ABSTRACT
The inhibition of the androgen receptor (AR) is an established strategy in prostate cancer (PCa) treatment until drug resistance develops either through mutations in the ligand-binding domain (LBD) portion of the receptor or its deletion. We previously identified a druggable pocket on the DNA binding domain (DBD) dimerization surface of the AR and reported several potent inhibitors that effectively disrupted DBD-DBD interactions and consequently demonstrated certain antineoplastic activity. Here we describe further development of small molecule inhibitors of AR DBD dimerization and provide their broad biological characterization. The developed compounds demonstrate improved activity in the mammalian two-hybrid assay, enhanced inhibition of AR-V7 transcriptional activity, and improved microsomal stability. These findings position us for the development of AR inhibitors with entirely novel mechanisms of action that would bypass most forms of PCa treatment resistance, including the truncation of the LBD of the AR.
Subject(s)
Androgen Receptor Antagonists/pharmacology , DNA, Neoplasm/metabolism , Gene Expression Regulation, Neoplastic/drug effects , Prostatic Neoplasms/drug therapy , Receptors, Androgen/chemistry , Small Molecule Libraries/pharmacology , Transcription, Genetic , Androgen Receptor Antagonists/chemistry , Computer Simulation , DNA, Neoplasm/antagonists & inhibitors , High-Throughput Screening Assays , Humans , Male , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/pathology , Protein Conformation , Protein Domains , Receptors, Androgen/genetics , Receptors, Androgen/metabolism , Small Molecule Libraries/chemistry , Tumor Cells, CulturedABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Cyperus rotundus L. (Cyperaceae) is a widespread herbal in China and widely used in Traditional Chinese Medicine for multiple effects such as anti-arthritic, anti-genotoxic, anti-mutagenic, anti-bacterial effects, and analgesic. α-Cyperone is an active compound in Cyperus rotundus and has analgesic effects, but the exact molecular mechanisms require further investigations. MATERIALS AND METHODS: Tumor-derived DNA isolated from Lewis cell lines was transfected into microglia, and analyzed for stimulator of interferon genes (STING) effects. The downstream protein, such as interferon regulatory factor 3 (IRF3) and p65 nuclear factor-κB (NF-κB) were treated with STING siRNA and 5,6-dimethyllxanthenone-4-acetic acid (DMXAA) in microglia. The α-Cyperone effect on microglia was also investigated. RESULTS: Tumor-derived DNA activate microglia by upregulation of STING and downstream proteins. STING siRNA was reduced to its downstream expression and neuroinflammation inhibition was caused by tumor-derived DNA. However, DMXAA reversed the STING siRNA effect and increased neuroinflammation. α-Cyperone takes inhibitory effects on tumor-derived DNA that trigger microglia by STING pathway. CONCLUSIONS: α-Cyperone inhibition by tumor-derived DNA activated microglial to neuroinflammation in STING signaling pathway.
Subject(s)
DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/genetics , Membrane Proteins/antagonists & inhibitors , Membrane Proteins/genetics , Microglia/drug effects , Naphthalenes/pharmacology , Animals , Carcinoma, Lewis Lung/drug therapy , Carcinoma, Lewis Lung/genetics , Dose-Response Relationship, Drug , Drugs, Chinese Herbal/pharmacology , Drugs, Chinese Herbal/therapeutic use , Mice , Microglia/physiology , Naphthalenes/therapeutic useABSTRACT
Phortress is an anticancer prodrug, which has active metabolite (5F-203) being potent agonist of the aryl hydrocarbon receptor (AhR). The 5F-203 switches on cytochrome P450 CYP1A1 gene expression and thus exhibits anticancer activity. In this study, it is aimed to obtain new phortress analogues by bioisosteric replacement of benzothiazole core in the structure to benzoxazole ring system. Synthesis of compounds (3a-3p) were performed according to literature methods. Their structures were elucidated by IR, 1H NMR, 13C NMR, 2D-NMR and HRMS spectroscopic methods. Cytotoxicity (MTT), inhibition of DNA synthesis and flow cytometric analysis assays were applied to determine anticancer activity of the compounds on colon (HT-29), breast (MCF7), lung (A549), liver (HepG2) and brain (C6) carcinoma cell types. When compared reference agent doxorubicin, compounds 3m and 3n displayed very attractive anticancer effect against carcinogenic cell lines. Due to structural similarity to phortress, biotransformation studies for 3m and 3n were examined by LCMS-IT-TOF system and probable metabolites of these compounds were determined. Induction potential of these compounds on CYP1A1/2 enzymes was also investigated to clarify possible mechanism of action. Interaction modes between CYP1A1 enzyme and compound 3n or its some metabolites were investigated by docking studies. In conclusion, findings of these study indicate that compounds 3m and 3n possess significant anticancer activity, probably with the same mechanism of action to Phortress.
Subject(s)
Antineoplastic Agents/pharmacology , Benzoxazoles/pharmacology , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Benzoxazoles/chemical synthesis , Benzoxazoles/chemistry , Cell Proliferation/drug effects , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/biosynthesis , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Humans , Molecular Docking Simulation , Molecular Structure , Structure-Activity Relationship , Tumor Cells, CulturedABSTRACT
BACKGROUND: A pentacyclic lupenane-type natural triterpenoid, betulin, has attracted attention in the field of medicinal chemistry since it exhibited a variety of biological activities, including anticancer activity. OBJECTIVE: The aim of this present work was to obtain derivatives of betulin through bacterial biotransformation and investigate its anticancer activity against A549, HepG2 and 5RP7 cancer cell lines. METHODS: Bacterial biotransformation studies were continued in an MBH broth medium for 7 days at 35oC. Anticancer activities of betulin against A549, HepG2 and 5RP7 cell lines were carried out using XTT assay, and their selectivity was determined using a healthy cell line of NIH/3T3. Cell proliferation ELISA, BRDU (colorimetric) assay was used for measuring proliferation in replicative cells in which DNA synthesis occurs. Flow cytometric analysis was used for measuring apoptotic cell percentages, caspase 3 activation and mitochondrial membrane potential. RESULTS: Bacterial biotransformation studies with 7 bacteria of Staphylococcus aureus ATCC 6538, Proteus vulgaris NRRL B-123, Bacillus subtilis NRRL B-4378, Streptomyces griseolus NRRL B-1062, Escherichia coli ATCC 8739, Staphylococcus aureus ATCC 43300 and Bacillus velezensis NRRL B-14580 produced no metabolite. In in vitro anticancer activity studies, betulin was found to exert anticancer activity against A549, HepG2 and 5RP7 cell lines with IC50 values of 207.7, 125.0 and 28.3 µg/mL, whereas SI values were found to be 30, 50 and 223, respectively. Early and late apoptotic percentages of betulin were found as 9.6, 12.1 and 85.4% on A549, HepG2 and 5RP7, respectively, while caspase 3 positive cell percentages were 2.3, 28.7 and 13.3% for IC50 concentrations. In addition, betulin caused G1 cell cycle arrest (49.5%) on 5RP7 cell line. CONCLUSION: The results have been shown that betulin activities against A549 and HepG2 cell lines were nonselective and limited its cytotoxic activity against healthy cells, but it is possible to say that it exerted selective activity against 5RP7 cell (28.33±1.53 µg/mL). Betulin effects on apoptosis were found to be dosedependent, while its effect on caspase 3 activation, mitochondrial membrane potential, and cell cycle arrest on G0/G1 phase was not dependent on doses. Therefore, betulin could be a good candidate for the treatment of H-ras active cancer types.
Subject(s)
Antineoplastic Agents/pharmacology , Triterpenes/pharmacology , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/metabolism , Apoptosis/drug effects , Bacillus/metabolism , Bacillus subtilis/metabolism , Biotransformation , Cell Line , Cell Proliferation/drug effects , Cell Survival/drug effects , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/biosynthesis , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Escherichia coli/metabolism , Humans , Mice , Molecular Conformation , Proteus vulgaris/metabolism , Staphylococcus aureus/metabolism , Streptomyces/metabolism , Structure-Activity Relationship , Triterpenes/chemistry , Triterpenes/metabolismABSTRACT
Although photodynamic therapy (PDT) has been clinically applied tumor hypoxia still greatly restricts the performance of this oxygen-dependent oncological treatment. The delivery of oxygen donors to tumor may produce excessive reactive oxygen species (ROS) and damage the peripheral tissues. Herein, we developed a strategy to solve the hypoxia issue by enhancing the lethality of ROS. Before PDT, the ROS-defensing system of the cancer cells was obstructed by an inhibitor to MTH1, which is a key for the remediation of ROS-caused DNA damage. As a result, both nuclei and mitochondrial DNA damages were increased, remarkably promoting cellular apoptosis. The therapeutic results demonstrated that the performance of PDT can be improved by the MTH1 inhibitor, leading to efficient cancer cell killing effect in the hypoxic tumor. This strategy makes better use of the limited oxygen, holding the promise to achieve satisfactory therapeutic effect by PDT without generating redundant cytotoxic ROS.
Subject(s)
Antineoplastic Agents/pharmacology , DNA Repair Enzymes/genetics , DNA, Neoplasm/genetics , Enzyme Inhibitors/pharmacology , Melanoma, Experimental/drug therapy , Phosphoric Monoester Hydrolases/genetics , Pyrimidines/pharmacology , Animals , Antineoplastic Agents/chemistry , Apoptosis/drug effects , Chlorophyllides , DNA Repair/drug effects , DNA Repair/genetics , DNA Repair Enzymes/antagonists & inhibitors , DNA Repair Enzymes/metabolism , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/metabolism , Drug Carriers/administration & dosage , Drug Carriers/chemical synthesis , Drug Compounding/methods , Enzyme Inhibitors/chemistry , Female , Gene Expression , HCT116 Cells , HeLa Cells , Humans , Light , MCF-7 Cells , Melanoma, Experimental/enzymology , Melanoma, Experimental/pathology , Mice, Nude , Nanoparticles/administration & dosage , Nanoparticles/chemistry , Phosphoric Monoester Hydrolases/antagonists & inhibitors , Phosphoric Monoester Hydrolases/metabolism , Photochemotherapy/methods , Photosensitizing Agents/chemistry , Photosensitizing Agents/pharmacokinetics , Porphyrins/chemistry , Porphyrins/pharmacokinetics , Pyrimidines/chemistry , Reactive Oxygen Species/metabolism , Tumor Burden/drug effects , Xenograft Model Antitumor AssaysABSTRACT
Ribonucleotide reductase (RNR) is an essential enzyme in DNA biogenesis and a target of several chemotherapeutics. Here, we investigate how anti-leukemic drugs (e.g., clofarabine [ClF]) that target one of the two subunits of RNR, RNR-α, affect non-canonical RNR-α functions. We discovered that these clinically approved RNR-inhibiting dATP-analogs inhibit growth by also targeting ZRANB3-a recently identified DNA synthesis promoter and nuclear-localized interactor of RNR-α. Remarkably, in early time points following drug treatment, ZRANB3 targeting accounted for most of the drug-induced DNA synthesis suppression and multiple cell types featuring ZRANB3 knockout/knockdown were resistant to these drugs. In addition, ZRANB3 plays a major role in regulating tumor invasion and H-rasG12V-promoted transformation in a manner dependent on the recently discovered interactome of RNR-α involving select cytosolic-/nuclear-localized protein players. The H-rasG12V-promoted transformation-which we show requires ZRANB3-supported DNA synthesis-was efficiently suppressed by ClF. Such overlooked mechanisms of action of approved drugs and a previously unappreciated example of non-oncogene addiction, which is suppressed by RNR-α, may advance cancer interventions.
Subject(s)
Antimetabolites, Antineoplastic/pharmacology , Cell Nucleus/drug effects , Clofarabine/pharmacology , DNA Helicases/antagonists & inhibitors , Ribonucleotide Reductases/antagonists & inhibitors , Signal Transduction/drug effects , Cell Line, Tumor , Cell Nucleus/metabolism , DNA Helicases/deficiency , DNA Helicases/metabolism , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/biosynthesis , HeLa Cells , Humans , Ribonucleotide Reductases/metabolismABSTRACT
We studied the effect of lipoxygenase inhibitors nordihydroguaiaretic acid (NDGA) and fungus Lecanicillum lecanii extract on lymphatic leukemia P388 cells. The cells grown in the abdominal cavity of DBA2 mice for 7 days were transferred into a nutrient medium. The effect of lipoxygenase inhibitors was evaluated by changes in cell number, trypan blue staining, nucleus damage, and changes in cell distribution by DNA content after 22-h incubation. NDGA and fungus extract induced apoptotic death of lymphatic leukemia cells, which was seen from nucleus damage and reduced DNA content in cells. IC50 for NDGA and fungus extract was 0.66 and 5.5 µg/ml, respectively.
Subject(s)
Antineoplastic Agents/pharmacology , Complex Mixtures/pharmacology , Cordyceps/chemistry , DNA, Neoplasm/antagonists & inhibitors , Lipoxygenase Inhibitors/pharmacology , Lymphocytes/drug effects , Masoprocol/pharmacology , Animals , Apoptosis/drug effects , Cell Nucleus/drug effects , Cell Nucleus/metabolism , Cell Nucleus/pathology , Cell Nucleus/ultrastructure , DNA, Neoplasm/biosynthesis , Humans , Inhibitory Concentration 50 , Lymphocytes/metabolism , Lymphocytes/pathology , Mice , Mice, Inbred DBA , Tumor Cells, CulturedABSTRACT
Four series of new hybrid molecules with 7-chloroquinoline and arylamidine moieties joined through the rigid -O- (groups I (2a-g) and II (5a-g)) or flexible -NH-CH2-CH2-O- (groups III (8a-g) and IV (10a-g)) linker were synthesized, and their DNA/RNA binding properties and cytotoxic activity were tested, against several human cancer lines. The compounds and their interaction with DNA and RNA were studied by UV-Vis and CD spectroscopy. The obtained results showed that the binding affinity of the investigated compounds increases proportionally with the increase of the length and number of groups able to form hydrogen bonds with ds-polynucleotides. Improvement of binding was additionally achieved by reduction of the structural rigidity of the investigated compounds, new hybrid compounds preferentially bind to ctDNA. For most of them the DNA/RNA grooves are dominant binding sites, except for the compounds from group II for which intercalation in polyA-polyU was the dominant binding mode. The antiproliferative effects were tested by the MTT test on normal (MDCK1), carcinoma (HeLa and CaCo2) and leukemia cell lines (Raji and K462). The GI50 values for all investigated compounds ranged from 5 to more than 100 × 10-6 mol dm-3. Carcinoma cells were more resistant to the investigated compounds than leukemia cells. The most effective compounds against leukemia cell lines were from group IV (10a-g), with GI50 values ranging from of 5 and 35 × 10-6 mol dm-3. The cell cycle arrest was investigated by flow cytometry and the obtained results indicate that the selected compounds, 2d, 2e, 8a, 10d, 10e, and 10f, induce changes in the cell cycle of treated cells, but the cycle phase distribution varies between them. A significant decrease in the number of cells in S phase (p < 0.001) was observed in all treated cells, but only 10d and 10f induce cell cycle arrest at G0/G1 phase, dominantly.
Subject(s)
Amidines/pharmacology , Antineoplastic Agents/pharmacology , DNA, Neoplasm/antagonists & inhibitors , Quinolines/pharmacology , RNA, Neoplasm/antagonists & inhibitors , Amidines/chemistry , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Binding Sites/drug effects , Cell Cycle Checkpoints/drug effects , Cell Line , Cell Proliferation/drug effects , DNA, Neoplasm/chemistry , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Humans , Molecular Structure , Quinolines/chemistry , RNA, Neoplasm/chemistry , Structure-Activity RelationshipABSTRACT
DNA-alkylating drugs continue to remain an important weapon in the arsenal against cancers. However, they typically suffer from several shortcomings because of the indiscriminate DNA damage that they cause and their inability to specifically target cancer cells. We have developed a strategy for overcoming the deficiencies in current DNA-alkylating chemotherapy drugs by designing a site-specific DNA-methylating agent that can target cancer cells because of its selective uptake via glucose transporters, which are overexpressed in most cancers. The design features of the molecule, its synthesis, its reactivity with DNA, and its toxicity in human glioblastoma cells are reported here. In this molecule, a glucosamine unit, which can facilitate uptake via glucose transporters, is conjugated to one end of a bispyrrole triamide unit, which is known to bind to the minor groove of DNA at A/T-rich regions. A methyl sulfonate moiety is tethered to the other end of the bispyrrole unit to serve as a DNA-methylating agent. This molecule produces exclusively N3-methyladenine adducts upon reaction with DNA and is an order of magnitude more toxic to treatment resistant human glioblastoma cells than streptozotocin is, a Food and Drug Administration-approved, glycoconjugated DNA-methylating drug. Cellular uptake studies using a fluorescent analogue of our molecule provide evidence of uptake via glucose transporters and localization within the nucleus of cells. These results demonstrate the feasibility of our strategy for developing more potent anticancer chemotherapeutics, while minimizing common side effects resulting from off-target damage.
Subject(s)
Antineoplastic Agents, Alkylating/chemical synthesis , DNA Adducts/biosynthesis , DNA, Neoplasm/antagonists & inhibitors , Glucose Transport Proteins, Facilitative/metabolism , Glycoconjugates/chemical synthesis , Neuroglia/drug effects , Adenine/analogs & derivatives , Adenine/chemistry , Adenine/metabolism , Alkanesulfonates/chemistry , Antineoplastic Agents, Alkylating/metabolism , Antineoplastic Agents, Alkylating/pharmacology , Biological Transport , Cell Line, Tumor , Cell Survival/drug effects , DNA Adducts/chemistry , DNA Damage , DNA Methylation , DNA, Neoplasm/chemistry , DNA, Neoplasm/metabolism , Gene Expression , Glucosamine/chemistry , Glucose Transport Proteins, Facilitative/genetics , Glycoconjugates/metabolism , Glycoconjugates/pharmacology , Humans , Molecular Dynamics Simulation , Molecular Targeted Therapy , Neuroglia/metabolism , Neuroglia/pathology , Nucleic Acid Conformation , Pyrroles/chemistry , Streptozocin/pharmacologyABSTRACT
DNA polymerase theta (pol θ) is encoded in the genomes of many eukaryotes, though not in fungi. Pol θ is encoded by the POLQ gene in mammalian cells. The C-terminal third of the protein is a family A DNA polymerase with additional insertion elements relative to prokaryotic homologs. The N-terminal third is a helicase-like domain with DNA-dependent ATPase activity. Pol θ is important in the repair of genomic double-strand breaks (DSBs) from many sources. These include breaks formed by ionizing radiation and topoisomerase inhibitors, breaks arising at stalled DNA replication forks, breaks introduced during diversification steps of the mammalian immune system, and DSB induced by CRISPR-Cas9. Pol θ participates in a route of DSB repair termed "alternative end-joining" (altEJ). AltEJ is independent of the DNA binding Ku protein complex and requires DNA end resection. Pol θ is able to mediate joining of two resected 3' ends harboring DNA sequence microhomology. "Signatures" of Pol θ action during altEJ are the frequent utilization of longer microhomologies, and the insertion of additional sequences at joining sites. The mechanism of end-joining employs the ability of Pol θ to tightly grasp a 3' terminus through unique contacts in the active site, allowing extension from minimally paired primers. Pol θ is involved in controlling the frequency of chromosome translocations and preserves genome integrity by limiting large deletions. It may also play a backup role in DNA base excision repair. POLQ is a member of a cluster of similarly upregulated genes that are strongly correlated with poor clinical outcome for breast cancer, ovarian cancer and other cancer types. Inhibition of pol θ is a compelling approach for combination therapy of radiosensitization.
Subject(s)
DNA End-Joining Repair , DNA, Neoplasm/genetics , DNA-Directed DNA Polymerase/genetics , DNA/genetics , Neoplasms/genetics , Recombinational DNA Repair , Animals , Antineoplastic Combined Chemotherapy Protocols , Base Pair Mismatch , Catalytic Domain , DNA/metabolism , DNA Breaks, Double-Stranded , DNA Mismatch Repair , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/metabolism , DNA-Directed DNA Polymerase/chemistry , DNA-Directed DNA Polymerase/metabolism , Enzyme Inhibitors , Gamma Rays , Humans , Models, Molecular , Neoplasms/enzymology , Neoplasms/pathology , Neoplasms/therapy , Treatment Outcome , DNA Polymerase thetaABSTRACT
DNA interstrand crosslinks (ICLs) covalently join the two strands of a DNA duplex and block essential processes such as DNA replication and transcription. Several important anti-tumor drugs such as cisplatin and nitrogen mustards exert their cytotoxicity by forming ICLs. However, multiple complex pathways repair ICLs and these are thought to contribute to the development of resistance towards ICL-inducing agents. While the understanding of many aspects of ICL repair is still rudimentary, studies in recent years have provided significant insights into the pathways of ICL repair. In this perspective we review the recent advances made in elucidating the mechanisms of ICL repair with a focus on the role of TLS polymerases. We describe the emerging models for how these enzymes contribute to and are regulated in ICL repair, discuss the key open questions and examine the implications for this pathway in anti-cancer therapy.
Subject(s)
DNA Repair , DNA, Neoplasm/genetics , DNA-Directed DNA Polymerase/genetics , DNA/genetics , Neoplasms/genetics , Animals , Antineoplastic Agents/therapeutic use , Cisplatin/therapeutic use , Cross-Linking Reagents/therapeutic use , DNA/antagonists & inhibitors , DNA/metabolism , DNA Replication/drug effects , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/metabolism , DNA-Directed DNA Polymerase/classification , DNA-Directed DNA Polymerase/metabolism , Drug Resistance, Neoplasm/genetics , Humans , Mechlorethamine/therapeutic use , Neoplasms/enzymology , Neoplasms/pathology , Neoplasms/therapy , Transcription, Genetic/drug effects , XenopusABSTRACT
DNA topoisomerase 2 (Top2) poisons, including common anticancer drugs etoposide and doxorubicin kill cancer cells by stabilizing covalent Top2-tyrosyl-DNA 5'-phosphodiester adducts and DNA double-strand breaks (DSBs). Proteolytic degradation of the covalently attached Top2 leaves a 5'-tyrosylated blocked termini which is removed by tyrosyl DNA phosphodiesterase 2 (TDP2), prior to DSB repair through non-homologous end joining (NHEJ). Thus, TDP2 confers resistance of tumor cells to Top2-poisons by repairing such covalent DNA-protein adducts, and its pharmacological inhibition could enhance the efficacy of Top2-poisons. We discovered NSC111041, a selective inhibitor of TDP2, by optimizing a high throughput screening (HTS) assay for TDP2's 5'-tyrosyl phosphodiesterase activity and subsequent validation studies. We found that NSC111041 inhibits TDP2's binding to DNA without getting intercalated into DNA and enhanced etoposide's cytotoxicity synergistically in TDP2-expressing cells but not in TDP2 depleted cells. Furthermore, NSC111041 enhanced formation of etoposide-induced γ-H2AX foci presumably by affecting DSB repair. Immuno-histochemical analysis showed higher TDP2 expression in a sub-set of different type of tumor tissues. These findings underscore the feasibility of clinical use of suitable TDP2 inhibitors in adjuvant therapy with Top2-poisons for a sub-set of cancer patients with high TDP2 expression.
Subject(s)
Aminoquinolines/pharmacology , DNA Repair/drug effects , DNA-Binding Proteins/antagonists & inhibitors , Etoposide/pharmacology , Nuclear Proteins/antagonists & inhibitors , Phosphodiesterase Inhibitors/pharmacology , Topoisomerase Inhibitors/pharmacology , Transcription Factors/antagonists & inhibitors , Antigens, Neoplasm/genetics , Antigens, Neoplasm/metabolism , Cell Line, Tumor , DNA Adducts/antagonists & inhibitors , DNA Adducts/metabolism , DNA Breaks, Double-Stranded , DNA Topoisomerases, Type II/genetics , DNA Topoisomerases, Type II/metabolism , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/genetics , DNA, Neoplasm/metabolism , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Drug Synergism , Epithelial Cells/drug effects , Epithelial Cells/enzymology , Epithelial Cells/pathology , Gene Expression Regulation, Neoplastic , High-Throughput Screening Assays , Histones/genetics , Histones/metabolism , Humans , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Phosphoric Diester Hydrolases , Poly-ADP-Ribose Binding Proteins , Protein Binding , Transcription Factors/genetics , Transcription Factors/metabolismABSTRACT
Many cancer chemotherapy agents act by targeting the DNA of cancer cells, causing substantial damage within their genome and causing them to undergo apoptosis. An effective DNA repair pathway in cancer cells can act in a reverse way by removing these drug-induced DNA lesions, allowing cancer cells to survive, grow and proliferate. In this context, DNA repair inhibitors opened a new avenue in cancer treatment, by blocking the DNA repair mechanisms from removing the chemotherapy-mediated DNA damage. In particular, the nucleotide excision repair (NER) involves more than thirty protein-protein interactions and removes DNA adducts caused by platinum-based chemotherapy. The excision repair cross-complementation group 1 (ERCC1)-xeroderma pigmentosum, complementation group A (XPA) protein (XPA-ERCC1) complex seems to be one of the most promising targets in this pathway. ERCC1 is over expressed in cancer cells and the only known cellular function so far for XPA is to recruit ERCC1 to the damaged point. Here, we build upon our recent advances in identifying inhibitors for this interaction and continue our efforts to rationally design more effective and potent regulators for the NER pathway. We employed in silico drug design techniques to: (1) identify compounds similar to the recently discovered inhibitors, but more effective at inhibiting the XPA-ERCC1 interactions, and (2) identify different scaffolds to develop novel lead compounds. Two known inhibitor structures have been used as starting points for two ligand/structure-hybrid virtual screening approaches. The findings described here form a milestone in discovering novel inhibitors for the NER pathway aiming at improving the efficacy of current platinum-based therapy, by modulating the XPA-ERCC1 interaction.
Subject(s)
Antineoplastic Agents/chemistry , DNA Repair/drug effects , DNA, Neoplasm/antagonists & inhibitors , DNA-Binding Proteins/antagonists & inhibitors , Drug Design , Endonucleases/antagonists & inhibitors , Small Molecule Libraries/chemistry , Xeroderma Pigmentosum Group A Protein/antagonists & inhibitors , Antineoplastic Agents/pharmacology , Antineoplastic Combined Chemotherapy Protocols , Binding Sites , Cisplatin/chemistry , Cisplatin/pharmacology , DNA Damage , DNA, Neoplasm/chemistry , DNA, Neoplasm/genetics , DNA-Binding Proteins/chemistry , DNA-Binding Proteins/genetics , Drug Discovery , Endonucleases/chemistry , Endonucleases/genetics , Gene Expression , Humans , Hydrogen Bonding , Molecular Docking Simulation , Molecular Dynamics Simulation , Protein Binding , Protein Interaction Domains and Motifs , Protein Structure, Secondary , Small Molecule Libraries/pharmacology , Static Electricity , Thermodynamics , Xeroderma Pigmentosum Group A Protein/chemistry , Xeroderma Pigmentosum Group A Protein/geneticsABSTRACT
Ovarian epithelial carcinoma is the leading cause of deaths from gynecologic malignancy. New reagents with therapeutic potentials against ovarian cancer, especially the drug-resistant cases, are required for better treatment of ovarian cancer patients. Epigenetic events such as changes in DNA methylation and histone modification, through their effects on DNA-protein interaction, chromatin conformation, and gene expression, affect cell function, cancer behavior, clinical manifestations, and outcomes. Previous studies have shown that histone deacetylase (HDAC) inhibitors have strong cytostatic and apoptotic activities in hematologic and some solid cancer cells. Oxamflatin, a compound containing the aromatic sulfonamide and hydroxamic acid groups, is known to be a potent HDAC inhibitor capable of inhibiting the growth of mouse and human cancer cell lines. In this study we found that oxamflatin in the nM range induced morphological changes in OVCAR-5 and SKOV-3 ovarian cancer cell lines. Treatment with oxamflatin also led to decreased cell viability. Moreover, results of BrdU incorporation assay, cell counting, and Ki-67 immunostaining indicated that oxamflatin was able to significantly inhibit DNA synthesis and cell proliferation. Using real-time PCR and Western blot analyses we demonstrated that oxamflatin was capable of downregulating the expression of c-Myc, CDK4, E2F1, and the phosphorylation levels of Rb protein, but upregulating p21. These findings pave the way to examine if oxamflatin along with or in combination with other reagents could deliver anticancer effects against ovarian cancers in vivo.
Subject(s)
Antineoplastic Agents/pharmacology , Epithelial Cells/drug effects , Gene Expression Regulation, Neoplastic , Histone Deacetylase Inhibitors/pharmacology , Histone Deacetylases/genetics , Hydroxamic Acids/pharmacology , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , Cyclin-Dependent Kinase 4/antagonists & inhibitors , Cyclin-Dependent Kinase 4/genetics , Cyclin-Dependent Kinase 4/metabolism , Cyclin-Dependent Kinase Inhibitor p21/agonists , Cyclin-Dependent Kinase Inhibitor p21/genetics , Cyclin-Dependent Kinase Inhibitor p21/metabolism , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/biosynthesis , DNA, Neoplasm/genetics , E2F1 Transcription Factor/antagonists & inhibitors , E2F1 Transcription Factor/genetics , E2F1 Transcription Factor/metabolism , Epigenesis, Genetic , Epithelial Cells/metabolism , Epithelial Cells/pathology , Female , Histone Deacetylases/metabolism , Humans , Ki-67 Antigen/genetics , Ki-67 Antigen/metabolism , Ovary/drug effects , Ovary/metabolism , Ovary/pathology , Proto-Oncogene Proteins c-myc/antagonists & inhibitors , Proto-Oncogene Proteins c-myc/genetics , Proto-Oncogene Proteins c-myc/metabolism , Retinoblastoma Protein/antagonists & inhibitors , Retinoblastoma Protein/genetics , Retinoblastoma Protein/metabolism , Signal TransductionABSTRACT
Tumor cell growth requires large iron quantities and the deprivation of this metal induced by synthetic metal chelators is therefore an attractive method for limiting the cancer cell proliferation. The antiproliferative effect of the Quilamine HQ1-44, a new iron chelator vectorized toward tumor cells by a polyamine chain, is related to its high selectivity for the Polyamine Transport System (PTS), allowing its preferential uptake by tumoral cells. The difference in PTS activation between healthy cells and tumor cells enables tumor cells to be targeted, whereas the strong dependence of these cells on iron ensures a secondary targeting. Here, we demonstrated in vitro that HQ1-44 inhibits DNA synthesis and cell proliferation of HCT116 cells by modulating the intracellular metabolism of both iron and polyamines. Moreover, in vivo, in xenografted athymic nude mice, we found that HQ1-44 was as effective as cis-platin in reducing HCT116 tumor growth, without its side effects. Furthermore, as suggested by in vitro data, the depletion in exogenous or endogenous polyamines, known to activate the PTS, dramatically enhanced the antitumor efficiency of HQ1-44. These data support the need for further studies to assess the value of HQ1-44 as an adjuvant treatment in cancer.
Subject(s)
Antineoplastic Agents/pharmacology , Colonic Neoplasms/drug therapy , DNA, Neoplasm/antagonists & inhibitors , Eflornithine/pharmacology , Iron Chelating Agents/pharmacology , Polyamines/antagonists & inhibitors , Animals , Biological Transport/drug effects , Cell Cycle/drug effects , Cell Survival/drug effects , Colonic Neoplasms/metabolism , Colonic Neoplasms/pathology , DNA, Neoplasm/biosynthesis , Female , HCT116 Cells , Humans , Mice , Mice, Nude , Molecular Targeted Therapy , Neoplasm Transplantation , Polyamines/metabolism , Transplantation, Heterologous , Tumor Burden/drug effectsABSTRACT
Protein families involved in chromatin-templated events are emerging as novel target classes in oncology and other disease areas. The ability to discover selective inhibitors against chromatin factors depends on the presence of structural features that are unique to the targeted sites. To evaluate challenges and opportunities toward the development of selective inhibitors, we calculated all pair wise structural distances between 575 structures from the protein databank representing 163 unique binding pockets found in protein domains that write, read or erase post-translational modifications on histones, DNA, and RNA. We find that the structural similarity of binding sites does not always follow the sequence similarity of protein domains. Our analysis reveals increased risks of activity across target-class for compounds competing with the cofactor of protein arginine methyltransferases, lysine acetyltransferases, and sirtuins, while exploiting the conformational plasticity of a protein target is a path toward selective inhibition. The structural diversity landscape of the epigenetics pocketome can be explored via an open-access graphic user interface at thesgc.org/epigenetics_pocketome.
Subject(s)
Antineoplastic Agents/chemistry , Epigenesis, Genetic , Histones/antagonists & inhibitors , Neoplasm Proteins/antagonists & inhibitors , Protein Processing, Post-Translational , Small Molecule Libraries/chemistry , Software , Acetyltransferases/antagonists & inhibitors , Acetyltransferases/chemistry , Acetyltransferases/genetics , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Binding Sites , Chromatin/chemistry , Chromatin/drug effects , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/chemistry , DNA, Neoplasm/genetics , Databases, Protein , Histones/chemistry , Histones/genetics , Humans , Internet , Ligands , Models, Molecular , Neoplasm Proteins/chemistry , Neoplasm Proteins/genetics , Neoplasms/chemistry , Neoplasms/drug therapy , Neoplasms/genetics , Neoplasms/pathology , Protein Binding , Protein Structure, Tertiary , Protein-Arginine N-Methyltransferases/antagonists & inhibitors , Protein-Arginine N-Methyltransferases/chemistry , Protein-Arginine N-Methyltransferases/genetics , RNA, Neoplasm/antagonists & inhibitors , RNA, Neoplasm/chemistry , RNA, Neoplasm/genetics , Sirtuins/antagonists & inhibitors , Sirtuins/chemistry , Sirtuins/genetics , Small Molecule Libraries/chemical synthesis , Small Molecule Libraries/pharmacologyABSTRACT
Cyclopropabenzaindoles (CBIs) are exquisitely potent cytotoxins which bind and alkylate in the minor groove of DNA. They are not selective for cancer cells, so prodrugs are required. CBIs can be formed at physiological pH by Winstein cyclisation of 1-chloromethyl-3-substituted-5-hydroxy-2,3-dihydrobenzo[e]indoles (5-OH-seco-CBIs). Corresponding 5-NH2-seco-CBIs should also undergo Winstein cyclisation similarly. A key triply orthogonally protected intermediate on the route to 5-NH2-seco-CBIs has been synthesised, via selective monotrifluoroacetylation of naphthalene-1,3-diamine, Boc protection, electrophilic iodination, selective allylation at the trifluoroacetamide and 5-exo radical ring-closure with TEMPO. This intermediate has potential for introduction of peptide prodrug masking units (deactivating the Winstein cyclisation and cytotoxicity), addition of diverse indole-amide side-chains (enhancing non-covalent binding prior to alkylation) and use of different leaving groups (replacing the usual chlorine, allowing tuning of the rate of Winstein cyclisation). This key intermediate was elaborated into a simple model 5-NH2-seco-CBI with a dimethylaminoethoxyindole side-chain. Conversion to a bio-reactive entity and the bioactivity of this system were confirmed through DNA-melting studies (ΔTm=13°C) and cytotoxicity against LNCaP human prostate cancer cells (IC50=18nM).
Subject(s)
Antineoplastic Agents, Alkylating/chemical synthesis , Cyclopropanes/chemical synthesis , DNA, Neoplasm/antagonists & inhibitors , Indoles/chemical synthesis , Prodrugs , Acetamides , Alkylation , Antineoplastic Agents, Alkylating/pharmacology , Cell Line, Tumor , Cell Survival/drug effects , Cyclic N-Oxides/chemistry , Cyclization , Cyclopropanes/pharmacology , DNA, Neoplasm/chemistry , Drug Design , Drug Screening Assays, Antitumor , Fluoroacetates , Humans , Hydrogen-Ion Concentration , Indoles/pharmacology , Models, Molecular , Naphthalenes/chemistry , Nucleic Acid Denaturation , Structure-Activity RelationshipABSTRACT
The synthesis, biological and molecular modeling evaluation of a series of macrocyclic naphthalene diimides is reported. The present investigation expands on the study of structure-activity relationships of prototype compound 2 by constraining the molecule into a macrocyclic structure with the aim of improving its G-quadruplex binding activity and selectivity. The new derivatives, compounds 4-7 carry spermidine- and spermine-like linkers while in compound 8 the inner basic nitrogen atoms of spermine have been replaced with oxygen atoms. The design strategy has led to potent compounds stabilizing both human telomeric (F21T) and c-KIT2 quadruplex sequences, and high selectivity for quadruplex in comparison to duplex DNA. Antiproliferative effects of the new derivatives 4-8 have been evaluated in a panel of cancer cell lines and all the tested compounds showed activity in the low micromolar or sub-micromolar range of concentrations. In order to rationalize the molecular basis of the DNA G-quadruplex versus duplex recognition preference, docking and molecular dynamics studies have been performed. The computational results support the observation that the main driving force in the recognition is due to electrostatic factors.
Subject(s)
Antineoplastic Agents/chemical synthesis , DNA, Neoplasm/antagonists & inhibitors , G-Quadruplexes , Imides/chemical synthesis , Naphthalenes/chemical synthesis , Antineoplastic Agents/pharmacology , Binding Sites , Cell Line, Tumor , Cell Survival/drug effects , DNA, Neoplasm/chemistry , Drug Design , Humans , Imides/pharmacology , Molecular Docking Simulation , Molecular Dynamics Simulation , Naphthalenes/pharmacology , Spermidine/chemistry , Spermine/chemistry , Static Electricity , Structure-Activity RelationshipABSTRACT
Introduction: Contrast induced nephropathy (CIN) is one of the complications of the use of intravascular contrast agents, being defined as a reduction of the glomerular filtration rate caused by the iodinated contrast. Most CIN data derive from the cardiovascular literature, which identified as the most consistent risk factors pre-existing chronic renal insufficiency and diabetes mellitus. However, these studies limit their conclusions to a more specific patient population. Computerized tomography as a cause of CIN has been studied less often. Objective: To report on the incidence of computerized tomography contrast induced nephropathy (CIN) in an inpatient population of a tertiary general hospital, identifying potentially avoidable risk factors. Methods: We performed a prospective cohort study with inpatients admitted at a tertiary hospital requiring contrast-induced CT. The primary outcome was the development of CIN, measure by the alteration of serum creatinine or glomerular filtration rate in 48 or 72 hours. Through clinical interview, we verified possible risk factors and preventive measures instituted by the medical team and their association with development of CIN. Results: Of a total of 410 patients, 35 (8.5%) developed CIN. There was a positive correlation between CIN and the presence of diabetes mellitus (OR = 2.15; 95%CI 1.35-4.06; p = 0.02), heart failure (OR = 2.23; 95%CI 1.18-8.8; p = 0.022), and renal failure (OR = 3.36; 95%CI 1.57- 7.17; p = 0.002) Conclusion: Incidence of CIN varies according to the population. Diabetes mellitus, heart failure and renal failure were independent risk factors for the development of CT-associated CIN. Further studies are needed to better understand and treat CT-associated CIN. .
Introdução: Nefropatia induzida por contraste (NIC) é consequência do uso de meios de contraste intravenoso, sendo definida como uma redução da taxa de filtração glomerular. A maioria dos dados de NIC são da literatura cardiovascular, que identificou como fatores de risco insuficiência renal crônica e diabetes. Entretanto, esses estudos limitam suas conclusões a uma população especifica de pacientes. Tomografia Computadorizada contrastada como causa de NIC foi menos estudada. Objetivo: Reportar incidência de NIC numa população de pacientes internados em hospital terciário submetidos à tomografia computadorizada com contraste, identificando possíveis fatores de risco evitáveis. Métodos: Realizamos um estudo de coorte prospectivo com pacientes internados em hospital terciário e que necessitaram de tomografia computadorizada com contraste. O desfecho primário foi desenvolvimento de NIC, verificado por meio da variação da creatinina sérica ou taxa de filtração glomerular em 48 ou 72 horas. Em entrevista clínica, verificamos possíveis fatores de risco, assim como medidas preventivas instituídas pela equipe médica e suas possíveis associações com desenvolvimento de NIC. Resultados: Do total de 410 pacientes, 35 (8,5%) desenvolveram NIC. Houve correlação positiva entre desenvolvimento de NIC e a presença de diabetes mellitus (OR = 2,15; 95%CI 1,35-4,06; p = 0,02), insuficiência cardíaca (OR = 2,23; 95%CI 1,18-8,8; p = 0,022), e insuficiência renal (OR = 3,36; 95%CI 1,57-7,17; p = 0,002). Conclusão: A incidência de NIC varia de acordo com a população. Diabetes, insuficiência cardíaca e insuficiência renal foram fatores de risco independentes para o desenvolvimento de NIC. Mais estudos são ...
Subject(s)
Humans , Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Carcinoma, Hepatocellular/drug therapy , Caspases/metabolism , Liver Neoplasms/drug therapy , Sulindac/analogs & derivatives , Sulindac/pharmacology , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Apoptosis/physiology , Carcinoma, Hepatocellular/enzymology , Carcinoma, Hepatocellular/pathology , Cell Division/drug effects , DNA, Neoplasm/antagonists & inhibitors , DNA, Neoplasm/biosynthesis , Dose-Response Relationship, Drug , Enzyme Activation/drug effects , Liver Neoplasms/enzymology , Liver Neoplasms/pathology , Sphingomyelin Phosphodiesterase/metabolism , Tumor Cells, CulturedABSTRACT
BACKGROUND: Glioblastoma multiforme (GBM), the most common and most aggressive type of primary adult brain tumour, responds poorly to conventional treatment. Temozolomide (TMZ) chemotherapy remains the most commonly used treatment, despite a large proportion of tumours displaying TMZ resistance. 60% of GBM tumours have unmethylated MGMT promoter regions, resulting in an overexpression of the DNA repair protein O6-methylguanine-DNA methyltransferase (MGMT), which is responsible for tumour resistance to TMZ chemotherapy. Tumours also often exhibit hyperactive PI3-kinase/mTOR signalling, which enables them to resynthesise proteins quickly. Since MGMT is a suicide protein that is degraded upon binding to and repairing TMZ-induced O6-methylguanine adducts, it has been hypothesized that inhibition of translation via the mTOR signalling pathway could generate a tumour-specific reduction in MGMT protein and increase TMZ sensitivity. METHODS: MGMT was monitored at the post-transcriptional, translational and protein levels, to determine what effect mTOR inhibition was having on MGMT protein expression in vitro. RESULTS: We show that inhibiting mTOR signalling is indeed associated with acute inhibition of protein synthesis. Western blots show that despite this, relative to loading control proteins, steady state levels of MGMT protein increased and MGMT mRNA was retained in heavy polysomes. Whilst TMZ treatment resulted in maintained MGMT protein levels, concomitant treatment of T98G cells with TMZ and KU0063794 resulted in increased MGMT protein levels without changes in total mRNA levels. CONCLUSIONS: These in vitro data suggest that, counterintuitively, mTOR inhibition may not be a useful adjunct to TMZ therapy and that more investigation is needed before applying mTOR inhibitors in a clinical setting.