Mechanism of Daunomycin Intercalation into DNA from Enhanced Sampling Simulations.

Daunomycin is a widely used anticancer drug, yet the mechanism underlying how it binds to DNA remains contested. 469 all-atom trajectories of daunomycin binding to the DNA oligonucleotide d(GCG CAC GTG CGC) were collected using weighted ensemble (WE)-enhanced sampling. Mechanistic insights were revealed through analysis of the ensemble of trajectories. Initially, the binding process involves a ubiquitous hydrogen bond between the DNA backbone and the NH3+ group on daunomycin. During the binding process, most trajectories exhibited similar structural changes to DNA, including DNA base pair rise, bending, and minor groove width changes. Variability within the ensemble of binding trajectories illuminates differences in the orientation of daunomycin as it initially intercalates; around 10% of trajectories needed minimal rearrangement from intercalation to reaching the fully bound configuration, whereas most needed an additional 1-5 ns to rearrange. The results here emphasize the utility of generating an ensemble of trajectories to discern biomolecular binding mechanisms.

Hyaluronic acid functionalized citric acid dendrimer/UiO-66-COOH as a stable and biocompatible platform for daunorubicin delivery.

This work aimed to prepare a new system for daunorubicin (DNR) delivery to improve therapeutic efficiency and decrease unwanted side effects. Typically, at first, a carboxylic acid functional group containing metal-organic framework (UiO-66-COOH) was synthesized in a simple way. Then, a third generation of citric acid dendrimer (CAD G3) was grown on it (UiO-66-COOH-CAD G3). Finally, the system was functionalized with pre-modified hyaluronic acid (UiO-66-COOH-CAD-HA). SEM analysis displayed that the synthesized particles have a spherical shape with an average particle size ranging from 260 to 280 nm. An increase in hydrodynamic diameter from 223 nm for UiO-66-COOH to 481 nm for UiO-66-COOH-CAD-HA is a sign of success in the performed reactions. Also, the average pore size was calculated at about 4.04 nm. The DNR loading efficiency of UiO-66-COOH-CAD-HA was evaluated at ∼74 % (DNR@UiO-66-COOH-CAD-HA). It was observed that the drug release rate at a lower pH is more than higher pH. The maximum hemolysis of <3 % means that the UiO-66-COOH-CAD-HA is hemocompatible. The use of DNR-loaded UiO-66-COOH-CAD-HA led to cell-killing of 77.9 % for MDA-MB 231. These results specified the great potential of UiO-66-COOH-CAD-HA for tumor drug delivery, so it could be proposed as a new carrier for anticancer agents to minimize adverse effects and improve therapeutic efficacy.

Nanoparticles loaded with Daunorubicin as an advanced tool for cancer therapy.

Nowadays, with the advent of cutting-edge technologies in the field of biotechnology, some highly advanced medical methods are introduced to treat cancers more efficiently. In the chemotherapy processes, anti-cancer drugs can be encapsulated in a stimuli-responsive coating which is capable of being functionalized by diverse ligands to increase the biocompatibility and control drug release behavior in a targeted drug delivery system. Nanoparticles (NPs) are playing an important role as nanocarriers in chemotherapy procedures, recently, numerous novel drug delivery systems have been studied which employed diverse types of NPs with remarkable structural features like porous nanocarriers with active and extended surface areas to enhance the drug loading and delivery efficacy. In this study, Daunorubicin (DAU) as an effective anti-cancer drug for treating various cancers introduced, and its application for novel drug delivery systems either as a single chemotherapy agent or co-delivery alongside other drugs with diverse NPs has been reviewed.

Spectroscopic, calorimetric and cytotoxicity studies on the combined binding of daunorubicin and acridine orange to a DNA tetrahedron.

Chemotherapy-phototherapy (CTPT) combination drugs co-loaded by targeted DNA nanostructures can achieve controlled drug delivery, reduce toxic side effects and overcome multidrug resistance. Herein, we constructed and characterized a DNA tetrahedral nanostructure (MUC1-TD) linked with the targeting aptamer MUC1. The interaction of daunorubicin (DAU)/acridine orange (AO) alone and in combination with MUC1-TD and the influence of the interaction on the cytotoxicity of the drugs were evaluated. Potassium ferrocyanide quenching analysis and DNA melting temperature assays were used to demonstrate the intercalative binding of DAU/AO to MUC1-TD. The interactions of DAU and/or AO with MUC1-TD were analyzed by fluorescence spectroscopy and differential scanning calorimetry. The number of binding sites, binding constant, entropy and enthalpy changes of the binding process were obtained. The binding strength and binding sites of DAU were higher than those of AO. The presence of AO in the ternary system weakened the binding of DAU to MUC1-TD. In vitro cytotoxicity studies demonstrated that the loading of MUC1-TD augmented the inhibitory effects of DAU and AO and the synergistic cytotoxic effects of DAU + AO on MCF-7 cells and MCF-7/ADR cells. Cell uptake studies showed that the loading of MUC1-TD was beneficial in promoting the apoptosis of MCF-7/ADR cells due to its enhanced targeting to the nucleus. This study has important guiding significance for the combined application of DAU and AO co-loaded by DNA nanostructures to overcome multidrug resistance.

In vitro biocompatibility evaluations of pH-sensitive Bi2MoO6/NH2-GO conjugated polyethylene glycol for release of daunorubicin in cancer therapy.

Here, a pH-sensitive biocompatible nanocarrier system is synthesized by the combination of Bi2MoO6 nanoparticles, NH2-graphene oxide (GO), and polyethylene glycol (PEG) for loading and delivery of daunorubicin (DNR) into breast cancer cells. DNR is loaded onto the nanocarrier surface via covalent bonding, exhibiting pH-sensitive behavior so that in acidic pH, nearly 86.85% of the drug is released, but in biological pH, only about 15% of the drug is released. The resulting Bi2MoO6/NH2-GO/PEG/DNR has a high drug loading content (33.29%) and encapsulation efficiency (99.75%). By examining the toxicity of the nanocarrier-loaded drug, no adverse effect is observed on healthy cells HUVEC, and the survival rate of cancer cells MCF-7 decreases with increasing the nanocarrier concentration. Moreover, the free drug is found to be more toxic than DNR attached to the nanocarrier. The complement activation (C3 and C4 levels), prothrombin time and activated partial thromboplastin time analyses also indicate its excellent blood compatibility. The hemolysis analysis (HRs),used to evaluate the nanocarrier compatibility. the results show that even in high concentrations(5-100 µg/ml), the percentage of hemolysis is below 1.8%, which indicates that the nanocarrier is safe to blood cells. These results evidence the therapeutic nature of the biocompatible Bi2MoO6/NH2-GO/PEG, proposing it as an efficient anticancer nanocarrier for drug delivery and other biomedical application purposes.

Graphene quantum dots based MnFe2O4@SiO2 magnetic nanostructure as a pH-sensitive fluorescence resonance energy transfer (FRET) system to enhance the anticancer effect of the drug.

Among the various methods of targeted drug delivery, magnetic nanoparticles been considered for a long time due to local drug delivery, reduction of side effects, and controlled drug release. In this work, fluorescence resonance energy transfer (FRET) system MnFe2O4@SiO2@ graphene quantum dots /DAU with 28.02 emu g-1 magnetism was prepared as pH-sensitive nanoplatform to enhance the anti-cancer effect of daunorubicin (DAU) drug (in the obtained FRET system, DAU act as acceptor molecule and graphene quantum dots act as donor molecule). The efficiency of the drug loaded on the nanoplatform in vitro is 78 %. DAU drug release from nanoplatform at pHs of 7.4 and 5.5 during 48 h is 21 % and 60 %, respectively. Release sensitive to pH facilitates the application of prepared nanoplatform for DAU delivery. The results of MTT-assay and annexin V-FITC/PI show that MnFe2O4@SiO2@ graphene quantum dots /DAU induces cell apoptosis by inhibiting the growth of more than 95 % of MCF-7 cells. Also, according to the results, it was found that MnFe2O4@SiO2@ graphene quantum dots /DAU can inhibit 66.65 % cell cycle in the sub-G1 phase. Therefore, due to the anti-cancer activity of MnFe2O4@SiO2@ graphene quantum dots /DAU, this biological nanoscale can be considered a candidate for drug delivery.

The effects of intermolecular interactions on the stability and in vitro drug release of daunorubicin/cytarabine co-loaded liposome.

Various studies were performed on the intermolecular interactions of daunorubicin (DNR) and cytarabine (Ara-C) co-loaded liposome to predict and elucidate its stability and in vitro drug release behavior. Langmuir monolayer and spectroscopy studies showed interactions between its components. The Langmuir monolayer study and blank liposomes stability study illustrated that interactions between lipids could affect their stability, and the DSPC/DSPG/Chol (7/2/1, mol%) mixed system tended to be thermodynamically and physicochemically stable. The interactions between daunorubicin and copper ions were then investigated by ultraviolet-visible (UV-vis) electronic absorption spectroscopy and circular dichroism (CD) spectroscopy, which revealed that the DNR-Cu complex was composed of daunorubicin and copper ions at a molar ratio of 1:1 or 1:2, and its solubility was related to the acidity of the solution. In vitro release experiment of liposomes with different copper gluconate contents illustrated that the interactions between drugs and copper ions were conducive to the retention and synergetic release of drugs. The stability and release studies of the DSPC/DSPG/Chol (7/2/1, mol%) co-loaded liposome illustrated that it had good storage and plasma stability, and the release behaviors of drugs were pH-related, i.e., drugs could be released faster under acidic condition. These studies indicated that intermolecular interactions could affect the stability and release behavior of the liposome, and a certain ratio of components could be conducive to its stability and synergistic release of drugs.

Development and Biochemical Characterization of Self-Immolative Linker Containing GnRH-III-Drug Conjugates.

The human gonadotropin releasing hormone (GnRH-I) and its sea lamprey analogue GnRH-III specifically bind to GnRH receptors on cancer cells and can be used as targeting moieties for targeted tumor therapy. Considering that the selective release of drugs in cancer cells is of high relevance, we were encouraged to develop cleavable, self-immolative GnRH-III-drug conjugates which consist of a p-aminobenzyloxycarbonlyl (PABC) spacer between a cathepsin B-cleavable dipeptide (Val-Ala, Val-Cit) and the classical anticancer drugs daunorubicin (Dau) and paclitaxel (PTX). Alongside these compounds, non-cleavable GnRH-III-drug conjugates were also synthesized, and all compounds were analyzed for their antiproliferative activity. The cleavable GnRH-III bioconjugates revealed a growth inhibitory effect on GnRH receptor-expressing A2780 ovarian cancer cells, while their activity was reduced on Panc-1 pancreatic cancer cells exhibiting a lower GnRH receptor level. Moreover, the antiproliferative activity of the non-cleavable counterparts was strongly reduced. Additionally, the efficient cleavage of the Val-Ala linker and the subsequent release of the drugs could be verified by lysosomal degradation studies, while radioligand binding studies ensured that the GnRH-III-drug conjugates bound to the GnRH receptor with high affinity. Our results underline the high value of GnRH-III-based homing devices and the application of cathepsin B-cleavable linker systems for the development of small molecule drug conjugates (SMDCs).

Adjusting the Structure of ß-Cyclodextrin to Improve Complexation of Anthraquinone-Derived Drugs.

ß-Cyclodextrin (CD) derivatives containing an aromatic triazole ring were studied as potential carriers of the following drugs containing an anthraquinone moiety: anthraquinone-2-sulfonic acid (AQ2S); anthraquinone-2-carboxylic acid (AQ2CA); and a common anthracycline, daunorubicin (DNR). UV-Vis and voltammetry measurements were carried out to determine the solubilities and association constants of the complexes formed, and the results revealed the unique properties of the chosen CDs as effective pH-dependent drug complexing agents. The association constants of the drug complexes with the CDs containing a triazole and lipoic acid (ßCDLip) or galactosamine (ßCDGAL), were significantly larger than that of the native ßCD. The AQ2CA and AQ2S drugs were poorly soluble, and their solubilities increased as a result of complex formation with ßCDLip and ßCDGAL ligands. AQ2CA and AQ2S are negatively charged at pH 7.4. Therefore, they were less prone to form an inclusion complex with the hydrophobic CD cavity than at pH 3 (characteristic of gastric juices) when protonated. The ßCDTriazole and ßCDGAL ligands were found to form weaker inclusion complexes with the positively charged drug DNR at an acidic pH (pH 5.5) than in a neutral medium (pH 7.4) in which the drug dissociates to its neutral, uncharged form. This pH dependence is favorable for antitumor applications.

Zwitterion-functionalized hollow mesoporous Prussian blue nanoparticles for targeted and synergetic chemo-photothermal treatment of acute myeloid leukemia.

Multifunctional drug delivery systems combining two or more therapies have a wide-range of potential for high efficacy tumor treatment. Herein, we designed a novel hollow mesoporous Prussian blue nanoparticles (HMPBs)-based platform for targeted and synergetic chemo-photothermal treatment of acute myeloid leukemia (AML). The HMPBs were first loaded with the anticancer drugs daunorubicin (DNR) and cytarabine (AraC), and were subsequently coated with polyethylenimine (PEI) through electrostatic adsorption. Then, zwitterionic sulfobetaine (ZS) and CXCR4 antagonist peptide E5 were modified onto the surface of the nanoparticles via covalent bonding to fabricate a nanoplatform (denoted as HMPBs(DNR + AraC)@PEI-ZS-E5). The nanoplatform showed excellent photothermal effects, superior photothermal stability, reduced nonspecific protein adsorption, efficient targeting capability, a constant hydrodynamic diameter and good biocompatibility. Additionally, a laser-responsive drug release pattern was observed. In vitro results indicated that the nanoplatform could achieve active targeting and remarkable chemo-photothermal synergetic therapeutic effects, showcasing its great potential in AML treatment.

Structural Characterization of Daunomycin-Peptide Conjugates by Various Tandem Mass Spectrometric Techniques.

The use of peptide-drug conjugates has generated wide interest as targeted antitumor therapeutics. The anthracycline antibiotic, daunomycin, is a widely used anticancer agent and it is often conjugated to different tumor homing peptides. However, comprehensive analytical characterization of these conjugates via tandem mass spectrometry (MS/MS) is challenging due to the lability of the O-glycosidic bond and the appearance of MS/MS fragment ions with little structural information. Therefore, we aimed to investigate the optimal fragmentation conditions that suppress the prevalent dissociation of the anthracycline drug and provide good sequence coverage. In this study, we comprehensively compared the performance of common fragmentation techniques, such as higher energy collisional dissociation (HCD), electron transfer dissociation (ETD), electron-transfer higher energy collisional dissociation (EThcD) and matrix-assisted laser desorption/ionization-tandem time-of-flight (MALDI-TOF/TOF) activation methods for the structural identification of synthetic daunomycin-peptide conjugates by high-resolution tandem mass spectrometry. Our results showed that peptide backbone fragmentation was inhibited by applying electron-based dissociation methods to conjugates, most possibly due to the "electron predator" effect of the daunomycin. We found that efficient HCD fragmentation was largely influenced by several factors, such as amino acid sequences, charge states and HCD energy. High energy HCD and MALDI-TOF/TOF combined with collision induced dissociation (CID) mode are the methods of choice to unambiguously assign the sequence, localize different conjugation sites and differentiate conjugate isomers.

Cancer-Targeted Controlled Delivery of Chemotherapeutic Anthracycline Derivatives Using Apoferritin Nanocage Carriers.

The interactions of chemotherapeutic drugs with nanocage protein apoferritin (APO) are the key features in the effective encapsulation and release of highly toxic drugs in APO-based controlled drug delivery systems. The encapsulation enables mitigating the drugs' side effects, collateral damage to healthy cells, and adverse immune reactions. Herein, the interactions of anthracycline drugs with APO were studied to assess the effect of drug lipophilicity on their encapsulation excess n and in vitro activity. Anthracycline drugs, including doxorubicin (DOX), epirubicin (EPI), daunorubicin (DAU), and idarubicin (IDA), with lipophilicity P from 0.8 to 15, were investigated. We have found that in addition to hydrogen-bonded supramolecular ensemble formation with n = 24, there are two other competing contributions that enable increasing n under strong polar interactions (APO(DOX)) or under strong hydrophobic interactions (APO(IDA) of the highest efficacy). The encapsulation/release processes were investigated using UV-Vis, fluorescence, circular dichroism, and FTIR spectroscopies. The in vitro cytotoxicity/growth inhibition tests and flow cytometry corroborate high apoptotic activity of APO(drugs) against targeted MDA-MB-231 adenocarcinoma and HeLa cells, and low activity against healthy MCF10A cells, demonstrating targeting ability of nanodrugs. A model for molecular interactions between anthracyclines and APO nanocarriers was developed, and the relationships derived compared with experimental results.

Lysine 268 adjacent to transmembrane helix 5 of hamster P-glycoprotein is the major photobinding site of iodomycin in CHO B30 cells.

P-glycoprotein (Pgp) detoxifies cells by exporting hundreds of chemically dissimilar hydrophobic and amphipathic compounds and is implicated in multidrug resistance (MDR) in the treatment of cancers. Photoaffinity labeling of plasma membrane vesicles of MDR CHO B30 cells with the anthracycline [125 I]-iodomycin, subsequent sequential cleavage with BNPS-skatol and endoproteinase Lys-C, and the Edman sequencing of the purified photoaffinity-labeled peptide identified the lysine residue at position 268 in the hamster Pgp primary sequence as the major photobinding site of iodomycin in CHO B30 cells. Lysine 268 is located adjacent to the cytosolic terminus of transmembrane 5. According to thermodynamic and kinetic analyses, this location should present the equilibrium binding site of ATP-free Pgp for daunomycin and iodomycin in B30 cells.

Comprehensive review for anticancer hybridized multitargeting HDAC inhibitors.

Despite the encouraging clinical progress of chemotherapeutic agents in cancer treatment, innovation and development of new effective anticancer candidates still represents a challenging endeavor. With 15 million death every year in 2030 according to the estimates, cancer has increased rising of an alarm as a real crisis for public health and health systems worldwide. Therefore, scientist began to introduce innovative solutions to control the cancer global health problem. One of the promising strategies in this issue is the multitarget or smart hybrids having two or more pharmacophores targeting cancer. These rationalized hybrid molecules have gained great interests in cancer treatment as they are capable to simultaneously inhibit more than cancer pathway or target without drug-drug interactions and with less side effects. A prime important example of these hybrids, the HDAC hybrid inhibitors or referred as multitargeting HDAC inhibitors. The ability of HDAC inhibitors to synergistically improve the efficacy of other anti-cancer drugs and moreover, the ease of HDAC inhibitors cap group modification prompt many medicinal chemists to innovate and develop new generation of HDAC hybrid inhibitors. Notably, and during this short period, there are four HDAC inhibitor hybrids have entered different phases of clinical trials for treatment of different types of blood and solid tumors, namely; CUDC-101, CUDC-907, Tinostamustine, and Domatinostat. This review shed light on the most recent hybrids of HDACIs with one or more other cancer target pharmacophore. The designed multitarget hybrids include topoisomerase inhibitors, kinase inhibitors, nitric oxide releasers, antiandrogens, FLT3 and JAC-2 inhibitors, PDE5-inhibitors, NAMPT-inhibitors, Protease inhibitors, BRD4-inhibitors and other targets. This review may help researchers in development and discovery of new horizons in cancer treatment.

Activation of Myosin Phosphatase by Epigallocatechin-Gallate Sensitizes THP-1 Leukemic Cells to Daunorubicin.

BACKGROUND: The Myosin Phosphatase (MP) holoenzyme is composed of a Protein Phosphatase type 1 (PP1) catalytic subunit and a regulatory subunit termed Myosin Phosphatase Target subunit 1 (MYPT1). Besides dephosphorylation of myosin, MP has been implicated in the control of cell proliferation via dephosphorylation and activation of the tumor suppressor gene products, retinoblastoma protein (pRb) and merlin. Inhibition of MP was shown to attenuate the drug-induced cell death of leukemic cells by chemotherapeutic agents, while activation of MP might have a sensitizing effect. OBJECTIVE: Recently, Epigallocatechin-Gallate (EGCG), a major component of green tea, was shown to activate MP by inducing the dephosphorylation of MYPT1 at phospho-Thr696 (MYPT1pT696), which might confer enhanced chemosensitivity to cancer cells. METHODS: THP-1 leukemic cells were treated with EGCG and Daunorubicin (DNR) and cell viability was analyzed. Phosphorylation of tumor suppressor proteins was detected by Western blotting. RESULTS: EGCG or DNR (at sub-lethal doses) alone had moderate effects on cell viability, while the combined treatment caused a significant decrease in the number of viable cells by enhancing apoptosis and decreasing proliferation. EGCG plus DNR decreased the phosphorylation level of MYPT1pT696, which was accompanied by prominent dephosphorylation of pRb. In addition, significant dephosphorylation of merlin was observed when EGCG and DNR were applied together. CONCLUSION: Our results suggest that EGCG-induced activation of MP might have a regulatory function in mediating the chemosensitivity of leukemic cells via dephosphorylation of tumor suppressor proteins.

Daunomycin delivery by ultrasmall graphene quantum dots to DNA duplexes: understanding the dynamics by resonance energy transfer.

Daunomycin (DN) is a natural product isolated from Streptomyces and it is widely used as a chemotherapeutic medication because of its antitumour properties. It is an anthracycline antibiotic that inhibits virus multiplication and shows activity against acute leukemia. This drug is either injected into a vein of the subject or typically delivered to cellular nuclei by polymeric or metallic nanoparticles and liposomal or proteinous substrates. The size of these delivering agents becomes a controlling factor affecting the interactions of this drug with nuclear DNA, where it intercalates. The fast-developing area of ultrasmall fluorescent graphene quantum dots (GQDs) provides a new platform for delivering DN, exploiting π-π stacking interactions between the planar anthracenyl moiety of the drug and the crystalline GQDs. The size of the GQDs might allow them to transport the medicine inside nuclei easily so that DN can intercalate between the DNA basal planes. Bringing DN close to the DNA duplex is essential, since the daunosamine residue of the drug binds to the minor groove, easing the intercalation of the anthracenyl moiety. The presented experimental analysis uses a fluorescence resonance energy transfer (FRET) mechanism to provide the details of the dynamics of the delivery of DN by GQDs and its subsequent uptake by the DNA duplex. The process is seen to be governed by differences in the binding constants and the non-interaction of the GQDs with DNA. This research provides vital input into understanding the mechanism of action of DN in greater detail, aiding its applicability in medical science.

Co-delivery of IR-768 and daunorubicin using mPEG-b-PLGA micelles for synergistic enhancement of combination therapy of melanoma.

Malignant melanoma is an emerging problem worldwide due to the high degree of lethalness. Its aggressiveness and the ability to metastasize along with the heterogeneity at the molecular and cellular levels, limit the overall therapeutic efficacy. Despite significant advances in melanoma treatment over the last decade, there is still a need for improved therapeutic modalities. Thus, we demonstrate here a combinatorial approach that targets multiple independent therapeutic pathways, in which polymeric micelles (PMs) were used as efficacious colloidal nanocarriers loaded with both daunorubicin (DRB) as a cytotoxic drug and IR-768 as a photosensitizer. This afforded the dual drug loaded delivery system IR-768 + DRB in PMs. The fabricated mPEG-b-PLGA micelles (hydrodynamic diameters ≈ 25 nm) had a relatively narrow size distribution (PdI > ca. 0.3) with uniform spherical shapes. CLSM study showed that mPEG-b-PLGA micelles were uptaken by mitochondria, which further contributed to excellent singlet oxygen generation capacity for PDT in A375 melanoma cells. Furthermore, the PMs were efficiently internalized by tested cells through endocytosis, resulting in much higher cellular uptake comparing to the free drug. As a result of these properties, IR-768 + DRB in PMs exhibited very potent and synergistically enhanced anticancer activity against A375 cells. Additionally, this combination approach allowed to reduce drug doses and provided low side effects towards normal HaCaT. This study indicates excellent properties of mPEG-b-PLGA micelles resulting in great therapeutic potential possessed by the developed nanoscale drug delivery system for combined chemo-photodynamic therapy of melanoma.

Profound Nanoscale Structural and Biomechanical Changes in DNA Helix upon Treatment with Anthracycline Drugs.

In our study, we describe the outcomes of the intercalation of different anthracycline antibiotics in double-stranded DNA at the nanoscale and single molecule level. Atomic force microscopy analysis revealed that intercalation results in significant elongation and thinning of dsDNA molecules. Additionally, using optical tweezers, we have shown that intercalation decreases the stiffness of DNA molecules, that results in greater susceptibility of dsDNA to break. Using DNA molecules with different GC/AT ratios, we checked whether anthracycline antibiotics show preference for GC-rich or AT-rich DNA fragments. We found that elongation, decrease in height and decrease in stiffness of dsDNA molecules was highest in GC-rich dsDNA, suggesting the preference of anthracycline antibiotics for GC pairs and GC-rich regions of DNA. This is important because such regions of genomes are enriched in DNA regulatory elements. By using three different anthracycline antibiotics, namely doxorubicin (DOX), epirubicin (EPI) and daunorubicin (DAU), we could compare their detrimental effects on DNA. Despite their analogical structure, anthracyclines differ in their effects on DNA molecules and GC-rich region preference. DOX had the strongest overall effect on the DNA topology, causing the largest elongation and decrease in height. On the other hand, EPI has the lowest preference for GC-rich dsDNA. Moreover, we demonstrated that the nanoscale perturbations in dsDNA topology are reflected by changes in the microscale properties of the cell, as even short exposition to doxorubicin resulted in an increase in nuclei stiffness, which can be due to aberration of the chromatin organization, upon intercalation of doxorubicin molecules.

Amphiphilic drug-peptide-polymer conjugates based on poly(ethylene glycol) and hyperbranched polyglycerol for epidermal growth factor receptor targeting: the effect of conjugate aggregation on in vitro activity.

Numerous peptide-drug conjugates have been developed over the years to enhance the specificity and selectivity of chemotherapeutic agents for tumour cells. In our present work, epidermal growth factor receptor targeting drug-peptide conjugates were prepared using GE11 and D4 peptides. To ensure the drug release, the cathepsin B labile GFLG spacer was incorporated between the targeting peptide and the drug molecule (daunomycin), which significantly increased the hydrophobicity and thereby decreased the water solubility of the conjugates. To overcome the solubility problem, drug-peptide-polymer conjugates with systematic structural variations were prepared, by linking poly(ethylene glycol) (PEG) or a well-defined amino-monofunctional hyperbranched polyglycerol (HbPG) directly or via a pentaglycine spacer to the targeting peptides. All the drug-peptide-polymer conjugates were water-soluble as confirmed by turbidimetric measurements. The results of the in vitro cell viability and cellular uptake measurements on HT-29 human colon adenocarcinoma cells proved that the HbPG and the PEG highly influenced the biological activity. The conjugation of the hydrophilic polymer resulted in the amphiphilic character of the conjugates, which led to self-aggregation and nanoparticle formation that decreased the cellular uptake above a specific aggregation concentration. On the other hand, the hydrodynamic volume and the different polymer chain topology of the linear PEG and the compact hyperbranched HbPG also played an important role in the biological activity. Therefore, in similar systems, the investigation of the colloidal properties is inevitable for the better understanding of the biological activity, which can reveal the structure-activity relationship of amphiphilic drug-peptide-polymer conjugates for efficient tumour targeting.