ABSTRACT
For decades prebiotic chemists have attempted to achieve replication of RNA under prebiotic conditions with only limited success. One of the long-recognized impediments to achieving true replication of a duplex (copying of both strands) is the so-called strand inhibition problem. Specifically, while the two strands of an RNA (or DNA) duplex can be separated by heating, upon cooling the strands of a duplex will reanneal before mononucleotide or oligonucleotide substrates can bind to the individual strands. Here we demonstrate that a class of plausible prebiotic solvents, when coupled with thermal cycling and varying levels of hydration, circumvents the strand inhibition problem, and allows multiple rounds of information transfer from both strands of a duplex (replication). Replication was achieved by simultaneous ligation of oligomers that bind to their templates with the aid of the solvents. The solvents used consisted of concentrated solutions of urea and acetamide in water (UAcW), components that were likely abundant on the early Earth. The UAcW solvent system favors the annealing of shorter strands over the re-annealing of long strands, thereby circumventing strand inhibition. We observed an improvement of DNA and RNA replication yields by a factor of 100× over aqueous buffer. Information transfer in the UAcW solvent system is robust, being achieved for a range of solvent component ratios, various drying conditions, and in the absence or presence of added salts.
Subject(s)
Acetamides/pharmacology , DNA/antagonists & inhibitors , RNA/antagonists & inhibitors , Urea/pharmacology , Acetamides/chemistry , DNA/metabolism , Nucleic Acid Conformation , RNA/metabolism , Solutions , Urea/chemistryABSTRACT
Androgen receptor (AR), a ligand-activated transcription factor, is a master regulator in the development and progress of prostate cancer (PCa). A major challenge for the clinically used AR antagonists is the rapid emergence of resistance induced by the mutations at AR ligand binding domain (LBD), and therefore the discovery of novel anti-AR therapeutics that can combat mutation-induced resistance is quite demanding. Therein, blocking the interaction between AR and DNA represents an innovative strategy. However, the hits confirmed targeting on it so far are all structurally based on a sole chemical scaffold. In this study, an integrated docking-based virtual screening (VS) strategy based on the crystal structure of the DNA binding domain (DBD) of AR was conducted to search for novel AR antagonists with new scaffolds and 2-(2-butyl-1,3-dioxoisoindoline-5-carboxamido)-4,5-dimethoxybenzoicacid (Cpd39) was identified as a potential hit, which was competent to block the binding of AR DBD to DNA and showed decent potency against AR transcriptional activity. Furthermore, Cpd39 was safe and capable of effectively inhibiting the proliferation of PCa cell lines (i.e., LNCaP, PC3, DU145, and 22RV1) and reducing the expression of the genes regulated by not only the full-length AR but also the splice variant AR-V7. The novel AR DBD-ARE blocker Cpd39 could serve as a starting point for the development of new therapeutics for castration-resistant PCa.
Subject(s)
Androgen Receptor Antagonists/pharmacology , DNA/antagonists & inhibitors , Drug Discovery , Molecular Docking Simulation , Receptors, Androgen/metabolism , Androgen Receptor Antagonists/chemistry , Binding Sites/drug effects , DNA/chemistry , Dose-Response Relationship, Drug , Drug Evaluation, Preclinical , Humans , Molecular Structure , Receptors, Androgen/chemistry , Structure-Activity RelationshipABSTRACT
Synchronous cell populations are commonly used for the analysis of various aspects of cellular metabolism at specific stages of the cell cycle. Cell synchronization at a chosen cell cycle stage is most frequently achieved by inhibition of specific metabolic pathway(s). In this respect, various protocols have been developed to synchronize cells in particular cell cycle stages. In this review, we provide an overview of the protocols for cell synchronization of mammalian cells based on the inhibition of synthesis of DNA building blocks-deoxynucleotides and/or inhibition of DNA synthesis. The mechanism of action, examples of their use, and advantages and disadvantages are described with the aim of providing a guide for the selection of suitable protocol for different studied situations.
Subject(s)
Cell Cycle , Cell Division , DNA Replication , DNA/biosynthesis , Animals , DNA/antagonists & inhibitors , HumansABSTRACT
Emerging evidence has revealed that oxidative damages of DNA correlate with the pathogenesis of some diseases, and numerous investigations have also suggested that supplementation of antioxidants is beneficial for keeping health by rectifying in vivo redox status. Here, we construct antioxidative dipeptides with the Ugi four-component reaction (comprising p-aminobenzyl alcohol, benzaldehyde, or vanillin, a series of antioxidative carboxylic acids and isocyanides as reagents) and then attempt to attach the dipeptides to [60]fullerene by the Bingel reaction. However, this endeavor does not lead to the amelioration of the radical-scavenging property because abilities of fullerenyl dipeptides to trap 2,2'-diphenyl-1-picrylhydrazyl and galvinoxyl radicals are still dependent upon the phenolic hydroxyl group in the dipeptide scaffold rather than upon the fullerenyl group. Alternatively, when the obtained fullerenyl dipeptides are evaluated in a peroxyl radical-induced oxidation of DNA, it is found that introducing a fullerene moiety into dipeptide enables antioxidative effect to be enhanced 20-30% because the fullerene moiety facilitates the corresponding dipeptide to intercalate with DNA strands, and thus, to increase the antioxidative efficacy. Our results suggest that connecting an antioxidative skeleton with the hydrophobic fullerene moiety might lead to a series of novel antioxidant hybrids applied for the inhibition of DNA oxidation.
Subject(s)
Antioxidants/pharmacology , DNA/antagonists & inhibitors , Dipeptides/pharmacology , Fullerenes/pharmacology , Antioxidants/chemistry , Biphenyl Compounds/antagonists & inhibitors , DNA/metabolism , Dipeptides/chemical synthesis , Dipeptides/chemistry , Fullerenes/chemistry , Molecular Structure , Oxidation-Reduction , Picrates/antagonists & inhibitorsABSTRACT
The NRF2 transcription factor is a key regulator in cellular oxidative stress response, and acts as a tumor suppressor. Aberrant activation of NRF2 has been implicated in promoting chemo-resistance, tumor growth, and metastasis by activating its downstream target genes. Hence, inhibition of NRF2 promises to be an attractive therapeutic strategy to suppress cell proliferation and enhance cell apoptosis in cancer. Direct targeting of NRF2 with small-molecules to discover protein-DNA interaction inhibitors is challenging as it is a largely intrinsically disordered protein. To discover molecules that bind to NRF2 at the DNA binding interface, we performed an NMR-based fragment screen against its DNA-binding domain. We discovered several weakly binding fragment hits that bind to a region overlapping with the DNA binding site. Using SAR by catalogue we developed an initial structure-activity relationship for the most interesting initial hit series. By combining NMR chemical shift perturbations and data-driven docking, binding poses which agreed with NMR information and the observed SAR were elucidated. The herein discovered NRF2 hits and proposed binding modes form the basis for future structure-based optimization campaigns on this important but to date 'undrugged' cancer driver.
Subject(s)
DNA/antagonists & inhibitors , NF-E2-Related Factor 2/antagonists & inhibitors , Protein Binding/drug effects , Small Molecule Libraries/chemistry , Binding Sites , DNA/metabolism , Humans , Molecular Docking Simulation , Molecular Structure , NF-E2-Related Factor 2/chemistry , NF-E2-Related Factor 2/metabolism , Nuclear Magnetic Resonance, Biomolecular , Protein Domains , Structure-Activity RelationshipABSTRACT
(1) Background: Chiral nanoparticular systems have recently emerged as a compelling platform for investigating stereospecific behavior at the nanoscopic level. We describe chiroselective supramolecular interactions that occur between DNA oligonucleotides and chiral polyurea nanocapsules. (2) Methods: We employ interfacial polyaddition reactions between toluene 2,4-diisocyanate and lysine enantiomers that occur in volatile oil-in-water nanoemulsions to synthesize hollow, solvent-free capsules with average sizes of approximately 300 nm and neutral surface potential. (3) Results: The resultant nanocapsules exhibit chiroptical activity and interact differentially with single stranded DNA oligonucleotides despite the lack of surface charge and, thus, the absence of significant electrostatic interactions. Preferential binding of DNA on D-polyurea nanocapsules compared to their L-counterparts is demonstrated by a fourfold increase in capsule size, a 50% higher rise in the absolute value of negative zeta potential (ζ-potential), and a three times lower free DNA concentration after equilibration with the excess of DNA. (4) Conclusions: We infer that the chirality of the novel polymeric nanocapsules affects their supramolecular interactions with DNA, possibly through modification of the surface morphology. These interactions can be exploited when developing carriers for gene therapy and theranostics. The resultant constructs are expected to be highly biocompatible due to their neutral potential and biodegradability of polyurea shells.
Subject(s)
DNA/chemistry , Drug Carriers/pharmacology , Nanocapsules/chemistry , Oligonucleotides/chemistry , Aptamers, Nucleotide/chemistry , DNA/antagonists & inhibitors , Drug Carriers/chemistry , Emulsions/chemistry , Emulsions/pharmacology , Humans , Oligonucleotides/genetics , Particle Size , Polymers/chemistryABSTRACT
There are few methods available for the rapid discovery of multitarget drugs. Herein, we describe the template-assisted, target-guided discovery of small molecules that recognize d(CTG) in the expanded d(CTG·CAG) sequence and its r(CUG) transcript that cause myotonic dystrophy type 1. A positive cross-selection was performed using a small library of 30 monomeric alkyne- and azide-containing ligands capable of producing >5000 possible di- and trimeric click products. The monomers were incubated with d(CTG)16 or r(CUG)16 under physiological conditions, and both sequences showed selectivity in the proximity-accelerated azide-alkyne [3+2] cycloaddition click reaction. The limited number of click products formed in both selections and the even smaller number of common products suggests that this method is a useful tool for the discovery of single-target and multitarget lead therapeutic agents.
Subject(s)
DNA/antagonists & inhibitors , Myotonic Dystrophy/drug therapy , Myotonic Dystrophy/genetics , RNA/antagonists & inhibitors , Small Molecule Libraries/pharmacology , Trinucleotide Repeat Expansion/drug effects , Cells, Cultured , DNA/genetics , DNA/metabolism , Humans , Myotonic Dystrophy/pathology , RNA/genetics , RNA/metabolism , Trinucleotide Repeat Expansion/geneticsABSTRACT
The aims of this study were to compare diagnostic value of anti-ribosomal P protein antibody (anti-P), anti-Smith antibody (anti-Sm), anti-double-stranded DNA antibody (anti-dsDNA), anti-nucleosome antibody (ANuA), and anti-histone antibody (AHA) for systemic lupus erythematosus (SLE) as well as explore the correlation between anti-P and SLE.A retrospective study was performed with 487 SLE patients, 235 non-SLE rheumatic diseases, and 124 healthy subjects from January 2015 to December 2018. Clinical manifestations, laboratory results and Systemic Lupus Erythematosus Disease Activity Index (SLEDAI)-2000 scores were analyzed between anti-P/+/ and anti-P/-/ patients. SPSS19.0 statistical software was used for data analysis.The sensitivities of anti-P, anti-Sm, anti-dsDNA, ANuA, and AHA in SLE were 31.6%, 20.7%, 45.0%, 27.9%, and 14.6%, and the specificities were 99.2%, 99.4%, 98.9%, 98.3%, and 96.7%, respectively. Only 27.9% of SLE had a single positive anti-P while the other 4 antibodies were all negative. There were significant differences in the age of onset, skin erythema, urinary protein, creatinine and serum IgG, IgM, C3, C4 between anti-P/+/ and anti-P/-/ patients (Pâ<â.05). When anti-Sjogren syndrome A antibody, anti-P were positive and anti-dsDNA was negative, the incidence of skin erythema was the highest (35.1%). Compared with anti-P/-/ patients, anti-P/+/ patients had higher SLEDAI scores (Pâ<â.001).Anti-P, anti-Sm, anti-dsDNA, ANuA, and AHA have high specificity but poor sensitivity in the diagnosis of SLE; combined detection can greatly improve the detection rate. Anti-P is more valuable in the diagnosis of SLE when other specific autoantibodies are negative. SLE patients with positive anti-P have an earlier onset age and are more prone to skin erythema, lupus nephritis as well as higher disease activity.
Subject(s)
Autoantibodies/blood , Lupus Erythematosus, Systemic/immunology , Membrane Transport Proteins/immunology , Ribosomal Proteins/immunology , Adult , Antibodies, Antinuclear/immunology , DNA/antagonists & inhibitors , DNA/metabolism , Erythema/immunology , Erythema/pathology , Female , Histones/antagonists & inhibitors , Histones/metabolism , Humans , Lupus Erythematosus, Systemic/diagnosis , Lupus Erythematosus, Systemic/pathology , Lupus Nephritis/immunology , Male , Middle Aged , Nucleosomes/metabolism , Retrospective Studies , Rheumatic Diseases/immunology , Sensitivity and Specificity , Skin Diseases/epidemiologyABSTRACT
Clustered regularly interspaced short palindromic repeats (CRISPR) together with their accompanying cas (CRISPR-associated) genes are found frequently in bacteria and archaea, serving to defend against invading foreign DNA, such as viral genomes. CRISPR-Cas systems provide a uniquely powerful defense because they can adapt to newly encountered genomes. The adaptive ability of these systems has been exploited, leading to their development as highly effective tools for genome editing. The widespread use of CRISPR-Cas systems has driven a need for methods to control their activity. This review focuses on anti-CRISPRs (Acrs), proteins produced by viruses and other mobile genetic elements that can potently inhibit CRISPR-Cas systems. Discovered in 2013, there are now 54 distinct families of these proteins described, and the functional mechanisms of more than a dozen have been characterized in molecular detail. The investigation of Acrs is leading to a variety of practical applications and is providing exciting new insight into the biology of CRISPR-Cas systems.
Subject(s)
CRISPR-Cas Systems/drug effects , Gene Editing/methods , Small Molecule Libraries/pharmacology , Viral Proteins/genetics , Viruses/genetics , Archaea/genetics , Archaea/immunology , Archaea/virology , Bacteria/genetics , Bacteria/immunology , Bacteria/virology , Bacterial Proteins/antagonists & inhibitors , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Biological Coevolution , CRISPR-Associated Proteins/antagonists & inhibitors , CRISPR-Associated Proteins/genetics , CRISPR-Associated Proteins/metabolism , DNA/antagonists & inhibitors , DNA/chemistry , DNA/genetics , DNA/metabolism , DNA Cleavage/drug effects , Endodeoxyribonucleases/antagonists & inhibitors , Endodeoxyribonucleases/genetics , Endodeoxyribonucleases/metabolism , Humans , Models, Molecular , Multigene Family , Protein Binding , Protein Multimerization/drug effects , RNA, Guide, Kinetoplastida/genetics , RNA, Guide, Kinetoplastida/metabolism , Small Molecule Libraries/chemistry , Small Molecule Libraries/metabolism , Viral Proteins/chemistry , Viral Proteins/metabolism , Viral Proteins/pharmacology , Viruses/metabolism , Viruses/pathogenicityABSTRACT
CRISPR-Cas adaptive immune systems protect bacteria and archaea against their invading genetic parasites, including bacteriophages/viruses and plasmids. In response to this immunity, many phages have anti-CRISPR (Acr) proteins that inhibit CRISPR-Cas targeting. To date, anti-CRISPR genes have primarily been discovered in phage or prophage genomes. Here, we uncovered acr loci on plasmids and other conjugative elements present in Firmicutes using the Listeria acrIIA1 gene as a marker. The four identified genes, found in Listeria, Enterococcus, Streptococcus and Staphylococcus genomes, can inhibit type II-A SpyCas9 or SauCas9, and are thus named acrIIA16-19. In Enterococcus faecalis, conjugation of a Cas9-targeted plasmid was enhanced by anti-CRISPRs derived from Enterococcus conjugative elements, highlighting a role for Acrs in the dissemination of plasmids. Reciprocal co-immunoprecipitation showed that each Acr protein interacts with Cas9, and Cas9-Acr complexes were unable to cleave DNA. Northern blotting suggests that these anti-CRISPRs manipulate single guide RNA length, loading or stability. Mirroring their activity in bacteria, AcrIIA16 and AcrIIA17 provide robust and highly potent broad-spectrum inhibition of distinct Cas9 proteins in human cells (for example, SpyCas9, SauCas9, SthCas9, NmeCas9 and CjeCas9). This work presents a focused analysis of non-phage Acr proteins, demonstrating a role in horizontal gene transfer bolstered by broad-spectrum CRISPR-Cas9 inhibition.
Subject(s)
CRISPR-Associated Protein 9/antagonists & inhibitors , CRISPR-Cas Systems , Gene Transfer, Horizontal , Plasmids/metabolism , RNA, Guide, Kinetoplastida/antagonists & inhibitors , CRISPR-Associated Protein 9/genetics , CRISPR-Associated Protein 9/metabolism , Clustered Regularly Interspaced Short Palindromic Repeats , Conjugation, Genetic , DNA/antagonists & inhibitors , DNA/genetics , DNA/metabolism , Enterococcus/genetics , Enterococcus/virology , HEK293 Cells , Humans , Listeria/genetics , Listeria/virology , Plasmids/chemistry , Protein Binding , RNA, Guide, Kinetoplastida/genetics , RNA, Guide, Kinetoplastida/metabolism , Staphylococcus/genetics , Staphylococcus/virology , Streptococcus/genetics , Streptococcus/virologyABSTRACT
3,6-Diethynyl-9,10-diethoxyphenanthrene (4) was synthesized from phenanthrene and employed in the synthesis of the binuclear gold(I) alkynyl complexes (R3P)Au(C≡C-3-[C14H6-9,10-diethoxy]-6-C≡C)Au(PR3) (R = Ph (5a), Cy (5b)). The diyne 4 and complexes 5a and 5b were characterized by NMR spectroscopy, mass spectrometry, and elemental analysis. UV-Vis spectroscopy studies of the metal complexes and precursor diyne show strong p à p* transitions in the near UV region that red shift by ca. 50 nm upon coordination at the gold centers. The emission spectrum of 4 shows an intense fluorescence band centered at 420 nm which red shifts, slightly upon coordination of 4 to gold. Binding studies of 4, 5a, and 5b against calf thymus DNA were carried out, revealing that 4, 5a, and 5b have >40% stronger binding affinities than the commonly used intercalating agent ethidium bromide. The molecular docking scores of 4, 5a, and 5b with B-DNA suggest a similar trend in behavior to that observed in the DNA-binding study. Unlike the ligand 4, promising anticancer properties for 5a and 5b were observed against several cell lines; the DNA binding capability of the precursor alkyne was maintained, and its anticancer efficacy enhanced by the gold centers. Such phenanthrenyl complexes could be promising candidates in certain biological applications because the two components (phenanthrenyl bridge and metal centers) can be altered independently to improve the targeting of the complex, as well as the biological and physicochemical properties.
Subject(s)
Antineoplastic Agents/chemistry , Coordination Complexes/pharmacology , DNA/chemistry , Neoplasms/drug therapy , Alkynes/chemical synthesis , Alkynes/chemistry , Alkynes/pharmacology , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cell Proliferation/drug effects , Coordination Complexes/chemical synthesis , Coordination Complexes/chemistry , DNA/antagonists & inhibitors , Drug Screening Assays, Antitumor , Ethidium/chemistry , Gold/chemistry , Humans , Ligands , Magnetic Resonance Spectroscopy , Molecular Docking Simulation , Neoplasms/chemistryABSTRACT
DNA forms not only the canonical duplex structure but also non-canonical structures. Most potential sequences that induce the formation of non-canonical structures are present in disease-related genes. Interestingly, biological reactions are inhibited or dysregulated by non-canonical structure formation in disease-related genes. To control biological reactions, methods for inducing the formation of non-canonical structures have been developed using small molecules and oligonucleotides. In this feature article, we review biological reactions such as replication, transcription, and reverse transcription controlled by non-canonical DNA structures formed by disease-related genes. Furthermore, we discuss recent studies aimed at developing methods for regulating these biological reactions using drugs targeting the DNA structure.
Subject(s)
DNA/antagonists & inhibitors , Pharmaceutical Preparations/chemistry , Animals , DNA/chemistry , DNA/metabolism , Humans , Nucleic Acid ConformationABSTRACT
The water-compatible optically pure metallohelices made by self-assembly of simple nonpeptidic organic components around Fe(II) ions are now recognized as a distinct subclass of helicates that exhibit similar architecture to some natural cationic antimicrobial peptides. Notably, a new series of metallohelices was recently shown to exhibit biological activity, displaying high, structure-dependent activity against bacteria. It is also important that, thanks to their properties, such metallohelices can exhibit specific interactions with biomacromolecules. Here, following our prior report on the metallohelices that have high, structure-dependent activity against bacteria, we investigated the interactions of the series of iron(II) metallohelices with DNA, which is a potential pharmacological target of this class of coordination compounds. The results obtained with the aid of biophysical and molecular biology methods show that the investigated metallohelices accumulate in eukaryotic cells and that a significant fraction of the metallohelices accumulates in the cell nucleus, allowing them to interact also with nuclear DNA. Additionally, we have demonstrated that some metallohelices have a high affinity to DNA and are able to condense/aggregate DNA molecules more efficiently than conventional DNA-condensing agents, such as polyamines. Moreover, this capability of the metallohelices correlates with their efficiency to inhibit DNA-related enzymatic activities, such as those connected with DNA transcription, catalysis of DNA relaxation by DNA topoisomerase I, and cleavage by restriction enzymes.
Subject(s)
Cell Nucleus/chemistry , DNA Topoisomerases, Type I/metabolism , DNA/antagonists & inhibitors , Ferrous Compounds/pharmacology , Topoisomerase I Inhibitors/pharmacology , Cell Nucleus/metabolism , DNA/genetics , DNA/metabolism , Ferrous Compounds/chemical synthesis , Ferrous Compounds/chemistry , HCT116 Cells , Humans , Molecular Structure , Optical Phenomena , Topoisomerase I Inhibitors/chemical synthesis , Topoisomerase I Inhibitors/chemistryABSTRACT
Introduction: The worldwide incidence of central nervous system (CNS) primary tumors is increasing. Most of the chemotherapeutic agents used for treating these cancer types induce DNA damage, and their activity is affected by the functional status of repair systems involved in the detection or correction of DNA lesions. Unfortunately, treatment of malignant high-grade tumors is still an unmet medical need.Areas covered: We summarize the action mechanisms of the main DNA inhibitors used for the treatment of brain tumors. In addition, studies on new agents or drug combinations investigated for this indication are reviewed, focusing our attention on clinical trials that in the last 3 years have been completed, terminated or are still recruiting patients.Expert opinion: Much still needs to be done to render aggressive CNS tumors curable or at least to transform them from lethal to chronic diseases, as it is possible for other cancer types. Drugs with improved penetration in the CNS, toxicity profile, and activity against primary and recurrent tumors are eagerly needed. Targeted agents with innovative mechanisms of action and ability to harness the cells of the tumor microenvironment against cancer cells represent a promising approach for improving the clinical outcome of CNS tumors.
Subject(s)
Antimetabolites, Antineoplastic/administration & dosage , Antineoplastic Agents/administration & dosage , Brain Neoplasms/drug therapy , DNA Damage/drug effects , Topoisomerase Inhibitors/administration & dosage , Animals , Antimetabolites, Antineoplastic/metabolism , Antineoplastic Agents/metabolism , Brain Neoplasms/diagnosis , Brain Neoplasms/metabolism , DNA/antagonists & inhibitors , DNA/metabolism , DNA Damage/physiology , Humans , Topoisomerase Inhibitors/metabolism , Tumor Microenvironment/drug effects , Tumor Microenvironment/physiologyABSTRACT
Deoxyribonucleoside triphosphates (dNTPs) are building blocks for the biosynthesis of DNA. Various modified dNTPs' analogs have synthesized by structural changes of nucleoside's susgar and nucleobases and employed for synthesis of modified DNA. A very few modified dNTPs have prepared from non-sugar nucleoside analogs. This report describes the synthesis of acyclic nucleoside triphosphate (NTP) analog from amino acid L-Serine as aminopropanolyl-thymine triphosphate (ap-TTP) and demonstrate its biochemical evaluation as enzymatic incorporation of ap-TTP into DNA with DNA polymerases with primer extension methods. Alanyl peptide nucleicacids (Ala-PNA) are the analogs of DNA which contains alanyl backbone. Aminopropanolyl - analogs are derivatives of alanyl back bone. Ap-TTP analog is nucleoside triphosphate analog derived from Ala-PNA. Importantly, this report also sheds light on the crystal packing arrangement of alaninyl thymine ester derivative in solid-state and reveals the formation of self-duplex assembly in anti-parallel fashion via reverse Watson-Crick hydrogen bonding and π-π interactions. Hence, ap-TTP is a useful analog which also generates the free amine functional group at the terminal of DNA oligonucleotide after incorporation.
Subject(s)
DNA/antagonists & inhibitors , Propranolol/pharmacology , DNA/biosynthesis , DNA-Directed DNA Polymerase/metabolism , Molecular Structure , Propranolol/chemical synthesis , Propranolol/chemistry , Thiamine/chemical synthesis , Thiamine/chemistry , Thiamine/pharmacologyABSTRACT
Antisense oligonucleotide (ASO)-induced cellular signaling pathway alteration is evolving as a promising therapeutic strategy for improving antitumor chemotherapy. However, the inherent instability and inefficiency of ASO delivery remain major hurdles for its application. Herein, we developed a self-assembled DNA nanosponge (DNS) for adsorption and clearance of intracellular miR-21. The densely packed DNA nanostructure is able to provide large amounts of repeated ASO copies for efficient capturing of miR-21 and inhibiting the miRNAs function in mammalian cells. The cell apoptosis-related protein expression (Bcl-2, Bax, and cleaved caspase-3/9) can be obviously interrupted with the delivery of DNS. Besides, we have shown that the DNS can efficiently carry Dox for chemotherapy and inducing tumor cell (MCF-7) apoptosis meanwhile has little affect to normal cells (Hs578 Bst). These polymeric DNS systems mimic the natural RNA circle-based miRNA sponges and have potential to be applied for specific and efficient regulation of gene expression in tumor cells for synergistic antitumor chemotherapy.
Subject(s)
Breast Neoplasms/drug therapy , DNA/genetics , MicroRNAs/genetics , Oligonucleotides, Antisense/pharmacology , Adsorption/drug effects , Animals , Apoptosis/drug effects , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Caspase 3/genetics , Cytoplasm/genetics , DNA/antagonists & inhibitors , Doxorubicin/chemistry , Doxorubicin/pharmacology , Female , Gene Expression Regulation, Neoplastic/drug effects , Humans , MCF-7 Cells , Mice , MicroRNAs/antagonists & inhibitors , Nanoparticles/chemistry , Oligonucleotides, Antisense/genetics , Proto-Oncogene Proteins c-bcl-2/genetics , RNA, Circular/genetics , Xenograft Model Antitumor AssaysABSTRACT
Here, we report the synthesis and characterization of four new aroyl-hydrazone derivatives L1 -L4 , and their structural as well as biological activities have been explored. In addition to docking with bovine serum albumin (BSA) and duplex DNA, the experimental results demonstrate the effective binding of L1 -L4 with BSA protein and calf thymus DNA (ct-DNA) which is in agreement with the docking results. Further biological activities of L1 -L4 have been examined through molecular docking with different proteins which are involved in the propagation of viral or cancer diseases. L1 shows best binding affinity with influenza A virus polymerase PB2 subunit (2VY7) with binding energy -11.42â kcal/mol and inhibition constant 4.23â nm, whereas L2 strongly bind with the hepatitis C virus NS5B polymerase (2WCX) with binding energy -10.47â kcal/mol and inhibition constant 21.06â nm. Ligand L3 binds strongly with TGF-beta receptor 1 (3FAA) and L4 with cancer-related EphA2 protein kinases (1MQB) with binding energy -10.61â kcal/mol, -10.02â kcal/mol and inhibition constant 16.67â nm and 45.41â nm, respectively. The binding energies of L1 -L4 are comparable with binding energies of their proven inhibitors. L1 , L3 and L4 can be considered as both 3FAA and 1MQB dual targeting anticancer agents, while L1 and L3 are both 2VY7 and 2WCX dual targeting antiviral agents. On the other side, L2 and L4 target only one virus related target (2WCX). Furthermore, the geometry optimizations of L1 -L4 were performed via density functional theory (DFT). Moreover, all four ligands (L1 -L4 ) were characterized by NMR, FT-IR, ESI-MS, elemental analysis and their molecular structures were validated by single crystal X-ray diffraction studies.
Subject(s)
Antineoplastic Agents/pharmacology , Antiviral Agents/pharmacology , DNA/antagonists & inhibitors , Drug Design , Hydrazones/pharmacology , Molecular Docking Simulation , Serum Albumin, Bovine/antagonists & inhibitors , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , Cattle , Cell Proliferation/drug effects , Crystallography, X-Ray , DNA/chemistry , Density Functional Theory , Drug Screening Assays, Antitumor , Hepacivirus/drug effects , Hydrazones/chemical synthesis , Hydrazones/chemistry , Influenza A virus/drug effects , Ligands , Microbial Sensitivity Tests , Molecular Structure , Serum Albumin, Bovine/chemistryABSTRACT
CC-115, a triazole-containing compound, is a dual mammalian target of rapamycin (mTOR)/DNA-dependent protein kinase (DNA-PK) inhibitor currently in clinical trials. To develop this compound further, we investigated factors that may affect cellular response to CC-115. Previously, fatty acid synthase (FASN) was shown to upregulate DNA-PK activity and contribute to drug resistance; therefore, we hypothesized that FASN may affect cellular response to CC-115. Instead, however, we showed that CC-115 is a substrate of ATP-binding cassette G2 (ABCG2), a member of the ATP-binding cassette transporter superfamily, and that expression of ABCG2, not FASN, affects the potency of CC-115. ABCG2 overexpression significantly increases resistance to CC-115. Inhibiting ABCG2 function, using small-molecule inhibitors, sensitizes cancer cells to CC-115. We also found that CC-115 may be a substrate of ABCB1, another known ABC protein that contributes to drug resistance. These findings suggest that expression of ABC transporters, including ABCB1 and ABCG2, may affect the outcome in clinical trials testing CC-115. Additionally, the data indicate that ABC transporters may be used as markers for future precision use of CC-115. SIGNIFICANCE STATEMENT: In this article, we report our findings on the potential mechanism of resistance to CC-115, a dual inhibitor of mTOR and DNA-PK currently in clinical trials. We show that CC-115 is a substrate of ABCG2 and can be recognized by ABCB1, which contributes to CC-115 resistance. These findings provide novel information and potential guidance on future clinical testing of CC-115.
Subject(s)
ATP Binding Cassette Transporter, Subfamily G, Member 2/metabolism , Drug Resistance/drug effects , Neoplasm Proteins/metabolism , Protein Kinase Inhibitors/pharmacology , Pyrazines/pharmacology , TOR Serine-Threonine Kinases/antagonists & inhibitors , TOR Serine-Threonine Kinases/metabolism , Triazoles/pharmacology , ATP Binding Cassette Transporter, Subfamily B/metabolism , Cell Survival/drug effects , Cell Survival/physiology , Clinical Trials as Topic/methods , DNA/antagonists & inhibitors , DNA/metabolism , Dose-Response Relationship, Drug , Drug Resistance/physiology , HEK293 Cells , Humans , MCF-7 Cells , Risk Factors , Substrate Specificity/drug effects , Substrate Specificity/physiologyABSTRACT
Antisense oligonucleotides (ASO), short single-stranded polymers based on DNA or RNA chemistries and synthesized in vitro, regulate gene expression by binding in a sequence-specific manner to an RNA target. The functional activity and selectivity in the action of ASOs largely depends on the combination of nitrogenous bases in a target sequence. This simple and natural property of nucleic acids provides an attractive route by which scientists can create different ASO-based techniques. Over the last 50 years, planned and realized applications in the field of antisense and nucleic acid nanotechnologies have produced astonishing results and posed new challenges for further developments, exemplifying the essence of the post-genomic era. Today the majority of ASOs are chemically modified and/or incorporated within nanoparticles to enhance their stability and cellular uptake. This review critically analyzes some successful cases using the antisense approach in medicine to address severe diseases, such as Duchenne muscular dystrophy and spinal muscular atrophy, and suggests some prospective directions for future research. We also examine in detail the elaboration of unmodified insect-specific DNA insecticides and RNA preparations in the areas of agriculture and forestry, a relatively new branch of ASO that allows circumvention of the use of non-selective chemical insecticides. When considering the variety of successful ASO modifications with an efficient signal-to-noise ratio of action, coupled with the affordability of in vitro oligonucleotide synthesis and post-synthesis procedures, we predict that the next half-century will produce a fruitful yield of tools created from effective ASO-based end products.
Subject(s)
Biological Control Agents/pharmacology , Moths/drug effects , Muscular Atrophy, Spinal/therapy , Muscular Dystrophy, Duchenne/therapy , Neuromuscular Agents/therapeutic use , Oligonucleotides, Antisense/genetics , Agriculture/methods , Animals , Biological Control Agents/chemical synthesis , Biological Control Agents/history , DNA/antagonists & inhibitors , DNA/genetics , DNA/metabolism , Forestry/methods , Gene Expression Regulation/drug effects , History, 20th Century , History, 21st Century , Humans , Larva/drug effects , Larva/genetics , Larva/metabolism , Moths/genetics , Moths/growth & development , Moths/metabolism , Muscular Atrophy, Spinal/genetics , Muscular Atrophy, Spinal/metabolism , Muscular Atrophy, Spinal/pathology , Muscular Dystrophy, Duchenne/genetics , Muscular Dystrophy, Duchenne/metabolism , Muscular Dystrophy, Duchenne/pathology , Nanoparticles/administration & dosage , Nanoparticles/chemistry , Neuromuscular Agents/chemical synthesis , Neuromuscular Agents/history , Oligonucleotides, Antisense/chemical synthesis , Oligonucleotides, Antisense/metabolism , RNA, Messenger/antagonists & inhibitors , RNA, Messenger/genetics , RNA, Messenger/metabolismABSTRACT
Culturing of bone marrow cells in serum-free RPMI-1640 medium for 24 h was accompanied by a decrease in the rate of [3H]-thymidine incorporation into DNA. Addition of native apolipoprotein A-I (apoA-I) or plasma LDL and HDL to the culture medium increased this parameter. In contrast to native apoA-I, its modified form decelerated DNA synthesis in bone marrow cells. A similar inhibitory effect of modified protein was observed in cultures of human embryonic kidney cells (HEK293) and in rapidly proliferating mouse macrophage cell line ANA-1. The only exclusion was human myeloid cell line U937: neither native nor modified apoA-I affected DNA synthesis in these cells. Thus, the regulatory effects of apoA-I are tissue-specific; this protein can produce either stimulatory or inhibitory effect on DNA biosynthesis in cells depending on its conformation.