Co-delivery Of Genes Research Articles

A cationic amino acid polymer nanocarrier synthesized in supercritical CO2 for co-delivery of drug and gene to cervical cancer cells

The present study was undertaken to investigate the ability of a drug curcumin-loaded polymer to inhibit the growth of cervical cancer cells by enhancing the anti-cancer efficiency of curcumin. We synthesized poly(methacryloyl beta-alanine) (PMBA) as a nanocarrier by radical polymerization in supercritical CO2. The results showed that the curcumin encapsulated and folic acid (FA)-treated PMBA (Poly@Cur-FA) for 24 h activated the reactive oxygen species-mediated programmed cell death machinery in HeLa cells. This remarkable effect of Poly@Cur-FA treatment was visualized using different fluorescent probes, which demonstrated that the Poly@Cur-FA treatment disrupted the cell membrane, as also supported by scanning electron microscopy observations. The effect of Poly@Cur-FA dispersion on the cells was observed under a transmission electron microscope. Further, the HeLa cells were treated with the polymer encapsulated curcumin and Bcl2 siRNA (Pol-Cur-siRNA) for 24 h, which effectively suppressed the Bcl2 and simulated the autophagic pathway. This co-delivery system was designed to inhibit curcumin efflux and can enhance the treatment efficacy by targeting multiple signaling pathways, including cell cycle, apoptotic, and autophagic pathways. Collectively, the Pol-Cur-siRNA system appears to offer an efficient combinational therapeutic strategy that might overcome the problems associated with the chemosensitivity against the standard synthetic anti-cancer drugs. To support the experimental data, an artificial neural network model was developed to foresee the drug and gene release behaviors.

Pre-transplantation of Bone Marrow Mesenchymal Stem Cells Amplifies the Therapeutic Effect of Ultrasound-Targeted Microbubble Destruction-Mediated Localized Combined Gene Therapy in Post-Myocardial Infarction Heart Failure Rats.

Although stem cell transplantation and single-gene therapy have been intensively discussed separately as treatments for myocardial infarction (MI) hearts and have exhibited ideal therapeutic efficiency in animal models, clinical trials turned out to be disappointing. Here, we deliver sarcoplasmic reticulum Ca2+-ATPase 2a (SERCA2a) and connexin 43 (Cx43) genes simultaneously via an ultrasound-targeted microbubble destruction (UTMD) approach to chronic MI hearts that have been pre-treated with bone marrow mesenchymal stem cells (BMSCs) to amplify cardiac repair. First, biotinylated microbubbles (BMBs) were fabricated, and biotinylated recombinant adenoviruses carrying the SERCA2a or Cx43 gene were conjugated to the surface of self-assembled BMBs to form SERCA2a-BMBs, Cx43-BMBs or dual gene-loaded BMBs. Then, the general characteristics of these bubbles, including particle size, concentration, contrast signal and gene loading capacity, were examined. Second, a rat myocardial infarction model was created by ligating the left anterior descending coronary artery and injecting BMSCs into the infarct and border zones. Four weeks later, co-delivery of SERCA2a and Cx43 genes to the infarcted heart were delivered together to the infarcted heart using the UTMD approach. Cardiac mechano-electrical function was determined 4 wk after gene transfection, and the infarcted hearts were collected for myocardial infarct size measurement and detection of expression of SERCA2a, Cx43 and cardiac-specific markers. Finally, to validate the role of BMSC transplantation, MI rats transplanted or not with BMSCs were transfected with SERCA2a and Cx43, and the cardiac mechano-electrical function of these two groups of rats was recorded and compared. General characteristics of the self-assembled gene-loaded BMBs were qualified, and the gene loading rate was satisfactory. The self-assembled gene-loaded BMBs were in microscale and exhibit satisfactory dual-gene loading capacity. High transfection efficiency was achieved under ultrasound irradiation in vitro. In addition, rats in which SERCA2a and Cx43 were overexpressed simultaneously had the best contractile function and electrical stability among all experimental groups. Immunofluorescence assay revealed that the levels of SERCA2a and/or Cx43 proteins were significantly elevated, especially in the border zone. Moreover, compared with rats that did not receive BMSCs, rats pre-treated with BMSCs have better mechano-electrical function after transfection with SERCA2a and Cx43. Collectively, we report a promising cardiac repair strategy for post-MI hearts that exploits the providential advantages of stem cell therapy and UTMD-mediated localized co-delivery of specific genes.

H2O2-responsive VEGF/NGF gene co-delivery nano-system achieves stable vascularization in ischemic hindlimbs

Peripheral vascular disease (PVD) is a common clinical manifestation of atherosclerosis. Vascular endothelial growth factor (VEGF) gene therapy is a promising approach for PVD treatment. However, due to single-gene therapy limitations and high H2O2 pathological microenvironment, VEGF gene therapy are not as expectations and its clinical application are limited. Synergistic effects of Nerve factors and vascular factors in angiogenesis have attracted attention in recent years. In this study, VEGF and nerve growth factor (NGF) genes co-delivery nanoparticles (VEGF/NGF-NPs) were prepared by using H2O2 responsive 6s-PLGA-Po-PEG as a carrier. 6s-PLGA-Po-PEG could react with H2O2 specifically due to the internal peroxalate bond. Angiogenic effects of VEGF/NGF-NPs has been evaluated in cells and hindlimb ischemia mice model. Results showed that VEGF/NGF-NPs promoted VEGF and NGF co-expression simultaneously, eliminated excessive H2O2, strengthened reactions between SH-SY5Ys and HUVECs, and finally enhanced migration, tube formation, proliferation and H2O2 damage resistance of HUVECs. VEGF/NGF-NPs also recovered blood perfusion, promoted the expression of VEGF, NGF, eNOS and NO, and enhanced vascular coverage of pericytes. Treatment effects of VEGF/NGF-NPs may related to VEGF/eNOS/NO pathway. Altogether, VEGF/NGF-NPs eliminated excessive H2O2 while achieving gene co-delivery, and promoted stable angiogenesis. It’s a promising way for PVD treatment by using VEGF/NGF-NPs.Graphical

Pharmacogenomics-Synergistic Strategies Using a Chimerical Peptide for Enhanced Chemotherapy Based on ROS and DNA Nanosystem

Co-delivery of gene and drug for synergistic therapy has provided a promising strategy to cure devastating diseases. The use of genomic predictors of response to cisplatin and pemetrexed can be incorporated into strategies to optimize therapy for advanced solid tumors. Here, a positive feedback strategy was utilized to amplify the concentration of intracellular reactive oxygen species (ROS) and a ROS-triggered self-accelerating drug release nanosystem (defined as T/D@RSMSNs) was demonstrated for enhanced tumor chemotherapy. It was found that in human breast cancer (MCF-7) cells, T/D@RSMSNs could not only release DOX and a-TOS initiatively, but also lead to increased concentration of intracellular ROS, which could be used as new trigger to cut away TK linkage and then in turn facilitate the further release of DOX for enhanced chemotherapy. Standard treatment for advanced non–small-cell lung cancer (NSCLC) includes the use of a platinum-based chemotherapy regimen. This novel ROS triggered self-accelerating drug release nanosystem with remarkably improved therapeutic effects could provide a general strategy to branch out the applications of existing ROS-responsive drug delivery systems (DDSs).

Construction of GSH-triggered cationic fluoropolymers as two-in-one nanoplatforms for combined chemo-gene therapy.

Combined chemo-gene therapy has become a promising approach for enhanced anti-cancer treatment efficacy. However, effective co-delivery of therapeutic genes and drugs into target cells and tissues remains a major challenge. In this work, a GSH-responsive cationic fluoropolymer PSSF was designed as a co-delivery platform and synthesized by introducing a perfluorinated chain into a low-molecular weight PEI-based cationic polymer through a disulfide bond. PSSF exhibits good ability for drug and gene loading, as well as fast drug release in a GSH-rich environment. Gene transfection assay revealed that PSSF could deliver both model genes and the p53 gene into tumor cells smoothly with good protein expression, while maintaining good biocompatibility. It was also demonstrated that PSSF could simultaneously deliver the p53 gene and DOX into the HeLa cells efficiently and realize fast release of DOX. In vitro and in vivo anti-tumor assays both demonstrated that the co-delivery system could inhibit tumor growth more effectively than individual gene or drug therapy. Histopathological analysis of major organs indicated negligible systemic toxicity of such synergistic therapy systems. This rationally designed co-delivery vector provides an effective platform for the development of gene-drug synergistic therapy.

Core-shell nanofibre scaffold mediated co-delivery of connexin-43 gene and histone deacetylase inhibitor for anticancer therapy

Connexin 43 (Cx43), a gap junction protein, also acts as a tumor suppressor, which reduces tumorigenicity. Cx43 expression sensitizes cancer cells towards anticancer drugs mainly due to the formation of functional gap junctions. 4-phenyl butyrate (4-PB), a histone deacetylase inhibitor (HDACi), has antitumor potential that is also reported to increase Cx43 expression and gap junction communication. In the present work, we have developed a core-shell nanofibrous scaffold loaded with the Cx43 gene and 4-PB. The nanofibrous scaffold has been characterized by spectroscopic and electron microscopic techniques. The nanofibrous scaffold mediated transfection of the Cx43 gene into Cx43 deficient breast cancer cells (MCF-7), followed by controlled and sustained release of 4-PB, exhibited anti-cell proliferative activity. The anticancer effects have been ascertained by cell staining, cell viability and flow cytometry-based assays. Thus, the novel core-shell nanofiber scaffold has shown enormous therapeutic translation ability via Cx-based enhancement of anticancer potential of HDACi.

Synthesis of L-DOPA conjugated doxorubicin-polyethylenimine nanocarrier and evaluation of its cytotoxicity on A375 and HepG2 cell lines

Objective(s): Polyethylenimine (PEI) is one of the most-extensively investigated cationic polymers for gene and drug delivery. Recently, great attention has been directed to design of carriers for co-delivery of nucleic acids and small molecules. These delivery systems are able to overcome the limitations of gene or drug delivery alone. The aim of this study is to prepare a targeted nano-carrier for co-delivery of doxorubicin (Dox) and gene using polyethylenimine. Materials and Methods: In order to prepare the ligand-containing polymer conjugates, succinic anhydride was conjugated onto the hydroxyl group of Dox through an ester bond following the protection of Dox amines by Fmoc. Drug-polymer conjugates were then coupled with L-DOPA in order to prepare the targeted nanocarriers to the cells through Large Amino Acid Transporter-1 (LAT-1). The PEI derivatives were characterized using 1H-NMR. The toxicity of conjugated polymer, Dox and PEI was assessed on HepG2 and A375 cell lines with different expression level of LAT-1 transporters using MTT assay. Results: The chemical structure of PEI conjugate was confirmed by 1H-NMR. The cytotoxicity measurement demonstrated a cell line-dependent toxicity profile at the concentrations tested in this study. It was shown that there was no significant difference in cell-induced toxicity between conjugated polymer and its parent form in A375 cell line while the cytotoxicity of conjugated polymer was significantly lower than the parent PEI in HepG2 cells.Conclusion: These results provide promising evidence for further evaluation of PEI conjugate for co-delivery of drug and gene via LAT-1 transporters.

Effects of nanoparticle-mediated Co-delivery of bFGF and VEGFA genes to deep burn wounds: An in vivo study

Deep burns are a common form of trauma worldwide, and they are hard to be cured in a short time and enhance psychological pressure of the patients. How to effectively promote the healing of wounds after burns is a continuing challenge currently faced by burn physicians. Various strategies of promoting wound healing of deep burns have been developed, including gene therapy and growth factor therapy. In this study, we developed a combined therapy using PLGA nanoparticles as carriers to deliver bFGF and VEGFA genes to promote healing of burn wounds. We first inserted the bFGF and VEGFA genes into pEGFP-N1 vectors and loaded the mixed generated plasmids into PLGA nanoparticles. Next, we injected the nanoparticle/plasmid complexes into the rats intracutaneously and found that the complexes were successfully transfected in vivo one week later. Finally, we injected the nanoparticle/plasmid complexes containing bFGF and VEGFA around burn wounds. We found that the percentage of wound healing of rats treated with nanoparticles/bFGF+ VEGFA plasmid complexes was higher than that of rats in the scald control group, and the early percentage of wound complete epithelialization was also higher. Therefore, combining gene therapy with nanoparticles may be an effective biological strategy for wound repair.

Multifunctional stimuli-responsive niosomal nanoparticles for co-delivery and co-administration of gene and bioactive compound: In vitro and in vivo studies

This study aims to optimize niosomes for carrying and delivering the anticancer agents and genes, simultaneously. We synthesized new cationic niosomal formulations containing Tween 80, Tween 60, cholesterol, and dioleoyl-3-trimethylammonium propane (DOTAP) as a platform to enhance transfection efficacy and stability. Curcumin as an anticancer drug was entrapped with high efficacy inside stable spherical niosomes with sufficient positive charges of about + 27 mV. Loading niosomal curcumins with microRNA-34a (miR-34a) decreased the surface charge to + 15 mV and enhanced the diameter to near 68 nm. The in vitro studies were performed to investigate the cytotoxicity, cellular uptake, and gene expression profiling of normal and cancer cells treated by free curcumin, free miR-34a, niosomal curcumin (NCur), niosomal miR-34a (NmiR), niosomal curcumin + miR-34a (NCur-miR) which is termed as co-delivery, and niosomal curcumin + niosomal miR-34a which is termed as co-administration. Results showed that co-delivery caused more cytotoxicity, uptake, and anticancer activity in cancer cells than in other groups. Most importantly, niosomal and free forms of curcumin and miR-34a showed less toxicity to normal human cells. Besides, the effect of these anticancer agents was studied on the 4 T1 xenografted Balb/C mouse tumor model. Co-delivery of curcumin and miR-34a to cancer models caused a higher tumor inhibition rate than other groups. Thus, a combined therapy of curcumin and miR-34a using the new cationic niosomal delivery can be recognized as a prominent strategy for more effective cancer treatments.

Synthesis and characterization of hybrid nanocarrier layered double hydroxide grafted by polyethylene glycol and gemcitabine

For the past few years, organic-inorganic hybrid nanocarriers have been widely explored for effective drug delivery and preferable disease treatments. In this article, hydrothermal method was utilized to prepare fine dispersed layered double hydroxide (Mg-Al LDH) suspension. Polyethylene glycol (PEG) was grafted on the surface of LDH lamella in order to improve the dispersibility of LDH. Besides, the anti-cancer drug gemcitabine was grafted on the surface of LDH lamellas through chemical grafting. Hence a novel new type of organic-inorganic hybrid drug delivery system LDH-mPEG-Gemcitabine was obtained. In addition, the siRNA was intercalated into the LDH interlamination by ion exchange method to realize drug and gene co-delivery. The loading capacity of LDH and LDH-mPEG-Gemcitabine was evaluated by agarose gel electrophoresis. The characterization by laser particle size analyzer, TEM, FT-IR, XRD, in vitro cell viability and in vitro drug release demonstrated that LDH-mPEG-Gemcitabine possessed fine dispersibility, uniform morphology and particle size, fine biocompatibility, ideal drug loading and releasing capacity and held great potential to be used as a desired co-delivery system for drug and gene.

Dual DNA Transfection Using 1,6-Hexanedithiol-Conjugated Maleimide-Functionalized PU-PEI600 For Gene Correction in a Patient iPSC-Derived Fabry Cardiomyopathy Model.

Non-viral gene delivery holds promises for treating inherited diseases. However, the limited cloning capacity of plasmids may hinder the co-delivery of distinct genes to the transfected cells. Previously, the conjugation of maleimide-functionalized polyurethane grafted with small molecular weight polyethylenimine (PU-PEI600-Mal) using 1,6-hexanedithiol (HDT) could promote the co-delivery and extensive co-expression of two different plasmids in target cells. Herein, we designed HDT-conjugated PU-PEI600-Mal for the simultaneous delivery of CRISPR/Cas9 components to achieve efficient gene correction in the induced pluripotent stem cell (iPSC)-derived model of Fabry cardiomyopathy (FC) harboring GLA IVS4 + 919 G > A mutation. This FC in vitro model recapitulated several clinical FC features, including cardiomyocyte hypertrophy and lysosomal globotriaosylceramide (Gb3) deposition. As evidenced by the expression of two reporter genes, GFP and mCherry, the addition of HDT conjugated two distinct PU-PEI600-Mal/DNA complexes and promoted the co-delivery of sgRNA/Cas9 and homology-directed repair DNA template into target cells to achieve an effective gene correction of IVS4 + 919 G > A mutation. PU-PEI600-Mal/DNA with or without HDT-mediated conjugation consistently showed neither the cytotoxicity nor an adverse effect on cardiac induction of transfected FC-iPSCs. After the gene correction and cardiac induction, disease features, including cardiomyocyte hypertrophy, the mis-regulated gene expressions, and Gb3 deposition, were remarkably rescued in the FC-iPSC-differentiated cardiomyocytes. Collectively, HDT-conjugated PU-PEI600-Mal-mediated dual DNA transfection system can be an ideal approach to improve the concurrent transfection of non-viral-based gene editing system in inherited diseases with specific mutations.

Enhanced antitumor efficacy of glutathione-responsive chitosan based nanoparticles through co-delivery of chemotherapeutics, genes, and immune agents

To achieve the co-delivery of chemotherapeutic drugs, genes, and immune agents in a single nanoparticulate system, p-mercaptobenzoic acid-grafted N, N, N-trimethyl chitosan nanoparticles (MT NPs) were successfully synthesized. Paclitaxel (PTX) was encapsulated into the hydrophobic core of the MT NPs, and meanwhile, survivin shRNA-expressing plasmid (iSur-pDNA) and recombinant human interleukin-2 (rhIL-2) were loaded onto the hydrophilic shell of the MT NPs. Owing to the redox-sensitiveness of MT NPs, a rapid release of PTX was triggered by the high concentration of glutathione. The synergistic effects of PTX (1.5 mg/kg), iSur-pDNA (1.875 mg/kg), and rhIL-2 (6 × 105 IU/kg) at a low dose endowed the MT/PTX/pDNA/rhIL-2 NPs with enhanced antitumor efficacies and improved tumor-induced immunosuppression. These results demonstrated that the co-delivery of PTX, iSur-pDNA, and rhIL-2 by the amphiphilic chitosan based NPs with redox-sensitiveness could be a promising strategy in the treatment of tumors.

Dual ATP/reduction-responsive polyplex to achieve the co-delivery of doxorubicin and miR-23b for the cancer treatment

Combination therapy based on the co-delivery of therapeutic genes and anti-cancer drugs has emerged as a promising approach in the cancer treatment, and stimuli-responsive delivery systems could further improve the therapeutic efficacy. Herein, an ATP aptamer and its complementary DNA were used to form Duplex into which doxorubicin (DOX) was loaded to construct DOX-Duplex, and then the lipoic acid-modified oligoethyleneimine (LA-OEI) was employed as a carrier to realize the co-delivery of DOX-Duplex and miR-23b. The ternary nanocomplex LA-OEI/miR-23b/DOX-Duplex showed excellent anti-proliferative effect by inducing the cell apoptosis via mitochondrial signaling pathway and arresting the cell cycle at S phase. Meanwhile, the co-delivery of DOX-Duplex and miR-23b could efficiently inhibit the metastasis of cancer cells by reducing the expression level of MMP-9. The favorable anti-tumor efficacy of ternary nanocomplex was attributed to the rapid drug release in response to intracellular ATP concentration and reduction conditions and the synergistic effect between DOX-Duplex and miR-23b. Thus, ATP aptamer and reduction-responsive polymer provided a convenient platform to construct dual stimuli-responsive systems for the co-delivery of gene and drug in the cancer treatment.

Engineering Precision Medicine to Enhance Graft-Versus-Lymphoma Activity: Hematopoietic Stem Cells Modified with Chimeric Antigen Receptors

Background: Gene-modified human hematopoietic stem cells (HSC) have been used to introduce genes to correct, prevent or treat diseases. Clinical trials using infusion of autologous T cells engineered with anti-CD19 chimeric antigen receptors (CAR) have demonstrated complete remissions even in chemotherapy-resistant malignancies, but the persistence of the effector cells is transient, limiting clinical efficacy and allowing the cancer to recur. Current trials of CAR T cells have recommended patients in remission post-T cell therapy to undergo hematopoietic stem cell transplantation (HSCT) due to the limited persistence of the CAR+ T cells. Seeking to foster persistence of the CAR-modified immune cells, we propose using gene modification of HSC to create persistent generation of multilineage immune effectors to directly target cancer cells. Concerns regarding malignant transformation, abnormal hematopoiesis and autoimmunity exist, making the co-delivery of a suicide gene a necessary safety measure. We evaluated a second-generation anti-CD19 chimeric antigen receptor (CAR) to human HSC in a humanized mouse model, co-delivered with a truncated epidermal growth factor receptor (EGFRt) as a suicide gene system.Significance: CAR+ HSC can be infused in the context of HSCT, when the chemotherapy-conditioning regimen favors engraftment of gene-modified cells, decreases residual tumor burden and decreases the chance of CAR immunogenicity. The prospect of modifying autologous cells to enhance GVL bears the possibility of decreased morbidity and mortality, a concern for specifically vulnerable populations such as children and elderly patients. In addition, this approach will offer alternative therapy for those without cell sources available for allogeneic HSCT, benefiting ethnic minorities.Methods: Third-generation self-inactivating lentiviral vectors were used to co-deliver an anti-CD19 CAR and EGFRt. In vitro, gene-modified HSC were differentiated into myeloid cells to allow transgene expression. Antibody-dependent cell-mediated cytotoxicity (ADCC) assay was used incubating target cells with leukocytes and monoclonal antibody cetuximab to determine percentage of surviving cells. In vivo, gene-modified HSC were engrafted into immunodeficient NSG mice treated or not with intraperitoneal cetuximab 1mg/mouse/day for 12 days. Persistence of gene-modified cells was assessed by flow cytometry, ddPCR, and PET imaging using 89Zr-Cetuximab.Results: Gene modification of HSC did not affect engraftment, proliferation or differentiation, at efficiency of 20-40%. Antigen specificity of myeloid, NK and T cells were successfully redirected against CD19. Ablation of gene-modified cells was significantly increased (p=0.01) in target cells expressing EGFRt after incubation with leukocytes and cetuximab 1µg/mL, compared to EGFRt+ cells without cetuximab, and non-transduced cells with or without cetuximab. This was seen at all effector-to-target ratios. Mice engrafted with CAR-modified HSC presented prolonged persistence of gene-modified cells in bone marrow, spleen, thymus and peripheral blood, and CAR+ cells elicited protection against CD19+ tumors, with inhibition or elimination of tumor development and survival advantage. Mice humanized with gene-modified HSC for the expression of the suicide gene had significant ablation of gene-modified cells after treatment with cetuximab (p=0.002). Remaining gene-modified cells were close to background on flow cytometry and within two logs of decrease of vector copy numbers by ddPCR in mouse tissues. PET imaging of humanized mice documented engraftment of EGFRt-expressing cells in bone marrow, spleen and liver, and confirmed ablation with an average decrease of 82.5% after cetuximab treatment.Conclusions: These results give proof of principle for persistence and anti-tumor activity of CAR-modified HSC regulated by suicide gene, and further studies are needed to enable clinical translation. Cetuximab ADCC of EGFRt-modified cells caused effective killing. Different ablation approaches, such as inducible caspase 9 or co-delivery of other inert cell markers should be evaluated. DisclosuresNo relevant conflicts of interest to declare.

Nuclear-targeted TAT-PZLL-D3 co-delivery DOX and p53 for chemotherapy resistance of SCLC

A deliver system based on a PZLL and PAMAM block copolymer (PZLL-D3) with TAT modified on the surface has been developed. It is designed to perform highly efficient nucleus-targeted gene and drug co-delivery. Confocal laser scanning microscopy (CLSM) revealed that more TAT-PZLL-D3 entered the nucleus than does PZLL-D3 alone. The TAT-modified vector serves as a gene and drug co-delivery system to achieve high gene transfection efficiency, high apoptosis and low viability in A549 cells.

Natural Polymers Decorated MOF-MXene Nanocarriers for Co-delivery of Doxorubicin/pCRISPR.

A one-pot and facile method with assistance of high gravity was applied for the synthesis of inorganic two-dimensional MOF-5 embedded MXene nanostructures. The innovative inorganic MXene/MOF-5 nanostructure was applied in co-delivery of drug and gene, and to increase its bioavailability and interaction with the pCRISPR, the nanomaterial was coated with alginate and chitosan. The polymer-coated nanosystems were fully characterized, and the sustained DOX delivery and comprehensive cytotoxicity studies were conducted on the HEK-293, PC12, HepG2, and HeLa cell lines, demonstrating acceptable and excellent cell viability at both very low (0.1 μg.mL-1) and high (10 μg·mL-1) concentrations. The chitosan-coated nanocarriers showed superior relative cell viability compared to others, more than 60% on average of relative cell viability in all of the cell lines. Then, alginate-coated nanocarriers ranked at second place on the higher relative cell viability, more than 50% on average for all of the cell lines. Also, MTT results showed a complete dose-dependence, and by increasing the time of treatment from 24 to 72 h, the relative cell viability decreased by a meaningful slope; however, this decrease was optimized by coating the nanocarrier with chitosan and alginate. The nanosystems were also tagged with pCRISPR to analyze the potential application in the co-delivery of drug/gene. CLSM images of the HEK-293 and HeLa cell lines unveiled successful delivery of pCRISPR into the cells, and the enhanced green fluorescent protein (EGFP) reached up to ca. 26% for the HeLa cell line. Also, a considerable drug payload of 35.7% was achieved, which would be because of the interactions between the nanocarrier and the doxorubicin. In this unprecedented report pertaining to the synthesis of MXene assisted by a MOF and high-gravity technique, the methodology and the optimized ensuing MXene/MOF-5 nanosystems can be further developed for the co-delivery of drug/gene in animal models.

Green surfactant-dendrimer aggreplexes: An ingenious way to launch dual attack on arch-enemy cancer

Combination therapy, which combines anti-cancer drugs with different oligonucleotides, have shown potential in cancer treatment. However, delivering a hydrophobic anti-cancer drug and a hydrophilic oligonucleotide simultaneously is a herculean task. This study takes advantage of interactions between histidine-lauric acid-based green surfactant and poly(amidoamine) dendrimers to achieve this aim. The green surfactant was synthesized by carbodiimide chemistry and characterized by FTIR, 1H-NMR, and mass spectroscopy. Further, green surfactant-dendrimer aggregates encapsulating DTX and complexing SIRT 1 shRNA i.e., “aggreplexes” were developed and characterized. The term “aggreplexes” signifies complexes which are formed between green-surfactant-dendrimer aggregates and SIRT-1 shRNA via electrostatic interaction. The aggreplexes displayed particle size of 262.33 ± 3.87 nm, PDI of 0.25 and entrapment efficiency of 70.56 %. The TEM images revealed spherical shape of aggreplexes with irregular outer surface and corroborated particle size obtained from zetasizer. The in-vitro release study revealed biphasic release patterns of DTX from aggreplexes and were compatible for intravenous administration. Further, aggreplexes augmented cellular uptake in MDA-MB-231 cells by ∼1.87-fold compared to free DTX. Also, EGFP expression revealed significantly higher transfection of aggreplexes compared to naked shRNA and Superfect™ complexes. Further, aggreplexes showed higher cytotoxicity in MDA-MB-231 cells and ∼4.16-fold reduction in IC50 value compared to free DTX. Finally, apoptosis-index observed in case of aggreplexes was ∼3.57-fold higher than free DTX. These novel aggreplexes showed increased drug loading capacity and superior gene transfection potential. Thus, they open new avenues for co-delivery of hydrophobic anti-cancer drugs and hydrophilic therapeutic genes for improving current standards of cancer therapy.

Co-delivery of IL-12 cytokine gene and cisplatin prodrug by a polymetformin-conjugated nanosystem for lung cancer chemo-gene treatment through chemotherapy sensitization and tumor microenvironment modulation

The combination of chemotherapy and gene therapy has been indicated as a promising approach for cancer therapy. However, this combination strategy is still faced a challenge by the lack of suitable carriers to co-loaded chemotherapeutic drug and gene into one single nanoplatform. In this study, a tumor-targeted HC/pIL-12/polyMET micelleplexes were developed for the co-loading and co-delivery of cisplatin (CDDP) and plasmid encoding interleukin-12 gene (pIL-12), which would be utilized to generate synergistic actions through chemotherapy sensitization and microenvironment modulation. The HC/pIL-12/polyMET exhibited desirable particle size, superior serum stability, effective intracellular CDDP release and pIL-12 transfection efficiency. More important, the HC/pIL-12/polyMET generated the enhanced LLC cell proliferation inhibition and apoptosis induction efficiency. The long-circulating HC/pIL-12/polyMET micelleplexes promoted the accumulation of CDDP and pIL-12 in tumor site, which resulted in significantly inhibiting the growth of lung cancer, and prolonging the overall survival of tumor-bearing mice. The underlying immune mechanism demonstrated the combination of CDDP and pIL-12 activated immune effector cells to release IFN-γ and induced M1-type differentiation of tumor-related macrophages, thereby generating synergistic chemoimmunotherapy effect. Taken together, this study may provide an effective strategy for drug/gene co-delivery and cancer chemoimmunotherapy. Statement of significanceChemoimmunotherapy has been indicated as an approach to improve efficacy of cancer therapy. Herein, a tumor-targeted micelleplexes (HC/pIL-12/polyMET) were developed for the co-delivery of cisplatin (CDDP) and plasmid encoding IL-12 gene (pIL-12), which can employ the synergistic effects through chemotherapy sensitization and microenvironment modulation. The HC/pIL-12/polyMET exhibited desirable particle size, superior serum stability, high gene transfection efficiency and antitumor activity on tumor cell proliferation inhibition and apoptosis induction. More importantly, the long-circulating HC/pIL-12/polyMET micelleplexes could effectively accumulate in tumor sites and then rapidly release the CDDP and pIL-12, significantly inhibit the growth of lung cancer. This strategy provides a new concept for chemo-gene combination with a strengthened overall therapeutic efficacy of chemoimmunotherapy.

Systemic gene delivery using lipid envelope systems and its potential in overcoming challenges

Nonviral vectors which offer a safer and versatile alternative to viral vectors have been developed to overcome problems caused by viral carriers. However, their transfection efficacy or level of expression is substantially lower than viral vectors. Among various nonviral gene vectors, lipid envelops systems are an ideal platform for the incorporation of safety and efficacy into a single delivery system. Emerging strategies for gene delivery using lipid-based delivery systems mainly aim at improving the transfection efficiency and potency while reducing toxicity, achieving prolonged release, cell-specific targeting, co-delivery of drug and gene. Earlier efforts to improve the transfection efficiency while overcoming the toxicity led to the need for preparing conjugates of lipids with polyamines. In this review, we highlight current lipidic vectors that have been developed for gene therapy, challenges, and their solutions.

Synthesis of Copolymers Polyethyleneimine-co-Polyphenylalanine as Gene and Drug Codelivery Carrier.

In this study, a series of hyperbranched copolymers polyethyleneimine-co-polyphenylalanine (PEI-co-PPhe) are synthesized by ring-opening polymerization with phenylalanine-N-carboxyanhydride as monomer and PEI-25k as initiator, using as a gene and drug codelivery carrier. Among them, PEI-co-PPhe (1:170) is selected out from transfection efficiency and cytotoxicity tests. Then, doxorubicin-cis-aconitic anhydride (CAD) and BCl2-shRNA (as a therapeutic gene) are coloaded into the PEI-co-PPhe carrier to form PEI-co-PPhe/Bcl2-shRNA/CAD complexes as a codeliver system. When the mass ratio of PEI-co-PPhe:Bcl2-shRNA:CAD is 5:1:1, the codeliver system has the most obvious synergistic therapeutic effect against B16F10 cells. Confirmed by confocal laser scanning microscope and flow cytometry, compared with drug and gene alone, the codeliver complexes can be endocytosed into B16F10 cells efficiently. As a result, the appropriate length of PPhe grafted on PEI will improve the gene transfer efficiency and decrease cytotoxicity, as well as effective codelivery of gene and drug into cancer cells to be a promising codelivery carrier for cancer therapy.