Recent Advances in Engineering Carriers for siRNA Delivery.

RNA interference (RNAi) technology has been a promising treatment strategy for combating intractable diseases. However, the applications of RNAi in clinical are hampered by extracellular and intracellular barriers. To overcome these barriers, various siRNA delivery systems have been developed in the past two decades. The first approved RNAi therapeutic, Patisiran (ONPATTRO) using lipids as the carrier, for the treatment of amyloidosis is one of the most important milestones. This has greatly encouraged researchers to work on creating new functional siRNA carriers. In this review, the recent advances in siRNA carriers consisting of lipids, polymers, and polymer-modified inorganic particles for cancer therapy are summarized. Representative examples are presented to show the structural design of the carriers in order to overcome the delivery hurdles associated with RNAi therapies. Finally, the existing challenges and future perspective for developing RNAi as a clinical modality will be discussed and proposed. It is believed that the addressed contributions in this review will promote the development of siRNA delivery systems for future clinical applications.

NIR-Cleavable and pH-Responsive Polymeric Yolk-Shell Nanoparticles for Controlled Drug Release.

Responsive drug release and low toxicity of drug carriers are important for designing controlled release systems. Here, a double functional diffractive o-nitrobenzyl, containing multiple electron-donating groups as a crosslinker and methacrylic acid (MAA) as a monomer, was used to decorate upconversion nanoparticles (UCNPs) to produce robust poly o-nitrobenzyl@UCNP nanocapsules using the distillation-precipitation polymerization and templating method. Poly o-nitrobenzyl@UCNP nanocapsules with a robust yolk-shell structure exhibited near-infrared (NIR) light-/pH-responsive properties. When the nanocapsules were exposed to 980 nm NIR irradiation, the loaded drug was efficiently released by altering the shell of the nanocapsules. The photodegradation kinetics of the poly o-nitrobenzyl@UCNP nanocapsules were studied. The anticancer drug, doxorubicin hydrochloride (DOX), was loaded at pH 8.0 with a loading efficiency of 13.2 wt %. The Baker-Lonsdale model was used to determine the diffusion coefficients under different release conditions to facilitate the design of dual-responsive drug release devices or systems. Additionally, cytotoxicity studies showed that the drug release of DOX could be efficiently triggered by NIR to kill cancer cells in a controlled manner.

Highly biocompatible chlorin e6-poly(dopamine) core-shell nanoparticles for enhanced cancer phototherapy.

Cancer is a life-threatening disease worldwide. Although several approaches, such as surgery, chemotherapy, and radiotherapy, have been proven effective for many patients in clinics, they usually suffer from drug resistance, severe toxic-side effects, patient discomfort, and sometimes, unsatisfactory efficacies. In recent years, phototherapy, as a less invasive but effective therapeutic method, has brought hope for cancer treatment. However, most reported photo-therapeutic agents are constructed using complex components with non-negligible toxicity risk, thus retarding the start of their clinical trials. To address this issue, herein, biocompatible photothermal/photodynamic dual-mode therapeutic nanoparticles (CBP NPs) were successfully designed and constructed based on the Food and Drug Administration (FDA)-approved ingredients, chlorin e6 (Ce6) and poly(dopamine) (PDA). Upon light irradiation, hyperthermia was induced and reactive oxygen species (ROS) were generated simultaneously by CBP NPs, contributing to synergistic phototherapy toward cancer. The in vitro and in vivo experiments have demonstrated well the antitumor effect of CBP NPs. More importantly, CBP NPs are completely harmless and degradable in vivo. Together, the CBP NPs developed by us are an ideal candidate for the enhanced phototherapy of tumors, which holds great potential for future clinical translation.

Recent advances in nanotechnology approaches for non-viral gene therapy.

Gene therapy has shown great potential in the treatment of many diseases by downregulating the expression of certain genes. The development of gene vectors as a vehicle for gene therapy has greatly facilitated the widespread clinical application of nucleic acid materials (DNA, mRNA, siRNA, and miRNA). Currently, both viral and non-viral vectors are used as delivery systems of nucleic acid materials for gene therapy. However, viral vector-based gene therapy has several limitations, including immunogenicity and carcinogenesis caused by the exogenous viral vectors. To address these issues, non-viral nanocarrier-based gene therapy has been explored for superior performance with enhanced gene stability, high treatment efficiency, improved tumor-targeting, and better biocompatibility. In this review, we discuss various non-viral vector-mediated gene therapy approaches using multifunctional biodegradable or non-biodegradable nanocarriers, including polymer-based nanoparticles, lipid-based nanoparticles, carbon nanotubes, gold nanoparticles (AuNPs), quantum dots (QDs), silica nanoparticles, metal-based nanoparticles and two-dimensional nanocarriers. Various strategies to construct non-viral nanocarriers based on their delivery efficiency of targeted genes will be introduced. Subsequently, we discuss the cellular uptake pathways of non-viral nanocarriers. In addition, multifunctional gene therapy based on non-viral nanocarriers is summarized, in which the gene therapy can be combined with other treatments, such as photothermal therapy (PTT), photodynamic therapy (PDT), immunotherapy and chemotherapy. We also provide a comprehensive discussion of the biological toxicity and safety of non-viral vector-based gene therapy. Finally, the present limitations and challenges of non-viral nanocarriers for gene therapy in future clinical research are discussed, to promote wider clinical applications of non-viral vector-based gene therapy.

Organic/Inorganic Self-Assembled Hybrid Nano-Architectures for Cancer Therapy Applications.

Since the conceptualization of nanomedicine, numerous nanostructure-mediated drug formulations have progressed into clinical trials for treating cancer. However, recent clinical trial results indicate such kind of drug formulations has a limited improvement on the antitumor efficacy. This is due to the biological barriers associated with those formulations, for example, circulation stability, extravasation efficiency in tumor, tumor penetration ability, and developed multi-drug resistance. When employing for nanomedicine formulations, pristine organic-based and inorganic-based nanostructures have their own limitations. Accordingly, organic/inorganic (O/I) hybrids have been developed to integrate the merits of both, and to minimize their intrinsic drawbacks. In this context, the recent development in O/I hybrids resulting from a self-assembly strategy will be introduced. Through such a strategy, organic and inorganic building blocks can be self-assembled via either chemical covalent bonds or physical interactions. Based on the self-assemble procedure, the hybridization of four organic building blocks including liposomes, micelles, dendrimers, and polymeric nanocapsules with five functional inorganic nanoparticles comprising gold nanostructures, magnetic nanoparticles, carbon-based materials, quantum dots, and silica nanoparticles will be highlighted. The recent progress of these O/I hybrids in advanced modalities for combating cancer, such as, therapeutic agent delivery, photothermal therapy, photodynamic therapy, and immunotherapy will be systematically reviewed.

Metal organic framework-coated gold nanorod as an on-demand drug delivery platform for chemo-photothermal cancer therapy.

Chemo-photothermal therapy based on nanoparticles has emerged as a promising strategy for cancer treatment. However, its therapeutic efficacy and application potential are largely subjected to the uncontrollability and biotoxicity of functional nanoplatforms. Herein, a novel biocompatible and biodegradable metal organic framework (MOF), which was constructed by growing crystalline zeolitic imidazolate framework-8 on gold nanoroad (Au@ZIF-8), was designed and fabricated for efficient drug loading and controlled release. Owing to the large surface area and guest-matching pore size of ZIF-8, doxorubicin (DOX) was successfully loaded into the Au@ZIF-8 with a high drug loading efficiency of ~ 37%. Under NIR light or weakly acidic environment, the ZIF-8 layer was quickly degraded, which resulted in an on-demand drug release in tumour site. More importantly, under the irradiation of near infrared (NIR) laser, highly efficient cancer treatment was achieved in both in vitro cell experiment and in vivo tumour-bearing nude mice experiment due to the synergistic effect of photothermal (PTT) therapy and chemotherapy. In addition, the in vivo study revealed the good biocompatibility of Au@ZIF-8. This work robustly suggested that Au@ZIF-8 could be further explored as a drug delivery system for chemo-photothermal synergistic therapy.

Manganese-doped near-infrared emitting nanocrystals for in vivo biomedical imaging.

Nanoprobes with multiple imaging modality have attracted a great deal of attention due to the capability of offering complementary information from each individual component. This work presents a hybrid approach to synthesize manganese doped near infrared (NIR) emitting quantum dots. The Mn-doping process was accomplished in aqueous phase followed by a phase transfer to organic phase for ZnS coating. This bimodal nanoprobe displayed high NIR luminescence quantum yield (~14%) and capability of magnetic resonance imaging (MRI) (1.44 mM-1 s-1). The RGD-targeted nanoprobes have been exploited for in vitro cell labelling, in vivo tumor targeting and lymph node mapping. In addition, no adverse toxic effects were observed, demonstrating the high biocompatibility of this nanoprobe.

Biodegradable charged polyester-based vectors (BCPVs) as an efficient non-viral transfection nanoagent for gene knockdown of the BCR-ABL hybrid oncogene in a human chronic myeloid leukemia cell line.

First-line therapy of chronic myelogenous leukemia (CML) has always involved the use of BCR-ABL tyrosine-kinase inhibitors which is associated with an abnormal chromosome called Philadelphia chromosome. Although the overall survival rate has been improved by the current therapeutic regime, the presence of resistance has resulted in limited efficacy. In this study, an RNA interference (RNAi)-based therapeutic regime is proposed with the aim to knockdown the BCR-ABL hybrid oncogene using small interfering RNA (siRNA). The siRNA transfection rates have usually been limited due to the declining contact probability among polyplexes and the non-adherent nature of leukemic cells. Our work aims at addressing this limitation by using a biodegradable charged polyester-based vector (BCPV) as a nanocarrier for the delivery of BCR-ABL-specific siRNA to the suspension culture of a K562 CML cell line. BCR-ABL siRNAs were encapsulated in the BCPVs by electrostatic force. Cell internalization was facilitated by the BCPV and assessed by confocal microscopy and flow cytometry. The regulation of the BCR-ABL level in K562 cells as a result of RNAi was analyzed by real-time polymerase chain reaction (RT-PCR). We observed that BCPV was able to form stable nanoplexes with siRNA molecules, even in the presence of fetal bovine serum (FBS), and successfully assisted in vitro siRNA transfection in the non-adherent K562 cells. As a consequence of downregulation of BCR-ABL, BCPV-siRNA nanoplexes inhibited cell proliferation and promoted cell apoptosis. All results were compared with a commercial transfection reagent, Lipofectamine2000™, which served as a positive control. More importantly, this class of non-viral vector exhibits biodegradable features and negligible cytotoxicity, thus providing a versatile platform to deliver siRNA to non-adherent leukemia cells with high transfection efficiency by effectively overcoming extra- and intra-cellular barriers. Due to the excellent in vitro transfection results from BCPV-siRNA, a newly developed biodegradable transfection agent, BCPV, is being probed for transfection performance in an animal model.

Near-infrared fluorescent peptide probes for imaging of tumor in vivo and their biotoxicity evaluation.

Optical imaging techniques are becoming increasingly urgent for the early detection and monitoring the progression of tumor development. However, tumor vasculature imaging has so far been largely unexplored because of the lack of suitable optical probes. In this study, we demonstrated the preparation of near-infrared (NIR) fluorescent RGD peptide probes for noninvasive imaging of tumor vasculature during tumor angiogenesis. The peptide optical probes combined the advantages of NIR emission and RGD peptide, which possesses minimal biological absorption and specially targets the integrin, which highly expressed on activated tumor endothelial cells. In vivo optical imaging of nude mice bearing pancreatic tumor showed that systemically delivered NIR probes enabled us to visualize the tumors at 24 hours post-injection. In addition, we have performed in vivo toxicity study on the prepared fluorescent RGD peptide probes formulation. The blood test results and histological analysis demonstrated that no obvious toxicity was found for the mice treated with RGD peptide probes for two weeks. These studies suggest that the NIR fluorescent peptide probes can be further designed and employed for ultrasensitive fluorescence imaging of angiogenic tumor vasculature, as well as imaging of other pathophysiological processes accompanied by activation of endothelial cells.

Cytotoxicity assessment of functionalized CdSe, CdTe and InP quantum dots in two human cancer cell models.

The toxicity of quantum dots (QDs) has been extensively studied over the past decade. Some common factors that originate the QD toxicity include releasing of heavy metal ions from degraded QDs and the generation of reactive oxygen species on the QD surface. In addition to these factors, we should also carefully examine other potential QD toxicity causes that will play crucial roles in impacting the overall biological system. In this contribution, we have performed cytotoxicity assessment of four types of QD formulations in two different human cancer cell models. The four types of QD formulations, namely, mercaptopropionic acid modified CdSe/CdS/ZnS QDs (CdSe-MPA), PEGylated phospholipid encapsulated CdSe/CdS/ZnS QDs (CdSe-Phos), PEGylated phospholipid encapsulated InP/ZnS QDs (InP-Phos) and Pluronic F127 encapsulated CdTe/ZnS QDs (CdTe-F127), are representatives for the commonly used QD formulations in biomedical applications. Both the core materials and the surface modifications have been taken into consideration as the key factors for the cytotoxicity assessment. Through side-by-side comparison and careful evaluations, we have found that the toxicity of QDs does not solely depend on a single factor in initiating the toxicity in biological system but rather it depends on a combination of elements from the particle formulations. More importantly, our toxicity assessment shows different cytotoxicity trend for all the prepared formulations tested on gastric adenocarcinoma (BGC-823) and neuroblastoma (SH-SY5Y) cell lines. We have further proposed that the cellular uptake of these nanocrystals plays an important role in determining the final faith of the toxicity impact of the formulation. The result here suggests that the toxicity of QDs is rather complex and it cannot be generalized under a few assumptions reported previously. We suggest that one have to evaluate the QD toxicity on a case to case basis and this indicates that standard procedures and comprehensive protocols are urgently needed to be developed and employed for fully assessing and understanding the origins of the toxicity arising from different QD formulations.

Well-defined diblock brush polymer-drug conjugates for sustained delivery of paclitaxel.

Using the 3(rd) generation Grubbs' catalyst as the initiator, diblock brush polymer drug conjugates (BPDCs) were synthesized by sequential ring-opening metathesis polymerization (ROMP) of a hydrophilic poly(ethylene glycol) (PEG)-based norbornene (NB)-functionalized macromonomer and a hydrophobic paclitaxel (PTXL)-based NB-functionalized monomer. These amphiphilic diblock BPDCs had well-defined structures, with narrow molecular weight distributions (Mw/Mn = 1.10-1.16). They self-assembled into multi-molecular nanostructures in aqueous solutions. Although the PTXL moieties were connected to the backbone with cycloacetal-based conjugation linkages, the cleavage of these linkages from the assemblies of diblock BPDCs was relatively slow and exhibited limited acid-sensitivity, indicating a significant influence of the macromolecular structure and assembly of BPDCs on their drug release behaviour. The cytotoxicity study not only showed that the diblock BPDCs are therapeutically effective against cancer cells, but also revealed a correlation between cytotoxicity and grafting structures of BPDCs. In summary, the results obtained in this work provide new insight into the structure-dependent properties of brush polymer-based drug delivery systems.

Biodegradable nanoparticle-mediated K-ras down regulation for pancreatic cancer gene therapy.

RNA interference (RNAi) targeting the K-ras oncogene mutation in pancreatic cancer mediated by small interfering RNA (siRNA) transfection is a very promising treatment. However, the rapid degradation and negative charge of naked siRNAs restrict their direct delivery into cells. In this contribution, we propose a safe and effective transmembrane transport nanocarrier formulation based on a newly developed biodegradable charged polyester-based vector (BCPV) for K-ras siRNA delivery into pancreatic cancer cells. Our results have shown that these biodegradable and biocompatible vectors are able to transfect siRNAs targeting mutant K-ras into MiaPaCa-2 cells with high transfection and knockdown efficiency. More importantly, the RNAi process initiated a cascade gene regulation of the downstream proteins of K-ras associated with cell proliferation, migration, invasion and apoptosis. We observed that after the mutant K-ras siRNA transfection, the growth, migration and invasion of the MiaPaCa-2 cells were significantly reduced; also, the apoptosis of the pancreatic cancer cells was promoted. Although in vivo testing data are limited, we propose that the BCPV based nanoparticle formulation could be a promising candidate as non-viral vectors for gene therapy in clinical settings.

Synthesis of pH-responsive chitosan nanocapsules for the controlled delivery of doxorubicin.

Well-defined chitosan nanocapsules (CSNCs) with tunable sizes were synthesized through the interfacial cross-linking of N-maleoyl-functionalized chitosan (MCS) in miniemulsions, and their application in the delivery of doxorubicin (Dox) was investigated. MCS was prepared by the amidation reaction of CS with maleic anhydride in water/DMSO at 65 °C for 20 h. Subsequently, thiol-ene cross-linking was conducted in oil-in-water miniemulsions at room temperature under UV irradiation for 1 h, using MCS as both a surfactant and precursor polymer, 1,4-butanediol bis(3-mercapto-propionate) as a cross-linker, and D-α-tocopheryl poly(ethylene glycol) 1000 succinate as a cosurfactant. With the increase in cosurfactant concentration in the reaction systems, the sizes of the resulting CSNCs decreased steadily. Dox-loaded CSNCs were readily prepared by in situ encapsulation of Dox during miniemulsion cross-linking. With acid-labile ß-thiopropionate cross-linkages, the Dox-loaded CSNCs demonstrated a faster release rate under acidic conditions. Relative to free Dox, Dox-loaded CSNCs exhibited enhanced cytotoxicity toward MCF-7 breast cancer cells without any noticeable cytotoxicity from empty CSNCs. The effective delivery of Dox to MCF-7 breast cancer cells via Dox-loaded CSNCs was also observed.

Polylactide-graft-doxorubicin nanoparticles with precisely controlled drug loading for pH-triggered drug delivery.

Nanoparticles (NPs) with high drug loading and pH-responsivity were prepared by nanoprecipitation of a hydrophobic polymer-drug conjugate (PDC). The PDC, polylactide-graft-doxorubicin (PLA-g-DOX), was synthesized by azide-alkyne click reaction to transform acetylene-functionalized PLA into PLA-graft-aldehyde (PLA-g-ALD), followed by DOX conjugation to form acid-sensitive Schiff base linkage between drug moieties and polymer scaffold. The DOX loading amount in PLA-g-DOX PDC was determined to be 32 wt % by (1)H NMR and UV-vis spectroscopies. PLA-g-DOX PDC was further used to prepare NPs with precisely controlled drug loading by nanoprecipitation in the presence of a PEGylated surfactant. The effects of organic solvent, PLA-g-DOX PDC concentration and PLA-g-DOX/surfactant mass ratio on size and size distribution of NPs were systematically examined based on analysis by dynamic light scattering (DLS) and transmission electron microscopy (TEM). NPs prepared under the optimal conditions exhibited well-defined spherical morphology with volume-average hydrodynamic diameter (Dh) around 100 nm. Due to the Schiff base conjugation linkage in PLA-g-DOX PDC, acid-sensitive drug release behavior of the NPs was observed. In vitro studies against MCF-7 breast cancer cells showed that the NPs can be readily taken up and result in enhanced therapeutic efficiency as compared to DOX·HCl, indicating their promising potential applications as anticancer nanomedicines.

Biodegradable cationic polymeric nanocapsules for overcoming multidrug resistance and enabling drug-gene co-delivery to cancer cells.

Having unique architectural features, cationic polymeric nanocapsules (NCs) with well-defined covalently stabilized biodegradable structures were generated as potentially universal and safe therapeutic nanocarriers. These NCs were synthesized from allyl-functionalized cationic polylactide (CPLA) by highly efficient UV-induced thiol-ene interfacial cross-linking in transparent miniemulsions. With tunable nanoscopic sizes, negligible cytotoxicity and remarkable degradability, they are able to encapsulate doxorubicin (Dox) with inner cavities and bind interleukin-8 (IL-8) small interfering RNA (siRNA) with cationic shells. The Dox-encapsulated NCs can effectively bypass the P-glycoprotein (Pgp)-mediated multidrug resistance of MCF7/ADR cancer cells, thereby resulting in increased intracellular drug concentration and reduced cell viability. In vitro studies also showed that the NCs loaded with Dox, IL-8 siRNA and both agents can be readily taken up by PC3 prostate cancer cells, resulting in a significant chemotherapeutic effect and/or IL-8 gene silencing.

Interleukin-8 gene silencing on pancreatic cancer cells using biodegradable polymer nanoplexes.

Pancreatic cancer is one of the deadliest cancers throughout the world with rarely efficient therapies currently available. Gene therapy on pancreatic cancer through small interfering RNA (siRNA)-based RNA interference (RNAi) has shown great potential and attracted much attention. However, due to the fragile nature of nucleic acid, the application of RNAi as a safe and efficient carrier faces great challenges. In this contribution, a self-assembly regime, which is based on well-defined cationic polylactides (CPLAs) with tertiary amine groups, has been used to encapsulate and protect siRNAs from fast degradation. CPLA is a safe and degradable formulation that allowed us to deliver siRNAs targeting the proangiogenic chemokine interleukin-8 (IL-8) to pancreatic cancer cells for gene therapy. Stable IL-8 siRNA-CPLA nanoplexes were successfully formed by electrostatic force and high gene transfection efficiencies were shown on two pancreatic cancer cell lines. We did not observe any cytotoxicity from these CPLAs over a large concentration range via cell viability evaluations. More importantly, the silencing of IL-8 gene expression significantly attenuated the proliferation of pancreatic cancer cells. Our preliminary results support the future development of gene therapy that might provide an effective and safe treatment approach towards pancreatic cancer.

Biomolecular recognition principles for bionanocombinatorics: an integrated approach to elucidate enthalpic and entropic factors.

Bionanocombinatorics is an emerging field that aims to use combinations of positionally encoded biomolecules and nanostructures to create materials and devices with unique properties or functions. The full potential of this new paradigm could be accessed by exploiting specific noncovalent interactions between diverse palettes of biomolecules and inorganic nanostructures. Advancement of this paradigm requires peptide sequences with desired binding characteristics that can be rationally designed, based upon fundamental, molecular-level understanding of biomolecule-inorganic nanoparticle interactions. Here, we introduce an integrated method for building this understanding using experimental measurements and advanced molecular simulation of the binding of peptide sequences to gold surfaces. From this integrated approach, the importance of entropically driven binding is quantitatively demonstrated, and the first design rules for creating both enthalpically and entropically driven nanomaterial-binding peptide sequences are developed. The approach presented here for gold is now being expanded in our laboratories to a range of inorganic nanomaterials and represents a key step toward establishing a bionanocombinatorics assembly paradigm based on noncovalent peptide-materials recognition.

Phospholipid micelle-based magneto-plasmonic nanoformulation for magnetic field-directed, imaging-guided photo-induced cancer therapy.

We present a magnetoplasmonic nanoplatform combining gold nanorods (GNR) and iron-oxide nanoparticles within phospholipid-based polymeric nanomicelles (PGRFe). The gold nanorods exhibit plasmon resonance absorbance at near infrared wavelengths to enable photoacoustic imaging and photothermal therapy, while the Fe3O4 nanoparticles enable magnetophoretic control of the nanoformulation. The fabricated nanoformulation can be directed and concentrated by an external magnetic field, which provides enhancement of a photoacoustic signal. Application of an external field also leads to enhanced uptake of the magnetoplasmonic formulation by cancer cells in vitro. Under laser irradiation at the wavelength of the GNR absorption peak, the PGRFe formulation efficiently generates plasmonic nanobubbles within cancer cells, as visualized by confocal microscopy, causing cell destruction. The combined magnetic and plasmonic functionalities of the nanoplatform enable magnetic field-directed, imaging-guided, enhanced photo-induced cancer therapy. FROM THE CLINICAL EDITOR: In this study, a nano-formulation of gold nanorods and iron oxide nanoparticles is presented using a phospholipid micelle-based delivery system for magnetic field-directed and imaging-guided photo-induced cancer therapy. The gold nanorods enable photoacoustic imaging and photothermal therapy, while the Fe3O4 nanoparticles enable magnetophoretic control of the formulation. This and similar systems could enable more precise and efficient cancer therapy, hopefully in the near future, after additional testing.

Biodegradable nanocapsules as siRNA carriers for mutant K-Ras gene silencing of human pancreatic carcinoma cells.

The application of small interfering RNA (siRNA)-based RNA interference (RNAi) for cancer gene therapy has attracted great attention. Gene therapy is a promising strategy for cancer treatment because it is relatively non-invasive and has a higher therapeutic specificity than chemotherapy. However, without the use of safe and efficient carriers, siRNAs cannot effectively penetrate the cell membranes and RNAi is impeded. In this work, cationic poly(lactic acid) (CPLA)-based degradable nanocapsules (NCs) are utilized as novel carriers of siRNA for effective gene silencing of pancreatic cancer cells. These CPLA-NCs can readily form nanoplexes with K-Ras siRNA and over 90% transfection efficiency is achieved using the nanoplexes. Cell viability studies show that the nanoparticles are highly biocompatible and non-toxic, indicating that CPLA-NC is a promising potential candidate for gene therapy in a clinical setting.