DNA Intercalating Research Articles

Integrated imaging and proteomics sensors detect proteome aggregation induced by platinum-based chemotherapy drugs in living cells and mice model

Proteome aggregation is challenging to analyze given its loss of defined 3-D structures to develop chemo- and bio-sensors. Herein, we present integrated chemical sensors to image the occurrence of proteome aggregation and dissect its proteomic composition. Via these combinative sensors, we systematically reveal for the first time platinum-based drugs can deteriorate proteome stability, compromise protein homeostasis (proteostasis) network, and consequently cause proteome aggregation. Fluorescent protein aggregation imaging sensors (AggStain and AggRetina) not only confirms these drugs generally induce intracellular proteome aggregation but also quantifies the compactness (polarity) heterogeneity inside these aggregates. Chemical proteomics sensor (AggLink) further captures and profiles the composition of aggregated proteome, which uncovers the fluctuations of proteostasis network and metabolic pathways. In mice model, proteome aggregation and oxidative stress are imaged in multi-organs with platinum enrichment. Together, we integrate imaging and proteomics sensors to provide an alternative mechanism-of-action for platinum-based drugs’ anti-cancer function in addition to their common DNA intercalation mechanism.

Hollow Polyethyleneimine Nanoparticles with Drug Loaded DNA for Chemotherapeutic Applications.

The next generation of anticancer agents are emerging from rationally designed nanostructured materials. This work involved the synthesis and characterization of novel hollow DNA-conjugated gold nanoparticles (DNA-AuNPs) for controlled drug delivery. Polyethyleneimine (PEI) was bound to AuNPs, forming polymer-shell nanoparticles. Dissolution of the gold core via iodine formed hollow core polymeric nanoparticles (HCPNPs) and a high density (85 molecules/particle) of DNA intercalated with daunorubicin was conjugated. Particles were spherical with an average diameter of 105.7 ± 17.3 nm and zeta potential of 20.4 ± 3.54 mV. We hypothesize the DNA backbone electrostatically condensed to the primary amines on the surface of the particle toroidally, weaving itself within the polymer shell. During the DNA intercalation process, increasing the ionic concentration and decreasing the amine/phosphate ratio 10-fold increased drug intercalation 64% and 61%, respectively, allowing us to determine the optimal method of particle synthesis. As intercalation sites increased with increasing DNA strand length, drug loading increased. An average of 874 ± 40.1 daunorubicin molecules were loaded per HCPNP. HCPNPs with drug intercalated DNA have strong potential to be clinically efficacious drug delivery vehicles due to the versatility of DNA and high drug loading capacities.

The crucial role of stability of intercalating agent for DNA binding studies in DMSO/water system

In recent years, extensive research has been directed towards understanding the interactions between various zinc complexes with DNA, specifically delving into their intercalation and binding behaviors. The binding of zinc complexes to DNA is particularly intriguing due to their distinctive intercalating capabilities. This study unveils a remarkable phenomenon observed with a specific Zn complex, ([B-Zn-N3], where B is a Schiff base ligand), during DNA intercalation investigations in the popular DMSO-Water binary solvent mixture. An unanticipated observation revealed time-dependent changes in the UV–visible absorption spectroscopic studies, coupled with the existence of an isosbestic point. This observation questions the stability of the intercalating agent itself during the intercalation process. The emergence of a decomposed product during the intercalation study has been confirmed through various analytical techniques, including CHN analysis, MALDI mass, XPS, Raman spectroscopy, and Powder XRD. The change in the chemical species on intercalation is further substantiated by theoretical studies, adding depth to our understanding of the intricate dynamics at play during DNA intercalation with the [B-Zn-N3] complex in the DMSO-Water system.

Ultrafast spectroscopy study of DNA photophysics after proflavine intercalation.

Proflavine (PF), an acridine DNA intercalating agent, has been widespread applied as an anti-microbial and topical antiseptic agent due to its ability to suppress DNA replication. On the other hand, various studies show that PF intercalation to DNA can increase photogenotoxicity and has potential chances to induce carcinomas of skin appendages. However, the effects of PF intercalation on the photophysical and photochemical properties of DNA have not been sufficiently explored. In this study, the excited state dynamics of the PF intercalated d(GC)9 • d(GC)9 and d(AT)9 • d(AT)9 DNA duplex are investigated in an aqueous buffer solution. Under 267nm excitation, we observed ultrafast charge transfer (CT) between PF and d(GC)9 • d(GC)9 duplex, generating a CT state with an order of magnitude longer lifetime compared to that of the intrinsic excited state reported for the d(GC)9 • d(GC)9 duplex. In contrast, no excited state interaction was detected between PF and d(AT)9 • d(AT)9. Nevertheless, a localized triplet state with a lifetime over 5 µs was identified in the PF-d(AT)9 • d(AT)9 duplex.

New imidazole-2-thiones linked to acenaphythylenone as dual DNA intercalators and topoisomerase II inhibitors: structural optimization, docking, and apoptosis studies

In this article, a new series of 2-((3,5-disubstituted-2-thioxo-imidazol-1-yl)imino)acenaphthylen-1(2H)-ones were synthesized. Imidazole-2-thione with acenaphthylen-one gave a hybrid scaffold that integrated key structural elements essential for DNA damage via direct DNA intercalation and inhibition of the topoisomerase II enzyme. All the synthesized compounds were screened to detect their DNA damage using a terbium fluorescent probe. Results demonstrated that 4-phenyl-imidazoles 5b and 5e in addition to 4-(4-chlorophenyl)imidazoles 5h and 5j would induce detectable potent damage in ctDNA. The four most potent compounds as DNA intercalators were further evaluated for their antiproliferative activity against HepG2, MCF-7 and HCT-116 utilizing the MTT assay. The highest anticancer activity was recorded with compounds 5b and 5h against the breast cancer cell line MCF-7 which were 1.5- and 3- folds more active than doxorubicin, respectively. Therefore, imidazole-2-thione tethered acenaphthylenone derivatives can be considered as promising scaffold for the development of effective dual DNA intercalators and topoisomerase II inhibitors.

Bacteria-mediated resistance of neutrophil extracellular traps to enzymatic degradation drives the formation of dental calculi.

Dental calculi can cause gingival bleeding and periodontitis, yet the mechanism underlying the formation of such mineral build-ups, and in particular the role of the local microenvironment, are unclear. Here we show that the formation of dental calculi involves bacteria in local mature biofilms converting the DNA in neutrophil extracellular traps (NETs) from being degradable by the enzyme DNase I to being degradation resistant, promoting the nucleation and growth of apatite. DNase I inhibited NET-induced mineralization in vitro and ex vivo, yet plasma DNases were ineffective at inhibiting ectopic mineralization in the oral cavity in rodents. The topical application of the DNA-intercalating agent chloroquine in rodents fed with a dental calculogenic diet reverted NET DNA to its degradable form, inhibiting the formation of calculi. Our findings may motivate therapeutic strategies for the reduction of the prevalence of the deposition of bacteria-driven calculi in the oral cavity.

Spectroscopic view on the interaction between the psoralen derivative amotosalen and DNA

Psoralens are eponymous for PUVA (psoralen plus UV-A radiation) therapy, which inter alia can be used to treat various skin diseases. Based on the same underlying mechanism of action, the synthetic psoralen amotosalen (AMO) is utilized in the pathogen reduction technology of the INTERCEPT® Blood System to inactivate pathogens in plasma and platelet components. The photophysical behavior of AMO in the absence of DNA is remarkably similar to that of the recently studied psoralen 4′-aminomethyl-4,5′,8-trimethylpsoralen (AMT). By means of steady-state and time-resolved spectroscopy, intercalation and photochemistry of AMO and synthetic DNA were studied. AMO intercalates with a higher affinity into A,T-only DNA (KD = 8.9 × 10−5 M) than into G,C-only DNA (KD = 6.9 × 10−4 M). AMO covalently photobinds to A,T-only DNA with a reaction quantum yield of ΦR = 0.11. Like AMT, it does not photoreact following intercalation into G,C-only DNA. Femto- and nanosecond transient absorption spectroscopy reveals the characteristic pattern of photobinding to A,T-only DNA. For AMO and G,C-only DNA, signatures of a photoinduced electron transfer are recorded.Graphical abstract

Discovery of indoleninyl-pyrazolo[3,4-b]pyridines as potent chemotherapeutic agents against colorectal cancer cells

A new series of indolenines decorated with pyrazolo[3,4-b]pyridines were designed and synthesized in up to 96% yield from the acid-catalyzed cyclocondensation of 1,3-dialdehydes with 3-aminopyrazoles. X-ray crystallography on a representative derivative, 5n, revealed two close to planar conformations whereby the N-atom of the pyridyl residue was syn or anti to the pyrrole-N atom in the two independent molecules of the asymmetric unit. The computational and DNA binding data suggest that 5n is a strong DNA intercalator with the results in agreement with its potent cytotoxicity against two colorectal cancer cell lines (HCT 116 and HT-29). In contrast to doxorubicin, compounds 5k-o have higher druggability (compliance to more criteria stated in Lipinski’s rule of five and Veber’s rule), higher bioavailability, and better medicinal chemistry properties, indicative of their potential application as chemotherapeutical agents.

Anticancer Tetrahydrocarbazoles: A Wide Journey from 2000 Till Date

Abstract: Tetrahydrocarbazoles (THCz) are widespread in numerous indole alkaloids and have been reported since time for exhibiting profound pharmacological potential. Many pharmaceuticals drugs have tetrahydrocarbazole nucleus in their structure e.g. vinca alkaloids (Vincristine, Vinblastine, Vinorelbine), Frovatriptan, (R)-Ramatroban, Ondansetron, etc. that are used in various multifactorial diseases. In this review article, the anticancer potential of tetrahydrocarbazole based derivatives has been covered, enumerating their vast journey from the year 2000 to 2021. Since the last twenty-one years, tetrahydrocarbazoles have been a matter of focus among researchers worldwide, whereby several novel tetrahydrocarbazole derivatives have been synthesized and reported for their anticancer potential against various cancer cell lines. Tetrahydrocarabzole and its derivatives have exhibited profound anticancer potential mediated via various cancer pathways like apoptosis, cell cycle arrest, microtubule inhibition, Nrf2 Modulators, DNA intercalators, pERK and pRb phosphorylation, VEGF (Vascular Endothelial Growth Factor) and TNF-α inhibition, TPSO (translocator protein), Histone Deacetylase (HDAC) Inhibitors also discussed. The present review entails the synthesis, SAR studies, and anticancer mechanism of tetrahydrocarbazoles derivatives reported in review literature till date, and would provide a strong database to the medicinal chemist world over in discovering newer potential anticancer agent against various types of cancer diseases.

Quantitative detection of chikungunya, Zika, and dengue viruses by one-step real-time PCR in different cell substrates.

Chikungunya (CHIKV), Zika (ZIKV), and dengue viruses (DENV) are vector-borne pathogens that cause emerging and re-emerging epidemics throughout tropical and subtropical countries. The symptomatology is similar among these viruses and frequently co-circulates in the same areas, making the diagnosis arduous. Although there are different methods for detecting and quantifying pathogens, real-time reverse transcription-polymerase chain reaction (real-time RT-qPCR) has become a leading technique for detecting viruses. However, the currently developed assays frequently involve probes and high-cost reagents, limiting access in low-income countries. Therefore, this study aims to design and evaluate a quantitative one-step RT-qPCR assay to detect CHIKV, ZIKV, and DENV with high specificity, reproducibility, and low cost in multiple cell substrates. We established a DNA intercalating green dye-based RT-qPCR test that targets nsP1 of CHIKV, and NS5 gene of ZIKV, and DENV for the amplification reaction. The assay exhibited a high specificity confirmed by the melting curve analysis. No cross-reactivity was observed between the three viruses or unspecific amplification of host RNA. The sensitivity of the reaction was evaluated for each virus assay, getting a limit of detection of one RNA copy per virus. Standard curves were constructed, obtaining a reaction efficiency of ~ 100%, a correlation coefficient (R2) of ~ 0.97, and a slope of -3.3. The coefficient of variation (CV) ranged from 0.02 to 1.43. In addition, the method was optimized for viral quantification and tested in Vero, BHK-21, C6/36, LULO, and the Aedes cell lines. Thus, the DNA intercalating green dye-based RT-qPCR assay was a highly specific, sensitive, reproducible, and effective method for detecting and quantifying CHIKV, ZIKV, and DENV in different cell substrates that could also be applied in clinical samples.

Multi-target rational design and synthesis of novel diphenyl-tethered pyrazolopyrimidines targeting EGFR and topoisomerase II with potential DNA intercalation and apoptosis induction

Herein, we envisioned the design and synthesis of novel pyrazolopyrimidines (confirmed by elemental analysis, 1H and 13C NMR, and mass spectra) as multitarget-directed drug candidates acting as EGFR/TOPO II inhibitors, DNA intercalators, and apoptosis inducers. The target diphenyl-tethered pyrazolopyrimidines were synthesized starting from the reaction of phenyl hydrazine and ethoxymethylenemalononitrile to give aminopyrazole-carbonitrile 2. The latter hydrolysis with NaOH and subsequent reaction with 4-chlorobenzaldhyde afforded the corresponding pyrazolo[3,4-d]pyrimidin-4-ol 4. Chlorination of 4 with POCl3 and sequential reaction with different amines afforded the target compounds in good yields (up to 73 %). The growth inhibition % of the new derivatives (6a-m) was investigated against different cancer and normal cells and the IC50 values of the most promising candidates were estimated for HNO97, MDA-MB-468, FaDu, and HeLa cancer cells. The frontier derivatives (6a, 6i, 6k, 6l, and 6m) were pursued for their EGFR inhibitory activity. Compound 6l decreased EGFR protein concentration by a 6.10-fold change, compared to imatinib as a reference standard. On the other side, compounds (6a, 6i, 6k, 6l, and 6m) underwent topoisomerase II (TOPO II) inhibitory assay. In particular, compounds 6a and 6l exhibited IC50s of 17.89 and 19.39 μM, respectively, surpassing etoposide with IC50 of 20.82 μM. Besides, the DNA fragmentation images described the great potential of both candidates 6a and 6l in inducing DNA degradation at lower concentrations compared to etoposide and doxorubicin. Moreover, compound 6l, with the most promising EGFR/TOPO II inhibition and DNA intercalation, was selected for further investigation for its apoptosis induction ability by measuring caspases 3, 7, 8, and 9, Bax, p53, MMP2, MMP9, and BCL-2 proteins. Additionally, molecular docking was used to explain the SAR results based on the differences in the molecular features of the investigated congeners and the target receptors' topology.

Autophagy-driven regulation of cisplatin response in human cancers: Exploring molecular and cell death dynamics

Despite the challenges posed by drug resistance and side effects, chemotherapy remains a pivotal strategy in cancer treatment. A key issue in this context is macroautophagy (commonly known as autophagy), a dysregulated cell death mechanism often observed during chemotherapy. Autophagy plays a cytoprotective role by maintaining cellular homeostasis and recycling organelles, and emerging evidence points to its significant role in promoting cancer progression. Cisplatin, a DNA-intercalating agent known for inducing cell death and cell cycle arrest, often encounters resistance in chemotherapy treatments. Recent studies have shown that autophagy can contribute to cisplatin resistance or insensitivity in tumor cells through various mechanisms. This resistance can be mediated by protective autophagy, which suppresses apoptosis. Additionally, autophagy-related changes in tumor cell metastasis, particularly the induction of Epithelial-Mesenchymal Transition (EMT), can also lead to cisplatin resistance. Nevertheless, pharmacological strategies targeting the regulation of autophagy and apoptosis offer promising avenues to enhance cisplatin sensitivity in cancer therapy. Notably, numerous non-coding RNAs have been identified as regulators of autophagy in the context of cisplatin chemotherapy. Thus, therapeutic targeting of autophagy or its associated pathways holds potential for restoring cisplatin sensitivity, highlighting an important direction for future clinical research.

Chromatin damage generated by DNA intercalators leads to degradation of RNA Polymerase II

Abstract In cancer therapy, DNA intercalators are mainly known for their capacity to kill cells by inducing DNA damage. Recently, several DNA intercalators have attracted much interest given their ability to inhibit RNA Polymerase I transcription (BMH-21), evict histones (Aclarubicin) or induce chromatin trapping of FACT (Curaxin CBL0137). Interestingly, these DNA intercalators lack the capacity to induce DNA damage while still retaining cytotoxic effects and stabilize p53. Herein, we report that these DNA intercalators impact chromatin biology by interfering with the chromatin stability of RNA polymerases I, II and III. These three compounds have the capacity to induce degradation of RNA polymerase II and they simultaneously enable the trapping of Topoisomerases TOP2A and TOP2B on the chromatin. In addition, BMH-21 also acts as a catalytic inhibitor of Topoisomerase II, resembling Aclarubicin. Moreover, BMH-21 induces chromatin trapping of the histone chaperone FACT and propels accumulation of Z-DNA and histone eviction, similarly to Aclarubicin and CBL0137. These DNA intercalators have a cumulative impact on general transcription machinery by inducing accumulation of topological defects and impacting nuclear chromatin. Therefore, their cytotoxic capabilities may be the result of compounding deleterious effects on chromatin homeostasis.

(Benz)imidazo[1,2-a]quinolinium Salts: Access via Unprecedented Regiospecific non-AAIPEX Strategy and Study of Their Tunable Properties.

An unprecedented non-AAIPEX protocol has been developed to access diverse monosubstituted cationic polycyclic heteroaromatic compounds (cPHACs) from the readily available azolium salts and phenacyl bromides via Ru(II)-catalyzed tandem annulation cum aromatization. This atom-economic protocol executes a range of intermediate steps e. g. double C-H activation, nucleophilic addition, annulation, and dehydration cum aromatization in one-pot manner under the generation of H2O as the sole byproduct. Moreover, the systematic tunability of photo-physical and electrochemical properties of these new class of cPHACs can be authenticated from the DFT calculated frontier molecular orbital energies that might be beneficial for their potential applications in optoelectronics and DNA intercalation.

Skin electroporation for transdermal drug delivery: Electrical measurements, numerical model and molecule delivery

Skin electroporation for drug delivery involves the application of Pulsed Electric Fields (PEFs) on the skin to disrupt its barrier function in a temporary and non-invasive manner, increasing the uptake of drugs. It represents a potential alternative to delivery methods that are invasive (e.g. injections) or limited. We have developed a drug delivery system comprising nanocomposite hydrogels which act as a reservoir for the drug and an electrode for applying electric pulses on the skin. In this study, we employed a multi-scale approach to investigate the drug delivery system on a mouse skin model, through electrical measurements, numerical modeling and fluorescence microscopy. The Electrical properties indicated a highly non-linear skin conductivity behavior and were used to fine-tune the simulations and study skin recovery after electroporation. Simulation of electric field distribution in the skin showed amplitudes in the range of reversible tissue electroporation (400–1200 V/cm), for 300 V PEF. Fluorescence microscopy revealed increased uptake of fluorescent molecules compared to the non-pulsed control. We reported two reversible electroporation domains for our configuration: (1) at 100 V PEF the first local transport regions appear in the extracellular lipids of the stratum corneum, demonstrated by a rapid increase in the skin's conductivity and an increased uptake of lucifer yellow, a small hydrophilic fluorophore and (2) at 300 V PEF, the first permeabilization of nucleated cells occurred, evidenced by the increased fluorescence of propidium iodide, a membrane-impermeable, DNA intercalating agent.

Mechanism of DNA Intercalation by Chloroquine Provides Insights into Toxicity.

Chloroquine has been used as a potent antimalarial, anticancer drug, and prophylactic. While chloroquine is known to interact with DNA, the details of DNA-ligand interactions have remained unclear. Here we characterize chloroquine-double-stranded DNA binding with four complementary approaches, including optical tweezers, atomic force microscopy, duplex DNA melting measurements, and isothermal titration calorimetry. We show that chloroquine intercalates into double stranded DNA (dsDNA) with a KD ~ 200 µM, and this binding is entropically driven. We propose that chloroquine-induced dsDNA intercalation, which happens in the same concentration range as its observed toxic effects on cells, is responsible for the drug's cytotoxicity.

Controlling Chiroptical Responses via Chemo-Mechanical Deformation of DNA Origami Structures.

DNA origami-based templates have been widely used to fabricate chiral plasmonic metamaterials due to their precise control of the placement of nanoparticles (NPs) in a desired configuration. However, achieving various chiroptical responses inevitably requires a change in the structure of DNA origami-based templates or binding sites on them, leading to the use of significantly different sets of DNA strands. Here, we propose an approach to controlling various chiroptical responses with a single DNA origami design using its chemo-mechanical deformation induced by DNA intercalators. The chiroptical response could be finely tuned by altering the concentration of intercalators only. The silver (Ag) enhancement was used to amplify the chiroptical signal by enlarging NPs and to maintain it by stiffening the template DNA structure. Furthermore, the sensitivity in the chiroptical signal change to the concentration of intercalators could be modulated by the type of intercalator, the mixture of two intercalators, and the stiffness of DNA origami structures. This approach would be useful in a variety of optical applications that require programmed spatial modification of chiroptical responses.

The Effect of UV-Vis Radiation on DNA Systems Containing the Photosensitizers Methylene Blue and Acridine Orange

As a vital biomolecule, DNA is known as a target of antineoplastic drugs for cancer therapy. These drugs can show different modes of interaction with DNA, with intercalation and groove binding being the most common types. The intercalation of anticancer drugs with DNA can lead to the disruption of its normal function, influencing cell proliferation. Methylene blue (MB) and acridine orange (AO) are examples of DNA-intercalating agents that have been studied for their application against some types of cancer, mainly for photodynamic therapy. In this work, the impact of light irradiation on these compounds in the absence and presence of DNA was analyzed by means of UV-vis spectroscopy. Bathochromic and hypochromic shifts were observed in the absorbance spectra, revealing the intercalation of the dyes with the DNA base pairs. Dyes with and without DNA present different profiles of photodegradation, whereby the dyes alone were more susceptible to degradation. This can be justified by the intercalation of the dyes on the DNA base pairs allowing the DNA molecule to partially hinder the molecules’ exposition and, therefore, reducing their degradation.

DNA-Targeted Complexes of Tc and Re for Biomedical Applications.

Due to their favorable chemical features, Re and Tc complexes have been widely used for the development of new therapeutic agents and imaging probes to solve problems of biomedical relevance. This review provides an update of the most relevant research efforts towards the development of novel cancer theranostic agents using Re and Tc-based compounds interacting with specific DNA structures. This includes a variety of homometallic complexes, namely those containing M(CO)3 (M=Re, Tc) moieties, that exhibit different modes of interaction with DNA, such as covalent binding, intercalation, groove binding or G-quadruplex DNA binding. Additionally, heterometallic complexes, designed to potentiate synergistic effects of different metal centers to improve DNA-targeting, cytotoxicity and fluorescence properties, are also reviewed. Particular attention is also given to 99m Tc- and 188 Re-labeled oligonucleotides that have been widely explored to develop imaging and therapeutic radiopharmaceuticals through the in vivo hybridization with a specific complementary DNA or RNA target sequence to provide useful molecular tools in precision medicine for cancer diagnosis and treatment. Finally, the need for further improvement of DNA-targeted Re and Tc-based compounds as potential therapeutic and diagnostic agents is highlighted, and future directions are discussed.

Iridium(III)-based minor groove binding complexes as DNA photocleavage agents.

Transition metal complexes containing the qtpy ligand (2':4,4'':4',4'''-quaterpyridyl) are known to be DNA intercalators or minor groove binders. In this study, new tricationic iridium(III) complexes of qtpy are reported. Both [Ir(bpy)2(qtpy)]3+1 and [Ir(phen)2(qtpy)]3+2 display good water solubility as chloride salts. The complexes possess high-energy excited states, which are quenched in the presence of duplex DNA and even by the mononucleotides guanosine monophosphate and adenosine monophosphate. Further studies reveal that although the complexes bind to quadruplex DNA, they display a preference for duplex structures, which are bound with an order of magnitude higher affinities than their isostructural dicationic RuII-analogues. Detailed molecular dynamics simulations confirm that the complexes are groove binders through the insertion of, predominantly, the qtpy ligand into the minor groove. Photoirradiation of 1 in the presence of plasmid DNA confirms that this class of complexes can function as synthetic photonucleases by cleaving DNA.