G4 Dendrimer Research Articles

Radical dendrimers: Synthesis, anti-tumor activity and enhanced cytoprotective performance of TEMPO free radical functionalized polyurethane dendrimers

Polyurethane dendrimers up to fourth generation, functionalized with stable 4-hydroxy-2, 2, 6, 6-tetramethylpiperidin-1-yloxyl (4-hydroxy TEMPO), were prepared via a single-step urethane interchange reaction and their structures were confirmed by NMR, FT-IR, MALDI-TOF MS and SEC-MALLS techniques. The terminal TEMPO radicals of the dendrimers exhibited a triplet with nitrogen hyperfine splitting constant of 14.4 G, at g = 2.0026 in the EPR spectrum and their paramagnetic nature were confirmed by vibrating sample magnetometer. The MTT assay results revealed that the G4 dendrimer had significant anti-tumor activity in A549 lung adenocarcinoma cells and this dendrimer was found not induced any cell death in normal L132 lung cells. Further, antioxidant effect of G4 dendrimer was evaluated in normal L132 lung cells; the in-vitro studies include cell morphological studies, measurement of total intracellular reactive oxygen species (ROS) using H2DCF dye and analysis of SOD, CAT, GSH and GPx. This study demonstrated the efficacy of the dendrimers through decrease in free radicals and confirmed their enhanced cytoprotective performance over N-acetyl cysteine standard used.

The Effect of Biotinylated PAMAM G3 Dendrimers Conjugated with COX-2 Inhibitor (celecoxib) and PPARγ Agonist (Fmoc-L-Leucine) on Human Normal Fibroblasts, Immortalized Keratinocytes and Glioma Cells in Vitro.

Glioblastoma multiforme (GBM) is the most malignant type of central nervous system tumor that is resistant to all currently used forms of therapy. Thus, more effective GBM treatment strategies are being investigated, including combined therapies with drugs that may cross the blood brain barrier (BBB). Another important issue considers the decrease of deleterious side effects of therapy. It has been shown that nanocarrier conjugates with biotin can penetrate BBB. In this study, biotinylated PAMAM G3 dendrimers substituted with the recognized anticancer agents cyclooxygenase-2 (COX-2) inhibitor celecoxib and peroxisome proliferator-activated receptor γ (PPARγ) agonist Fmoc-L-Leucine (G3-BCL) were tested in vitro on human cell lines with different p53 status: glioblastoma (U-118 MG), normal fibroblasts (BJ) and immortalized keratinocytes (HaCaT). G3-BCL penetrated efficiently into the lysosomal and mitochondrial compartments of U-118 MG cells and induced death of U-118 MG cells via apoptosis and inhibited proliferation and migration at low IC50 = 1.25 µM concentration, considerably lower than either drug applied alone. Comparison of the effects of G3-BCL on expression of COX-2 and PPARγ protein and PGE2 production of three different investigated cell line phenotypes revealed that the anti-glioma effect of the conjugate was realized by other mechanisms other than influencing PPAR-γ expression and regardless of p53 cell status, it was dependent on COX-2 protein level and high PGE2 production. Similar G3-BCL cytotoxicity was seen in normal fibroblasts (IC50 = 1.29 µM) and higher resistance in HaCaT cells (IC50 = 4.49 µM). Thus, G3-BCL might be a good candidate for the targeted, local glioma therapy with limited site effects.

Synthesis and Different Effects of Biotinylated PAMAM G3 Dendrimer Substituted with Nimesulide in Human Normal Fibroblasts and Squamous Carcinoma Cells

Squamous cell carcinoma (SCC) remains a main cause of mortality in patients with neck and head cancers, with poor prognosis and increased prevalence despite of available therapies. Recent studies have identified a role of cyclooxygenases, particularly inducible isoform cyclooxygenase-2 (COX-2) and its metabolite prostaglandin E2 (PGE2) in cancer cell proliferation, and its inhibition become a target for control of cancer development, particularly in the view of recognized additive or synergic action of COX-2 inhibitors with other forms of therapy. Nimesulide (N), the selective COX-2 inhibitor, inhibits growth and proliferation of various types of cancer cells by COX-2 dependent and independent mechanisms. In the presented study, the conjugates of biotinylated third generation poly(amidoamine) dendrimer (PAMAM) with covalently linked 18 (G3B18N) and 31 (G3B31N) nimesulide residues were synthesized and characterized by NMR spectroscopy. Biological properties of conjugates were evaluated, including cytotoxicity, proliferation, and caspase 3/7 activities in relation to COX-2/PGE2 axis signaling in human normal fibroblast (BJ) and squamous cell carcinoma (SCC-15). Both conjugates exerted a selective cytotoxicity against SCC-15 as compared with BJ cells at low 1.25–10 µM concentration range and their action in cancer cells was over 250-fold stronger than nimesulide alone. Conjugates overcome apoptosis resistance and sensitized SCC-15 cells to the apoptotic death independently of COX-2/PGE2 axis. In normal human fibroblasts the same concentrations of G3B31N conjugate were less effective in inhibition of proliferation and induction of apoptosis, as measured by caspase 3/7 activity in a manner depending on increase of PGE2 production by either COX-1/COX-2.

Tumor Uptake of Triazine Dendrimers Decorated with Four, Sixteen, and Sixty-Four PSMA-Targeted Ligands: Passive versus Active Tumor Targeting.

Various glutamate urea ligands have displayed high affinities to prostate specific membrane antigen (PSMA), which is highly overexpressed in prostate and other cancer sites. The multivalent versions of small PSMA-targeted molecules are known to be even more efficiently bound to the receptor. Here, we employ a well-known urea-based ligand, 2-[3-(1,3-dicarboxypropyl)-ureido] pentanedioic acid (DUPA) and triazine dendrimers in order to study the effect of molecular size on multivalent targeting in prostate cancer. The synthetic route starts with the preparation of a dichlorotriazine bearing DUPA in 67% overall yield over five steps. This dichlorotriazine reacts with G1, G3, and G5 triazine dendrimers bearing a 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) group for 64Cu-labeling at the core to afford poly(monochlorotriazine) intermediates. Addition of 4-aminomethylpiperidine (4-AMP) and the following deprotection produce the target compounds, G1-(DUPA)4, G3-(DUPA)16, and G5-(DUPA)64. These targets include 4/16/64 DUPA groups on the surface and a DOTA group at the core, respectively. In vitro cell assay using PC3-PIP (PSMA positive) and PC3-FLU (PSMA negative) cells reveals that G1-(DUPA)4 has the highest PC3-PIP to PC3-FLU uptake ratio (10-fold) through the PSMA-mediated specific uptake. While G5-(DUPA)64 displayed approximately 12 times higher binding affinity (IC50 23.6 nM) to PC3-PIP cells than G1-(DUPA)4 (IC50 282.3 nM) as evaluated in a competitive binding assay, the G5 dendrimer also showed high non-specific binding to PC3-FLU cells. In vivo uptake of the 64Cu-labeled dendrimers was also evaluated in severe combined inmmunodeficient (SCID) mice bearing PC3-PIP and PC3-FLU xenografts on each shoulder, respectively. Interestingly, quantitative imaging analysis of positron emission tomograph (PET) displayed the lowest tumor uptake in PC3-PIP cells for the midsize dendrimer G3-(DUPA)16 (19.4 kDa) (0.66 ± 0.15%ID/g at 1 h. p.i., 0.64 ± 0.11%ID/g at 4 h. p.i., and 0.67 ± 0.08%ID/g at 24 h. p.i.). Through the specific binding of G1-(DUPA)4 to PSMA, the smallest dendrimer (5.1 kDa) demonstrated the highest PC3-PIP to muscle and PC3-PIP to PC3-FLU uptake ratios (17.7 ± 5.5 and 6.7 ± 3.0 at 4 h p.i., respectively). In addition, the enhanced permeability and retention (EPR) effect appeared to be an overwhelming factor for tumor uptake of the largest dendrimer G5-(DUPA)64 as the uptake was at a similar level irrelevant to the PSMA expression.

Calorimetric study of siloxane dendrimer of the third generation with trimethylsilyl terminal groups

The molar heat capacity of siloxane dendrimer of the third generation with trimethylsilyl terminal groups G3[OSi(CH3)3]24 was determined by precise adiabatic calorimetry and differential scanning calorimetry over the temperature range T = (6–570) K for the first time. The low-temperature structural anomaly and the glass transition were observed in the above temperature range, and the standard thermodynamic characteristics of the revealed transformations were determined and analyzed. The fundamental thermodynamic functions such as the enthalpy [H°(T) − H°(0)], the entropy [S°(T) − S°(0)], and the Gibbs energy [G°(T) − H°(0)] were calculated for the range from T → 0 to 570 K based on the experimentally determined molar heat capacity of the investigated compound. The standard entropy of formation ΔfS° of dendrimer G3[OSi(CH3)3]24 was evaluated at T = 298.15 K. The thermal stability of the studied compound was investigated by thermogravimetric analysis. The standard thermodynamic properties of siloxane dendrimer G3[OSi(CH3)3]24 were compared and discussed with the previously reported data for the studied G3 carbosilane dendrimers with different functional terminal groups on the surface layer.

Analysis of Dimer Impurity in Polyamidoamine Dendrimer Solutions by Small-angle Neutron Scattering

Dimer impurity in the solution of a generation five (G5) polyamidoamine (PAMAM) dendrimer has been investigated by small-angle neutron scattering (SANS). The existence of dimer impurity in dendrimer solution was evidenced by indirect Fourier transform (IFT) analysis of the SANS data, in which the maximum dimension of particles in solution was found to be about twice the diameter of G5 dendrimer. We then developed an analytical model which accounts for the scattering contribution from both dendrimer monomer and dimer. The experimental data were well fitted by using the established model. The results showed that the amount of dimer impurities is significant for the measured three batches of G5 PAMAM dendrimers.

Modified Carboxyl-Terminated PAMAM Dendrimers as Great Cytocompatible Nano-Based Drug Delivery System.

Polyamidoamine (PAMAM) dendrimers are extensively researched as potential drug delivery system thanks to their desirable features such as controlled and stable structures, and ease of functionalization onto their surface active groups. However, there have been concerns about the toxicity of full generation dendrimers and risks of premature clearance from circulation, along with other physical drawbacks presented in previous formulations, including large particle sizes and low drug loading efficiency. In our study, carboxyl-terminated PAMAM dendrimer G3.5 was grafted with poly (ethylene glycol) methyl ether (mPEG) to be employed as a nano-based drug delivery system with great cytocompatibility for the delivery of carboplatin (CPT), a widely prescribed anticancer drug with strong side effects so that the drug will be effectively entrapped and not exhibit uncontrolled outflow from the open structure of unmodified PAMAM G3.5. The particles formed were spherical in shape and had the optimal size range (around 36 nm) that accommodates high drug entrapment efficiency. Surface charge was also determined to be almost neutral and the system was cytocompatible. In vitro release patterns over 24 h showed a prolonged CPT release compared to free drug, which correlated to the cytotoxicity assay on malignant cell lines showing the lack of anticancer effect of CPT/mPEG-G3.5 compared with CPT.

Generation 2 (G2) – Generation 4 (G4) PAMAM dendrimers disrupt key plasma coagulation parameters

The aim of the research was to evaluate the effects of G2 – G4 PAMAM dendrimers on basic plasma haemostasis parameters (Partially Activated Thrombin Time (APTT), Prothrombin Time (PT), Thrombin Time (TT)) as well as the activity of factor X, antithrombin III (AT), protein C and plasmin. Furthermore, tissue factor (TF) synthesis in endothelial cells and viability of smooth muscle cells in the presence of PAMAM dendrimers was investigated.APTT, PT and TT were performed according to the available commercial methods. The activity of factor X was conducted based on deficient plasma factor X. Protein C, AT and plasmin activity were measured spectrophotometrically using chromogenic substrates. Intracellular TF production in human umbilical vein endothelial cells (HUVECs) was measured using immunohistochemical method. Viability of Human Aortal Smooth Muscle cells (hAoSMCs) was established using WST-1 assay.PAMAM dendrimers decreased activity of factor X, and concomitantly prolonged PT and APTT. We also demonstrated shortened TT and increased fibrinogen concentrations in plasma treated with G4 PAMAM dendrimers, suggesting formation of fibrinogen aggregates. G2 – G4 PAMAM dendrimers decreased the activity of both naturally occurring anticoagulants AT and protein C. G2 and G3 PAMAM dendrimers did not affect the proteolytic reaction with plasmin. PAMAM dendrimers were found not to trigger TF production in undisturbed endothelial cells.PAMAM dendrimers, depending on the concentration and generation decreased viability of AoSMCs.The results presented within the current study suggest complex but mostly undesirable effect of G2 – G4 PAMAM dendrimers on plasma haemostasis and underscore the need for further in-depth research.

Evaluation of the localization and biological effects of PAMAM G3 dendrimer-biotin/pyridoxal conjugate as HaCaT keratinocyte targeted nanocarrier.

Recognition of the molecular mechanisms of keratinocyte participation in normal skin homeostasis and in pathogenesis may lead to creation of more effective tools for topical application of cosmetics, cosmeceutics and drugs to a particular location within the skin for prevention and therapy of many skin disorders and diseases. For this purpose, the PAMAM G3 dendrimer with amide linkages of 9 biotin molecules and 10 molecules of pyridoxal phosphate (BC-PAMAM) was constructed, and its biological properties and cellular uptake and localization were investigated in the HaCaT keratinocytes. BC-PAMAM is nontoxic for HaCaT cells, as estimated by two assays (Neutral Red and tetrazolium salt reduction, XTT), and revealed low apoptosis induction at up to 50 µM concentration. Fluorescent labeled BC-PAMAM accumulates in HaCaT cells with high efficiency in a concentration-dependent manner. Its mitochondrial localization, estimated as Mander's colocalization coefficient, is substantially lower than the native PAMAM, and that correlates with its cytotoxicity. The only undesirable, but significant inhibitory effect on cell mobility, evaluated by the wound healing test, was observed at 10 µM BC-PAMAM. The important anti-inflammatory action of BC-PAMAM was clearly documented by decreased production of total IL-1α, assayed with an ELISA test with unstimulated and stimulated by bacterial antigens (LPS and GroEL) HaCaT cells. Thus, it is expected that the biotin pyridoxal phosphate conjugated PAMAM may be considered as a potential carrier for safe delivery of vitamins and drugs into the epidermis.

Dendrimeric Nanoparticles for Two-Photon Photodynamic Therapy and Imaging: Synthesis, Photophysical Properties, Innocuousness in Daylight and Cytotoxicity under Two-Photon Irradiation in the NIR.

The synthesis and the photophysical properties of a new class of fully organic monodisperse nanoparticles for combined two-photon imaging and photodynamic therapy are described. The design of such nanoparticles is based on the covalent immobilization of a dedicated quadrupolar dye that combines excellent two-photon absorbing (2PA) properties, fluorescence and singlet oxygen generation ability, in a phosphorous-based dendrimeric architecture. First, a bifunctional quadrupolar dye bearing two different grafting moieties, a phenol function and an aldehyde function, was synthesized. It was then covalently grafted through its phenol function to a phosphorus-based dendrimer scaffold of generation 1. The remaining aldehyde functions were then used to continue the dendrimer synthesis up to generation 2, introducing finally 24 water-solubilizing triethyleneglycol chains at its periphery. A dendrimer confining 12 photoactive quadrupolar units in its inner scaffold and showing water solubility was thus obtained. Interestingly, the G1 and G2 dendrimers retain some fluorescence as well as significant singlet oxygen production efficiencies while they were found to show very high 2PA cross-sections in a broad range of the NIR biological spectral window. Hydrophilic dendrimer G2 was tested in vitro on breast cancer cells, first in one- and two-photon microscopy, which allowed for visualization of their cell internalization, then in two-photon photodynamic therapy. While being nontoxic in the dark and, more importantly, under exposure to daylight, dendrimer G2 proved to be a very efficient cell-death inducer only under two-photon irradiation in the NIR.

Synthesis and evaluation of polyethylene glycol- and folic acid-conjugated polyamidoamine G4 dendrimer as nanocarrier

Synthesis of folic-acid mediated targeting nanocarrier is one of the most important and effective approaches in drug delivery. In this study, firstly, the surface of polyamidoamine G4 (PAMAM G4) dendrimers was modified with polyethylene glycol (PEG) to reduce dendrimer toxicity and was engineered with folic acid (FA) to increase nanocomplex targeting of the mice myoblast C2C12 cell lines. Both these synthetic phases were characterized via fourier transform infrared spectroscopy (FT-IR), Proton nuclear magnetic resonance (1H NMR), ultraviolet–visible spectrophotometry (UV–Vis), and differential scanning calorimetry (DSC) analysis. Secondly, 5-fluorouracil (5-FU), as a chemotherapeutic agent, was loaded in the nanocarrier to therapeutic goals and was characterized. In vitro drug release was also assessed and an excellent controlled release with good release efficiency was reported. The size, diameter, surface morphology, and surface charge of these synthesized nanocomplexes were ascertained with the scanning electron microscopy (SEM), dynamic light scattering (DLS) and zeta potential analyser, respectively, and PAMAM G4 positive charges eliminated. Finally, our nanocomplex was tested on the HT-29 human colorectal cancer and mice myoblast C2C12 normal cell lines and cell viability was evaluated with an 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) assay. Good cancer cell inhibition and cell growth promotion effects of the nanocomplexes have been indicated in this research study.

PEGylated poly(amidoamine) dendrimers-based drug loading vehicles for delivering carboplatin in treatment of various cancerous cells

The fourth generation (G4) of poly(amidoamine) dendrimer was PEGylated and then physically incorporated with carboplatin to develop the new drug delivery carrier. The proton nuclear magnetic resonance (1H NMR) and Fourier-transform infrared spectroscopy (FTIR) were utilized to confirm the successful synthesis and incorporation. The PEGylation leads to increase the particle dimension of G4 dendrimer to 4.04 to 30.67 nm that was still in the effective range for killing cancerous cells. The cytotoxicity on L929 fibroblasts demonstrated that the PEGylation of G4 could eliminate the toxic caused by amine surrounding groups of G4. Three cancerous cells including the cervical cancer cell line (HeLa), the adenocarcinomic human alveolar basal epithelial cells (A549), and the human breast cancer cell line (MCF7) were treated with G4-PEG@CAR as a function of CAR concentrations. The results of resazurin test and live/dead assay indicated that the killing effect of G4-PEG@CAR on three representative cells were achieved and increased following the enhancement of CAR concentration. Furthermore, the better effectiveness of G4-PEG@CAR was achieved on Hela.

Preparation and optimization of lidocaine transferosomal gel containing permeation enhancers: a promising approach for enhancement of skin permeation.

AimTo develop the topical gel containing transferosomal lidocaine as alternative to painful local anesthetic injection.Materials and methodsThe transfersomes were prepared by film hydration technique using soybean phosphatidylcholine and cholesterol. The prepared transfersomes were evaluated for the morphology, drug loading, %EE, particle size and in vitro release. The transferosomal gel of lidocaine was prepared using HPMC k15 as gelling agent and propylene glycol, dimethyl sulfoxide (DMSO), and polyamidoamine dendrimer third generation (PAMAM G3) solutions were used as permeation enhancer. The formulated gels were evaluated for pH, viscosity, drug content and ex-vivo permeation of the gel. The analgesic effect of the formulation was tested using tail flick test.ResultsThe transfersomes showed that transfersomes (F4) had the highest entrapment efficiency (%EE) approaching 79.87±2.35, low particle size 179.5 nm, and zeta potential of −43.5±4.74 mV. According to the rat tail flick test, the AUC0–90 minutes of the control formulation (lidocaine solution, A) was 352.32±5.87 seconds minutes. While the maximum AUC0–90 minutes value was found to be 570.5±6.81 seconds minutes for gel formulation (F) containing transfersomal lidocaine with PAMAM G3 dendrimer as permeation enhancer. In this case, the local anesthetic efficacy was increased by 1.62-folds as compared to control formulation.ConclusionFrom the present study, it can be concluded that the topical gel loaded with transfersomal lidocaine shows enhanced skin permeation effect along with increase in local anesthetic action of lidocaine.

Cyclohexadienone core 3,6-di-tert-butylcarbazole decorated triazole bridged dendrimers: synthesis, photophysical and electrochemical properties and application as an additive in dye-sensitized solar cells

An enhanced efficiency of 9.01% was realized for G2 dendrimer based DSSC.

Rotaxane-branched dendrimers with aggregation-induced emission behavior

By introducing a 9,10-distyrylanthrance (DSA) moiety as the core, we demonstrated the synthesis of a new family of rotaxane-branched dendrimers G1–G3 through the controllable divergent approach.

Inherent anti-HIV activity of biocompatible anionic citrate-PEG-citrate dendrimer.

The development of new combinations to empower better protection against HIV infection is particularly important. Anionic polymers can block HIV infection. In the current study, first generation (G1) and second generation (G2) novel water-soluble anionic citrate-PEG-citrate dendrimers were synthesized and characterized with Fourier-transform infrared spectroscopy (FTIR), nuclear magnetic resonance spectroscopy (NMR), and dynamic light scattering (DLS) methods. After the biocompatibility of the G2 dendrimer was determined, its antiviral activity was evaluated. This function may contribute to the peripheral groups of this dendrimer (carboxylate group). In order to measure the inhibitory effect of G2 on HIV infection, both pre-treatment (treated with G2 dendrimer before HIV infection) and co-treatment (simultaneously treated with G2 dendrimer and HIV infection) were used in vitro. The results showed the good synthesis of the G2 dendrimer, and the dendrimer showed antiviral properties (ICC50:0.4mM) and low toxicity (CC50:0.6mM) at high concentrations. A strong inhibitory effect was found when the co-treatment approach was used. This study achieved promising results which encourage the use of G2 dendrimers as anti-HIV agents.

Properties of Bioconjugates of Streptokinase with Anionic Polyamidoamine Dendrimers of Various Generations

Covalent conjugates of streptokinase (SK) with polyamidoamine (PAMAM) dendrimers G1.5, G2.5, and G3.5 (SK–G1.5, SK–G2.5, and SK–G3.5) with the protein–polymer molar ratios of (1: 1), (1: 5), and (1: 10) were obtained and their properties were studied as compared to the properties of free SK. It was shown that the initial rates of formation of the modified Pm. SK complex, activation of plasminogen, and lysis of the plasma clot under the action of SK–dendrimer conjugates decreased with increasing number of bound dendrimers (from 1 to 10) and increased with increasing dendrimer generation (from G1.5 up to G3.5). Conjugates SK–G3.5 (1: 1) and (1: 5) were the most active compared to other conjugates. It was found that the catalytic efficiency of plasminogen activation (kPg/KPg) by conjugates SK–G3.5 (1: 1) (0.15 μM–1 min–1) and SK–G3.5 (1: 5) (0.12 μM–1 min–1) was comparable to the efficiency of free SK (0.18 μM–1 min–1). Probably, small in size, soft, and easily deformable dendrimers G1.5 and G2.5 are able to penetrate into the internal shielded cavities of the native SK molecule and there modify amino groups that are important for the effective formation of the Pm · SK complex. By contrast, the larger and more rigid molecule of dendrimer G3.5 modifies, mainly, exposed lysine residues in the SK molecule, without affecting the latent internal lysines. Conjugates SK–G3.5 (1: 1) and (1: 5), which had the maximum activator activity, retained up to 85% of thrombolytic activity compared to the activity of free SK. In addition, due to modification of the exposed lysines—most sensitive to proteolysis in the SK molecule—with dendrimer G3.5, which has the highest density of negative charge on its surface, SK–G3.5 (1: 1) and (1: 5) conjugates were more stable in plasma and caused less exhaustion of plasma levels of plasminogen, α2-antiplasmin, and fibrinogen than free SK in vitro. Thus, thrombolytic activity of the SK–dendrimer conjugates depends on the degree of modification of the amino groups of SK, size, stiffness, and density of the negative charge on the surface of the PAMAM dendrimer. Conjugates SK–G3.5 (1: 1) and (1: 5) are potential candidates for the development of a new thrombolytic agent.

Pentafluoropropionic Anhydride Functionalized PAMAM Dendrimer as miRNA Delivery Reagent

Poly(amidoamine) (PAMAM) dendrimers are good candidates for nucleic acid delivery with their well-defined characteristics. MicroRNA (miRNA) mediated regulation of biological process are also active area of investigation. Fibroblast cells, such as MRC-5, are one of the cell lines used in biological researches due to their hard to transfect nature. In this two-staged study, cystamine core G5 PAMAM dendrimers were synthesized and modified with pentafluoropropionic anhydride (PA) and subsequently tested as miRNA delivery reagent on MRC-5 cells. Effect of fluorination against to naked G5 dendrimer over transfection efficiency was also investigated by molecular docking and quantitative structure-activity relationship (QSAR) calculations. Structural characterization of the synthesized dendrimers was verified by spectroscopic techniques. Gel retardation assay, particle size and transmission electron microscopy results demonstrated polyplex formation ability of fluorinated dendrimers with miRNA at nanoscale level. Zeta potential values indicated non-aggregation and increased stability of the polyplexes. Prepared polyplexes with fluorinated dendrimer showed over 90% cell viability and transfection efficiency. In silico calculations confirmed the stable complexation with miRNA and smooth penetration capability into the cell.

Biotinylated PAMAM G3 dendrimer conjugated with celecoxib and/or Fmoc-l-Leucine and its cytotoxicity for normal and cancer human cell lines

Tumors still remain one of the main causes of mortality due to the lack of effective anti-cancer therapy. Recently it has been shown, that overexpression of inducible cyclooxygenase-2 (COX-2) and decrease of peroxisome proliferator-activated receptor γ (PPARγ) expression accompany many malignances, therefore, it has been proposed, that COX-2 inhibitors and PPARγ agonists are potential candidates for anticancer therapy and their synergistic, antineoplastic action has been described. In the present study a COX-2 inhibitor (celecoxib) and/or PPARγ agonist (Fmoc-l-Leucine) were conjugated with the biotinylated G3 PAMAM dendrimer to form a three different constructs targeted to cells with increased biotin uptake. All conjugates were characterized by the NMR spectroscopy. Investigation of three types of human cells: normal skin fibroblasts (BJ), immortalized keratinocytes (HaCaT) and cancer lines: glioblastoma (U-118 MG) and squamous cell carcinoma (SCC-15) revealed similar biotin labeled ATTO590 accumulation (after 24 h), except for SCC-15 with significantly lower loading. Constitutive expression of COX-2 protein was confirmed in all tested cells with significantly higher levels (2–2.5 times) in both cancer lines.Comparison of cytotoxicity of the new synthetized dendrimers clearly documented the highest cytotoxicity of the G31B16C15L dendrimer conjugated with both drugs (1: 1) as compared with drugs alone and single conjugates. Additive effects of construct with both compounds were shown for fibroblasts and both cancer cell lines in the order BJ > U-118 MG > SCC-15 with IC50 in the range: 0.69, 1.44 and 2.22 μM, respectively and lowest cytotoxicity in HaCaT cells (IC50 = 2.88). Our results showed, that biotinylated G3 PAMAM dendrimers substituted with COX-2 inhibitor, celecoxib, and PPARγ agonist, Fmoc-l-Leucine (1:1) may be a good candidate for local therapy of glioblastoma but not a skin cancer. Since the effect of PPARγ agonists on COX-2 expression vary depending upon the cell type, specificity of used agonist and the presence of other environmental factors, it is necessary to carefully evaluate the response of chosen drugs on the target cells.

Evaluation of Amino-Functional Polyester Dendrimers Based on Bis-MPA as Nonviral Vectors for siRNA Delivery.

Herein, we present the first evaluation of cationic dendrimers based on 2,2-bis(methylol)propionic acid (bis-MPA) as nonviral vectors for transfection of short interfering RNA (siRNA) in cell cultures. The study encompassed dendrimers of generation one to four (G1–G4), modified to bear 6–48 amino end-groups, where the G2–G4 proved to be capable of siRNA complexation and protection against RNase-mediated degradation. The dendrimers were nontoxic to astrocytes, glioma (C6), and glioblastoma (U87), while G3 and G4 exhibited concentration dependent toxicity towards primary neurons. The G2 showed no toxicity to primary neurons at any of the tested concentrations. Fluorescence microscopy experiments suggested that the dendrimers are highly efficient at endo-lysosomal escape since fluorescently labeled dendrimers were localized specifically in mitochondria, and diffuse cytosolic distribution of fluorescent siRNA complexed by dendrimers was observed. This is a desired feature for intracellular drug delivery, since the endocytic pathway otherwise transfers the drugs into lysosomes where they can be degraded without reaching their intended target. siRNA-transfection was successful in C6 and U87 cell lines using the G3 and G4 dendrimers followed by a decrease of approximately 20% of target protein p42-MAPK expression.