands, which was confirmed by anomalous diffraction data, suggests that they interact with one another, with their anionic charges neutralized in their bound state. The structure also showed the tail of the enzyme to be fully disordered, indicating that simultaneous binding of two Pi molecules with a bisphosphonate cannot induce the tail-closing conformational change in hFPPS. Examination of homologous FPPSs suggested that this ligand-dependent tail closure is only conserved in the mammalian proteins. The prevalence of Pi-bound hFPPS structures in the PDB raises a question regarding the in vivo relevance of Pi binding to the function of the enzyme.


Subject(s)
Aminopyridines/chemistry , Diphosphonates/chemistry , Geranyltranstransferase/chemistry , Phosphates/chemistry , Amino Acid Sequence , Catalytic Domain , Crystallography, X-Ray , Humans , Models, Molecular , Molecular Sequence Data , Protein Binding
10.
BMC Struct Biol ; 12: 32, 2012 Dec 12.
Article in English | MEDLINE | ID: mdl-23234314

ABSTRACT

BACKGROUND: Human farnesyl pyrophosphate synthase (FPPS) controls intracellular levels of farnesyl pyrophosphate, which is essential for various biological processes. Bisphosphonate inhibitors of human FPPS are valuable therapeutics for the treatment of bone-resorption disorders and have also demonstrated efficacy in multiple tumor types. Inhibition of human FPPS by bisphosphonates in vivo is thought to involve closing of the enzyme's C-terminal tail induced by the binding of the second substrate isopentenyl pyrophosphate (IPP). This conformational change, which occurs through a yet unclear mechanism, seals off the enzyme's active site from the solvent environment and is essential for catalysis. The crystal structure of human FPPS in complex with a novel bisphosphonate YS0470 and in the absence of a second substrate showed partial ordering of the tail in the closed conformation. RESULTS: We have determined crystal structures of human FPPS in ternary complex with YS0470 and the secondary ligands inorganic phosphate (Pi), inorganic pyrophosphate (PPi), and IPP. Binding of PPi or IPP to the enzyme-inhibitor complex, but not that of Pi, resulted in full ordering of the C-terminal tail, which is most notably characterized by the anchoring of the R351 side chain to the main frame of the enzyme. Isothermal titration calorimetry experiments demonstrated that PPi binds more tightly to the enzyme-inhibitor complex than IPP, and differential scanning fluorometry experiments confirmed that Pi binding does not induce the tail ordering. Structure analysis identified a cascade of conformational changes required for the C-terminal tail rigidification involving Y349, F238, and Q242. The residues K57 and N59 upon PPi/IPP binding undergo subtler conformational changes, which may initiate this cascade. CONCLUSIONS: In human FPPS, Y349 functions as a safety switch that prevents any futile C-terminal closure and is locked in the "off" position in the absence of bound IPP. Q242 plays the role of a gatekeeper and directly controls the anchoring of R351 side chain. The interactions between the residues K57 and N59 and those upstream and downstream of Y349 are likely responsible for the switch activation. The findings of this study can be exploited for structure-guided optimization of existing inhibitors as well as development of new pharmacophores.


Subject(s)
Catalytic Domain , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Geranyltranstransferase/antagonists & inhibitors , Geranyltranstransferase/chemistry , Models, Molecular , Crystallography, X-Ray , Diphosphates/chemistry , Diphosphates/metabolism , Drug Design , Hemiterpenes/chemistry , Hemiterpenes/metabolism , Humans , Ligands , Organophosphorus Compounds/chemistry , Organophosphorus Compounds/metabolism , Protein Binding/drug effects , Protein Structure, Tertiary , Static Electricity
11.
Bioorg Med Chem ; 20(18): 5583-91, 2012 Sep 15.
Article in English | MEDLINE | ID: mdl-22884353

ABSTRACT

Nitrogen-containing bisphosphonates (N-BPs) are potent active site inhibitors of the human farnesyl pyrophosphate synthase (hFPPS) and valuable human therapeutics for the treatment of bone-related malignancies. N-BPs are also useful in combination chemotherapy for patients with breast, prostate and multiple myeloma cancers. A structure-based approach was employed in order to design inhibitors that exhibit higher lipophilicity and better occupancy for the GPP sub-pocket of hFPPS than the current therapeutic drugs. These novel analogs were designed to bind deeper into the GPP sub-pocket by displacing the side chains of the 'capping' residue Phe 113 and engaging in favorable π-interactions with the side chain of Phe112.


Subject(s)
Diphosphonates/pharmacology , Drug Design , Enzyme Inhibitors/pharmacology , Geranyltranstransferase/antagonists & inhibitors , Catalytic Domain/drug effects , Diphosphonates/chemical synthesis , Diphosphonates/chemistry , Dose-Response Relationship, Drug , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Geranyltranstransferase/metabolism , Humans , Models, Molecular , Molecular Structure , Structure-Activity Relationship
12.
J Med Chem ; 55(7): 3201-15, 2012 Apr 12.
Article in English | MEDLINE | ID: mdl-22390415

ABSTRACT

Human farnesyl pyrophosphate synthase (hFPPS) controls intracellular levels of FPP and post-translational prenylation of small GTPase proteins, which are essential for cell signaling and cell proliferation. Clinical investigations provide evidence that N-BP inhibitors of hFPPS are disease modifying agents that improve survival of multiple myeloma (MM) patients via mechanisms unrelated to their skeletal effects. A new series of N-BPs was designed that interact with a larger portion of the GPP subpocket, as compared to the current therapeutic drugs, and rigidify the (364)KRRK(367) tail of hFPPS in the closed conformation in the absence of IPP. An analogue of this series was used to demonstrate inhibition of the intended biological target, resulting in apoptosis and down-regulation of ERK phosphorylation in human MM cell lines.


Subject(s)
Antineoplastic Agents/chemical synthesis , Apoptosis/drug effects , Diphosphonates/chemical synthesis , Extracellular Signal-Regulated MAP Kinases/metabolism , Geranyltranstransferase/antagonists & inhibitors , Multiple Myeloma/pathology , Aminopyridines/chemical synthesis , Aminopyridines/chemistry , Aminopyridines/pharmacology , Aniline Compounds/chemical synthesis , Aniline Compounds/chemistry , Aniline Compounds/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Catalytic Domain , Cell Survival/drug effects , Crystallography, X-Ray , Diphosphonates/chemistry , Diphosphonates/pharmacology , Drug Design , Hemiterpenes/chemistry , Humans , Models, Molecular , Multiple Myeloma/metabolism , Organophosphorus Compounds/chemistry , Phosphorylation , Protein Conformation , Small Molecule Libraries , Stereoisomerism , Structure-Activity Relationship , Tumor Cells, Cultured
14.
J Med Chem ; 51(23): 7428-41, 2008 Dec 11.
Article in English | MEDLINE | ID: mdl-19053781

ABSTRACT

This report describes the synthesis of four novel paclitaxel based prodrugs with glycan conjugation (1-4). Glycans were conjugated using an ester or ether bond as the linker between 2'-paclitaxel and the 2'-glucose or glucuronic acid moiety. These prodrugs showed good water solubility and selective cytotoxicity against cancer cell lines, but showed reduced toxicity toward normal cell lines and cancer cell lines with low expression levels of GLUTs. The ester conjugated prodrug 1 showed the most cytotoxicity among the prodrugs examined and could be transported into cells via GLUTs. Fluorescent and confocal microscopy demonstrated that targeted cells exhibited morphological changes in tubulin and chromosomal alterations that were similar to those observed with paclitaxel treatment. Therefore, these glycan-based prodrugs may be good drug candidates for cancer therapy, and the glycan conjugation approach is an alternative method to enhance the targeted delivery of other drugs to cancer cells that overexpress GLUTs.


Subject(s)
Drug Delivery Systems , Glucose Transport Proteins, Facilitative/metabolism , Paclitaxel/pharmacology , Polysaccharides/chemistry , Prodrugs/pharmacology , Animals , CHO Cells , Cell Line , Cell Proliferation/drug effects , Cricetinae , Cricetulus , Dose-Response Relationship, Drug , Drug Design , Drug Evaluation, Preclinical , Humans , Molecular Structure , Paclitaxel/chemistry , Paclitaxel/metabolism , Prodrugs/chemistry , Prodrugs/metabolism , Stereoisomerism
15.
J Proteome Res ; 7(4): 1379-87, 2008 Apr.
Article in English | MEDLINE | ID: mdl-18290608

ABSTRACT

Tumor secreted substances (secretome), including extracellular matrix (ECM) components, act as mediators of tumor-host communication in the breast tumor microenvironment. Proteomic analysis has emphasized the value of the secretome as a source of prospective markers and drug targets for the treatment of breast cancers. Utilizing bioinformatics, our recent studies revealed global changes in protein expression after the activation of ECM-mediated signaling in breast cancer cells. A newly designed technique integrating a capillary ultrafiltration (CUF) probe with mass spectrometry was demonstrated to dynamically sample and identify in vivo and pure secretome from the tumor microenvironment. Such in vivo profiling of breast cancer secretomes may facilitate the development of novel drugs specifically targeting secretome.


Subject(s)
Breast Neoplasms/metabolism , Extracellular Space/metabolism , Proteins/metabolism , Proteomics/methods , Apoptosis/drug effects , Breast Neoplasms/drug therapy , Breast Neoplasms/pathology , Extracellular Matrix/chemistry , Extracellular Matrix/metabolism , Female , Humans , Integrins/antagonists & inhibitors , Models, Biological , Proteins/analysis
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