Selective Anticervical Cancer Injectable and Self-Healable Hydrogel Platforms Constructed of Drug-Loaded Cross-Linkable Unimolecular Micelles in a Single and Combination Therapy.

In the face of severe side effects of systemic chemotherapy used in cervical cancer, topical selective drug carriers with long-lasting effects are being sought. Hydrogels are suitable platforms, but their use is problematic in the case of delivery of hydrophobic drugs with anticancer activity. Herein, hydrogels constructed of unimolecular micelles displaying enhanced solubilization of aromatic lipophilic bioactive compounds are presented. Star-shaped poly(benzyl glycidyl ether)-block-poly(glycidyl glycerol ether) with an aryl-enriched core show high encapsulation capacity of poor water-soluble nifuratel and clotrimazole. Nifuratel attained selectivity against cervical cancer cells, whereas clotrimazole preserved its original selectivity. The combination of unimolecular micelles loaded with both drugs provided synergism; however, they were still selective against cervical cancer cells. The cross-linking of drug-loaded unimolecular micelles via dynamic boronic esters provided injectable and self-healable hydrogel drug carriers also displaying synergistic anticancer activity, suitable for intravaginal administration and assuring the effective coverage of the afflicted tissue area and efficient tissue permeability with hydrophobic bioactive compounds. Here, we show that the combination of star-shaped polyether amphiphiles and boronic ester cross-linking chemistry provides a new strategy for obtaining hydrogel platforms suitable for efficient hydrophobic drug delivery.

Advancing Antiamyloidogenic Activity by Fine-Tuning Macromolecular Topology.

Amyloid ß peptide can aggregate into thin ß-sheet fibrils or plaques deposited on the extracellular matrix, which is the hallmark of Alzheimer's disease. Multifunctional macromolecular structures play an important role in inhibiting the aggregate formation of amyloidogenic materials and thus are promising candidates with antiamyloidogenic characteristics for the development of next-generation therapeutics. In this study, we evaluate how small differences in the dendritic topology of these structures influence their antiamyloidogenic activity by the comparison of "perfectly dendritic" and "pseudodendritic" macromolecules, both decorated with mannose units. Their compactness, the position of surface units, and the size of glyco-architectures influence their antiamyloidogenic activity against Aß 40, a major component of amyloid plaques. For the advanced analysis of the aggregation of the Aß peptide, we introduce asymmetric flow field flow fractionation as a suitable method for the quantification of large and delicate structures. This alternative method focuses on the quantification of complex aggregates of Aß 40 and glycodendrimer/glyco-pseudodendrimer over different time intervals of incubation, showing a good correlation to ThT assay and CD spectroscopy results. Kinetic studies of the second-generation glyco-pseudodendrimer revealed maximum inhibition of Aß 40 aggregates, verified with atomic force microscopy. The second-generation glyco-pseudodendrimer shows the best antiamyloidogenic properties confirming that macromolecular conformation in combination with optimal functional group distribution is the key to its performance. These molecular properties were validated and confirmed by molecular dynamics simulation.

Dendrimersomes: Biomedical applications.

In recent years, dendrimer-based vesicles, known as dendrimersomes, have garnered significant attention as highly promising alternatives to lipid vesicles in a variety of biomedical applications. Dendrimersomes offer several advantages, including relatively straightforward synthesis, non-immunogenic properties, stability in circulation, and minimal size variability. These vesicles are composed of Janus dendrimers, which are polymers characterized by two dendritic wedges with different terminal groups - hydrophilic and hydrophobic. This dendrimer structure enables the self-assembly of dendrimersomes. The purpose of this highlight is to provide an overview of recent advancements achieved through the utilization of biomimetic dendrimersomes in various biomedical applications such as drug and nucleic acid delivery.

Is acriflavine an efficient co-drug in chemotherapy?

Cancer is a global health problem being the second worldwide cause of deaths right after cardiovascular diseases. The main methods of cancer treatment involve surgery, radiation and chemotherapy with an emphasis on the latter. Thus development of nanochemistry and nanomedicine in a search for more effective and safer cancer treatment is an important area of current research. Below, we present interaction of doxorubicin and acriflavine and the cytotoxicity of these drug nano-complexes towards cervical cancer (HeLa) cells. Experimental results obtained from NMR measurements and fluorescence spectroscopy show that the drugs' interaction was due to van der Waals forces, formation of hydrogen bonds and π-π stacking. Quantum molecular simulations confirmed the experimental results with regard to existing π-π stacking. Additionally it was shown that, at the level of theory applied (DFT, triple zeta basis set), the stacking interactions comprise the most preferable interactions (the lowest ΔG ca. -12 kcal mol-1) both between the molecules forming the acriflavine system and between the other component - another drug (doxorubicin) dimer. Biological tests performed on HeLa cells showed high cytotoxicity of the complexes, comparable to free drugs (ACF and DOX), both after 24 and 48 hours of incubation. For non-cancerous cells, a statistically significant difference in the cytotoxicity of drugs and complexes was observed in the case of a short incubation period. The results of the uptake study showed significantly more efficient cellular uptake of acriflavine than doxorubicin, whether administered alone or in combination with an anthracycline. The mechanism determining the selective uptake of acriflavine and ACF : DOX complexes towards non-cancer and cancer cells should be better understood in the future, as it may be of key importance in the design of complexes with toxic anti-cancer drugs.

Cross-linkable star-hyperbranched unimolecular micelles for the enhancement of the anticancer activity of clotrimazole.

Clotrimazole, a hydrophobic drug routinely used in the treatment of vaginal candidiasis, also shows antitumor activity. However, its use in chemotherapy has been unsuccessful to date due to its low solubility in aqueous media. In this work, new unimolecular micelles based on polyether star-hyperbranched carriers of clotrimazole are presented that can enhance solubility, and consequently the bioavailability, of clotrimazole in water. The amphiphilic constructs consisting of a hydrophobic poly(n-alkyl epoxide) core and hydrophilic corona of hyperbranched polyglycidol were synthesized in a three-step anionic ring-opening polymerization of epoxy monomers. The synthesis of such copolymers, however, was only possible by incorporating a linker to facilitate the elongation of the hydrophobic core with glycidol. Unimolecular micelles-clotrimazole formulations displayed significantly increased activity against human cervical cancer HeLa cells compared to the free drug, along with a weak effect on the viability of the normal dermal microvascular endothelium cells HMEC1. This selective activity of clotrimazole on cancer cells with little effect on normal cells was a result of the fact that clotrimazole targets the Warburg effect in cancer cells. Flow cytometric analysis revealed that the encapsulated clotrimazole significantly blocks the progression of the HeLa cycle in the G0/G1 phase and induces apoptosis. In addition, the ability of the synthesized amphiphilic constructs to form a dynamic hydrogel was demonstrated. Such a gel facilitates the delivery of drug-loaded single-molecule micelles to the affected area, where they can form a continuous, self-healing layer.

Self-Healable, Injectable Hydrogel with Enhanced Clotrimazole Solubilization as a Potential Therapeutic Platform for Gynecology.

Injectable, self-healing hydrogels with enhanced solubilization of hydrophobic drugs are urgently needed for antimicrobial intravaginal therapies. Here, we report the first hydrogel systems constructed of dynamic boronic esters cross-linking unimolecular micelles, which are a reservoir of antifungal hydrophobic drug molecules. The selective hydrophobization of hyperbranched polyglycidol with phenyl units in the core via ester or urethane bonds enabled the solubilization of clotrimazole, a water-insoluble drug of broad antifungal properties. The encapsulation efficiency of clotrimazole increases with the degree of the HbPGL core modification; however, the encapsulation is more favorable in the case of urethane derivatives. In addition, the rate of clotrimazole release was lower from HbPGL hydrophobized via urethane bonds than with ester linkages. In this work, we also revealed that the hydrophobization degree of HbPGL significantly influences the rheological properties of its hydrogels with poly(acrylamide-ran-2-acrylamidephenylboronic acid). The elastic strength of networks (GN) and the thermal stability of hydrogels increased along with the degree of HbPGL core hydrophobization. The degradation of the hydrogel constructed of the neat HbPGL was observed at approx. 40 °C, whereas the hydrogels constructed on HbPGL, where the monohydroxyl units were modified above 30 mol %, were stable above 50 °C. Moreover, the flow and self-healing ability of hydrogels were gradually decreased due to the reduced dynamics of macromolecules in the network as an effect of increased hydrophobicity. The changes in the rheological properties of hydrogels resulted from the engagement of phenyl units into the intermolecular hydrophobic interactions, which besides boronic esters constituted additional cross-links. This study demonstrates that the HbPGL core hydrophobized with phenyl units at 30 mol % degrees via urethane linkages is optimal in respect of the drug encapsulation efficiency and rheological properties including both self-healable and injectable behavior. This work is important because of a proper selection of a building component for the construction of a therapeutic hydrogel platform dedicated to the intravaginal delivery of hydrophobic drugs.

Star-Shaped Poly(furfuryl glycidyl ether)-Block-Poly(glyceryl glycerol ether) as an Efficient Agent for the Enhancement of Nifuratel Solubility and for the Formation of Injectable and Self-Healable Hydrogel Platforms for the Gynaecological Therapies.

In this paper, we present novel well-defined unimolecular micelles constructed a on poly(furfuryl glycidyl ether) core and highly hydrophilic poly(glyceryl glycerol ether) shell, PFGE-b-PGGE. The copolymer was synthesized via anionic ring-opening polymerization of furfuryl glycidyl ether and (1,2-isopropylidene glyceryl) glycidyl ether, respectively. MTT assay revealed that the copolymer is non-cytotoxic against human cervical cancer endothelial (HeLa) cells. The copolymer thanks to furan moieties in its core is capable of encapsulation of nifuratel, a hydrophobic nitrofuran derivative, which is a drug applied in the gynaecology therapies that shows a broad antimicroorganism spectrum. The study shows high loading capacity of the copolymer, i.e., 146 mg of nifuratel per 1 g of copolymer. The load unimolecular micelles were characterized using DLS and TEM microscopy and compared with the reference glyceryl glycerol ether homopolymer sample. The presence of numerous 1,2-diol moieties in the shell of PFGE-b-PGG macromolecules enabled the formation of reversible cross-links with 2-acrylamidephenylboronic acid-based polyacrylamide. The obtained hydrogels were both injectable and self-healable, which was confirmed with a rheological study.

Recent Advances in Preclinical Research Using PAMAM Dendrimers for Cancer Gene Therapy.

Carriers of genetic material are divided into vectors of viral and non-viral origin. Viral carriers are already successfully used in experimental gene therapies, but despite advantages such as their high transfection efficiency and the wide knowledge of their practical potential, the remaining disadvantages, namely, their low capacity and complex manufacturing process, based on biological systems, are major limitations prior to their broad implementation in the clinical setting. The application of non-viral carriers in gene therapy is one of the available approaches. Poly(amidoamine) (PAMAM) dendrimers are repetitively branched, three-dimensional molecules, made of amide and amine subunits, possessing unique physiochemical properties. Surface and internal modifications improve their physicochemical properties, enabling the increase in cellular specificity and transfection efficiency and a reduction in cytotoxicity toward healthy cells. During the last 10 years of research on PAMAM dendrimers, three modification strategies have commonly been used: (1) surface modification with functional groups; (2) hybrid vector formation; (3) creation of supramolecular self-assemblies. This review describes and summarizes recent studies exploring the development of PAMAM dendrimers in anticancer gene therapies, evaluating the advantages and disadvantages of the modification approaches and the nanomedicine regulatory issues preventing their translation into the clinical setting, and highlighting important areas for further development and possible steps that seem promising in terms of development of PAMAM as a carrier of genetic material.

Synthesis and Shaping of Core-Shell Tecto Dendrimers for Biomedical Applications.

In recent years, the use of poly(amidoamine) (PAMAM) dendrimers of different generations as building blocks or reactive modules to construct core-shell tecto dendrimers (CSTDs) that are superior to the performance of single-generation dendrimers has received great attention in the field of biomedical applications. The CSTDs are always based on high-generation dendrimers as the core and low-generation dendrimers as the shell; not only do they have excellent properties similar to single high-generation dendrimers, but they also have overcome some of the shortcomings (e.g., limited drug loading capacity or enhanced permeability and retention effect due to small size) of single-generation dendrimers in biomedical applications. Herein, the recent advances of CSTDs synthesized by different approaches as nanoplatforms for different biomedical applications, particularly for chemotherapy, gene delivery, and combination therapy, as well as biological imaging, are summarized. In addition, the current challenges and future perspectives of CSTDs are also discussed.

Hydrophilic Polyhedral Oligomeric Silsesquioxane, POSS(OH)32, as a Complexing Nanocarrier for Doxorubicin and Daunorubicin.

A novel strategy, recently developed by us, to use polyhedral oligomeric silsesquioxanes (POSS) as an anti-cancer drug carrier is presented. Anthracycline:POSS complexes were prepared by simple co-addition of doxorubicin (DOX) or daunorubicin (DAU) with hydrophilic POSS(OH)32. Co-delivery of POSS and anthracyclines led to higher anti-cancer activity towards HeLa (cervical cancer endothelial) and MCF-7 (human breast adenocarcinoma) cell lines. The obtained supramolecular hybrid complexes were characterised by nuclear magnetic resonance (NMR) spectroscopy (nuclear Overhauser effect spectroscopy [NOESY] and homonuclear correlation spectroscopy [COSY]), Fourier transform infrared spectroscopy (FTIR), and dynamic light scattering (DLS). The two-dimensional (2D) NOESY spectra of the complexes showed the cross-correlation peaks for hydroxyl groups of POSS (~4.3-4.8 ppm) with OH groups of DOX and DAU. FTIR showed that hydroxyl group of POSS can interact with amine and hydroxyl groups of DOX and DAU. The viability of HeLa and MCF-7 was analysed with the MTT assay to evaluate the cytotoxicity of free DOX and DAU and the relevant complexes with POSS at different molar ratios. At a low DOX concentration (2.5 µM), for molar ratios 1:1, 1:4, and 1:8 (POSS:DOX), the complexes showed two and three times higher cytotoxicity towards HeLa and MCF-7 cells, respectively, than DOX itself after both 24- and 48-h incubation. The 1 µM concentration for a 1:4 POSS:DOX molecular ratio and the 2.5 µM concentration for all complexes were more toxic towards MCF-7 cells than free DOX after 48-h incubation. In the case of POSS:DAU complexes, there was higher toxicity than that of free drug after 48-h incubation. It can be concluded that the formation of non-covalent complexes increases toxicity of anthracycline drugs towards Hela and MCF-7 cells. The novel complexes are inexpensive to prepare and more effective than free drugs at low systemic toxicity.

Influence of Free Fatty Acids on Lipid Membrane-Nisin Interaction.

The influence of free fatty acids (FFAs) on the nisin-membrane interaction was investigated through micro-DSC and fluorescence spectroscopy. A simple but informative model membrane was prepared (5.7 DMPC:3.8 DPPS:0.5 DOPC molar ratio) by considering the presence of different phospholipid headgroups in charge and size and different phospholipid tails in length and unsaturation level, allowing the discrimination of the combined interaction of nisin and FFAs with the single phospholipid constituents. The effects of six FFAs on membrane stability were evaluated, namely two saturated FFAs (palmitic acid and stearic acid), two monounsaturated FFAs (cis-unsaturated oleic acid and trans-unsaturated elaidic acid) and two cis-polyunsaturated FFAs (ω-6 linoleic acid and ω-3 docosahexaenoic acid). The results permitted assessment of a thermodynamic picture of such interactions which indicates that the peptide-membrane interaction does not overlook the presence of FFAs within the lipid bilayer since both FFAs and nisin are able to selectively promote thermodynamic phase separations as well as a general lipid reorganization within the host membrane. Furthermore, the magnitude of the effects may be different depending on the FFA chemical structure as well as the membrane lipid composition.

In Search of a Phosphorus Dendrimer-Based Carrier of Rose Bengal: Tyramine Linker Limits Fluorescent and Phototoxic Properties of a Photosensitizer.

Photodynamic therapy (PDT) is a skin cancer treatment alternative to chemotherapy and radiotherapy. This method exploits three elements: a phototoxic compound (photosensitizer), light source and oxygen. Upon irradiation by light of a specific wavelength, the photosensitizer generates reactive oxygen species triggering the cascade of reactions leading to cell death. The positive therapeutic effect of PDT may be limited due to low solubility, low tumor specificity and inefficient cellular uptake of photosensitizers. A promising approach to overcome these obstacles involves the use of nanocarrier systems. The aim of this initial study was to determine the potential of the application of phosphorus dendrimers as carriers of a photosensitizer-rose bengal (RB). The primary goal involved the synthesis and in vitro studies of covalent drug-dendrimer conjugates. Our approach allowed us to obtain RB-dendrimer conjugates with the use of tyramine as an aromatic linker between the carrier and the drug. The compounds were characterized by FT-IR, 1H NMR, 13C NMR, 31P NMR, size and zeta potential measurements and spectrofluorimetric analysis. The dialysis to check the drug release from the conjugate, flow cytometry to specify intracellular uptake, and singlet oxygen generation assay were also applied. Finally, we used MTT assay to determine the biological activity of the tested compounds. The results of our experiments indicate that the conjugation of RB to phosphorus dendrimers via the tyramine linker decreases photodynamic activity of RB.

Synthesis, Characterization, and In Vitro Studies of an Reactive Oxygen Species (ROS)-Responsive Methoxy Polyethylene Glycol-Thioketal-Melphalan Prodrug for Glioblastoma Treatment.

Glioblastoma (GBM) is the most frequent and aggressive primary tumor of the brain and averages a life expectancy in diagnosed patients of only 15 months. Hence, more effective therapies against this malignancy are urgently needed. Several diseases, including cancer, are featured by high levels of reactive oxygen species (ROS), which are possible GBM hallmarks to target or benefit from. Therefore, the covalent linkage of drugs to ROS-responsive molecules can be exploited aiming for a selective drug release within relevant pathological environments. In this work, we designed a new ROS-responsive prodrug by using Melphalan (MPH) covalently coupled with methoxy polyethylene glycol (mPEG) through a ROS-cleavable group thioketal (TK), demonstrating the capacity to self-assembly into nanosized micelles. Full chemical-physical characterization was conducted on the polymeric-prodrug and proper controls, along with in vitro cytotoxicity assayed on different GBM cell lines and "healthy" astrocyte cells confirming the absence of any cytotoxicity of the prodrug on healthy cells (i.e. astrocytes). These results were compared with the non-ROS responsive counterpart, underlining the anti-tumoral activity of ROS-responsive compared to the non-ROS-responsive prodrug on GBM cells expressing high levels of ROS. On the other hand, the combination treatment with this ROS-responsive prodrug and X-ray irradiation on human GBM cells resulted in an increase of the antitumoral effect, and this might be connected to radiotherapy. Hence, these results represent a starting point for a rationale design of innovative and tailored ROS-responsive prodrugs to be used in GBM therapy and in combination with radiotherapy.

Poly(lysine) Dendrimers Form Complexes with siRNA and Provide Its Efficient Uptake by Myeloid Cells: Model Studies for Therapeutic Nucleic Acid Delivery.

The disruption of the cellular pathways of protein biosynthesis through the mechanism of RNA interference has been recognized as a tool of great diagnostic and therapeutic significance. However, in order to fully exploit the potential of this phenomenon, efficient and safe carriers capable of overcoming extra- and intracellular barriers and delivering siRNA to the target cells are needed. Recently, attention has focused on the possibility of the application of multifunctional nanoparticles, dendrimers, as potential delivery devices for siRNA. The aim of the present work was to evaluate the formation of dendriplexes using novel poly(lysine) dendrimers (containing lysine and arginine or histidine residues in their structure), and to verify the hypothesis that the use of these polymers may allow an efficient method of siRNA transfer into the cells in vitro to be obtained. The fluorescence polarization studies, as well as zeta potential and hydrodynamic diameter measurements were used to characterize the dendrimer:siRNA complexes. The cytotoxicity of dendrimers and dendriplexes was evaluated with the resazurin-based assay. Using the flow cytometry technique, the efficiency of siRNA transport to the myeloid cells was determined. This approach allowed us to determine the properties and optimal molar ratios of dendrimer:siRNA complexes, as well as to demonstrate that poly(lysine) dendrimers may serve as efficient carriers of genetic material, being much more effective than the commercially available transfection agent Lipofectamine 2000. This outcome provides the basis for further research on the application of poly(lysine) dendrimers as carriers for nucleic acids in the field of gene therapy.

Application of new lysine-based peptide dendrimers D3K2 and D3G2 for gene delivery: Specific cytotoxicity to cancer cells and transfection in vitro.

In order to enhance intracellular uptake and accumulation of therapeutic nucleic acids for improved gene therapy methods, numerous delivery vectors have been elaborated. Based on their origin, gene carriers are generally classified as viral or non-viral vectors. Due to their significantly reduced immunogenicity and highly optimized methods of synthesis, nanoparticles (especially those imitating natural biomolecules) constitute a promising alternative for virus-based delivery devices. Thus, we set out to develop innovative peptide dendrimers for clinical application as transfection agents and gene carriers. In the present work we describe the synthesis of two novel lysine-based dendritic macromolecules (D3K2 and D3G2) and their initial characterization for cytotoxicity/genotoxicity and transfection potential in two human cell line models: cervix adenocarcinoma (HeLa) and microvascular endothelial (HMEC-1). This approach allowed us to identify more cationic D3K2 as potent delivery agent, being able to increase intracellular accumulation of large nucleic acid molecules such as plasmids. Moreover, the dendrimers exhibited specific cytotoxicity towards cancer cell line without showing significant toxic effects on normal cells. These observations are promising prognosis for future clinical application of this type of nanoparticles.

Molecular Mechanisms of Antitumor Activity of PAMAM Dendrimer Conjugates with Anticancer Drugs and a Monoclonal Antibody.

Taxanes are considered fundamental drugs in the treatment of breast cancer, but despite the similarities, docetaxel (doc) and paclitaxel (ptx) work differently. For this reason, it is interesting to identify mechanisms of antitumor activity of PAMAM dendrimer conjugates that carry docetaxel or paclitaxel and monoclonal antibody trastuzumab, specifically targeted to cells which overexpressed HER-2. For this purpose, the impact on the level of reactive oxygen species, the mitochondrial membrane potential, cell cycle distribution and the activity of caspases-3/7, -8 and -9 of PAMAM-doc-trastuzumab and PAMAM-ptx-trastuzumab conjugates was determined and compared with free docetaxel and paclitaxel toward HER-2-positive (SKBR-3) and negative (MCF-7) human breast cancer cell lines. Moreover, apoptosis and necrosis were studied using flow cytometry and confocal microscopy, respectively. Our studies show the complexity of the potential mechanism of cytotoxic action of PAMAM-drug-trastuzumab conjugates that should be sought as a resultant of oxidative stress, mitochondrial activation of the caspase cascade and the HER-2 receptor blockade.

Multicomponent Conjugates of Anticancer Drugs and Monoclonal Antibody with PAMAM Dendrimers to Increase Efficacy of HER-2 Positive Breast Cancer Therapy.

PURPOSE: Conjugation of nanocarriers with antibodies that bind to specific membrane receptors that are overexpressed in cancer cells enables targeted delivery. In the present study, we developed and synthesised two PAMAM dendrimer-trastuzumab conjugates that carried docetaxel or paclitaxel, specifically targeted to cells which overexpressed HER-2. METHODS: The 1H NMR, 13C NMR, FTIR and RP-HPLC were used to analyse the characteristics of the products and assess their purity. The toxicity of PAMAM-trastuzumab, PAMAM-doc-trastuzumab and PAMAM-ptx-trastuzumab conjugates was determined using MTT assay and compared with free trastuzumab, docetaxel and paclitaxel toward HER-2-positive (SKBR-3) and negative (MCF-7) human breast cancer cell lines. The cellular uptake and internal localisation were studied using flow cytometry and confocal microscopy, respectively. RESULTS: The PAMAM-drug-trastuzumab conjugates in particular showed extremely high toxicity toward the HER-2-positive SKBR-3 cells and very low toxicity towards to HER-2-negative MCF-7 cells. As expected, the HER-2-positive SKBR-3 cell line accumulated trastuzumab from both conjugates rapidly; but surprisingly, although a large amount of PAMAM-ptx-trastuzumab conjugate was observed in the HER-2-negative MCF-7 cells. Confocal microscopy confirmed the intracellular localisation of analysed compounds. The key result of fluorescent imaging was the identification of strong selective binding of the PAMAM-doc-trastuzumab conjugate with HER-2-positive SKBR-3 cells only. CONCLUSIONS: Our results confirm the high selectivity of PAMAM-doc-trastuzumab and PAMAM-ptx-trastuzumab conjugates for HER-2-positive cells, and demonstrate the utility of trastuzumab as a targeting agent. Therefore, the analysed conjugates present an promising approach for the improvement of efficacy of targeted delivery of anticancer drugs such as docetaxel or paclitaxel.

Sugar Modification Enhances Cytotoxic Activity of PAMAM-Doxorubicin Conjugate in Glucose-Deprived MCF-7 Cells - Possible Role of GLUT1 Transporter.

PURPOSE: In order to overcome the obstacles and side effects of classical chemotherapy, numerous studies have been performed to develop the treatment based on targeted transport of active compounds directly to the site of action. Since tumor cells are featured with intensified glucose metabolism, we set out to develop innovative, glucose-modified PAMAM dendrimer for the delivery of doxorubicin to breast cancer cells. METHODS: PAMAM-dox-glc conjugate was synthesized and characterized by 1H NMR, FT-IR, size and zeta potential measurements. The drug release rate from conjugate was evaluated by dialysis under different pH conditions. The expression level of GLUT family receptors in cells cultured in full and glucose-deprived medium was evaluated by quantitative real-time RT-PCR and flow cytometry. The cytotoxicity of conjugate in presence or absence of GLUT1 inhibitors was determined by MTT assay. RESULTS: We showed that PAMAM-dox-glc conjugate exhibits pH-dependent drug release and increased cytotoxic activity compared to free drug in cells cultured in medium without glucose. Further, we proved that these cells overexpress transporters of GLUT family. The toxic effect of conjugate was eliminated by the application of specific GLUT1 inhibitors. CONCLUSION: Our findings revealed that the glucose moiety plays a crucial role in the recognition of cells with high expression of GLUT receptors. By selectively blocking GLUT1 transporter we showed its importance for the cytotoxic activity of PAMAM-dox-glc conjugate. These results suggest that PAMAM-glucose formulations may constitute an efficient platform for the specific delivery of anticancer drugs to tumor cells overexpressing transporters of GLUT family.

Cytotoxicity of Dendrimers.

Drug delivery systems are molecular platforms in which an active compound is packed into or loaded on a biocompatible nanoparticle. Such a solution improves the activity of the applied drug or decreases its side effects. Dendrimers are promising molecular platforms for drug delivery due to their unique properties. These macromolecules are known for their defined size, shape, and molecular weight, as well as their monodispersity, the presence of the void space, tailorable structure, internalization by cells, selectivity toward cells and intracellular components, protection of guest molecules, and controllable release of the cargo. Dendrimers were tested as carriers of various molecules and, simultaneously, their toxicity was examined using different cell lines. It was discovered that, in general, dendrimer cytotoxicity depended on the generation, the number of surface groups, and the nature of terminal moieties (anionic, neutral, or cationic). Higher cytotoxicity occurred for higher-generation dendrimers and for dendrimers with positive charges on the surface. In order to decrease the cytotoxicity of dendrimers, scientists started to introduce different chemical modifications on the periphery of the nanomolecule. Dendrimers grafted with polyethylene glycol (PEG), acetyl groups, carbohydrates, and other moieties did not affect cell viability, or did so only slightly, while still maintaining other advantageous properties. Dendrimers clearly have great potential for wide utilization as drug and gene carriers. Moreover, some dendrimers have biological properties per se, being anti-fungal, anti-bacterial, or toxic to cancer cells without affecting normal cells. Therefore, intrinsic cytotoxicity is a comprehensive problem and should be considered individually depending on the potential destination of the nanoparticle.

Pyrrolidone-modified PAMAM dendrimers enhance anti-inflammatory potential of indomethacin in vitro.

The therapeutic effect of indomethacin, a water-insoluble non-steroidal anti-inflammatory drug, requires its efficient transport through cellular membranes and accumulation inside the target cells. The application of dendritic polymers has been proposed for the improvement of the drug's solubility and intracellular delivery. In this study we evaluated the anti-inflammatory potential of novel, highly-biocompatible 4-carbomethoxypyrrolidone-coated PAMAM dendrimers loaded with indomethacin. Our results indicate that complexation with dendrimers do not hamper the inhibitory action of indomethacin towards cyclooxygenases. Drug-dendrimer formulations exhibited improved anti-inflammatory activity in in vitro-cultured cellular models, showing enhanced inhibition of prostaglandin secretion and significantly decreased expression of NF-κB marker genes compared to free drug.