bims-novged 2023-01-22 papers

bims-novged

Biomed News

on Non-viral vectors for gene delivery

Issue of 2023–01–22
27 papers selected by
the Merkel lab, Ludwig-Maximilians University

Regulation of Pro-Inflammatory Macrophage Polarization via Lipid Nanoparticles Mediated Delivery of Anti-Prostaglandin-E2 siRNA.
Self-Assembled Oleic Acid-Modified Polyallylamines for Improved siRNA Transfection Efficiency and Lower Cytotoxicity.
Iterative Design of Ionizable Lipids for Intramuscular mRNA Delivery.
Development of hyaluronic acid-anchored polycaprolactone nanoparticles for efficient delivery of PLK1 siRNA to breast cancer.
Efficient Transfection via an Unexpected Mechanism by Near Neutral Polypiperazines with Tailored Response to Endosomal pH.
Lipid nanoparticle-mediated mRNA delivery in lung fibrosis.
Ligand-tethered lipid nanoparticles for targeted RNA delivery to treat liver fibrosis.
Endosomal escape of nucleic acids from extracellular vesicles mediates functional therapeutic delivery.
Suppression of lung cancer malignancy by micellized siRNA through cell cycle arrest.
Matrices Activated with Messenger RNA.
Liver cancer cells as the model for developing liver-targeted RNAi therapeutics.
A novel heterologous receptor-binding domain dodecamer universal mRNA vaccine against SARS-CoV-2 variants.
Nucleic acid drug vectors for diagnosis and treatment of brain diseases.
Gene, RNA, and ASO-based therapeutic approaches in Cystic Fibrosis.
Histidine-based coordinative polymers for efficient intracellular protein delivery via enhanced protein binding, cellular uptake, and endosomal escape.
Synergistic effect of p53 gene/DOX intracellular delivery and P-gp inhibition by pullulan thiomers on cancer cells: in vitro and in vivo evaluations.
Endosomal escape of RNA therapeutics: How do we solve this rate-limiting problem?
Surface Engineering of HEK293 Cell-Derived Extracellular Vesicles for Improved Pharmacokinetic Profile and Targeted Delivery of IL-12 for the Treatment of Hepatocellular Carcinoma.
Surface-modified CuO nanoparticles for photocatalysis and highly efficient energy storage devices.
Polyethyleneimine-Based Drug Delivery Systems for Cancer Theranostics.
Evaluation of the Cytotoxicity of Cationic Polymers on Glioblastoma Cancer Stem Cells.
CRISPR/Cas9 therapeutics: progress and prospects.
Valency and affinity control of aptamer-conjugated nanoparticles for selective cancer cell targeting.
Multifunctional nanoparticle for cancer therapy.
Cancer cell membrane coated PLGA nanoparticles as biomimetic drug delivery system for improved cancer therapy.
The Challenges and Prospects of p53-Based Therapies in Ovarian Cancer.
Comparison of DNA targeting CRISPR editors in human cells.

Curr Issues Mol Biol. 2022 Dec 20. 45(1): 1-11

Regulation of Pro-Inflammatory Macrophage Polarization via Lipid Nanoparticles Mediated Delivery of Anti-Prostaglandin-E2 siRNA.

Ahmad Almatroudi, Mohammed A Alsahli, Mansoor Ali Syed, Amjad Ali Khan, Arshad Husain Rahmani.

  Pro-inflammatory macrophage polarization is crucial in acute inflammatory diseases like Acute lung injury (ALI), and acute respiratory distress syndrome (ARDS). Prostaglandin E2 (PGE2) is believed to promote inflammation in such cases. Therefore, our study aimed to deliver anti-prostaglandin E synthase 2 small interfering RNA antibodies (anti-PGE2-siRNA) through lipid nanoparticles (LNPs) in RAW264.7 (The murine macrophage cell line) to find a possible cure to the acute inflammatory diseases. LNPs were synthesized by using thin layer evaporation method and were characterized by dynamic light scattering (DLS), Zeta potential, SEM and TEM analysis. The obtained NPs were spherical with an average size of 73 nm and zeta potential +29mV. MTT assay revealed that these NPs were non-toxic in nature. Gel retardation assay displayed 5:2 ratio of siRNA and NPs as the best siRNA:LNPs ratio for the delivery of siRNA into cells. After siRNA delivery by using LNPs, real time gene expression analysis revealed significant decrease in the expression of PGE2. Western blot results confirmed that silencing of PGE2 gene influence inducible nitric oxide synthase (iNOS) and interlukin-1β (1L-1β), markers involved in pro-inflammatory macrophage polarization. Our study revealed that LNPs synthesized in present study can be one of the effective methods to deliver anti-PGE2-siRNA to control pro-inflammatory macrophage polarization for the treatment of acute inflammatory response.

Keywords:  inflammation; lipid nano particles; macrophage polarization; prostaglandin E2; siRNA

DOI:  https://doi.org/10.3390/cimb45010001
ACS Appl Bio Mater. 2023 Jan 16.

Self-Assembled Oleic Acid-Modified Polyallylamines for Improved siRNA Transfection Efficiency and Lower Cytotoxicity.

Cristian Salvador, Patrizia Andreozzi, Gabriela Romero, Iraida Loinaz, Damien Dupin, Sergio E Moya.

  Small interference RNA (siRNA) is a tool for gene modulation, which can silence any gene involved in genetic disorders. The potential of this therapeutic tool is hampered by RNA instability in the blood stream and difficulties to reach the cytosol. Polyamine-based nanoparticles play an important role in gene delivery. Polyallylamine hydrochloride (PAH) is a polycation displaying primary amines that can be easily chemically modified to match the balance between cell viability and siRNA transfection. In this work, PAH has been covalently functionalized with oleic acid at different molar ratios by carbodiimide chemistry. The substituted polymers form polyplexes that keep positive surface charge and fully encapsulate siRNA. Oleic acid substitution improves cell viability in the pulmonary cell line A549. Moreover, 6 and 14% of oleic acid substitution show an improvement in siRNA transfection efficiency. CD47 is a ubiquitous protein which acts as "don't eat me signal." SIRPα protein of macrophages recognizes CD47, leading to tumor cell phagocytosis by macrophages. By knocking down CD47 with siRNA, cancer cells become vulnerable to be eliminated by the immune system. PAH-oleic acid substitutes show high efficacy in silencing the CD47 protein, making them a potential candidate for immunotherapy.

Keywords:  CD47; oleic acid; polyallylamine hydrochloride; polymeric nanoparticles; siRNA delivery

DOI:  https://doi.org/10.1021/acsabm.2c00845
J Am Chem Soc. 2023 Jan 18.

Iterative Design of Ionizable Lipids for Intramuscular mRNA Delivery.

Grayson Tilstra, Julien Couture-Senécal, Yan Ming Anson Lau, Alanna M Manning, Daniel S M Wong, Wanda W Janaeska, Titobioluwa A Wuraola, Janice Pang, Omar F Khan.

Lipid nanoparticles (LNPs) are the most clinically advanced delivery vehicles for RNA and have enabled the development of RNA-based drugs such as the mRNA COVID-19 vaccines. Functional delivery of mRNA by an LNP greatly depends on the inclusion of an ionizable lipid, and small changes to these lipid structures can significantly improve delivery. However, the structure-function relationships between ionizable lipids and mRNA delivery are poorly understood, especially for LNPs administered intramuscularly. Here, we show that the iterative design of a novel series of ionizable lipids generates key structure-activity relationships and enables the optimization of chemically distinct lipids with efficacy that is on-par with the current state of the art. We find that the combination of ionizable lipids comprising an ethanolamine core and LNPs with an apparent pKa between 6.6 and 6.9 maximizes intramuscular mRNA delivery. Furthermore, we report a nonlinear relationship between the lipid-to-mRNA mass ratio and protein expression, suggesting that a critical mass ratio exists for LNPs and may depend on ionizable lipid structure. Our findings add to the mechanistic understanding of ionizable lipids and demonstrate that hydrogen bonding, ionization behavior, and lipid-to-mRNA mass ratio are key design parameters affecting intramuscular mRNA delivery. We validate these insights by applying them to the rational design of new ionizable lipids. Overall, our iterative design strategy efficiently generates potent ionizable lipids. This hypothesis-driven method reveals structure-activity relationships that lay the foundation for the optimization of ionizable lipids in future LNP-RNA drugs. We foresee that this design strategy can be extended to other optimization parameters beyond intramuscular expression.

DOI: https://doi.org/10.1021/jacs.2c10670
Drug Deliv Transl Res. 2023 Jan 15.

Development of hyaluronic acid-anchored polycaprolactone nanoparticles for efficient delivery of PLK1 siRNA to breast cancer.

Dolly Jain, Awesh K Yadav.

  PlK1 has a significant role in the development of breast cancer. Thus, silencing of PlK1 gene may arrest the growth of breast cancer. However, the in vivo stability of PlK1 siRNA after injection remains a challenge to target the specific site. The delivery of siPlK1 RNA via viral vector and amine group-terminated dendrimer is associated with immune reaction and cellular cytotoxicity. Thus, in the present study, hyaluronic acid-functionalized and -thiolated polycaprolactone nanoparticles (SH-HPP NPs) were developed for enhancing the targeting capabilities of siRNA towards human breast cancer cells. NPs displayed size in the range of 180-217 nm, and with sustain and pH-dependent release of siRNA up to 120 h. The in vitro treatments with siRNA-containing NPs showed the high number of necrotic cells and the cell cycle arrest at the G2/M phase. The gene expression analysis depicts the decrease of endogenous PLK1 siRNA expression on MCF-7 cells upon PLK1 NPs treatment. In vitro cytotoxicity experiments demonstrated effective anticancer properties against MCF-7. Finally, in vivo results showed that substantial tumor inhibition was achieved with PLK1 siRNA-containing SH-HPP NPs in comparison of the control group. Hence, HPP NPs have enormous potential for the selective delivery of siRNA, i.e., breast cancer cells.

Keywords:  Breast cancer; CD44 receptors; Gene silencing; Hyaluronic acid; Nanoparticles; PLK1 siRNA

DOI:  https://doi.org/10.1007/s13346-022-01288-2
Macromol Biosci. 2023 Jan 19. e2200517

Efficient Transfection via an Unexpected Mechanism by Near Neutral Polypiperazines with Tailored Response to Endosomal pH.

Franziska Hausig-Punke, Gregor Dekevic, Fabian H Sobotta, Jana I Solomun, Friederike Richter, Denise Salzig, Anja Traeger, Johannes C Brendel.

  Cationic pH-responsive polymers promise to overcome critical challenges in cellular delivery. Ideally, the polymers become selectively charged along the endosomal pathway disturbing only the local membrane and avoiding unintended interactions or cytotoxic side effects at physiological conditions. Polypiperazines represent a novel, hydrophilic class of pH-responsive polymers whose response can be tuned within the relevant pH range (5 - 7.4). We discovered that the polypiperazines are effectively binding plasmid DNA (pDNA) and demonstrate high efficiency in transfection. By design of experiments (DoE), a wide parameter space (pDNA and polymer concentration) was screened to identify the range of effective concentrations for transfection. An isopropyl modified polypiperazine was highly efficient over a wide range of concentrations outperforming linear polyethylenimine (l-PEI, 25 kDa) in regions of low N*/P ratios. A quantitative polymerase chain reaction (qPCR) surprisingly revealed that the pDNA within the piperazine-based polyplexes can be amplified in contrast to polyplexes based on l-PEI. The pDNA must therefore be more accessible and bound differently than for other known transfection polymers. Considering the various opportunities to further optimize their structure, polypiperazines represent a promising platform for designing effective soluble polymeric vectors, which are charge-neutral at physiological conditions. This article is protected by copyright. All rights reserved.

Keywords:  design of experiments; endosomal escape; gene delivery; pDNA binding; polyplex characterization; structure-activity relationship

DOI:  https://doi.org/10.1002/mabi.202200517
Eur J Pharm Sci. 2023 Jan 12. pii: S0928-0987(23)00001-5. [Epub ahead of print] 106370

Lipid nanoparticle-mediated mRNA delivery in lung fibrosis.

Matteo Massaro, Suhong Wu, Gherardo Baudo, Haoran Liu, Scott Collum, Hyunho Lee, Cinzia Stigliano, Victor Segura-Ibarra, Harry Karmouty-Quintana, Elvin Blanco.

  mRNA delivery enables the specific synthesis of proteins with therapeutic potential, representing a powerful strategy in diseases lacking efficacious pharmacotherapies. Idiopathic pulmonary fibrosis (IPF) is a chronic lung disease characterized by excessive extracellular matrix (ECM) deposition and subsequent alveolar remodeling. Alveolar epithelial type 2 cells (AEC2) and fibroblasts represent important targets in IPF given their role in initiating and driving aberrant wound healing responses that lead to excessive ECM deposition. Our objective was to examine a lipid nanoparticle (LNP)-based mRNA construct as a viable strategy to target alveolar epithelial cells and fibroblasts in IPF. mRNA-containing LNPs measuring ∼34 nm had high encapsulation efficiency, protected mRNA from degradation, and exhibited sustained release kinetics. eGFP mRNA LNP transfection in human primary cells proved dose- and time-dependent in vitro. In a bleomycin mouse model of lung fibrosis, luciferase mRNA LNPs administered intratracheally led to site-specific lung accumulation. Importantly, bioluminescence signal was detected in lungs as early as 2 h after delivery, with signal still evident at 48 h. Of note, LNPs were found associated with AEC2 and fibroblasts in vivo. Findings highlight the potential for pulmonary delivery of mRNA in IPF, opening therapeutic avenues aimed at halting and potentially reversing disease progression.

Keywords:  Lipid nanoparticles (LNPs); alveolar epithelial cells; gene delivery; idiopathic pulmonary fibrosis; lung fibroblasts; mRNA

DOI:  https://doi.org/10.1016/j.ejps.2023.106370
Nat Commun. 2023 Jan 17. 14(1): 75

Ligand-tethered lipid nanoparticles for targeted RNA delivery to treat liver fibrosis.

Xuexiang Han, Ningqiang Gong, Lulu Xue, Margaret M Billingsley, Rakan El-Mayta, Sarah J Shepherd, Mohamad-Gabriel Alameh, Drew Weissman, Michael J Mitchell.

Lipid nanoparticle-mediated RNA delivery holds great potential to treat various liver diseases. However, targeted delivery of RNA therapeutics to activated liver-resident fibroblasts for liver fibrosis treatment remains challenging. Here, we develop a combinatorial library of anisamide ligand-tethered lipidoids (AA-lipidoids) using a one-pot, two-step modular synthetic method and adopt a two-round screening strategy to identify AA-lipidoids with both high potency and selectivity to deliver RNA payloads to activated fibroblasts. The lead AA-lipidoid AA-T3A-C12 mediates greater RNA delivery and transfection of activated fibroblasts than its analog without anisamide and the FDA-approved MC3 ionizable lipid. In a preclinical model of liver fibrosis, AA-T3A-C12 enables ~65% silencing of heat shock protein 47, a therapeutic target primarily expressed by activated fibroblasts, which is 2-fold more potent than MC3, leading to significantly reduced collagen deposition and liver fibrosis. These results demonstrate the potential of AA-lipidoids for targeted RNA delivery to activated fibroblasts. Furthermore, these synthetic methods and screening strategies open a new avenue to develop and discover potent lipidoids with targeting properties, which can potentially enable RNA delivery to a range of cell and tissue types that are challenging to access using traditional lipid nanoparticle formulations.

DOI: https://doi.org/10.1038/s41467-022-35637-z
Pharmacol Res. 2023 Jan 16. pii: S1043-6618(23)00021-X. [Epub ahead of print]188 106665

Endosomal escape of nucleic acids from extracellular vesicles mediates functional therapeutic delivery.

Tuan Thach Pham, Huan Chen, Phuong Hoang Diem Nguyen, Migara Kavishka Jayasinghe, Anh Hong Le, Minh Tn Le.

  Extracellular vesicles hold great promise as a drug delivery platform for RNA-based therapeutics. However, there is a lack of experimental evidence for the intracellular trafficking of nucleic acid cargos, specifically, whether they are capable of escaping from the endolysosomal confinement in the recipient cells to be released into the cytosol and hence, interact with their cytoplasmic targets. Here, we demonstrated how red blood cell-derived extracellular vesicles (RBCEVs) release their therapeutic RNA/DNA cargos at specific intracellular compartments characteristic of late endosomes and lysosomes. The released cargos were functional and capable of knocking down genes of interest in recipient cells, resulting in tumor suppression in vitro and in an acute myeloid leukemia murine model without causing significant toxicity. Notably, surface functionalization of RBCEVs with an anti-human CXCR4 antibody facilitated their specific uptake by CXCR4+ leukemic cells, leading to enhanced gene silencing efficiency. Our results provide insights into the cellular uptake mechanisms and endosomal escape routes of nucleic acid cargos delivered by RBCEVs which have important implications for further improvements of the RBCEV-based delivery system.

Keywords:  Cancer; Drug delivery; Endosomal escape; Extracellular vesicles; RNA; Therapeutics

DOI:  https://doi.org/10.1016/j.phrs.2023.106665
Adv Healthc Mater. 2023 Jan 16. e2202358

Suppression of lung cancer malignancy by micellized siRNA through cell cycle arrest.

Haejoo Kim, In-Ho Jeong, Yeol Kyo Choi, Yeon Kyung Lee, Eunyoung Moon, Yang Hoon Huh, Wonpil Im, Jun-O Jin, Minseok Kwak, Peter Chang-Whan Lee.

  UBA6-specific E2 conjugation enzyme 1 (USE1) was frequently overexpressed in lung cancer patients. Moreover, the critical role of USE1 in the progression of human lung cancer was also indicated. As the next step, we aimed to develop USE1-targeted therapeutic agents based on RNA interference (RNAi). In this study, we introduce a lipid-modified DNA carrier, namely U4T, which consists of four consecutive dodec-1-ynyluracil (U) nucleobases to increase the cell permeability of siRNA targeting of USE1. The U4Ts aggregate to form micelles, and the USE1-silencing siRNA-incorporated soft spherical nucleic acid aggregate (siSNA) can be created simply through base-pairing with siRNA. Treatment with siSNA was effective in suppressing tumor growth in vivo as well as cell proliferation, migration, and invasion of lung cancer cells. Furthermore, siSNA inhibited tumor cell growth by inducing cell cycle arrest in the G1 phase and apoptosis. Thus, we confirmed the anti-tumor efficacy of siSNA in lung cancer cell lines and that siSNA possesses effective cell penetrating ability without using cationic transfection moieties. Collectively, these results suggest that siSNA could be applied to the clinical application of RNAi-based therapeutics for lung cancer treatment. This article is protected by copyright. All rights reserved.

Keywords:  USE1; gene therapy; lipid-DNAs; nanomedicine; self-assembly

DOI:  https://doi.org/10.1002/adhm.202202358
J Funct Biomater. 2023 Jan 15. pii: 48. [Epub ahead of print]14(1):

Matrices Activated with Messenger RNA.

Raquel Martinez-Campelo, Marcos Garcia-Fuentes.

  Over two decades of preclinical and clinical experience have confirmed that gene therapy-activated matrices are potent tools for sustained gene modulation at the implantation area. Matrices activated with messenger RNA (mRNA) are the latest development in the area, and they promise an ideal combination of efficiency and safety. Indeed, implanted mRNA-activated matrices allow a sustained delivery of mRNA and the continuous production of therapeutic proteins in situ. In addition, they are particularly interesting to generate proteins acting on intracellular targets, as the translated protein can directly exert its therapeutic function. Still, mRNA-activated matrices are incipient technologies with a limited number of published records, and much is still to be understood before their successful implementation. Indeed, the design parameters of mRNA-activated matrices are crucial for their performance, as they affect mRNA stability, device immunogenicity, translation efficiency, and the duration of the therapy. Critical design factors include matrix composition and its mesh size, mRNA chemical modification and sequence, and the characteristics of the nanocarriers used for mRNA delivery. This review aims to provide some background relevant to these technologies and to summarize both the design space for mRNA-activated matrices and the current knowledge regarding their pharmaceutical performance. Furthermore, we will discuss potential applications of mRNA-activated matrices, mainly focusing on tissue engineering and immunomodulation.

Keywords:  bone regeneration; immunomodulation; mRNA delivery; mRNA-activated matrix; tissue engineering

DOI:  https://doi.org/10.3390/jfb14010048
Biochem Biophys Res Commun. 2023 Jan 06. pii: S0006-291X(23)00019-0. [Epub ahead of print]644 85-94

Liver cancer cells as the model for developing liver-targeted RNAi therapeutics.

Beibei Hou, Linhui Qin, Linfeng Huang.

  RNAi is a sequence-specific gene regulation mechanism that involves small interfering RNAs (siRNAs). RNAi therapeutic has become a new class of precision medicine and has shown great potential in treating liver-associated diseases, especially metabolic diseases. To facilitate the development of liver-targeted RNAi therapeutics in cell model, we surveyed a panel of liver cancer cell lines for the expression of genes implicated in RNAi therapeutics including the asialoglycoprotein receptor (ASGR) and metabolic disease associated genes PCSK9, ANGPTL3, CIDEB, and LDLR. A high-content screen assay based on lipid droplet staining confirmed the involvement of PCSK9, ANGPTL3, and CIDEB in lipid metabolism in selected liver cancer cell lines. Several liver cancer cell lines have high levels of ASGR1 expression, which is required for liver-specific uptake of GalNAc-conjugated siRNA, a clinically approved siRNA delivery platform. Using an EGFP reporter system, we demonstrated Hep G2 can be used to evaluate gene knockdown efficiency of GalNAc-siRNA. Our findings pave the way for using liver cancer cells as a convenient model system for the identification and testing of siRNA drug candidate genes and for studying ASGR-mediated GalNAc-siRNA delivery in liver.

Keywords:  ANGPTL3; ASGR; CIDEB; GalNAc; Hepatocellular carcinoma; PCSK9; RNAi; siRNA

DOI:  https://doi.org/10.1016/j.bbrc.2023.01.007
Acta Pharm Sin B. 2023 Jan 12.

A novel heterologous receptor-binding domain dodecamer universal mRNA vaccine against SARS-CoV-2 variants.

Shugang Qin, Hai Huang, Wen Xiao, Kepan Chen, Xi He, Xiaoshan Tang, Zhiying Huang, Yupei Zhang, Xing Duan, Na Fan, Qian Zheng, Min Wu, Guangwen Lu, Yuqan Wei, Xiawei Wei, Xiangrong Song.

  There are currently approximately 4,000 mutations in the SARS-CoV-2 S protein gene and emerging SARS-CoV-2 variants continue to spread rapidly worldwide. Universal vaccines with high efficacy and safety urgently need to be developed to prevent SARS-CoV-2 variants pandemic. Here, we described a novel self-assembling universal mRNA vaccine containing a heterologous receptor-binding domain (HRBD)-based dodecamer (HRBDdodecamer) against SARS-CoV-2 variants, including Alpha (B.1.1.7), Beta (B.1.351), Gamma (B.1.1.28.1), Delta (B.1.617.2) and Omicron (B.1.1.529). HRBD containing four heterologous RBD (Delta, Beta, Gamma, and Wild-type) can form a stable dodecameric conformation under T4 trimerization tag (Flodon, FD). The HRBDdodecamer -encoding mRNA was then encapsulated into the newly-constructed LNPs consisting of a novel ionizable lipid (4N4T). The obtained universal mRNA vaccine (4N4T-HRBDdodecamer) presented higher efficiency in mRNA transfection and expression than the approved ALC-0315 LNPs, initiating potent immune protection against the immune escape of SARS-CoV-2 caused by evolutionary mutation. These findings demonstrated the first evidence that structure-based antigen design and mRNA delivery carrier optimization may facilitate the development of effective universal mRNA vaccines to tackle SARS-CoV-2 variants pandemic.

Keywords:  Antigen design; Heterologous RBD; Immune escape; Ionizable lipids; Lipid nanoparticles; RBD dodecamer; SARS-CoV-2 variants pandemic; Universal mRNA vaccines

DOI:  https://doi.org/10.1016/j.apsb.2023.01.010
Signal Transduct Target Ther. 2023 Jan 17. 8(1): 39

Nucleic acid drug vectors for diagnosis and treatment of brain diseases.

Zhi-Guo Lu, Jie Shen, Jun Yang, Jing-Wen Wang, Rui-Chen Zhao, Tian-Lu Zhang, Jing Guo, Xin Zhang.

Nucleic acid drugs have the advantages of rich target selection, simple in design, good and enduring effect. They have been demonstrated to have irreplaceable superiority in brain disease treatment, while vectors are a decisive factor in therapeutic efficacy. Strict physiological barriers, such as degradation and clearance in circulation, blood-brain barrier, cellular uptake, endosome/lysosome barriers, release, obstruct the delivery of nucleic acid drugs to the brain by the vectors. Nucleic acid drugs against a single target are inefficient in treating brain diseases of complex pathogenesis. Differences between individual patients lead to severe uncertainties in brain disease treatment with nucleic acid drugs. In this Review, we briefly summarize the classification of nucleic acid drugs. Next, we discuss physiological barriers during drug delivery and universal coping strategies and introduce the application methods of these universal strategies to nucleic acid drug vectors. Subsequently, we explore nucleic acid drug-based multidrug regimens for the combination treatment of brain diseases and the construction of the corresponding vectors. In the following, we address the feasibility of patient stratification and personalized therapy through diagnostic information from medical imaging and the manner of introducing contrast agents into vectors. Finally, we take a perspective on the future feasibility and remaining challenges of vector-based integrated diagnosis and gene therapy for brain diseases.

DOI: https://doi.org/10.1038/s41392-022-01298-z
J Cyst Fibros. 2023 Jan 17. pii: S1569-1993(22)01436-9. [Epub ahead of print]

Gene, RNA, and ASO-based therapeutic approaches in Cystic Fibrosis.

Normand E Allaire, Uta Griesenbach, Batsheva Kerem, John D Lueck, Noemie Stanleigh, Yifat S Oren.

  Most people with Cystic Fibrosis (PwCF) harbor Cystic Fibrosis Transmembrane Conductance (CFTR) mutations that respond to highly effective CFTR modulators (HEM); however, a small fraction of non-responsive variants will require alternative approaches for treatment. Furthermore, the long-term goal to develop a cure for CF will require novel therapeutic strategies. Nucleic acid-based approaches offer the potential to address all CF-causing mutations and possibly a cure for all PwCF. In this minireview, we discuss current knowledge, recent progress, and critical questions surrounding the topic of Gene-, RNA-, and ASO-based therapies for the treatment of Cystic Fibrosis (CF).

Keywords:  ASO; Cystic Fibrosis; Delivery; Gene; RNA; Therapeutic

DOI:  https://doi.org/10.1016/j.jcf.2022.12.016
Biomater Sci. 2023 Jan 17.

Histidine-based coordinative polymers for efficient intracellular protein delivery via enhanced protein binding, cellular uptake, and endosomal escape.

Changyuan Chen, Peng Gao, Hui Wang, Yiyun Cheng, Jia Lv.

Polymers are one of the most promising protein delivery carriers; however, their applications are hindered by low delivery efficacy owing to their undesirable performance in protein binding, cellular uptake and endosomal escape. Here, we designed a series of histidine-based coordinative polymers for efficient intracellular protein delivery. Coordination of metal ions such as Ni2+, Zn2+, and Cu2+ with histidine residues on a polymer greatly improved its performance in protein binding, complex stability, cellular uptake and endosomal escape, therefore achieving highly improved protein delivery efficacy. Among the coordinative polymers, the Zn2+-coordinated one exhibited the highest cellular uptake, while the Cu2+-coordinated one exhibited the highest endosomal escape. The Ni2+-coordinated polymer formed large-sized aggregates with cargo proteins and showed insufficient protein release after endocytosis. The results obtained in this study provided new insight into the development of coordinative polymer-based protein delivery systems.

DOI: https://doi.org/10.1039/d2bm01541b
J Mater Chem B. 2023 Jan 19.

Synergistic effect of p53 gene/DOX intracellular delivery and P-gp inhibition by pullulan thiomers on cancer cells: in vitro and in vivo evaluations.

Priya S S, Rekha M R.

Numerous reports emphasize the inverse relationship between the mutant p53 protein and P-glycoprotein overexpression, which adversely affects the chemosensitivity of cancer cells. In this study, the cationised pullulan polysaccharide was conjugated with dithiobutyric acid (PPDBA) for the intracellular delivery of doxorubicin and the p53 gene. The transfection efficiency of PPDBA using the apoptotic gene p53 and its ability to modulate efflux pumps in the presence and absence of glutathione and the subsequent drug retention were studied in different cell lines. The percentage cell death mediated by the PPDBA/p53 nanoplex (4 : 1 ratio) was 59%, and by DOX alone a 50% cell death was attained at 3.13 μM in C6 cells, but the percentage cell death mediated by PPDBA/p53 (4 : 1) in combination with 1 μM DOX was as high as 98%. The effect of PPDBA II/p53/DOX nanoplexes on the mouse tumor model was evaluated in BALB/c mice which demonstrated good efficacy when compared with the drug or gene alone.

DOI: https://doi.org/10.1039/d2tb01770a
RNA. 2023 Jan 20. pii: rna.079507.122. [Epub ahead of print]

Endosomal escape of RNA therapeutics: How do we solve this rate-limiting problem?

Steven F Dowdy.

  With over 15 FDA approved drugs on the market and numerous ongoing clinical trials, RNA therapeutics, such as small interfering RNAs (siRNAs) and antisense oligonucleotides (ASOs), have shown great potential to treat human disease. Their mechanism of action is based entirely on the sequence of validated disease-causing genes without the prerequisite knowledge of protein structure, activity or cellular location. In contrast to small molecule therapeutics that passively diffuse across the cell membrane's lipid bilayer, RNA therapeutics are too large, too charged and/or too hydrophilic to passively diffuse across the cellular membrane and instead are taken up into cells by endocytosis. However, endosomes are also composed of a lipid bilayer barrier that results in endosomal capture and retention of 99% of RNA therapeutics with 1% or less entering the cytoplasm. Although this very low level of endosomal escape has proven sufficient for liver and some CNS disorders, it is insufficient for the vast majority of extra-hepatic diseases. Unfortunately, there are currently no acceptable solutions to the endosomal escape problem. Consequently, before RNA therapeutics can be used to treat widespread human disease, the rate-limiting delivery problem of endosomal escape must be solved in a non-toxic manner.

Keywords:  ASOs; RNA therapeutics; delivery; endosomal escape; siRNAs

DOI:  https://doi.org/10.1261/rna.079507.122
Int J Nanomedicine. 2023 ;18 209-223

Surface Engineering of HEK293 Cell-Derived Extracellular Vesicles for Improved Pharmacokinetic Profile and Targeted Delivery of IL-12 for the Treatment of Hepatocellular Carcinoma.

Jing Zhang, Haijing Song, Yanan Dong, Ganghui Li, Jun Li, Qizhe Cai, Shoujun Yuan, Yi Wang, Haifeng Song.

   Background: Extracellular vesicles (EVs) are considered a promising drug delivery platform. Naïve EVs face numerous issues that limit their applications, such as fast clearance, hepatic accumulations, and a lack of target-specific tropism. We aimed to explore a series of surface engineering approaches to: 1) reduce the non-specific adhesion of EVs, and 2) improve their enrichment in the target tissue. As a proof-of-concept, we investigated the therapeutic potentials of a multi-modal EVs system carrying a tumor-specific nanobody and the immuno-stimulant interleukin-12 (IL12) using in vivo models of hepatocellular carcinoma.
Methods: The major cell adhesion molecule on the HEK293-derived EVs, integrin β1 (ITGB1), was knocked out (KO) by CRISPR/Cas9-mediated gene editing, followed by deglycosylation to generate ITGB1-Deg EVs for the subsequent pharmacokinetic and biodistribution analyses. ITGB1-Deg EVs were further loaded with glypican-3 (GPC3)-specific nanobody (HN3) and mouse single-chain IL12 (mscIL12) to generate ITGB1-mscIL12+HN3+Deg EVs, for evaluation of tumor tropism and therapeutic potential in a mice model of hepatocellular carcinoma.
Results: Removal of ITGB1 led to the broad suppression of integrins on the EVs surface, resulting in a decrease in cellular uptake. Deglycosylation of ITGB1- EVs gave rise to inhibition of the EVs uptake by activated RAW264.7 cells. ITGB1 removal did not significantly alter the pharmacokinetic behaviors of HEK293-EVs, whereas the ITGB1-Deg EVs exhibited enhanced systemic exposure with reduced hepatic accumulation. Loading of HN3 conferred the ITGB1-Deg EVs with tumor-specific tropism for both subcutaneous and metastasized tumors in mice. The ITGB1-mscIL12+HN3+Deg EVs activated mouse splenocytes with high potency. Systemic administration of the EVs with the equivalent dose of 1.5µg/kg of exosomal IL12 achieved satisfactory tumor growth inhibition and good tolerability.
Conclusion: The combinatorial approach of EVs surface engineering conferred HEK293-EVs with reduced non-specific clearance and enhanced tumor targeting efficacy, which constituted an efficient delivery platform for critical cancer therapeutics like IL12.

Keywords:  drug delivery; exosome; glycan; glycosylation

DOI:  https://doi.org/10.2147/IJN.S388916
Environ Sci Pollut Res Int. 2023 Jan 19.

Surface-modified CuO nanoparticles for photocatalysis and highly efficient energy storage devices.

Anup Pandith, Gururaj Kudur Jayaprakash, Zeid A ALOthman.

  Herein we report multifunctional surface-modified CuO nanomaterials were used to fulfill escalating needs in the electrochemical energy storage system and to achieve efficient photocatalysts for the degradation of AR88 organic dye. Due to the atom economy, ease of synthesis, high capacitance, observable electrochemical responsiveness, and low bandgap in CuO-based nanomaterials, its active surface was modified through cationic surfactant CTAB. Surface-modified nanoparticles were characterized using various characterization techniques such as XRD, DRS, FESEM, and TEM. Intriguingly the synthesized materials demonstrated a capacitance of 133 F/g with a long-term charge-discharge cycle of 2000 cycles. In addition, at pH 11, the material also exhibited a superior dye degradation performance under the UV lamp by showing 94.8% AR88 degradation at a catalyst concentration of 1.0 g/L. Hence, we believe this concept would provide novel insights into the preparation of the simplest and cheaper multifunctional materials for next-generation energy storage and photocatalytic applications.

Keywords:  AR88; AR88 dye; CTAB; CuO nanocomposite; Photocatalysts; Supercapacitors

DOI:  https://doi.org/10.1007/s11356-023-25131-4
J Funct Biomater. 2022 Dec 23. pii: 12. [Epub ahead of print]14(1):

Polyethyleneimine-Based Drug Delivery Systems for Cancer Theranostics.

Chong Zhao, Benqing Zhou.

  With the development of nanotechnology, various types of polymer-based drug delivery systems have been designed for biomedical applications. Polymer-based drug delivery systems with desirable biocompatibility can be efficiently delivered to tumor sites with passive or targeted effects and combined with other therapeutic and imaging agents for cancer theranostics. As an effective vehicle for drug and gene delivery, polyethyleneimine (PEI) has been extensively studied due to its rich surface amines and excellent water solubility. In this work, we summarize the surface modifications of PEI to enhance biocompatibility and functionalization. Additionally, the synthesis of PEI-based nanoparticles is discussed. We further review the applications of PEI-based drug delivery systems in cancer treatment, cancer imaging, and cancer theranostics. Finally, we thoroughly consider the outlook and challenges relating to PEI-based drug delivery systems.

Keywords:  cancer imaging; cancer theranostics; cancer treatment; drug delivery; polyethylenimine

DOI:  https://doi.org/10.3390/jfb14010012
J Funct Biomater. 2022 Dec 28. pii: 17. [Epub ahead of print]14(1):

Evaluation of the Cytotoxicity of Cationic Polymers on Glioblastoma Cancer Stem Cells.

Conor McCartin, Juliette Blumberger, Candice Dussouillez, Patricia Fernandez de Larrinoa, Monique Dontenwill, Christel Herold-Mende, Philippe Lavalle, Béatrice Heurtault, Stéphane Bellemin-Laponnaz, Sylvie Fournel, Antoine Kichler.

  Cationic polymers such as polyethylenimine (PEI) have found a pervasive place in laboratories across the world as gene delivery agents. However, their applications are not limited to this role, having found a place as delivery agents for drugs, in complexes known as polymer-drug conjugates (PDCs). Yet a potentially underexplored domain of research is in their inherent potential as anti-cancer therapeutic agents, which has been indicated by several studies. Even more interesting is the recent observation that certain polycations may present a significantly greater toxicity towards the clinically important cancer stem cell (CSC) niche than towards more differentiated bulk tumour cells. These cells, which possess the stem-like characteristics of self-renewal and differentiation, are highly implicated in cancer drug resistance, tumour recurrence and poor clinical prognosis. The search for compounds which may target and eliminate these cells is thus of great research interest. As such, the observation in our previous study on a PEI-based PDC which showed a considerably higher toxicity of PEI towards glioblastoma CSCs (GSCs) than on more differentiated glioma (U87) cells led us to investigate other cationic polymers for a similar effect. The evaluation of the toxicity of a range of different types of polycations, and an investigation into the potential source of GSC's sensitivity to such compounds is thus described.

Keywords:  cancer stem cells; cationic polymers; cytotoxicity; glioblastoma; polyethylenimine

DOI:  https://doi.org/10.3390/jfb14010017
Signal Transduct Target Ther. 2023 Jan 16. 8(1): 36

CRISPR/Cas9 therapeutics: progress and prospects.

Tianxiang Li, Yanyan Yang, Hongzhao Qi, Weigang Cui, Lin Zhang, Xiuxiu Fu, Xiangqin He, Meixin Liu, Pei-Feng Li, Tao Yu.

Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) gene-editing technology is the ideal tool of the future for treating diseases by permanently correcting deleterious base mutations or disrupting disease-causing genes with great precision and efficiency. A variety of efficient Cas9 variants and derivatives have been developed to cope with the complex genomic changes that occur during diseases. However, strategies to effectively deliver the CRISPR system to diseased cells in vivo are currently lacking, and nonviral vectors with target recognition functions may be the focus of future research. Pathological and physiological changes resulting from disease onset are expected to serve as identifying factors for targeted delivery or targets for gene editing. Diseases are both varied and complex, and the choice of appropriate gene-editing methods and delivery vectors for different diseases is important. Meanwhile, there are still many potential challenges identified when targeting delivery of CRISPR/Cas9 technology for disease treatment. This paper reviews the current developments in three aspects, namely, gene-editing type, delivery vector, and disease characteristics. Additionally, this paper summarizes successful examples of clinical trials and finally describes possible problems associated with current CRISPR applications.

DOI: https://doi.org/10.1038/s41392-023-01309-7
J Control Release. 2023 Jan 12. pii: S0168-3659(23)00011-1. [Epub ahead of print]

Valency and affinity control of aptamer-conjugated nanoparticles for selective cancer cell targeting.

Laura Woythe, David Porciani, Tessa Harzing, Stijn van Veen, Donald H Burke, Lorenzo Albertazzi.

  Nanoparticles (NPs) are commonly functionalized using targeting ligands to drive their selective uptake in cells of interest. Typical target cell types are cancer cells, which often overexpress distinct surface receptors that can be exploited for NP therapeutics. However, these targeted receptors are also moderately expressed in healthy cells, leading to unwanted off-tumor toxicities. Multivalent interactions between NP ligands and cell receptors have been investigated to increase the targeting selectivity towards cancer cells due to their non-linear response to receptor density. However, to exploit the multivalent effect, multiple variables have to be considered such as NP valency, ligand affinity, and cell receptor density. Here, we synthesize a panel of aptamer-functionalized silica-supported lipid bilayers (SSLB) to study the effect of valency, aptamer affinity, and epidermal growth factor receptor (EGFR) density on targeting specificity and selectivity. We show that there is an evident interplay among those parameters that can be tuned to increase SSLB selectivity towards high-density EGFR cells and reduce accumulation at non-tumor tissues. Specifically, the combination of high-affinity aptamers and low valency SSLBs leads to increased high-EGFR cell selectivity. These insights provide a better understanding of the multivalent interactions of NPs with cells and bring the nanomedicine field a step closer to the rational design of cancer nanotherapeutics.

Keywords:  Aptamer avidity and affinity; Multivalency; Nanoparticle targeting; Silica-supported lipid bilayers; Tumor targeting

DOI:  https://doi.org/10.1016/j.jconrel.2023.01.008
MedComm (2020). 2023 Feb;4(1): e187

Multifunctional nanoparticle for cancer therapy.

Yan Gao, Kaiyu Wang, Jin Zhang, Xingmei Duan, Qiu Sun, Ke Men.

  Cancer is a complex disease associated with a combination of abnormal physiological process and exhibiting dysfunctions in multiple systems. To provide effective treatment and diagnosis for cancer, current treatment strategies simultaneously focus on various tumor targets. Based on the rapid development of nanotechnology, nanocarriers have been shown to exhibit excellent potential for cancer therapy. Compared with nanoparticles with single functions, multifunctional nanoparticles are believed to be more aggressive and potent in the context of tumor targeting. However, the development of multifunctional nanoparticles is not simply an upgraded version of the original function, but involves a sophisticated system with a proper backbone, optimized modification sites, simple preparation method, and efficient function integration. Despite this, many well-designed multifunctional nanoparticles with promising therapeutic potential have emerged recently. Here, to give a detailed understanding and analyzation of the currently developed multifunctional nanoparticles, their platform structures with organic or inorganic backbones were systemically generalized. We emphasized on the functionalization and modification strategies, which provide additional functions to the nanoparticle. We also discussed the application combination strategies that were involved in the development of nanoformulations with functional crosstalk. This review thus provides an overview of the construction strategies and application advances of multifunctional nanoparticles.

Keywords:  backbone; cancer; modification; multifunctional nanoparticles

DOI:  https://doi.org/10.1002/mco2.187
Colloids Surf B Biointerfaces. 2023 Jan 06. pii: S0927-7765(23)00009-7. [Epub ahead of print]222 113131

Cancer cell membrane coated PLGA nanoparticles as biomimetic drug delivery system for improved cancer therapy.

Yue Li, Junfang Ke, Hongxin Jia, Jungang Ren, Li Wang, Zhiqiang Zhang, Chen Wang.

  Based on the immune escape and homologous adhesion ability of cancer cells, a drug delivery system (DDS) could overcome the dilemma of immune clearance and non-specific binding by coating the cancer cell membrane (CCM). In this study, a biomimetic DDS based on CCM and poly lactic acid-glycolic acid (PLGA) nanoparticles was successfully constructed for tumor active and homologous targeting therapy. The doped CCM on the surface of the nanoparticle enabled the DDS to achieve immune escape and had an affinity for tumor tissues. The cellular uptake and in vivo distribution tests showed a superior cellular affinity of CCM coated PLGA nanoparticles (CCMNPs) than that of PLGA nanoparticles (PLGANPs). All of those results proved that CCMNPs endowed with drug-loaded nanoparticles had the abilities of immune escape and homologous targeting through the biological functional proteins retained on the coated CCM. In addition, the tumor inhibition rate of CCMNPs in tumor-bearing nude mice was 1.3 and 2.0-fold compared to PLGANPs and PTX injection, which showed the capacity to efficiently and accurately deliver drugs to cancer sites improved the therapeutic effect of tumor and achieved accurately targeted therapy.

Keywords:  Biomimetic; Cancer cell membrane; Drug delivery system; Nanoparticle

DOI:  https://doi.org/10.1016/j.colsurfb.2023.113131
Biomolecules. 2023 Jan 12. pii: 159. [Epub ahead of print]13(1):

The Challenges and Prospects of p53-Based Therapies in Ovarian Cancer.

Bryce Wallis, Katherine Redd Bowman, Phong Lu, Carol S Lim.

  It has been well established that mutations in the tumor suppressor gene, p53, occur readily in a vast majority of cancer tumors, including ovarian cancer. Typically diagnosed in stages three or four, ovarian cancer is the fifth leading cause of death in women, despite accounting for only 2.5% of all female malignancies. The overall 5-year survival rate for ovarian cancer is around 47%; however, this drops to an abysmal 29% for the most common type of ovarian cancer, high-grade serous ovarian carcinoma (HGSOC). HGSOC has upwards of 96% of cases expressing mutations in p53. Therefore, wild-type (WT) p53 and p53-based therapies have been explored as treatment options via a plethora of drug delivery vehicles including nanoparticles, viruses, polymers, and liposomes. However, previous p53 therapeutics have faced many challenges, which have resulted in their limited translational success to date. This review highlights a selection of these historical p53-targeted therapeutics for ovarian cancer, why they failed, and what the future could hold for a new generation of this class of therapies.

Keywords:  HGSOC; gene delivery; gene therapy; ovarian cancer; p53 therapies

DOI:  https://doi.org/10.3390/biom13010159
Cell Biosci. 2023 Jan 16. 13(1): 11

Comparison of DNA targeting CRISPR editors in human cells.

Hongxin Huang, Weiqi Lv, Jinhe Li, Guanjie Huang, Zhihong Tan, Yongfei Hu, Shufeng Ma, Xin Zhang, Linxuan Huang, Ying Lin.

   BACKGROUND: Profiling and comparing the performance of current widely used DNA targeting CRISPR systems provide the basic information for the gene-editing toolkit and can be a useful resource for this field. In the current study, we made a parallel comparison between the recently reported miniature Cas12f1 (Un1Cas12f1 and AsCas12f1) and the widely used Cas12a and Cas9 nucleases in mammalian cells.
RESULTS: We found that as a CRISPRa activator, Un1Cas12f1 could induce gene expression with a comparable level to that of Cas12a and Cas9, while as a DNA cleavage editor, Cas12f1 exhibited similar properties to Cas12a, like high specificity and dominantly induced deletions over insertions, but with less activity. In contrast, wild-type SpCas9 showed the highest activity, lowest specificity, and induced balanced deletions and insertions. Thus, Cas12f1 is recommended for gene-activation-based applications, Cas12a is for therapy applications, and wild-type Cas9 is for in vitro and animal investigations.
CONCLUSION: The comparison provided the editing properties of the widely used DNA-targeting CRISPR systems in the gene-editing field.

Keywords:  CRISPR-Cas system; Cas12f1 nuclease; Comparison; DNA targeting; Specificity

DOI:  https://doi.org/10.1186/s13578-023-00958-z