Low Molecular Weight Polyethylenimine Research Articles

Development of poly(ethyleneimine) grafted amphiphilic copolymers: Evaluation of their cytotoxicity and ability to complex DNA

Poly(ethyleneimine) (PEI) is one of the most widely used cationic polymers for gene delivery. The high molecular weight polymer, which is commercially available, is highly efficient but also very cytotoxic. The reduction in charge density by using nonlinear architectures based on low molecular weight (LMW) PEI is a promising approach to produce safer DNA-vectors. Herein, a group of cationic graft copolymers with different composition containing a hydrophobic biocompatible backbone and LMW linear PEI (lPEI) grafts obtained by ring opening polymerization and click chemistry was studied. The self-assembly and DNA complexation behavior of these materials was analyzed by the gel retardation assay, zeta potential measurements, and dynamic light scattering. The copolymers formed positively charged particles in water with average sizes between 270 and 377 nm. After they were added to DNA in serum-free medium, these particles acquired negative/near-neutral charges and increased in size depending on the N/P ratio. All copolymers showed reduced cytotoxicity compared to the 25 kDa lPEI used as reference, but the transfection efficiency was reduced. This result suggested that the cationic segments were too small to fully condense the DNA and promote cellular uptake, even with the use of several grafts and the introduction of hydrophobic domains. The trends found in this research showed that a higher degree of hydrophobicity and a higher grafting density can enhance the interaction between the copolymers and DNA. These trends could direct further structural modifications in the search for effective and safe vectors based on this polycation.

Removing free-floating oil from water using cationic polymers/surfactant-modified silica

Abstract The purpose of this work was to evaluate the efficiency of oil sorption of silica particles modified by three different types of cationic polymers and a cationic surfactant. Low-molecular-weight polyethyleneimine (LPEI), high-molecular-weight polyethyleneimine (HPEI), polydiallyldimethylammonium chloride (PDM), and cetyltrimethylammonium bromide (CTAB) were used to modify the silica particles and then compared their performances for oil removal. The scanning electron microscope and zeta potential measurements were used to analyze the surface characteristics of unmodified and modified silica particles. Adsorptions of motor oil and palm oil on the modified silica particles have been investigated under various parameters such as the silica particle size, the oil concentration, the polymer/surfactant concentrations, and the pH. The results have shown that the modified silica particles enhanced the oil sorption ability by approximately 10–20 times depending on the size of silica particles, pH, and the type of polymer/surfactant used when compared with the unmodified silica particles. The highest palm oil adsorption values of LPEI-silica, HPEI-silica, PDM-silica, CTAB-silica, and unmodified silica were 2.40, 2.10, 1.95, 1.50, and 0.15 g/gsilica, respectively. Moreover, the oil sorption of the modified silica particles was increased by approximately 30–50% for the smallest-sized silica particles.

Therapeutic delivery of siRNA with polymeric carriers to down-regulate STAT5A expression in high-risk B-cell acute lymphoblastic leukemia (B-ALL).

Overexpression and persistent activation of STAT5 play an important role in the development and progression of acute lymphoblastic leukemia (ALL), the most common pediatric cancer. Small interfering RNA (siRNA)-mediated downregulation of STAT5 represents a promising therapeutic approach for ALL to overcome the limitations of current treatment modalities such as high relapse rates and poor prognosis. However, to effectively transport siRNA molecules to target cells, development of potent carriers is of utmost importance to surpass hurdles of delivery. In this study, we investigated the use of lipopolymers as non-viral delivery systems derived from low molecular weight polyethylenimines (PEI) substituted with lauric acid (Lau), linoleic acid (LA) and stearic acid (StA) to deliver siRNA molecules to ALL cell lines and primary samples. Among the lipid-substituted polymers explored, Lau- and LA-substituted PEI displayed excellent siRNA delivery to SUP-B15 and RS4;11 cells. STAT5A gene expression was downregulated (36–92%) in SUP-B15 and (32%) in RS4;11 cells using the polymeric delivery systems, which consequently reduced cell growth and inhibited the formation of colonies in ALL cells. With regard to ALL primary cells, siRNA-mediated STAT5A gene silencing was observed in four of eight patient cells using our leading polymeric delivery system, 1.2PEI-Lau8, accompanied by the significant reduction in colony formation in three of eight patients. In both BCR-ABL positive and negative groups, three of five patients demonstrated marked cell growth inhibition in both MTT and trypan blue exclusion assays using 1.2PEI-Lau8/siRNA complexes in comparison with their control siRNA groups. Three patient samples did not show any positive results with our delivery systems. Differential therapeutic responses to siRNA therapy observed in different patients could result from variable genetic profiles and patient-to-patient variability in delivery. This study supports the potential of siRNA therapy and the designed lipopolymers as a delivery system in ALL therapy.

Benign prostatic hyperplasia therapy through liquisolid technology composed of polymer-layered nanocomposites based on silicate that contain babassu oil and copaiba oil-resin

In a previous article by our research group, the association of babassu oil (BBS) and copaiba oil-resin (COPA) in a nanosystem was verified as a potential phytotherapic alternative for treatment and prophylaxis of benign prostatic hyperplasia (BPH). Since few studies have examined their association with nanosystems, we developed polymeric nanocomposites of lamellar silicates containing BBS and COPA to potentially enhance their anti-hyperplastic activity through a process that would be amenable to larger scale production. Organoclay-based-nanocomposites were prepared by an intercalation method in a polyethyleneimine (PEI) solution of low or high molar weight polymer and BBS alone or together with COPA, aiming to develop a technology capable of transforming liquid or oily active pharmaceutical ingredients into suitable powders for compression. A nanocomposite of Viscogel S7® containing BBS-COPA with low molecular weight PEI, presented the highest intercalation efficiency values of BBS (83.08 ± 0.10%) and COPA (70.14 ± 0.88%) along with the highest values of basal spacing (22.12 ± 0.87 Å) and the nanocomposite presented particle diameter suitable for obtaining compactable solids. The produced tablets presented an average weight and hardness within established limits and a rapid dissolution profile in simulated gastric and enteric fluids for both oils. The anti-hyperplasic effect of the BBS-COPA-nanocomposite led to a reduction of the prostate weight (4.53 ± 2.07%), which was similar to results obtained with Finasteride (5.23 ± 2.09%). The histopathological patterns of prostrates from animals treated with the nanocomposite closely matched those observed with control animals. In conclusion, the newly developed nanosystem showed promising potential to reduce the pathological process of BPH and could serve as a natural alternative for treatment.

A diaminoethane motif bearing low molecular weight polymer as a new nucleic acid delivery agent

Among polymer-based gene delivery systems, poly(ethylene imine) (PEI) stands out as an effective polycation. However, the toxic effects of PEI especially at higher molecular weights limit its usage. Although the effects of PEI's architecture and molecular weight on gene delivery is controversial in literature, low molecular weight PEI appears to be efficient at transfection while having lower toxicity. Herein, as an alternative to low molecular weight, linear PEI, a methacrylate polymer bearing diamimoethane motifs, poly(2-((2-aminoethyl)amino)ethyl methacrylate) (P(AEAEMA)), was evaluated in vitro as a new nucleic acid delivery agent. P(AEAEMA) (8 kDa) showed low toxicity on Skov-3-luc and NIH/3T3 cell lines at polymer concentrations where PEI (8 kDa) was highly toxic. P(AEAEMA) could efficiently form complexes with siRNA at an N/P ratio of 2 as shown by gel electrophoresis. The diameter of P(AEAEMA)-siRNA complexes was found to be significantly lower than PEI-siRNA complexes almost at all tested N/P ratios. P(AEAEMA) could improve the stability of siRNA in serum containing media by protecting the siRNA against serum nucleases. siRNA and pDNA transfection efficiency of P(AEAEMA) on luciferase expressing Skov-3-luc cell line and HEK 293T cell line, respectively was found to be comparable to well-known nucleic acid carrier, PEI. The transfection efficiency of both P(AEAEMA) and PEI was found to be cell-type-dependent. None of the polymers were able to transfect MDA-MB-231 cells with siRNA or pDNA.

Investigation of water-insoluble hydrophobic polyethylenimines as RNAi vehicles in chronic myeloid leukemia therapy.

The discovery of RNA interference (RNAi) more than two decades ago opened avenues for avant-garde cancer treatments that possess the ability to evade issues hampering current chemotherapeutic strategies, owing to its specific gene sequence-driven mechanism of action. A potent short interfering RNA (siRNA) delivery vehicle designed to overcome physiological barriers is imperative for successful RNAi therapy. For this purpose, this study explored the characteristics and therapeutic efficacy of low-molecular weight (MW) polyethylenimine (PEI) with high cholesterol substitution, yielding water-insoluble polymers, in chronic myeloid leukemia (CML) K562 cells. A strong impact of cholesterol grafting on the physicochemical attributes of the resultant polymers and their corresponding complexes with siRNA was observed, with the siRNA binding capacity of polymers increasing and complex dissociation sensitivity decreasing with increase in cholesterol content of the polymers. The modified polymer complexes were significantly smaller in size and possessed higher cationic charge compared to the parent polymer. The interaction with anionic heparan sulfate preoteoglycans present on the cell surface was significant in cellular uptake of the complexes. The therapeutic efficacy of siRNA/polymer complexes was reflected in their ability to effectively silence the reporter green fluorescent protein gene and endogenous CML oncogene BCR-ABL as well as significantly inhibit colony formation by K562 cells post BCR-ABL silencing. The results of this study demonstrated beneficial effects of high levels of hydrophobic substitution on low MW PEI on their functional performance bestowing them the potential to be potent RNAi agents for CML therapy.

Manipulating the solution environment to control the surface roughness of elastin-based polymer coatings

Elastin-like polypeptides (ELP) have been used as a genetically-engineered, biocompatible substitute for elastin. Cell culture coatings prepared using ELP conjugated to low molecular weight polyethyleneimine (PEI) entices cells to form three-dimensional cellular aggregates that mimic their in vivo counterparts. This study seeks to control the deposition of the ELP and ELP-PEI molecules to control the roughness of the final coatings. The two polymers were coated onto three different substrates (glass, polystyrene, tissue-culture polystyrene) and the solution environment was altered by changing the polymer concentration (0.5, 1.0, 1.5 mg/mL) and/or salt concentration (None, 0.2 M phosphate buffered saline) for a total of 36 conditions. Atomic force microscopy (AFM) was used to measure the average roughness (Ra) of the samples and found that ELP coated samples had a higher Ra than their ELP-PEI counterparts. The coatings were tested for stability by performing cell culture media changes every three days for 11 days. AFM showed that the average roughness of the tested samples increased with each media change. To address this, the surfaces were crosslinked using hexamethyl diisocyanate, which minimized the change in surface roughness even when subjected to an intense sonication process. This study provides parameters to achieve elastin-based coatings with controlled roughness that can be used to support stable, long-term in vitro cell culture.

Development of Covalent Chitosan-Polyethylenimine Derivatives as Gene Delivery Vehicle: Synthesis, Characterization, and Evaluation.

There is an increasing interest in cationic polymers as important constituents of non-viral gene delivery vectors. In the present study, we developed a versatile synthetic route for the production of covalent polymeric conjugates consisting of water-soluble depolymerized chitosan (dCS; MW 6–9 kDa) and low molecular weight polyethylenimine (PEI; 2.5 kDa linear, 1.8 kDa branched). dCS-PEI derivatives were evaluated based on their physicochemical properties, including purity, covalent bonding, solubility in aqueous media, ability for DNA condensation, and colloidal stability of the resulting polyplexes. They were complexed with non-integrating DNA vectors coding for reporter genes by simple admixing and assessed in vitro using liver-derived HuH-7 cells for their transfection efficiency and cytotoxicity. Using a rational screening cascade, a lead compound was selected (dCS-Suc-LPEI-14) displaying the best balance of biocompatibility, cytotoxicity, and transfection efficiency. Scale-up and in vivo evaluation in wild-type mice allowed for a direct comparison with a commercially available non-viral delivery vector (in vivo-jetPEI). Hepatic expression of the reporter gene luciferase resulted in liver-specific bioluminescence, upon intrabiliary infusion of the chitosan-based polyplexes, which exceeded the signal of the in vivo jetPEI reference formulation by a factor of 10. We conclude that the novel chitosan-derivative dCS-Suc-LPEI-14 shows promise and potential as an efficient polymeric conjugate for non-viral in vivo gene therapy.

Polyethyleneimine as a surface activator for low surface energy substrates bonded with cyanoacrylate adhesives

The modification of surface properties of polypropylene (PP) substrates has attracted much attention in fundamental science and industrial applications. Primer treatment is one of the most simple and facile surface modification methods. This study used polyethyleneimine (PEI) as a surface activator for bonding polypropylene substrates with ethyl cyanoacrylate (ECA) adhesive. We have understood the effect of various factors such as the open time, solvent composition, optimum concentration and molecular weight of the primer. The bonding strength was maximum when the substrates were bonded immediately on priming. It was also observed that the solvent composition plays a minor role in the development of bond strength. The bond strength of substrates coated with a primer having a PEI concentration as low as 0.05% showed better results and was, in fact, three times better than the unprimed ones. It was also concluded that a low molecular weight PEI performs better than one having a higher molecular weight. DSC results confirmed that an improvement in strength is due to an increase in the molecular weight of the ECA polymer, which occurs in the presence of PEI molecules.

Polyethylenimine-modified graphitic carbon nitride nanosheets: a label-free Raman traceable siRNA delivery system.

Since the nanotoxicity of gene delivery carriers has raised world-wide concerns, it is important to trace their intracellular performance, for example via uptake visualization. Here, we develop a novel ultrathin graphitic carbon nitride (g-C3N4) composite nanosystem for label-free Raman-traceable small interfering RNA (siRNA) delivery. Through low molecular weight polyethylenimine (PEI) modifications, these nanosystems can obtain siRNA loading capabilities. The lateral size of the PEI-g-C3N4 composite is around 100-150 nm with a thickness of nearly 0.6 nm. The designed label-free delivery system could avoid possible obstacles associated with artificial labels and it shows cytotoxicity toward cancer cells and good biocompatibility in normal human cells. The label-free PEI-g-C3N4 gene nanocarrier can be directly traced via Raman microscopy, which makes it suitable for intracellular visualization. Intracellular uptake of the self-fluorescent g-C3N4 nanosheets can also be traced via fluorescence imaging. The PEI modified g-C3N4 ultrathin nanosheets possess gene delivery capacity together with unique dual-traceable Raman and fluorescence features. Raman traces not only have higher specificity than fluorescence ones but they can also avoid background noises. Thus, they may replace widely implemented fluorescence tracing. This work could provide a label-free traceable platform for investigating the intracellular performances of gene delivery nanosystems.

Zn(ii)-Dipicolylamine analogues with amphiphilic side chains endow low molecular weight PEI with high transfection performance.

To investigate the effect of amphiphilic balance of Zn(ii)-dipicolylamine analogues on the transfection process, we fabricated a series of Zn(ii)-dipicolylamine functional modules (DDAC-Rs) with different hydrophilic-phobic side chains to modify low molecular weight PEI (Zn-DP-Rs) by the Michael addition reaction. Zn-DP-Rs with hydrophilic terminal hydroxy group side chains demonstrate superior overall performance compared to those of hydrophobic alkyl side chains. In terms of the influence of the chain lengths in DDAC-Rs, from Zn-DP-A/OH-3 to Zn-DP-A/OH-5, the corresponding transfection efficiency shows an upward trend as the lengths increase. However, decreasing efficacy is observed with further increase in the length of side chains. In addition, the Zn-DP-Rs with amphiphilic side chains show prominent performance in every respect, highlighting the significance of balance in the amphipathy of side chains in DDAC-Rs. This work is of great significance for the development of polycationic gene carrier materials with excellent performance.

Treatment of Colon Cancer by Degradable rrPPC Nano-Conjugates Delivered STAT3 siRNA.

BackgroundDrugs that work based on the mechanism of RNA interference have shown strong potential in cancer gene therapy. Although significant progress has been made in small interfering RNA (siRNA) design and manufacturing, ideal delivery system remains a limitation for the development of siRNA-based drugs. Particularly, it is necessary to focus on parameters including delivery efficiency, stability, and safety when developing siRNA formulations for cancer therapy.MethodsIn this work, a novel degradable siRNA delivery system cRGD-R9-PEG-PEI-Cholesterol (rrPPC) was synthesized based on low molecular weight polyethyleneimine (PEI). Functional groups including cholesterol, cell penetrating peptides (CPPs), and poly(ethylene oxide) were introduced to PEI backbone to attain enhanced transfection efficiency and biocompatibility.ResultsThe synthesized rrPPC was dispersed as nanoparticles in water with an average size of 195 nm and 41.9 mV in potential. rrPPC nanoparticles could efficiently deliver siRNA into C26 clone cancer cells and trigger caveolae-mediated pathway during transmembrane transportation. By loading the signal transducer and activator of transcription 3 (STAT3) targeting siRNA, rrPPC/STAT3 siRNA (rrPPC/siSTAT3) complex demonstrated strong anti-cancer effects in multiple colon cancer models following local delivery. In addition, intravenous (IV) injection of rrPPC/siSTAT3 complex efficiently suppressed lung metastasis tumor progression with ideal in vivo safety.ConclusionOur results provide evidence that rrPPC nanoparticles constitute a potential candidate vector for siRNA-based colon cancer gene therapy.

Nanoparticle-complexed antimiRs for inhibiting tumor growth and metastasis in prostate carcinoma and melanoma

BackgroundMiRNAs act as negative regulators of gene expression through target mRNA degradation or inhibition of its translation. In cancer, several miRNAs are upregulated and play crucial roles in tumorigenesis, making the inhibition of these oncomiRs an interesting therapeutic approach. This can be achieved by directly complementary single-stranded anti-miRNA oligonucleotides (antimiRs). A major bottleneck in antimiR therapy, however, is their efficient delivery. The nanoparticle formation with polyethylenimine (PEI) may be particularly promising, based on the PEI’s ability to electrostatically interact with oligonucleotides. This leads to their protection and supports delivery. In the present study, we explore for the first time PEI for antimiR formulation and delivery. We use the branched low molecular weight PEI F25-LMW for the complexation of different antimiRs, and analyse tumor- and metastasis-inhibitory effects of PEI/antimiR complexes in different tumor models.ResultsIn prostate carcinoma, transfection of antimiRs against miR-375 and miR-141 leads to tumor cell inhibition in 2D- and 3D-models. More importantly, an in vivo tumor therapy study in prostate carcinoma xenografts reveals anti-tumor effects of the PEI/antimiR complexes. In advanced melanoma and metastasis, we identify by a microRNA screen miR-150 as a particularly relevant oncomiR candidate, and validate this result in vitro and in vivo. Again, the systemic application of PEI/antimiR complexes inhibiting this miRNA, or the previously described antimiR-638, leads to profound tumor growth inhibition. These effects are associated with the upregulation of direct miRNA target genes. In a melanoma metastasis mouse model, anti-metastatic effects of PEI/antimiR treatment are observed as well.ConclusionsWe thus describe PEI-based complexes as efficient platform for antimiR therapy, as determined in two different tumor entities using in vivo models of tumor growth or metastasis. Our study also highlights the therapeutic relevance of miR-375, miR-141, miR-150 and miR-638 as target miRNAs for antimiR-mediated inhibition.

CSIG-11. TARGETING PD-L1 IN GLIOBLASTOMA USING NANOPARTICLE-BASED GENE EDITING

Abstract Glioblastoma multiforme (GBM) is an astrocyte derived brain tumor with very poor prognosis, usually with a less than one year survival rate. Immunotherapy has shown promising therapeutic potentials in research and clinical application, however, GBM associated immunosuppressive microenvironment creates a significant barrier for effective anti-GBM immune responses. The immune checkpoint PD-1/PD-L1 pathways play a critical role in tumor-induced immunosupression to evade immune surveillance. Gene-editing tools targeting PD-1/PD-L1 pathway have gained increased research attention, however the major challenge is how to effectively deliver gene-editing tools without causing adverse effects for clinical translation. We have developed a nanoparticle (NP) delivery system from a low molecular weight PEI lipid shell and a PLGA core that can package PD-L1 gRNA-CRISPR/Cas9 plasmid to transfect human U87 glioma cells overexpressing PD-L1. PD-L1 gRNA-CRISPR/Cas9 plasmid is constructed by inserting single guide targeting PD-L1 sequence into GFP CRISPR/Cas9 plasmid for visualizing transfection efficacy. NP is labelled with Rhodamine 6G to monitor cellular uptake and trafficking. Fluorescence microscopy shows human U87 glioma cells can quickly uptake PD-L1 GFP-CRISPR/Cas9 plasmid-NPs within 2 hours as indicated by the Rhodamine 6G. GFP expression was obtained after 48 hours of transfection and maintained up to 7 days without causing toxicity. Western blot analysis confirmed successful knockdown of PD-L1 in U87 cells. These findings reveal that NPs made by a cationic branched PEI lipid shell and a PLGA core are non-toxic and efficient in delivering CRISPR/Cas9 system to U87 cells. Editing of pathological gene in human glioma cells with PD-L1 GFP-CRISPR/Cas9 plasmid using NPs as a delivery system may provide a novel immunotherapy platform to treat GBM.

Supramolecular self-assemblies of engineered polyethylenimines as multifunctional nanostructures for DNA transportation with excellent antimicrobial activity

In this study, indole-3-butanoic acid (IBA), a biologically and environmentally safe entity, has been grafted onto low and high molecular weight (1.8 and 25 kDa) polyethylenimines (PEI) mainly through primary amines to obtain amphiphilic indole-3-butanoyl-polyethylenimines (IBPs). Two series of IBPs (IBP1.8 and IBP25) were prepared which, on self-assembly in aqueous medium, yielded multifunctional nanomicellar structures (IBP1.8 and IBP25) capable of transporting genetic material in vitro and exhibiting other biological activities. Physicochemical characterization showed the size of IBP1.8 and IBP25 nanostructures in the range of ~332–234 nm and ~283–166 nm, respectively, with zeta potential varying from ~+29–17 mV and ~+37–25 mV. DNA release assay demonstrated higher release of plasmid DNA from IBP nanostructures as compared to native PEIs. Cytotoxicity showed a decreasing pattern with increasing degree of grafting of IBA onto PEIs making these nanostructures non-toxic. pDNA complexes of these nanostructures (both IBPs1.8 and IBPs25) displayed considerably higher transfection efficiency, however, IBP1.8/pDNA complexes performed much better (~7–9 folds) as compared to native PEI/pDNA and Lipofectamine/pDNA complexes on mammalian cells. CLSM analysis revealed that these complexes entered nucleus in sufficient amounts suggesting higher uptake and efficient internalization of the complexes. Besides, these supramolecular nanostructures not only exhibited excellent antimicrobial potential (MIC ~49–100 µg/ml) against clinical as well as resistant pathogenic strains but also found to possess antioxidant property. Overall, the projected low molecular weight PEI-based vectors could serve as more effective multifunctional nanomaterials having promising potential for future gene therapy applications with capability to provide protection against other bacterial infections.

Low molecular weight polyethyleneimine modified by 2-aminoimidazole achieving excellent gene transfection efficiency

The transfection efficiency and cytotoxicity of polyethyleneimine (PEI), as a non-viral vector, increase with molecular weight. Low molecular weight PEI, such as branched PEI1800 (Mw ~ 1800 kDa), has negligible transfection efficiency but low cytotoxicity. Given the low efficiency of PEI1800, a variety of modifications were explored. A structure-integrated group of guanidyl and imidazolyl, 2-aminoimidazole was designed (Scheme 1). To verify the molecular interaction between 2-aminoimidazole and gene, molecular docking experiment, 1H NMR, FTIR and isothermal titration calorimetry (ITC) measurement were carried out. Studies confirmed that 2-aminoimidazole form double hydrogen bonds with phosphate group, which enhanced the thermodynamic interaction between 2-aminoimidazole and gene. Hence, we synthesized 2-aminoimidazole modified PEI1800, and as controls, guanidyl and imidazolyl modified PEI1800 were synthesized respectively. As expected, the results showed that 2-aminoimidazole modified PEI1800 could condense DNA into a higher positively charged complex with 116 ~ 120 nm in diameter and improve the cellular uptake. EGFP assay and luciferase assay showed that 2-aminoimidazole modified PEI1800 exhibited much better transfection than PEI1800, guanidyl modified PEI1800, imidazolyl modified PEI1800 both in HEK 293 T cells and SGC-7901 cells. Besides, MTT assay showed 2-aminoimidazole modified PEI1800 exhibited better biocompatibility than PEI25 K. These results highlight the critical role that 2-aminoimidazole played in non-viral gene delivery.

“Building-block crosslinking” micelles for enhancing cellular transfection of biocompatible polycations

Incorporating functional ligands and biodegradable bonds into biocompatible low-molecular-weight (LMW) polymers, such as 1.8 kDa poly(ethylenimine) (PEI 1.8k), is a common strategy to improve the properties of LMW polymers including biosafety and delivery efficacy. This study demonstrates the hypothesis that introducing different functional ligands and linked reductive disulfides in PEI 1.8k will achieve superior siRNA transfection efficiency. By incorporating PEI-X (X represents cholesterol (Ch), heptafluorobutyric anhydride (HFBA, F) and 4-carboxyphenyl-boronic acid (PBA)) functional ligands into PEI 1.8k and subsequently crosslinking with each other via disulfide bond links, reductive-responsive PEI-X-SS-X-PEI copolymers were constructed to enhance the cellular transfection via the synergistic effect of the high affinity of Ch, F and PBA to cell membranes and the disulfide reduction triggered intracellular disassembly of micelles and subsequent siRNA release. Extraordinarily, ternary Ch-SS-F-SS-PBA micelles exhibited the strongest siRNA transfection efficiencies in in vitro cell experiments and in vivo animal experiments due to the coordination of enhanced serum stability, promoted cell uptake and endosomal escape, and cell targeting ability. This strategy of constructed multifunctional polymer here we called “building-block crosslinking” showed a simple and smart way to synthesize new materials. Also this strategy of constructing ligands-directed reduction-sensitive micelles improves the transfection efficiency of LMW PEI and provides a valuable insight to develop novel gene delivery systems.

Low molecular weight polyethyleneimine conjugated guar gum for targeted gene delivery to triple negative breast cancer

The study emphasized on the development of an efficient, receptor-targeted non-viral gene delivery vehicle for gene therapy of triple negative breast cancer (TNBC). Here, naturally abundant guar gum based non-viral carrier was developed through conjugating by low molecular weight polyethylenimine (LPEI) (GNP) using napthalic anhydride coupling agent and characterized them by FT-IR, 1H NMR, XRD and UV spectrophotometer. The carrier was found to be cytocompatible as revealed by MTT assay against MDA-MB-231 and HeLa cell lines and excellent blood compatibility till the concentration of 200 μg/ml. In addition to these, the carrier exhibited excellent gene binding capability and formed spherical shaped complexes. The carrier showed very high in vitro transfection efficiency in TNBC cell (MDA-MB-231) compared to lipofectamine 2000 (LF2k) which could be justified by its high buffering capacity. Therefore, GNP may be an attractive non-viral gene carrier for gene therapy of TNBC in future.

A Co-Delivery System of Curcumin and p53 for Enhancing the Sensitivity of Drug-Resistant Ovarian Cancer Cells to Cisplatin.

In order to enhance the sensitivity of drug-resistant ovarian cancer cells to cisplatin (DDP), a co-delivery system was designed for simultaneous delivery of curcumin (CUR) and p53 DNA. Firstly, the bifunctional peptide K14 composed of tumor targeting peptide (tLyP-1) and nuclear localization signal (NLS) was synthesized. A nonviral carrier (PEI-K14) was synthesized by cross-linking low molecular weight polyethyleneimine (PEI) with K14. Then, CUR was coupled to PEI-K14 by matrix metalloproteinase 9 (MMP9)-cleavable peptide to prepare CUR-PEI-K14. A co-delivery system, named CUR-PEI-K14/p53, was obtained by CUR-PEI-K14 and p53 self-assembly. Furthermore, the physicochemical properties and gene transfection efficiency were evaluated. Finally, ovarian cancer cisplatin-resistant (SKOV3-DDP) cells were selected to evaluate the effect of CUR-PEI-K14/p53 on enhancing the sensitivity of drug-resistant cells to DDP. The CUR-PEI-K14/DNA complexes appeared uniformly dispersed and spherical. The particle size was around 20–150 nm and the zeta potential was around 18–37 mV. It had good stability, high transfection efficiency, and low cytotoxicity. CUR-PEI-K14/p53 could significantly increase the sensitivity of SKOV3-DDP cells to DDP, and this effect was better as combined with DDP. The sensitizing effect might be related to the upregulation of p53 messenger RNA (mRNA), the downregulation of P-glycoprotein (P-gp) mRNA, and the upregulation of BCL2-Associated X (bax) mRNA. CUR-PEI-K14/p53 can be used as an effective strategy to enhance the sensitivity of drug-resistant ovarian cancer cells to DDP.

Novel fluorinated polycationic delivery of anti-VEGF siRNA for tumor therapy

Small-interfering RNA (siRNA) can specifically silence disease gene expression, bringing hope for the effective treatment of gene-related diseases. However, its use in vivo is limited due to the lack of efficient carriers. Therefore, it is necessary to construct carriers with high efficiency, low toxicity and serum stability. As a promising polycation carrier, polyethylenimine (PEI) can be further modified with a fluorine-containing alkyl chain that brings hydrophobic and oleophobic characteristics to its surface. In this study, low molecular weight PEI 1.8 kDa was selected and fluorinated through an anhydride reaction, and the product was named PEIF. In vitro experiments have shown that PEIF/siRNA polyplexes have suitable and stable particle size and potential, compress nucleic acids at a very low w/w ratio, and have the ability to effectively silence specific genes with low cytotoxicity. Compared with the polyplexes prepared using PEI 25 kDa, the PEIF/siRNA polyplexes were more stable in serum, showed a better antiangiogenic ability in in vivo experiments, and had a better tumor inhibition effect. The above results indicate that fluoropolymers based on the fluorine effect have great potential as gene delivery carriers for tumor therapy.