ABSTRACT
Inhibiting the PD-1/PD-L1 protein-protein interaction is a key immunotherapy for cancer. Antibodies dominate the clinical space but are costly, with limited applicability and immune side effects. We developed a photo-controlled azobenzene peptide that selectively inhibits the PD-1/PD-L1 interaction when in the cis isomer only. Activity is demonstrated in in vitro and cellular assays.
Subject(s)
Azo Compounds , B7-H1 Antigen , Light , Peptides , Programmed Cell Death 1 Receptor , Azo Compounds/chemistry , Azo Compounds/pharmacology , Humans , Peptides/chemistry , Peptides/pharmacology , Programmed Cell Death 1 Receptor/antagonists & inhibitors , Programmed Cell Death 1 Receptor/metabolism , B7-H1 Antigen/antagonists & inhibitors , B7-H1 Antigen/metabolism , Protein BindingABSTRACT
The synthesis of the ethyl ester analogue of the ultrapotent antitumour antibiotic seco-duocarmycin SA has been achieved in eleven linear steps from commercially available starting materials. The DSA alkylation subunit can be made in ten linear steps from the same precursor. The route involves C-H activation at the equivalent of the C7 position on indole leading to a borylated intermediate 9 that is stable enough for peptide coupling reactions but can be easily converted to the free hydroxyl analogue.
Subject(s)
Duocarmycins , Indoles , Iridium , Catalysis , Indoles/chemistry , Indoles/chemical synthesis , Iridium/chemistry , Molecular Structure , Pyrroles/chemistry , Pyrroles/chemical synthesis , Boron Compounds/chemistry , Boron Compounds/chemical synthesisABSTRACT
Suramin was the first effective drug for the treatment of human African sleeping sickness. Structural analogues of the trypanocide have previously been shown to be potent inhibitors of several enzymes. Therefore, four suramin analogues lacking the methyl group on the intermediate rings and with different regiochemistry of the naphthalenetrisulphonic acid groups and the phenyl rings were tested to establish whether they exhibited improved antiproliferative activity against bloodstream forms of Trypanosomes brucei compared to the parent compound. The four analogues exhibited low trypanocidal activity and weak inhibition of the antitrypanosomal activity of suramin in competition experiments. This indicates that the strong trypanocidal activity of suramin is most likely due to the presence of methyl groups on its intermediate rings and to the specific regiochemistry of naphthalenetrisulphonic acid groups. These two structural features are also likely to be important for the inhibition mechanism of suramin because DNA distribution and nucleus/kinetoplast configuration analyses suggest that the analogues inhibit mitosis while suramin inhibits cytokinesis.
Subject(s)
Suramin , Trypanocidal Agents , Trypanosoma brucei brucei , Suramin/pharmacology , Suramin/chemistry , Trypanocidal Agents/pharmacology , Trypanocidal Agents/chemistry , Trypanosoma brucei brucei/drug effects , Animals , Structure-Activity Relationship , DNA, Protozoan/drug effects , DNA, Kinetoplast/drug effects , Mice , Mitosis/drug effects , Trypanosomiasis, African/drug therapy , Trypanosomiasis, African/parasitologyABSTRACT
Cancer metastasis is the cause of up to 90 % of cancer related mortality. The CXCR4 receptor and its cognate ligand, CXCL12, have major roles in enabling cancer metastasis and consequently, the CXCR4 receptor has become an attractive therapeutic target for the prevention of metastasis. Despite this, CXCR4 antagonists have had limited success in clinical trials due to cellular toxicity and poor stability and efficacy. In this study, we developed a novel, competitive CXCR4 antagonist (IS4) that through copper-catalysed-azide-alkyne-cycloaddition can be clicked to other chemical moieties such as fluorescent dyes (IS4-FAM) for CXCR4-based imaging. We determined that these CXCR4 antagonists were non-toxic and could be used to specifically label the CXCR4 receptor. Furthermore, IS4 and IS4-FAM inhibited CXCL12-stimulated cancer cell migration and Ca2+ release in both adherent and suspension cell lines with similar or improved potency as compared to two literature CXCR4 antagonists. Our results highlight the potential of IS4 and IS4-FAM as research tools and as potent CXCR4 antagonists for the prevention of metastasis.
Subject(s)
Chemokine CXCL12 , Receptors, CXCR4 , Humans , Cell Line, Tumor , Chemokine CXCL12/metabolism , Signal Transduction , Cell Movement , Fluorescent Dyes , Neoplasm Metastasis/prevention & controlABSTRACT
Osteoarthritis is a chronic degenerative joint disease affecting millions of people worldwide, with no disease-modifying drugs currently available to treat the disease. Tissue inhibitor of metalloproteinases 3 (TIMP-3) is a potential therapeutic target in osteoarthritis because of its ability to inhibit the catabolic metalloproteinases that drive joint damage by degrading the cartilage extracellular matrix. We previously found that suramin inhibits cartilage degradation through its ability to block endocytosis and intracellular degradation of TIMP-3 by low-density lipoprotein receptor-related protein 1 (LRP1), and analysis of commercially available suramin analogues indicated the importance of the 1,3,5-trisulfonic acid substitutions on the terminal naphthalene rings for this activity. Here we describe synthesis and structure-activity relationship analysis of additional suramin analogues using ex vivo models of TIMP-3 trafficking and cartilage degradation. This showed that 1,3,6-trisulfonic acid substitution of the terminal naphthalene rings was also effective, and that the protective activity of suramin analogues depended on the presence of a rigid phenyl-containing central region, with para/para substitution of these phenyl rings being most favourable. Truncated analogues lost protective activity. The physicochemical characteristics of suramin and its analogues indicate that approaches such as intra-articular injection would be required to develop them for therapeutic use.
Subject(s)
Osteoarthritis , Tissue Inhibitor of Metalloproteinase-3 , Humans , Tissue Inhibitor of Metalloproteinase-3/metabolism , Tissue Inhibitor of Metalloproteinase-3/pharmacology , Tissue Inhibitor of Metalloproteinase-3/therapeutic use , Suramin/pharmacology , Suramin/metabolism , Suramin/therapeutic use , Cartilage/metabolism , Osteoarthritis/drug therapy , Osteoarthritis/metabolism , Metalloproteases/metabolism , Metalloproteases/pharmacology , Metalloproteases/therapeutic useABSTRACT
DNA has been a key target for cancer therapy, with a range of compounds able to bind and either impair its processing or induce damage. Targeting DNA with small molecules in a truly sequence specific way, to impair gene specific processes, remains out of reach. The ability of DNA to assume different structures from the classical double helix allows access to more specific ligand binding modes and, potentially, to new avenues of treatment. In this review, we illustrate the small molecules that have been reported to bind to three- and four-way junctions.
Subject(s)
DNA , DNA/chemistry , Nucleic Acid ConformationABSTRACT
The P2X4 receptor is a ligand-gated ion channel activated by extracellular ATP. P2X4 activity is associated with neuropathic pain, vasodilation, and pulmonary secretion and is therefore of therapeutic interest. The structure-activity relationship of P2X4 antagonists is poorly understood. Here we elucidate the structure-activity of 5-(3-bromophenyl)-1,3-dihydro-2H-benzofuro[3,2-e]-1,4-diazepin-2-one (5-BDBD) at human P2X4 by combining pharmacology, electrophysiology, molecular modeling, and medicinal chemistry. 5-BDBD antagonized P2X4 in a noncompetitive manner but lacked effect at human P2X2. Molecular modeling and site-directed mutagenesis suggested an allosteric binding site for 5-BDBD located between two subunits in the body region of P2X4, with M109, F178, Y300, and I312 on one subunit and R301 on the neighboring subunit as key residues involved in antagonist binding. The bromine group of 5-BDBD was redundant for the antagonist activity of 5-BDBD, although an interaction between the carbonyl group of 5-BDBD and R301 in P2X4 was associated with 5-BDBD activity. 5-BDBD could inhibit the closed channel but poorly inhibited the channel in the open/desensitizing state. We hypothesize that this is due to constriction of the allosteric site after transition from closed to open channel state. We propose that M109, F178, Y300, R301, and I312 are key residues for 5-BDBD binding; provide a structural explanation of how they contribute to 5-BDBD antagonism; and highlight that the limited action of 5-BDBD on open versus closed channels is due to a conformational change in the allosteric site. SIGNIFICANCE STATEMENT: Activity of P2X4 receptor is associated with neuropathic pain, inflammation, and vasodilatation. Molecular information regarding small-molecule interaction with P2X4 is very limited. Here, this study provides a structural explanation for the action of the small-molecule antagonist 5-BDBD at the human P2X4 receptor.
Subject(s)
Benzodiazepinones/chemistry , Benzodiazepinones/metabolism , Purinergic P2X Receptor Antagonists/chemistry , Purinergic P2X Receptor Antagonists/metabolism , Receptors, Purinergic P2X4/chemistry , Receptors, Purinergic P2X4/metabolism , Allosteric Regulation/drug effects , Allosteric Regulation/physiology , Benzodiazepinones/pharmacology , HEK293 Cells , Humans , Molecular Dynamics Simulation , Protein Structure, Secondary , Protein Structure, Tertiary , Purinergic P2X Receptor Antagonists/pharmacologyABSTRACT
BACKGROUND AND PURPOSE: P2X4 is a ligand-gated cation channel activated by extracellular ATP involved in neuropathic pain, inflammation and arterial tone. EXPERIMENTAL APPROACH: Natural products were screened against human or mouse P2X4 activity using fura-2 loaded 1321N1 cells for measurement of intracellular Ca2+ responses. Whole-cell currents were measured by patch clamp. Human primary macrophage chemokine release was used to assess effect of taspine on inflammatory cell function. An enzymatic assay was performed to assess the effect of taspine on recombinant PI3-kinase. KEY RESULTS: A natural product screen identified taspine as an inhibitor of human P2X4 activity. Taspine inhibits human and mouse P2X4-mediated Ca2+ influx in 1321N1 cells expressing receptors but lacked activity at human P2X2, P2X3, P2X2/3 and P2X7 receptors. Taspine inhibited the maximal response at human and mouse P2X4 but effective on ATP potency. Taspine has a slow onset rate (~15 min for half-maximal inhibition), irreversible over 30 min of washout. Taspine inhibits P2X4-mediated Ca2+ signalling in mouse BV-2 microglia cells and human primary macrophage. Taspine inhibited P2X4-mediated CXCL5 secretion in human primary macrophage. Taspine reversed ivermectin-induced potentiation of P2X4 currents in 1321N1 stably expressing cells. The PI3-kinase inhibitor LY294002 mimicked the properties of taspine on P2X4-mediated Ca2+ influx and whole-cell currents. Taspine directly inhibited the enzymatic activity of recombinant PI3-kinase in a competitive manner. CONCLUSION AND IMPLICATIONS: Taspine is a novel natural product P2X4 receptor inhibitor, mediating its effect through PI3-kinase inhibition rather than receptor antagonism. Taspine can inhibit the pro-inflammatory signalling by P2X4 in human primary macrophage.
Subject(s)
Biological Products , Receptors, Purinergic P2X4 , Adenosine Triphosphate , Alkaloids , Animals , Mice , Phosphatidylinositol 3-Kinase , Phosphatidylinositol 3-Kinases , Receptors, Purinergic P2X7ABSTRACT
The duocarmycins belong to a class of agent which has great potential for use in cancer therapy. Their exquisite potency means they are too toxic for systemic use, and targeted approaches are required to unlock their clinical potential. In this study, we have explored seco-OH-chloromethylindoline (CI) duocarmycin-based bioprecursors for their potential for cytochrome P450 (CYP)-mediated cancer cell kill. We report on synthetic and biological explorations of racemic seco-CI-MI, where MI is a 5-methoxy indole motif, and dehydroxylated analogues. We show up to a 10-fold bioactivation of de-OH CI-MI and a fluoro bioprecursor analogue in CYP1A1-transfected cells. Using CYP bactosomes, we also demonstrate that CYP1A2 but not CYP1B1 or CYP3A4 has propensity for potentiating these compounds, indicating preference for CYP1A bioactivation.
Subject(s)
Antineoplastic Agents/pharmacology , Cytochrome P-450 Enzyme Inhibitors/pharmacology , Cytochrome P-450 Enzyme System/metabolism , Duocarmycins/pharmacology , Indoles/pharmacology , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Cell Line, Tumor , Cell Proliferation/drug effects , Cytochrome P-450 Enzyme Inhibitors/chemical synthesis , Cytochrome P-450 Enzyme Inhibitors/chemistry , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Duocarmycins/chemical synthesis , Duocarmycins/chemistry , Humans , Indoles/chemical synthesis , Indoles/chemistry , Molecular Structure , Structure-Activity RelationshipABSTRACT
Gold nanoparticles (AuNPs) are of increasing interest for their unique properties and their biocompatability, minimal toxicity, multivalency and size tunability make them exciting drug carriers. The functionalisaton of AuNPs with targeting moieties allows for their selective delivery to cancers, with antibodies, proteins, peptides, aptamers, carbohydrates and small molecules all exploited. Here, we review the recent advances in targeted-AuNPs for the treatment of cancer, with a particular focus on these classes of targeting ligands. We highlight the benefits and potential drawbacks of each ligand class and propose directions in which the field could grow.
Subject(s)
Antineoplastic Agents/therapeutic use , Gold/chemistry , Metal Nanoparticles/chemistry , Neoplasms/drug therapy , Antineoplastic Agents/chemistry , Drug Carriers/chemistry , Humans , LigandsABSTRACT
Solid-phase synthesis allowed the rapid generation of a peptide-drug conjugate. A peptide targeting the Thomsen-Friedenreich antigen (TFα) was conjugated to the alkylating subunit of the potent cytotoxin duocarmycin SA. The compound, containing a cathepsin B cleavable linker, was shown to be active and selective against TFα expressing tumor cell lines.
Subject(s)
Antigens, Tumor-Associated, Carbohydrate/drug effects , Antineoplastic Agents/pharmacology , Duocarmycins/chemistry , Peptides/chemistry , Amino Acid Sequence , Antineoplastic Agents/chemistry , Cell Line, Tumor , Cell Proliferation/drug effects , HumansABSTRACT
Gold nanoparticles, covalently functionalised with the photosensitiser C11Pc and PEG, were actively targeted towards epidermal growth factor receptor overexpressing cancers using the peptide FITC-ßAAEYLRK. Selective phototoxicity was observed at nanomolar concentrations with minimal dark toxicity.
ABSTRACT
The development of protein-protein interaction (PPI) inhibitors with therapeutic value is of increasing importance as the first clinical agent has now been approved, but PPIs remain difficult targets for the development of small molecule ligands. This article describes a highly efficient approach to the development of inhibitors of the p53/hDMX or hDM2 interaction that involves the design of small molecules in silico based upon a peptide/protein structure. The process for molecule design, starting from a virtual library of just over 1200 fragments, led to the eventual synthesis of twenty compounds, of which ten bound to either hDM2, hDMX or both in in vitro binding assays. This 50% success rate is extremely efficient compared to traditional high throughput screening. The identification of two selective hDMX inhibitors from twenty compounds highlights this efficiency as, to date, only two other hDMX-selective agents exist in the literature. Preliminary biological studies show that 20% of the compounds identified have cellular activity and activate downstream pathways associated with p53 activation.
ABSTRACT
In a proof-of-concept study, solid phase synthesis allowed the rapid generation of a small molecule drug conjugate in which the glutamate carboxypeptidase II (GCPII) targeting small molecule DUPA was conjugated to the alkylating subunit of the potent cytotoxin duocarmycin SA. The targeted SMDC contained a cathepsin B cleavable linker, which was shown to be active and selective against cathepsin B over-expressing and GCPII-expressing tumour cell lines.
ABSTRACT
(C^N) and (C^N^C) cyclometalated Au(iii) represent a highly promising class of potential anticancer agents. We report here the synthesis of seven new cyclometalated Au(iii) complexes with five of them bearing an acridine moiety attached via (N^O) or (N^N) chelates, acyclic amino carbenes (AAC) and N-heterocyclic carbenes (NHC). The antiproliferative properties of the different complexes were evaluated in vitro on a panel of cancer cells including leukaemia, lung and breast cancer cells. We observed a trend between the cytotoxicity and the intracellular gold uptake of some representative compounds of the series. Some of the acridine-decorated complexes were demonstrated to interact with ds-DNA using FRET-melting techniques.
Subject(s)
Acridines/chemistry , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Organogold Compounds/chemical synthesis , Organogold Compounds/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/metabolism , Cell Line, Tumor , Cell Proliferation/drug effects , Chemistry Techniques, Synthetic , DNA/metabolism , Drug Screening Assays, Antitumor , Glutathione/metabolism , Humans , Models, Molecular , Molecular Conformation , Organogold Compounds/chemistry , Organogold Compounds/metabolismABSTRACT
A series of cationic mixed cyclometallated (C^N)Au(iii) dithiocarbamate complexes has been synthesized in good yields [HC^N = 2-(p-t-butylphenyl)pyridine]. The crystal structure of [(C^N)AuS2CNEt2]PF6 (3) has been determined. The cytotoxic properties of the new complexes have been evaluated in vitro against a panel of human cancer cell lines and healthy cells and compared with a neutral mixed (C^C)Au(iii) dithiocarbamate complex (C^C = 4,4'-di-t-butylbiphenyl-2,2'-diyl). The complexes appeared to be susceptible to reduction by glutathione but were stable in the presence of N-acetyl cysteine. The potential mechanism of action of this class of compounds has been investigated by measuring the intracellular uptake of some selected complexes, by determining their interactions with higher order DNA structures, and by assessing the ability to inhibit thioredoxin reductase. The complexes proved unable to induce the formation of reactive oxygen species. The investigations add to the picture of the possible mode of action of this class of complexes.
Subject(s)
Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Cell Proliferation/drug effects , Coordination Complexes/chemistry , Gold Compounds/pharmacology , Gold/chemistry , Thiocarbamates/pharmacology , Drug Screening Assays, Antitumor , Gold Compounds/chemistry , Humans , Tumor Cells, CulturedABSTRACT
The Nrf2/Keap1 interaction is a target in the development of new therapeutic agents, where inhibition of the interaction activates Nrf2 and leads to the generation of downstream anti-inflammatory effects. Peptides that mimic the ß-turn in the Keap1 active site and are constrained by a disulfide bridge have high affinity for Keap1 but no intracellular activity. The introduction of a perfluoroalkyl-bridging group to constrain the peptides, coupled with a glutamic acid to proline replacement leads to a new peptide with a Ki of 6.1â¯nM for the Nrf2/Keap1 binding interaction, although this does not translate into intracellular activity.
Subject(s)
Fluorocarbons/pharmacology , Kelch-Like ECH-Associated Protein 1/antagonists & inhibitors , Macrocyclic Compounds/pharmacology , NF-E2-Related Factor 2/antagonists & inhibitors , Peptides/pharmacology , Dose-Response Relationship, Drug , Fluorocarbons/chemistry , Humans , Kelch-Like ECH-Associated Protein 1/metabolism , Macrocyclic Compounds/chemical synthesis , Macrocyclic Compounds/chemistry , Molecular Structure , NF-E2-Related Factor 2/metabolism , Peptides/chemistry , Structure-Activity RelationshipABSTRACT
A series of cyclometallated gold(iii) complexes supported by pyrazine-based (C^Npz^C)-type pincer ligands were synthesized via two different pathways. Nucleophilic attack on the isocyanide complex [(C^Npz^C)Au(C[triple bond, length as m-dash]NC6H3Me2-2,6)]SbF6 (2) gave [(C^Npz^C)Au(ACC)]SbF6 complexes with aniline (4·SbF6), adamantylamine (5), glycine ethyl ester (6), alanine methyl ester (7), valine methyl ester (8), phenylglycine methyl ester (9) and methionine methyl ester (10) substituents (ACC = acyclic carbene). The pathway via isocyanide insertion into gold-amide bonds was also investigated; e.g. the reaction of xylyl isocyanide with (C^Npz^C)AuNHPh followed by protonation with HBF4·OEt2 gave the acyclic carbene complex 4·BF4. To the best of our knowledge compounds 6-10 represent the first examples of gold(iii) acyclic carbene complexes bearing amino acid functions. The compounds provide a versatile platform for the study of the anti-proliferative properties of gold(iii) complexes. Tests against human adenoma-type lung cancer cells identified 5, 6, 7 and 10 as particularly promising and demonstrate the synthetic flexibility of acyclic carbene complexes and the potential of that class of compounds for anticancer applications. Compared to cisplatin, amino ester-containing ACC complexes showed improved selectivity for MCF-7 breast cancer cells over that for healthy fibroblasts.
Subject(s)
Antineoplastic Agents/chemical synthesis , Coordination Complexes/chemistry , Gold/chemistry , Methane/analogs & derivatives , Pyrazines/chemistry , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Carbon/chemistry , Cell Line , Cell Proliferation/drug effects , Cell Survival/drug effects , Coordination Complexes/chemical synthesis , Coordination Complexes/pharmacology , Crystallography, X-Ray , Drug Screening Assays, Antitumor , Glutathione/chemistry , Glutathione/metabolism , Humans , Ligands , MCF-7 Cells , Methane/chemistry , Molecular Conformation , Nitrogen/chemistry , Reactive Oxygen Species/metabolismABSTRACT
The synthesis of a series of cyclometalated gold(III) complexes supported by pyrazine-based (C^N^C)-type pincer ligands is reported, including the crystal structure of a cationic example. The compounds provide a new platform for the study of antiproliferative properties of gold(III) complexes. Seven complexes were tested: the neutral series (C^Npz^C)AuX [X = Cl (1), 6-thioguanine (4), C≡CPh (5), SPh (6)] and an ionic series that included the N-methyl complex [(C^NpzMe^C)AuCl]BF4 (7) and the N-heterocyclic carbene complexes [(C^Npz^C)AuL]+ with L = 1,3-dimethylbenzimidazol-2-ylidene (2) or 1,3,7,9-tetramethylxanthin-8-ylidene (3). Tests against human leukemia cells identified 1, 2, 3, and 4 as particularly promising, whereas protecting the noncoordinated N atom on the pyrazine ring by methylation (as in 7) reduced the cytotoxicity. Complex 2 proved to be the most effective of the entire series against the HL60 leukemia, MCF-7 breast cancer, and A549 lung cancer cell lines, with IC50 values down to submicromolar levels, associated with a lower toxicity toward healthy human lung fibroblast cells. The benzimidazolylidene complex 2 accumulated more effectively in human lung cancer cells than its caffeine-based analogue 3 and the gold(III) chloride 1. Compound 2 proved to be unaffected by glutathione under physiological conditions for periods of up to 6 days and stabilizes the DNA G-quadruplex and i-motif structures; the latter is the first such report for gold compounds. We also show the first evidence of inhibition of MDM2-p53 protein-protein interactions by a gold-based compound and identified the binding mode of the compound with MDM2 using saturation transfer difference NMR spectroscopy combined with docking calculations.
Subject(s)
Antineoplastic Agents/pharmacology , Methane/analogs & derivatives , Organogold Compounds/pharmacology , Pyrazines/pharmacology , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Cell Line, Tumor , Cell Proliferation/drug effects , Crystallography, X-Ray , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Fluorescence Polarization , Humans , Ligands , Methane/chemistry , Methane/pharmacology , Models, Molecular , Molecular Structure , Organogold Compounds/chemical synthesis , Organogold Compounds/chemistry , Pyrazines/chemistry , Structure-Activity RelationshipABSTRACT
Inhibitors of the p53-MDM2 protein-protein interaction are emerging as a new and validated approach to treating cancer. Herein, we describe the synthesis and inhibitory evaluation of a series of isoquinolin-1-one analogues, and highlight the utility of an initial growth-rates saturation-transfer difference (STD) NMR approach supported by protein-ligand docking to investigate p53-MDM2 inhibition. The approach is illustrated by the study of compound 1, providing key insights into the binding mode of this kind of MDM2 ligands and, more importantly, readily unveiling the previously proposed three-finger pharmacophore requirement for p53-MDM2 inhibition.