Polybrene Research Articles

Macrocyclic Neutralizer to Polybrene via Direct Host-Guest Complexation.

Hexadimethrine bromide (HB), a synthetic polycationic species, was introduced to clinical practice as a heparin antidote and recently used in gene therapy. However, HB causes various complications such as severe red blood cells (RBCs) aggregation and tissue damage. Herein, we have synthesized a water-soluble quaterphen[3]arene containing multiple sulfonate moieties (SQP3) as a novel macrocyclic neutralizer to reverse HB via direct host-guest complexation. SQP3 exhibited a robust binding affinity toward HB with a considerably high association constant of (4.73 ± 0.61) × 107 M-1. Co-dosed with 1 equiv of SQP3, HB-induced RBCs aggregation and blood coagulation could be effectively reversed. In vitro cellular assay verified that complexation of HB with SQP3 significantly decreased reactive oxygen species production, thereby suppressing cell apoptosis. In vivo neutralization efficacy studies demonstrated that HB/SQP3 was capable of alleviating related organic damage caused by HB and improving the survival rate of HB-treated mice from 20 to 100%.

Adaptive and Ultrahigh‐Affinity Recognition in Water by Sulfated Conjugated Corral[5]arene

AbstractThe pursuit of synthetic receptors with high binding affinities has long been a central focus in supramolecular chemistry, driven by their significant practical relevance in various fields. Despite the numerous synthetic receptors that have been developed, most exhibit binding affinities in the micromolar range or lower. Only a few exceptional receptors achieve binding affinities exceeding 109 M−1, and their substrate scopes remain rather limited. In this context, we introduce SC[5]A, a conjugated corral‐shaped macrocycle functionalized with ten sulfate groups. Owing to its deep one‐dimensional confined hydrophobic cavity and multiple sulfate groups, SC[5]A displays an extraordinarily high binding strength of up to 1011 M−1 towards several size‐matched, rod‐shaped organic dications in water. Besides, its conformation exhibits good adaptability, allowing it to encapsulate a wide range of other guests with diverse molecular sizes, shapes, and functionalities, exhibiting relatively strong affinities (Ka=106–108 M−1). Additionally, we've explored the preliminary application of SC[5]A in alleviating blood coagulation induced by hexadimethrine bromide in vitro and in vivo. Therefore, the combination of ultrahigh binding affinities (towards complementary guests) and adaptive recognition capability (towards a wide range of functional guests) of SC[5]A positions it as exceptionally valuable for numerous practical applications.

Adaptive and Ultrahigh-Affinity Recognition in Water by Sulfated Conjugated Corral[5]arene.

The pursuit of synthetic receptors with high binding affinities has long been a central focus in supramolecular chemistry, driven by their significant practical relevance in various fields. Despite the numerous synthetic receptors that have been developed, most exhibit binding affinities in the micromolar range or lower. Only a few exceptional receptors achieve binding affinities exceeding 109 M-1 , and their substrate scopes remain rather limited. In this context, we introduce SC[5]A, a conjugated corral-shaped macrocycle functionalized with ten sulfate groups. Owing to its deep one-dimensional confined hydrophobic cavity and multiple sulfate groups, SC[5]A displays an extraordinarily high binding strength of up to 1011 M-1 towards several size-matched, rod-shaped organic dications in water. Besides, its conformation exhibits good adaptability, allowing it to encapsulate a wide range of other guests with diverse molecular sizes, shapes, and functionalities, exhibiting relatively strong affinities (Ka =106 -108 M-1 ). Additionally, we've explored the preliminary application of SC[5]A in alleviating blood coagulation induced by hexadimethrine bromide in vitro and in vivo. Therefore, the combination of ultrahigh binding affinities (towards complementary guests) and adaptive recognition capability (towards a wide range of functional guests) of SC[5]A positions it as exceptionally valuable for numerous practical applications.

The Effect of Polybrene on an aPTT-based Lupus Anticoagulant Test in the Plasma of Patients Receiving Unfractionated Heparin

Objective: Heparins can interfere with lupus anticoagulant (LA) testing, especially when the detection assays do not contain a heparin neutralizer. In this study we evaluated whether hexadimethrine bromide commonly known as polybrene was able to neutralize the effects of unfractionated heparin (UFH) on activated partial thromboplastin time (aPTT)-based LA determination. Material and Methods: The influence of polybrene on the aPTT-based LA testing results of 14 patients receiving UFH therapy was studied. Measurements were performed using Dade Actin FSL (aPTT-LA screen) and Dade Actin FS (aPTT-LA confirm) reagents on a Sysmex CS-1600 analyzer. The aPTT-LA screen/confirm ratios before and after the polybrene treatments were compared. Results: The UFH treatment affected the aPTT-LA screen/confirm ratio in four patterns, 1) no coagulation (NC code), 2) false positive, 3) false low, and 4) unchanged interpretation. After adding polybrene in the patient plasmas, the aPTT-LA screen/confirm ratios returned nearly to baseline without false positive or negative results. Conclusion: The best practice for LA testing should be done outside anticoagulant therapies, but if necessary, the tests can be performed by adding the proper concentration of polybrene.

Two-step micelle-to-solvent stacking of arsenic species from foods in permanently coated tubing for capillary electrophoresis

An effective and sensitive two-step stacking of arsenic species by electrokinetic injection and micelle-to-solvent stacking (MSS) using zwitterionic surfactant 3-(N,N-dimethylpalmitylammonio) propane sulfonate was successfully developed in a co-electro-osmotic flow (co-EOF) capillary zone electrophoresis (CZE). The fused silica capillaries were coated with 1% hexadimethrine bromide solution to satisfy the condition of the same EOF direction between the test anions and capillary. The background solution was 100 mM borax buffer (pH 9.2). The proposed method was performed by a CZE system equipped with a diode array detector, and the detection wavelength was monitored at 192 nm. The separation voltage was −20 kV. The group of micelles and target analytes was stacked by field-enhanced sample injection at −10 kV for 180 s. The second stacking step used 60% MeOH to serve as an organic solvent where the analytes were brought by the micelles to the MSS boundary. The enrichment factors of As(Ⅴ), As(Ⅲ), disodium methyl arsonate hexahydrate (MMA), and dimethylarsinic acid (DMA) were 1230, 840, 3820, and 1450, respectively, compared to typical injections in CZE. The limits of detection (S/N = 3) ranged from 0.382 to 0.911 ng mL−1. The intracapillary repeatabilities (%RSD, n = 3) were 0.5–1.0% for migration time and 0.3–0.7% for peak areas. The technology of two-step stacking by MSS in co-EOF CZE together with a simple extraction procedure for As(Ⅴ), As(Ⅲ), MMA and DMA were demonstrated in kelp and rice samples.

Inhibiting the growth of melanoma cells via hTERT gene editing using CRISPR-dCas9-dnmt3a system

CRISPR-Cas9 gene-editing technology has pushed the boundaries of genetic modification. The principle of this method is based on the purposeful defense system of DNA degradation and will be one of the most powerful instruments for gene editing shortly. The purpose of this study was to evaluate the capability of this approach to manage melanoma cells. The present study used EF1a-hsaCas9-U6-gRNA as a hybrid vector of sgRNA and Cas9 for the transfection of A-375 melanoma cells. Transfection efficiency was enhanced by examining the two concentrations of 4 and 8 µg/mL of hexadimethrine bromide (trade name Polybrene). The existence of Cas9 in transfected cells was detected by flow cytometry. The expression level of the metabisulfite-modified hTERT gene was measured by real-time PCR technique. The presence of telomerase in cells was determined by flow cytometry and western blotting analysis. The hTERT gene promoter methylation was also evaluated by HRM assay. Finally, the induction of apoptosis in transfected A375 cells was assessed using flow cytometry. The results showed that the presence of gRNA significantly increased the transfection efficiency (up to about 7.75 times higher). The hTERT expression levels in A-375 cells were significantly decreased at different concentrations of Polybrene (in a dose-dependent manner) and various amounts of transfection (P < 0.05). The expression of hTERT in basal cells was not significantly different from the group transfected without gRNA (P˃0.05) but was significantly higher than the group transfected with gRNA (P < 0.05). The results of flow cytometry and western blotting analysis showed a decrease in hTERT level compared to cells transfected without gRNA as well as basal cells. The methylation of hTERT gene promoter in the cells transfected with gRNA at a concentration of 80 μg/mL in the presence of both 4 μg/mL and 8 μg/mL of Polybrene was significantly increased compared to those transfected without sRNA (P < 0.05). The flow cytometry results indicated no significant difference in the induction of apoptosis in the transfected cells compared to the basal cells (P < 0.05). Evidence suggests that the designed CRISPR/Cas9 system reduces the expression of the hTERT gene and telomerase presence, thereby inhibiting the growth of melanoma cells.

Comparison of Polybrene DAT and Conventional DAT in Cases of Suspected DAT Negative Autoimmune Hemolytic Anemia (AIHA)

Abstract Introduction/Objective A negative Direct Antiglobulin Test (DAT) is seen in approximately 1-5% of patients presenting with signs and symptoms of immune hemolysis. A well-performed ‘standard’ DAT detects ~100-500 molecules of bound IgG per red cell. A more sensitive polybrene DAT may be helpful in order to confirm a diagnosis of AIHA in patients whose samples have tested negative via standard DAT. However, there has been little reported on utility of the polybrene DAT in evaluating such cases with modern DAT reagents. We hypothesized that the polybrene DAT would not contribute substantially to the analysis of Coombs-negative hemolytic anemia (C-NHA), based primarily on our anecdotal observation. As such, we undertook a study to assess results of polybrene DAT in cases evaluated for possible (C-NHA). Methods/Case Report Two study sites were used for data analysis (Yale-New Haven Hospital, Site A, and VA Connecticut, Site B) over a five year period (2016-2021). During this time, standard DATs were performed at both study sites by the tube method using polyspecific antiglobulin and, if positive, reflex to anti-IgG and -C3. For cases of suspected C-NHA (which are reported to Blood Bank via a consult mechanism), conventioned DAT- samples are referred to our regional immunohematology laboratory (American Red Cross, Farmington, CT) for manual hexadimethrine bromide (Polybrene) DAT. Polybrene DAT is reported as negative with two sources of polyspecific AHG. Subsets of patients also underwent acid elution studies (Gamma ELU-KITII) as part of a C-NHA algorithm developed during the study period. Results of standard DAT, acId elution, and polybrene DAT were extracted from Blood Bank electronic records at both study sites. Results (if a Case Study enter NA) Evaluation for C-NHA was performed in 32 patients/cases over the study period. Amongst these individuals, 96.8% (31/32) underwent polybrene DAT assessment and none (0%; 0/31) demonstrated a positive polybrene DAT result. Notably, acid elution studies were performed in 90.6% (29/32) of traditional DAT negative cases. Of these, 10.3% (3/29) had reactive eluates. Conclusion Performance of the polybrene DAT appeared to be of no value in the assessment of suspected C-NHA. Given that nearly 10% of individuals with a negative conventional DAT had a positive acid elution, this testing step appears to be of greater value in potentially identifying an autoantibody in suspected cases of C-NHA.

Production and Use of Gesicles for Nucleic Acid Delivery.

Over-expression of the vesicular stomatitis virus glycoprotein (VSVG) in mammalian cells can induce the formation of VSVG-pseudotyped vesicles (named "gesicles") from membrane budding. Its use as a nucleic acid delivery tool is still poorly documented. Naked-plasmid DNA can be delivered in animal cells with gesicles in presence of hexadimethrine bromide (polybrene). However, little is known about gesicle manufacturing process and conditions to obtain successful nucleic acid delivery. In this study, gesicles production process using polyethylenimine (PEI)-transfected HEK293 cells was developed by defining the VSVG-plasmid concentration, the DNA:PEI mass ratio, and the time of gesicle harvest. Furthermore, parameters described in the literature relevant for nucleic acid delivery such as (i) component concentrations in assembly mixture, (ii) component addition order, (iii) incubation time, and (iv) polybrene concentration were tested by assessing the transfection capacity of the gesicles complexed with a green fluorescent protein (GFP)-coding plasmid. Interestingly, freezing/thawing cycles and storage at + 4°C, - 20°C, and - 80°C did not reduce gesicles' ability to transfer plasmid DNA. Transfection efficiency of 55% and 22% was obtained for HeLa cells and for hard-to-transfect cells such as human myoblasts, respectively. For the first time, gesicles were used for delivery of a large plasmid (18-kb) with 42% of efficiency and for enhanced green fluorescent protein (eGFP) gene silencing with siRNA (up to 60%). In conclusion, gesicles represent attractive bioreagents with great potential to deliver nucleic acids in mammalian cells.

Study of in vitro thrombin generation after neutralization of heparin.

Thrombin generation (TG) documentshypercoagulability. TG in platelet-poor plasma is exquisitely sensitive toheparins, which thus mustbe neutralized before testing. Heparinase and hexadimethrine bromide (polybrene) have been used for that purpose, but their effectsper seon TG have been poorly studied so far. (i) TG was studied in commercial normal pooled plasma (NPP; CryoCheck® , Cryopep) in absence or presence of neutralizing agents. (ii) NPP was spiked with increasing concentrations of unfractionated heparin (UFH; up to 1.0IU/mL) or low-molecular-weight heparin (LMWH; enoxaparin up to 1.2IU/mL) and TG studied after incubation of heparinase (Hepzyme® ; 15minutes) or polybrene (0.025mg/mL; 10minutes). (i) With ThromboScreen reagent to initiate TG, addition of heparinase was associated with increased peak, whereas polybrene caused lengthening of lag time and time to peak, compared with nonsupplemented NPP. (ii) With polybrene, TG was completely restored over the whole range of UFH and LMWH studied. By contrast, heparinase failed to fully restore TG in presence of UFH concentrations ≥0.8IU/mL or LMWH concentrations ≥1.0IU/mL. Those effects were matched with detectable tiny residual amounts of non-neutralized heparin (as assessed with an anti-Xa assay) and were less pronounced with a higher picomolar concentration of tissue factor (DrugScreen reagent). Polybrene fully restored TG of heparinized plasma at the expense of an alteration of TG, pointing to the need to use adapted reference ranges. Heparinase failed to do so in presence of high concentrations of both heparins.

Analysis of intact proteins with capillary zone electrophoresis coupled to mass spectromery using uncoated and coated capillaries

Although, in general, the application of coated capillaries is recommended for the separation of intact proteins, bare silica capillary is still the most often used capillary due to its simplicity and cheapness. In this work, the performance of bare fused silica capillary for intact protein analysis was compared to that of different (dynamically coated polybrene (PB) and permanently coated linear polyacrylamide (LPA)) coated capillaries using capillary zone electrophoresis - mass spectrometry (CZE-MS). In cases where low pH (pH=1.8) was used in bare silica capillaries, good precision (0.56-0.78 RSD% and 1.7-6.5 RSD% for migration times and peak areas, respectively), minimal adsorption and separation efficiency (N= 27 000/m - 322 000/m) similar to or even better than those obtained with the coated capillaries (created by an intricate multi-step process) was achieved. The PB and the LPA capillaries demonstrated their slightly better resolving power in terms of separating the different forms/variants of the same protein (e.g., hemoglobin subunits). Among the studied capillaries the one with LPA coating showed the most stable separations in the long term (n=25: 0.18-0.49 RSD% and 3.1-4.9 RSD% for migration times and peak areas, respectively). For the separation of a few proteins or even a larger number of proteins in biological samples (e.g., snake venom) the application of the simple and cheap bare fused silica capillary can be considered as an efficient choice.

A surface charge governed nanofluidic diode based on a single polydimethylsiloxane (PDMS) nanochannel

HypothesisNanofluidic diodes have attracted intense attention recently. Commonly used materials to design these devices are membrane-based short nanopores and aligned Carbon nanotube bundles. It is highly desirable and very challenging to develop a nanofluidic diode based on a single PDMS nanochannel which is easier to be introduced into an integrated electronic system on a chip. Layer-by-layer (LBL) deposition of charged polyelectrolytes can change the size and surface properties of PDMS nanochannels that provides new possibilities to develop high-performance nanofluidic based on PDMS nanochannels. ExperimentsA novel design of nanofluidic diode is presented by controlling the surface charges and sizes of single PDMS nanochannels by surface modification using polyelectrolytes. Polybrene (PB) and Dextran sulfate (DS) are used to reduce the PDMS nanochannel size to meet the requirement of ion gating by LBL method and generate opposite surface charges at the ends of nanochannels. The parameters of such a nanofluidic diode are investigated systematically. FindingsThis nanofluidic diode developed in this work has high effective current rectification performance. The rectification ratio can be as high as 218 which is the best ever reported in PB/DS modified nanochannels. This rectification ratio reduces with high voltage frequency and ionic concentration whereas increases in shorter nanochannels.

Effective separation of chromium species in technological solutions using amino-immobilized silica prior to their determination

Amino-immobilized (poly(4,9-dioxadodecane-1,12-guanidine, polydiallyldimethylammonium, hexadimethrin bromide, polyhexamethylene guanidine) silicas were proposed for chromium speciation for the first time. Adsorbents surface was characterized by TGA-DSC, FT-IR, CHN, XRD and SEM analysis. Polyamines were strongly fixed on the silica surface and were not washed off with solutions of 3М HNO3 and 20 g L−1 NaCl. Аmino-immobilized silica quantitatively removed (R ≥ 99%) Cr(VI) from solutions at pH 4–7. Cr(III) was not recovered in this pH range, which makes it possible to separate Cr(VI) from Cr(III). The separation factor (КCr(VI)/Cr(III)) was ≥ 1∙104. Silica-based adsorbents layer-by-layer immobilized with polyamines and 2-(1,8-dihydroxy-3,6-disulfo-2-naphthylazo)benzenearsonic acid were proposed for quantitative removal of Cr(III) from aqueous solutions with pH 4–6 at 90 °C. A system of sequentially connected columns filled with selective adsorbents was used to separate the chromium species in stream at рН= 5 and a flow rate of 1 mL min−1. Chromium was determined after its elution with 5 mL of 2 M HNO3 at a flow rate of 1 mL min−1 using ICP-OES or ICP-MS. The pre-concentration factors for Cr(VI) and Cr(III) was 60. A two-column system was used for chromium speciation in technological solutions. The efficiency of chromium speciation was confirmed by state standard procedure.

Determination of gastrodin activity inhibition on acetylcholinesterase by capillary electrophoresis

Alzheimer's disease (AD) is the most common cause of dementia in elderly individuals. Currently, acetylcholinesterase inhibitors (AChEI) are the most effective clinical treatment for AD. AChEIs in natural products may have therapeutic potential and should be screened for use in AD treatment. The authors describe a simple and reliable method for AChEI screening by capillary electrophoresis (CE). A hexadimethrine bromide (HDB) solution was pushed into a capillary (0.015 MPa×10 s) and incubated for 5 min. The capillary was flushed with deionized water for 5 min to remove free HDB, followed by plugging with an acetylcholinesterase (AChE) solution. After a 5 min incubation, the AChE was immobilized on the positively charged coating by ion binding, and the micro-reactor was created. The substrate solution, acetylthiocholine iodide (AThC), was injected into the capillary and incubated in the micro-reactor for 1 min. The unreacted substrate and the enzymolysis product were separated by CE. Gastrodin, an important component of Gastrodia elata, can inhibit AChE activity. After a certain amount of gastrodin was spiked into the substance solution, the peak area of the product decreased. Greater peak area reduction indicated stronger inhibition of AChEI. We observed good reproducibility of the product peak, with relative standard deviation (RSD) values less than 5.3%. The micro-reactor can be reused up to 300 times, which greatly improves efficiency. When the concentration of gastrodin was 5.24 μmol/L, the inhibition rate of AChE reached 64.8%. The IC50 of gastrodin was (2.26±0.14) μmol/L (R2=0.9983), which was consistent with the result of traditional UV method (2.09±0.18 μmol/L). If the function of the micro-reactor deteriorates, it can be conveniently renewed by flushing the column to remove the enzyme and repeating the AChE immobilization protocol. The proposed method is simple, efficient, and low cost, and can be used to screen AChEI from natural products, thus contributing to the improvement of AD treatment.

Simultaneous IСP-MS determination of trace metals in natural water and snow after their preconcentration on novel adsorbent based on Al2O3 impregnated with Alizarin Complexone

ABSTRACTA simple and available method for on-line preparation of the adsorbent and solid phase extraction procedure for multi-element determination by inductively coupled plasma mass spectrometry (ICP-MS) has been proposed. The adsorbent (Al2O3-PB-AC) was synthesised by sequential impregnation of Al2O3 with Polybrene (PB) and Alizarin Complexone (AC). Cd(II), Cr(III), Ni(II), Pb(II) and Zn(II) were quantitatively recovered using Al2O3-PВ-AC from aqueous solutions at pH 6.5 and at a flow rate of 1.0 mL min−1. All the elements can easily be eluted by 3.0 mL 0.5 M HNO3. A mini-column packed with 0.100 g Al2O3-PB-AC retained all elements quantitatively from up to 50 mL multi-element solution with an enrichment factor of 16.7. The relative standard deviation for five replicate determinations was 3.5%, 7.3%, 6.2%, 7.4% and 4.7% for Cd(II), Cr(III), Ni(II), Pb(II) and Zn(II), respectively, for determination of 5 μg L−1 level. The developed method was applied for SPE-ICP-MS determination of Cd, Cr, Ni, Pb, Zn in snow and river waters.

Extracellular nucleic acid scavenging rescues rats from sulfur mustard analog-induced lung injury and mortality.

Sulfur mustard (SM) is a highly toxic war chemical that causes significant morbidity and mortality and lacks any effective therapy. Rats exposed to aerosolized CEES (2-chloroethyl ethyl sulfide; 10% in ethanol), an analog of SM, developed acute respiratory distress syndrome (ARDS), which is characterized by increased inflammation, hypoxemia and impaired gas exchange. We observed elevated levels of extracellular nucleic acids (eNA) in the bronchoalveolar lavage fluid (BALF) of CEES-exposed animals. eNA can induce inflammation, coagulation and barrier dysfunction. Treatment with hexadimethrine bromide (HDMBr; 10mg/kg), an eNA neutralizing agent, 2h post-exposure, reduced lung injury, inhibited disruption of alveolar-capillary barrier, improved blood oxygenation (PaO2/FiO2 ratio), thus reversing ARDS symptoms. HDMBr treatment also reduced lung inflammation in the CEES-exposed animals by decreasing IL-6, IL-1A, CXCL-1 and CCL-2 mRNA levels in lung tissues and HMGB1 protein in BALF. Furthermore, HDMBr treatment also reduced levels of lung tissue factor and plasminogen activator inhibitor-1 indicating reduction in clot formation and increased fibrinolysis. Fibrin was reduced in BALF of the HDMBr-treated animals. This was further confirmed by histology that revealed diminished airway fibrin, epithelial sloughing and hyaline membrane in the lungs of HDMBr-treated animals. HDMBr completely rescued the CEES-associated mortality 12h post-exposure when the survival rate in CEES-only group was just 50%. Experimental eNA treatment of cells caused increased inflammation that was reversed by HDMBr. These results demonstrate a role of eNA in the pathogenesis of CEES/SM-induced injury and that its neutralization can serve as a potential therapeutic approach in treating SM toxicity.

Application of polybrene to increase transfection efficiency of lentiviral vector in bone marrow derived dendritic cells

Objective To investigate the toxicity and optimal concentration of polybrene on lentiviral vector transduction of bone marrow derived dendritic cells (BMDCs). Methods The BMDCs suspensions in mice were prepared and randomly divided into experimental and control groups. The BMDCs in experimental groups were treated with different concentrations of polybrene, and those in control groups were added with phosphate buffer (PBS). Cell counting kit-8 (CCK-8) assay and annexin V-fluoresceine isothiocyanate (FITC)/propidium iodide (PI) staining were performed to detect cells viability and apoptosis. Add to BMDCs suspensions were added with lentiviral vector [multiplicity of infection (MOI)=10] and the mixture was divided into experimental and control groups. The mixture in experimental groups was treated with different concentrations of polybrene, and that in control groups was given PBS. The expression of green fluorescent protein (GFP) was observed by fluorescence microscope every day. After the BMDCs were stimulated to mature, the GFP expression was detected using flow cytometry. The t test was used for comparison between two groups. Results The number of living cells in experimental group was significantly less than in control group when polybrene ≥11 mg/L [The absorbance (A) values were 0.881±0.007 vs. 1.031±0.017, t=7.832, P<0.05]. The GFP expression in experimental groups was significantly higher than in control groups. The highest GFP expression was observed when polybrene at 8 mg/L. Conclusion Polybrane shows dose-dependent toxicity on BMDCs, and in the safe range of concentration, polybrane can significantly increase the transfection efficacy of lentiviral vector to BMDCs and the optimal concentration is 8 mg/L. Key words: Polybrene; Lentiviral vector; Bone marrow derived dendritic cells

Aqueous solubility of kinase inhibitors: II the effect of hexadimethrine bromide on the dovitinib/γ-cyclodextrin complexation

Dovitinib is a small molecule kinase inhibitor (KI) that is practically insoluble in water but demonstrates high antitumor activity. The aqueous solubility of dovitinib can be improved through formation of water-soluble γ-cyclodextrin (γCD) complexes. However, the very low intrinsic solubility hammers the complex formation. The aim of the present study was to investigate dovitinib binary and ternary complexes with both γCD and hexadimethrine bromide (HDMBr) in both aqueous solution and the solid state. The phase-solubility diagrams of dovitinib were of Bs-type. Addition of HDMBr to the complexation media, that is formation of ternary dovitinib/γCD/HDMBr complex, resulted in four-fold increase in the complexation efficacy (CE). The enhanced CE can decrease the amount of γCD needed to solubilize given amount of dovitinib. The solid dovitinib complexes were characterized by differential scanning calorimetry (DSC), X-ray powder diffraction (XRPD), Fourier transform infrared spectroscopy (FTIR), and in vitro dissolution. The dissolution test showed that γCD did stabilize supersaturated dovitinib solution.

Aqueous solubility of kinase inhibitors: I the effect of hydrophilic polymers on their γ-cyclodextrin solubilization

Most small-molecule protein kinase inhibitors (KIs) are neutral or weakly basic lipophilic compounds with very poor solubility in aqueous media that decreases their ability to permeate biological membranes from an aqueous exterior and consequently bestow them with low bioavailability after oral and topical administration. Cyclodextrin (CD) complexation can increase water-solubility of poorly soluble drugs without affecting their ability to permeate biological membranes. The aim of the present study was to enhance aqueous solubility of six KIs that is axitinib, cediranib, dovitinib, motesanib, pazopanib and regorafenib, through formation of drug/γ-cyclodextrin (γCD)/polymer ternary complexes. The complexes were prepared in an aqueous environment by heating technique. The phase-solubility profiles showed formation of binary KI/γCD complexes of Bs-type. γCD alone had significant solubilizing effect on dovitinib (stability constant (K1:1) = 684 M−1 and complexation efficacy (CE) = 0.011). The aqueous solubility of the binary KI/γCD complexes was pH-dependent. Hexadimethrine bromide (HDMBr) was the most effective polymer of the ones tested. The ternary complexes formed nano-sized aggregates with diameter between 200 and 400 nm. Osmolarity and pH were also monitored. TEM study showed formation of complex aggregates in aqueous media.

Screening of inhibitors against histone demethylation jumonji domain-containing protein 3 by capillary electrophoresis

Jumonji domain-containing proteins (JMJDs) play an important role in the epigenetic regulation of gene expression. Aberrant regulation of histone modification has been observed in the progression of a variety of diseases, such as neurological disorders and cancer. Therefore, discovery of selective modulators of JMJDs is very attractive in new drug discovery. Herein, a simple capillary electrophoresis (CE) method was developed for screening of inhibitors against JMJD3. A known JMJD3 inhibitor GSK-J1, 5-carboxyfluorescein labeled substrate peptide with an amino acid sequence of KAPRKQLATKAARK(me3)SAPATGG (truncated from histone H3), as well as a small chemical library composed of 37 purified natural compounds and 30 natural extracts were used for method development and validation. The separation of substrate from its demethylated product was achieved by addition of polycation hexadimethrine bromide (HDB) in the running buffer. The enzyme activity was thus assayed accurately through separating the demethylated product from the substrate and then measuring the peak area of the product. The enzyme inhibition can be read out by comparing the peak area of the demethylated product obtained in the present of inhibitors and that of the negative control in the absence of any inhibitor. The merit of the method is proved by discovering two new JMJD3 inhibitors: salvianic acid A and puerarin 6′’-O-xyloside.

Vortex generation in electroosmotic flow in a straight polydimethylsiloxane microchannel with different polybrene modified-to-unmodified section length ratios

Previous studies on electrokinetically generated vortices generally involve using relatively complicated heterogeneous surface charge patterns (with zeta potentials of different polarities). In this paper, vortex formation in a straight channel with zeta potentials of two different values, but the same polarity was investigated. Particularly, the effects of the length ratio of the polybrene (PB)-modified section to the unmodified section on vortex formation in electroosmotic flow in a straight polydimethylsiloxane (PDMS) microchannel were studied numerically and experimentally. The numerical results show that for the 5% PB-modified channel (with a zeta potential of about − 5 mV), a vortex will be formed when the length ratio of the modified section to the unmodified section was larger than 4. Such results were experimentally verified with tracing particles. The critical length ratio decreases with the decrease in the absolute value of the zeta potential of PB-modified section and increases with the increased channel width. The results presented in this paper are valuable for understanding vortex formation in a straight channel which is partially modified with PB.