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1.
Int J Mol Sci ; 22(11)2021 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-34199901

RESUMO

The establishment of CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) technology for eukaryotic gene editing opened up new avenues not only for the analysis of gene function but also for therapeutic interventions. While the original methodology allowed for targeted gene disruption, recent technological advancements yielded a rich assortment of tools to modify genes and gene expression in various ways. Currently, clinical applications of this technology fell short of expectations mainly due to problems with the efficient and safe delivery of CRISPR/Cas9 components to living organisms. The targeted in vivo delivery of therapeutic nucleic acids and proteins remain technically challenging and further limitations emerge, for instance, by unwanted off-target effects, immune reactions, toxicity, or rapid degradation of the transfer vehicles. One approach that might overcome many of these limitations employs extracellular vesicles as intercellular delivery devices. In this review, we first introduce the CRISPR/Cas9 system and its latest advancements, outline major applications, and summarize the current state of the art technology using exosomes or microvesicles for transporting CRISPR/Cas9 constituents into eukaryotic cells.


Assuntos
Sistemas CRISPR-Cas , Vesículas Extracelulares/genética , Edição de Genes , Técnicas de Transferência de Genes , Terapia Genética/métodos , Animais , Humanos
2.
Nat Commun ; 12(1): 4251, 2021 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-34253733

RESUMO

Aromatic L-amino acid decarboxylase (AADC) deficiency is a rare genetic disorder characterized by deficient synthesis of dopamine and serotonin. It presents in early infancy, and causes severe developmental disability and lifelong motor, behavioral, and autonomic symptoms including oculogyric crises (OGC), sleep disorder, and mood disturbance. We investigated the safety and efficacy of delivery of a viral vector expressing AADC (AAV2-hAADC) to the midbrain in children with AADC deficiency (ClinicalTrials.gov Identifier NCT02852213). Seven (7) children, aged 4-9 years underwent convection-enhanced delivery (CED) of AAV2-hAADC to the bilateral substantia nigra (SN) and ventral tegmental area (VTA) (total infusion volume: 80 µL per hemisphere) in 2 dose cohorts: 1.3 × 1011 vg (n = 3), and 4.2 × 1011 vg (n = 4). Primary aims were to demonstrate the safety of the procedure and document biomarker evidence of restoration of brain AADC activity. Secondary aims were to assess clinical improvement in symptoms and motor function. Direct bilateral infusion of AAV2-hAADC was safe, well-tolerated and achieved target coverage of 98% and 70% of the SN and VTA, respectively. Dopamine metabolism was increased in all subjects and FDOPA uptake was enhanced within the midbrain and the striatum. OGC resolved completely in 6 of 7 subjects by Month 3 post-surgery. Twelve (12) months after surgery, 6/7 subjects gained normal head control and 4/7 could sit independently. At 18 months, 2 subjects could walk with 2-hand support. Both the primary and secondary endpoints of the study were met. Midbrain gene delivery in children with AADC deficiency is feasible and safe, and leads to clinical improvements in symptoms and motor function.


Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/genética , Erros Inatos do Metabolismo dos Aminoácidos/terapia , Descarboxilases de Aminoácido-L-Aromático/deficiência , Dependovirus/genética , Neurônios Dopaminérgicos/metabolismo , Técnicas de Transferência de Genes , Terapia Genética , Imageamento por Ressonância Magnética , Mesencéfalo/patologia , Erros Inatos do Metabolismo dos Aminoácidos/líquido cefalorraquidiano , Erros Inatos do Metabolismo dos Aminoácidos/fisiopatologia , Descarboxilases de Aminoácido-L-Aromático/líquido cefalorraquidiano , Descarboxilases de Aminoácido-L-Aromático/genética , Criança , Pré-Escolar , Discinesias/fisiopatologia , Feminino , Terapia Genética/efeitos adversos , Humanos , Masculino , Metaboloma , Atividade Motora , Neurotransmissores/líquido cefalorraquidiano , Neurotransmissores/metabolismo , Fatores de Tempo
3.
World J Gastroenterol ; 27(24): 3568-3580, 2021 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-34239270

RESUMO

Pancreatic cancer is one of the highest and in fact, unchanged mortality-associated tumor, with an exceptionally low survival rate due to its challenging diagnostic approach. So far, its treatment is based on a combination of approaches (such as surgical resection with or rarely without chemotherapeutic agents), but with finite limits. Thus, looking for additional space to improve pancreatic tumorigenesis therapeutic approach, research has focused on gene therapy with unexpectedly growing horizons not only for the treatment of inoperable pancreatic disease, but also for its early stages. In vivo gene delivery viral vectors, despite few disadvantages (possible immunogenicity, toxicity, mutagenicity, or high cost), could be one of the most efficient cancer gene therapeutic strategies for clinical application due to their superiority compared with other systems (ex vivo delivery strategies). Their dominance consists of simple preparation, easy operation and a wide range of functions. Adenoviruses are one of the most common used vectors, inducing strong immune as well as inflammatory reactions. Oncolytic virotherapy, using the above mentioned in vivo viral vectors, is one of the most promising non-pathogenic, highly-selective cytotoxic anti-cancer therapy using anti-cancer agents with high anti-tumor potency and strong oncolytic effect. There have been a variety of targeted therapeutic and pre-clinical strategies tested for gene therapy in pancreatic cancer such as gene-editing systems (e.g., clustered regularly interspaced palindromic repeats-Cas9), RNA interference technology (e.g., microRNAs, short hairpin RNA or small interfering RNA), adoptive immunotherapy and vaccination (e.g., chimeric antigen receptor T-cell therapy) with encouraging results.


Assuntos
Terapia Viral Oncolítica , Neoplasias Pancreáticas , Edição de Genes , Técnicas de Transferência de Genes , Terapia Genética , Humanos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/terapia
4.
Int J Nanomedicine ; 16: 4391-4407, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34234433

RESUMO

Background: Gold nanocages have been widely used as multifunctional platforms for drug and gene delivery, as well as photothermal agents for cancer therapy. However, their potential as gene delivery systems for cancer treatment has been reported in combination with chemotherapeutics and photothermal therapy, but not in isolation so far. The purpose of this work was to investigate whether the conjugation of gold nanocages with the cancer targeting ligand lactoferrin, polyethylene glycol and polyethylenimine could lead to enhanced transfection efficiency on prostate cancer cells in vitro, without assistance of external stimulation. Methods: Novel lactoferrin-bearing gold nanocages conjugated to polyethylenimine and polyethylene glycol have been synthesized and characterized. Their transfection efficacy and cytotoxicity were assessed on PC-3 prostate cancer cell line following complexation with a plasmid DNA. Results: Lactoferrin-bearing gold nanocages, alone or conjugated with polyethylenimine and polyethylene glycol, were able to condense DNA at conjugate:DNA weight ratios 5:1 and higher. Among all gold conjugates, the highest gene expression was obtained following treatment with gold complex conjugated with polyethylenimine and lactoferrin, at weight ratio 40:1, which was 1.71-fold higher than with polyethylenimine. This might be due to the increased DNA cellular uptake observed with this conjugate, by up to 8.65-fold in comparison with naked DNA. Conclusion: Lactoferrin-bearing gold nanocages conjugates are highly promising gene delivery systems to prostate cancer cells.


Assuntos
Portadores de Fármacos/química , Técnicas de Transferência de Genes , Ouro/química , Lactoferrina/química , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , DNA/administração & dosagem , DNA/química , DNA/genética , Terapia Genética , Humanos , Masculino , Plasmídeos/genética , Polietilenoglicóis/química , Polietilenoimina/química , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/terapia , Transfecção
5.
Int J Mol Sci ; 22(12)2021 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-34203739

RESUMO

It is without any doubt that precision medicine therapeutic strategies targeting neurodegenerative disorders are currently witnessing the spectacular rise of newly designed approaches based on the use of viral vectors as Trojan horses for the controlled release of a given genetic payload. Among the different types of viral vectors, adeno-associated viruses (AAVs) rank as the ones most commonly used for the purposes of either disease modeling or for therapeutic strategies. Here, we reviewed the current literature dealing with the use of AAVs within the field of Parkinson's disease with the aim to provide neuroscientists with the advice and background required when facing a choice on which AAV might be best suited for addressing a given experimental challenge. Accordingly, here we will be summarizing some insights on different AAV serotypes, and which would be the most appropriate AAV delivery route. Next, the use of AAVs for modeling synucleinopathies is highlighted, providing potential readers with a landscape view of ongoing pre-clinical and clinical initiatives pushing forward AAV-based therapeutic approaches for Parkinson's disease and related synucleinopathies.


Assuntos
Pesquisa Biomédica , Dependovirus/genética , Vetores Genéticos/uso terapêutico , Doença de Parkinson/genética , Animais , Modelos Animais de Doenças , Técnicas de Transferência de Genes , Humanos
6.
Int J Mol Sci ; 22(11)2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-34205995

RESUMO

The increasing spread of multidrug-resistant pathogenic bacteria is one of the major threats to public health worldwide. Bacteria can acquire antibiotic resistance and virulence genes through horizontal gene transfer (HGT). A novel horizontal gene transfer mechanism mediated by outer membrane vesicles (OMVs) has been recently identified. OMVs are rounded nanostructures released during their growth by Gram-negative bacteria. Biologically active toxins and virulence factors are often entrapped within these vesicles that behave as molecular carriers. Recently, OMVs have been reported to contain DNA molecules, but little is known about the vesicle packaging, release, and transfer mechanisms. The present review highlights the role of OMVs in HGT processes in Gram-negative bacteria.


Assuntos
Proteínas da Membrana Bacteriana Externa/genética , Membrana Externa Bacteriana/metabolismo , Técnicas de Transferência de Genes , Transferência Genética Horizontal/genética , Bactérias/genética , Bactérias/patogenicidade , Humanos , Fatores de Virulência/genética , Fatores de Virulência/metabolismo
7.
Int J Mol Sci ; 22(13)2021 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-34209222

RESUMO

Hyaluronan (HA) is a natural glycosaminoglycan present in many tissues of all vertebrates. HA has various biological functions, which are dependent on its molar mass. High-molar-mass HA has anti-angiogenic, immunosuppressive and anti-inflammatory properties, while low-molar-mass HA has opposite effects. HA has also antioxidative properties, however on the other hand it can be readily degraded by reactive oxygen species. For many years it has been used in treatment of osteoarthritis, cosmetics and in ophthalmology. In the last years there has been a growing interest of HA to also be applied in other fields of medicine such as skin wound healing, tissue engineering, dentistry and gene delivery. In this review we summarize information on modes of HA administration, properties and effects of HA in various fields of medicine including recent progress in the investigation of HA.


Assuntos
Cosméticos , Técnicas de Transferência de Genes , Ácido Hialurônico , Osteoartrite/tratamento farmacológico , Engenharia Tecidual , Cicatrização/efeitos dos fármacos , Cosméticos/química , Cosméticos/uso terapêutico , Humanos , Ácido Hialurônico/química , Ácido Hialurônico/uso terapêutico
8.
Mater Sci Eng C Mater Biol Appl ; 126: 112161, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34082966

RESUMO

Nowadays, the nanoparticle-based delivery approach is becoming more and more attractive in gene therapy due to its low toxicity and immunogenicity, sufficient packaging capacity, targeting, and straightforward, low-cost, large-scale good manufacturing practice (GMP) production. A number of research works focusing on multilayer structures have explored different factors and parameters that can affect the delivery efficiency of pDNA. However, there are no systematic studies on the performance of these structures for enhanced gene delivery regarding the gene loading methods, the use of additional organic components and cell/particle incubation conditions. Here, we conducted a detailed analysis of different parameters such as (i) strategy for loading pDNA into carriers, (ii) incorporating both pDNA and organic additives within one carrier and (iii) variation of cell/particle incubation conditions, to evaluate their influence on the efficiency of pDNA delivery with multilayer structures consisting of inorganic cores and polymer layers. Our results reveal that an appropriate combination of all these parameters leads to the development of optimized protocols for high transfection efficiency, compared to the non-optimized process (> 70% vs. < 7%), and shows a good safety profile. In conclusion, we provide the proof-of-principle that these multilayer structures with the developed parameters are a promising non-viral platform for an efficient delivery of nucleic acids.


Assuntos
DNA , Técnicas de Transferência de Genes , Terapia Genética , Tamanho da Partícula , Plasmídeos/genética , Transfecção
9.
Int J Nanomedicine ; 16: 3741-3754, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34113099

RESUMO

Introduction: The reprogramming of induced cardiomyocytes (iCMs) is of particular significance in regenerative medicine; however, it remains a great challenge to fabricate an efficient and safe gene delivery system to induce reprogramming of iCMs for therapeutic applications in heart injury. Here, we report branched polyethyleneimine (BP) coated nitrogen-enriched carbon dots (BP-NCDs) as highly efficient nanocarriers loaded with microRNAs-combo (BP-NCDs/MC) for cardiac reprogramming. Methods: The BP-NCDs nanocarriers were prepared and characterized by several analytical techniques. Results: The BP-NCDs nanocarriers showed good microRNAs-combo binding affinity, negligible cytotoxicity, and long-term microRNAs expression. Importantly, BP-NCDs/MC nanocomplexes led to the efficient direct reprogramming of fibroblasts into iCMs without genomic integration and resulting in effective recovery of cardiac function after myocardial infarction (MI). Conclusion: This study offers a novel strategy to provide safe and effective microRNAs-delivery nanoplatforms based on carbon dots for promising cardiac regeneration and disease therapy.


Assuntos
Carbono/química , Técnicas de Reprogramação Celular/métodos , Fibroblastos/citologia , Técnicas de Transferência de Genes , MicroRNAs/administração & dosagem , Infarto do Miocárdio/terapia , Miócitos Cardíacos/citologia , Animais , Reprogramação Celular , Fibroblastos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Infarto do Miocárdio/genética , Infarto do Miocárdio/patologia , Miócitos Cardíacos/metabolismo , Medicina Regenerativa
10.
Int J Biol Macromol ; 183: 2055-2073, 2021 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-34087309

RESUMO

Gene therapy encompasses the transfer of exogenous genetic materials into the patient's target cells to treat or prevent diseases. Nevertheless, the transfer of genetic material into desired cells is challenging and often requires specialized tools or delivery systems. For the past 40 years, scientists are mainly pursuing various viruses as gene delivery vectors, and the overall progress has been slow and far from the expectation. As an alternative, nonviral vectors have gained substantial attention due to their several advantages, including superior safety profile, enhanced payload capacity, and stealth abilities. Since nonviral vectors encounter multiple extra- and intra-cellular barriers limiting the transfer of genetic payload into the target cell nucleus, we have discussed these barriers in detail for this review. A direct approach, utilizing physical methods like electroporation, sonoporation, gene gun, eliminate the requirement for a specific carrier for gene delivery. In contrast, chemical methods of gene transfer exploit natural or synthetic compounds as carriers to increase cellular targeting and gene therapy effectiveness. We have also emphasized the recent advancements aimed at enhancing the current nonviral approaches. Therefore, in this review, we have focused on discussing the current evolving state of nonviral gene delivery systems and their future perspectives.


Assuntos
Técnicas de Transferência de Genes , Terapia Genética , Lipídeos/química , Polímeros/química , Animais , Vacinas contra COVID-19/genética , Vacinas contra COVID-19/uso terapêutico , Técnicas de Transferência de Genes/efeitos adversos , Terapia Genética/efeitos adversos , Humanos , RNA Mensageiro/genética , RNA Mensageiro/uso terapêutico
11.
Colloids Surf B Biointerfaces ; 205: 111918, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34144325

RESUMO

How to overcome multiple obstacles to achieve the efficient and safe delivery of therapeutic genes is still the key to gene therapy. To address this issue, a cationic carrier consisting of polyamide-amine (HPAA-peptide-Fc) modified by an enzyme-responsive polypeptide as the core and hyperbranched polyglycerol derivative (CD-HPG) as the shell was synthesized by self-assembly. The obtained HPAA-peptide-HPG could form the compact nanocomplex with siPlk1, thus confirming the stable load of genes and subsequent targeted gene delivery. And the nanogenes could significantly induce apoptotic effect via the down-expression of Plk1 protein in breast cancer cells. Moreover, compared to polyethylenimine, HPAA-peptide-HPG exhibited superior biocompatibility through hemolysis and cell viability assays because of the shielding function of CD-HPG, thereby being beneficial to increasing the circulation time of the complex when administrated in vivo. Such an efficient and safe gene delivery complex (HPAA-peptide-HPG) presents a good example of rational design of cationic supramolecular vesicles for stimulus-responsive siRNA transport, which should be encouraged in cancer gene therapy.


Assuntos
Neoplasias da Mama , Nanopartículas , Neoplasias da Mama/genética , Feminino , Técnicas de Transferência de Genes , Homicídio , Humanos , RNA Interferente Pequeno/genética
12.
Nat Commun ; 12(1): 3568, 2021 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-34117255

RESUMO

Bone marrow (BM) chimeric mice are a valuable tool in the field of immunology, with the genetic manipulation of donor cells widely used to study gene function under physiological and pathological settings. To date, however, BM chimera protocols require myeloablative conditioning of recipient mice, which dramatically alters steady-state hematopoiesis. Additionally, most protocols use fluorescence-activated cell sorting (FACS) of hematopoietic stem/progenitor cells (HSPCs) for ex vivo genetic manipulation. Here, we describe our development of cell culture techniques for the enrichment of functional HSPCs from mouse BM without the use of FACS purification. Furthermore, the large number of HSPCs derived from these cultures generate BM chimeric mice without irradiation. These HSPC cultures can also be genetically manipulated by viral transduction, to allow for doxycycline-inducible transgene expression in donor-derived immune cells within non-conditioned immunocompetent recipients. This technique is therefore expected to overcome current limitations in mouse transplantation models.


Assuntos
Transplante de Medula Óssea , Medula Óssea/metabolismo , Quimera/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Animais , Células da Medula Óssea , Técnicas de Transferência de Genes , Engenharia Genética , Terapia Genética , Hematopoese , Transplante de Células-Tronco Hematopoéticas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Quimeras de Transplante
13.
Biomater Sci ; 9(11): 4127-4138, 2021 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-33954320

RESUMO

Ultrasound-activated microbubble destruction is a promising platform for gene delivery due to the low toxicity, non-invasiveness, and high specificity. However, the gene transfection efficiency is still low, especially for suspension cells. It is desirable to develop a universal gene delivery tool that overcomes the drawbacks existing in ultrasound-mediated methods. Here, we present a three-dimensional acoustic field-based conformal transfection (AFCT) system by designing a Sono-hole that can fit the three-dimensional acoustic field to maximally utilize the acoustic energy from bubble cavitation, thus greatly promoting the gene delivery efficiency. Surprisingly, compared with the traditional two-dimensional transfection system, the gene transfection efficiency of the AFCT system increased by more than 3 times, achieving nearly 30%. The parameters including acoustic pressure, duration, duty cycle, DNA concentrations, and bubble kinds were optimized to obtain higher gene transfection. In conclusion, our study provides an effective ultrasound-based gene delivery approach for gene transfection, especially for suspension-cultured cells.


Assuntos
Microbolhas , Ultrassom , Acústica , Técnicas de Transferência de Genes , Transfecção
14.
Nat Commun ; 12(1): 2928, 2021 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-34006888

RESUMO

Small interfering RNA (siRNA) is an effective therapeutic to regulate the expression of target genes in vitro and in vivo. Constructing a siRNA delivery system with high serum stability, especially responsive to endogenous stimuli, remains technically challenging. Herein we develop anti-degradation Y-shaped backbone-rigidified triangular DNA bricks with sticky ends (sticky-YTDBs) and tile them onto a siRNA-packaged gold nanoparticle in a programmed fashion, forming a multi-functional three-dimensional (3D) DNA shell. After aptamers are arranged on the exterior surface, a biocompatible siRNA-encapsulated core/shell nanoparticle, siRNA/Ap-CS, is achieved. SiRNAs are internally encapsulated in a 3D DNA shell and are thus protected from enzymatic degradation by the outermost layer of YTDB. The siRNAs can be released by endogenous miRNA and execute gene silencing within tumor cells, causing cell apoptosis higher than Lipo3000/siRNA formulation. In vivo treatment shows that tumor growth is completely (100%) inhibited, demonstrating unique opportunities for next-generation anticancer-drug carriers for targeted cancer therapies.


Assuntos
DNA/química , Técnicas de Transferência de Genes , Ouro/química , Nanopartículas Metálicas/química , Neoplasias/genética , RNA Interferente Pequeno/genética , Células A549 , Animais , DNA/genética , Inativação Gênica , Células HeLa , Humanos , Células MCF-7 , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias/metabolismo , Neoplasias/terapia , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/farmacocinética , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
15.
Biomacromolecules ; 22(6): 2436-2450, 2021 06 14.
Artigo em Inglês | MEDLINE | ID: mdl-34009977

RESUMO

Carbon dots (CDs) and G4-G6 (polyamidoamine)PAMAM-NH2 dendrimers were self-assembled to produce CDs@PAMAM nanohybrids for transfection and bioimaging purposes. CDs were synthesized by the hydrothermal method, using ascorbic acid as a starting precursor and characterized by transmission electron microscopy, UV-Vis, and fluorescence (in solution and solid-state) techniques. CDs were electrostatically combined with PAMAM dendrimers at room temperature, and the UV-Vis, fluorescence, and NMR spectroscopies were used to confirm the self-assembly. When compared to pristine CDs, nanohybrids were more photostable, resisting high acidic and basic pH. Moreover, they were considerably internalized by cells, as assessed by flow cytometry and fluorescence microscopy, and, when excited, displayed multi-color emission easily quantified and visualized. These nanoscale hybrids, coined hybridplexes, can condense pDNA and transfecting cells successfully, particularly the G5 CDs@PAMAM nanohybrids. In summary, CDs prepared in mild and smooth lab conditions, showing good optical properties, were used to prepare elegantly CDs@PAMAM nanohybrids with promising biomedical applications.


Assuntos
Dendrímeros , Carbono , Técnicas de Transferência de Genes , Transfecção
16.
Int J Nanomedicine ; 16: 2419-2441, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33814908

RESUMO

Lignin is an abundant renewable natural biopolymer. Moreover, a significant development in lignin pretreatment and processing technologies has opened a new window to explore lignin and lignin-based bionanomaterials. In the last decade, lignin has been widely explored in different applications such as drug and gene delivery, tissue engineering, food science, water purification, biofuels, environmental, pharmaceuticals, nutraceutical, catalysis, and other interesting low-value-added energy applications. The complex nature and antioxidant, antimicrobial, and biocompatibility of lignin attracted its use in various biomedical applications because of ease of functionalization, availability of diverse functional sites, tunable physicochemical and mechanical properties. In addition to it, its diverse properties such as reactivity towards oxygen radical, metal chelation, renewable nature, biodegradability, favorable interaction with cells, nature to mimic the extracellular environment, and ease of nanoparticles preparation make it a very interesting material for biomedical use. Tremendous progress has been made in drug delivery and tissue engineering in recent years. However, still, it remains challenging to identify an ideal and compatible nanomaterial for biomedical applications. In this review, recent progress of lignin towards biomedical applications especially in drug delivery and in tissue engineering along with challenges, future possibilities have been comprehensively reviewed.


Assuntos
Sistemas de Liberação de Medicamentos , Técnicas de Transferência de Genes , Lignina/química , Engenharia Tecidual , Animais , Biomassa , Humanos , Nanopartículas/química , Nanopartículas/ultraestrutura
17.
Int J Mol Sci ; 22(6)2021 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-33805602

RESUMO

Carriers of genetic material are divided into vectors of viral and non-viral origin. Viral carriers are already successfully used in experimental gene therapies, but despite advantages such as their high transfection efficiency and the wide knowledge of their practical potential, the remaining disadvantages, namely, their low capacity and complex manufacturing process, based on biological systems, are major limitations prior to their broad implementation in the clinical setting. The application of non-viral carriers in gene therapy is one of the available approaches. Poly(amidoamine) (PAMAM) dendrimers are repetitively branched, three-dimensional molecules, made of amide and amine subunits, possessing unique physiochemical properties. Surface and internal modifications improve their physicochemical properties, enabling the increase in cellular specificity and transfection efficiency and a reduction in cytotoxicity toward healthy cells. During the last 10 years of research on PAMAM dendrimers, three modification strategies have commonly been used: (1) surface modification with functional groups; (2) hybrid vector formation; (3) creation of supramolecular self-assemblies. This review describes and summarizes recent studies exploring the development of PAMAM dendrimers in anticancer gene therapies, evaluating the advantages and disadvantages of the modification approaches and the nanomedicine regulatory issues preventing their translation into the clinical setting, and highlighting important areas for further development and possible steps that seem promising in terms of development of PAMAM as a carrier of genetic material.


Assuntos
Dendrímeros/síntese química , Regulação Neoplásica da Expressão Gênica , Técnicas de Transferência de Genes , Terapia Genética/métodos , Proteínas de Neoplasias/antagonistas & inibidores , Neoplasias/terapia , Materiais Biocompatíveis/administração & dosagem , Materiais Biocompatíveis/síntese química , Dendrímeros/administração & dosagem , Regulamentação Governamental , Humanos , MicroRNAs/administração & dosagem , MicroRNAs/genética , MicroRNAs/metabolismo , Nanomedicina/legislação & jurisprudência , Nanomedicina/métodos , Nanopartículas/administração & dosagem , Nanopartículas/química , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Oligonucleotídeos Antissenso/administração & dosagem , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo , Plasmídeos/administração & dosagem , Plasmídeos/química , Plasmídeos/metabolismo , RNA Mensageiro/administração & dosagem , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Propriedades de Superfície
18.
Int J Mol Sci ; 22(9)2021 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-33926102

RESUMO

Inherited retinal dystrophies (IRDs) are a group of rare eye diseases caused by gene mutations that result in the degradation of cone and rod photoreceptors or the retinal pigment epithelium. Retinal degradation progress is often irreversible, with clinical manifestations including color or night blindness, peripheral visual defects and subsequent vision loss. Thus, gene therapies that restore functional retinal proteins by either replenishing unmutated genes or truncating mutated genes are needed. Coincidentally, the eye's accessibility and immune-privileged status along with major advances in gene identification and gene delivery systems heralded gene therapies for IRDs. Among these clinical trials, voretigene neparvovec-rzyl (Luxturna), an adeno-associated virus vector-based gene therapy drug, was approved by the FDA for treating patients with confirmed biallelic RPE65 mutation-associated Leber Congenital Amaurosis (LCA) in 2017. This review includes current IRD gene therapy clinical trials and further summarizes preclinical studies and therapeutic strategies for LCA, including adeno-associated virus-based gene augmentation therapy, 11-cis-retinal replacement, RNA-based antisense oligonucleotide therapy and CRISPR-Cas9 gene-editing therapy. Understanding the gene therapy development for LCA may accelerate and predict the potential hurdles of future therapeutics translation. It may also serve as the template for the research and development of treatment for other IRDs.


Assuntos
Amaurose Congênita de Leber/genética , Distrofias Retinianas/genética , Distrofias Retinianas/terapia , Dependovirus/genética , Proteínas do Olho/genética , Técnicas de Transferência de Genes , Terapia Genética , Vetores Genéticos , Humanos , Amaurose Congênita de Leber/terapia , Mutação , RNA , Retina/efeitos dos fármacos , Retina/metabolismo , Células Fotorreceptoras Retinianas Cones/efeitos dos fármacos , Células Fotorreceptoras Retinianas Cones/metabolismo
19.
Biomaterials ; 272: 120780, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33813260

RESUMO

There is growing interest in PEGylation of cationic polymeric vehicles for gene delivery in order to improve vehicle stability and reduce toxicity, but little is known about the effects of PEG coatings on transfection. We used a polymer from the poly(amine-co-ester) (PACE) family blended with PEG-conjugated PACE at different ratios in order to explore the effects of polyplex PEGylation on the transfection efficiency of plasmid DNA, mRNA, and siRNA in vitro and mRNA in vivo. We discovered that concentrations of PACE-PEG as low as 0.25% by weight improved polyplex stability but also inhibited transfection in vitro. In vivo, the effect of PACE-PEG incorporation on mRNA transfection varied by delivery route; the addition of PACE-PEG improved local delivery to the lung, but PEGylation had little effect on intravenous systemic delivery. By both delivery routes, transfection was inhibited at concentrations higher than 5 wt% PACE-PEG. These results demonstrate that excess PEGylation can be detrimental to vehicle function, and suggest that PEGylation of cationic vehicles must be optimized by PEG content, cargo type, and delivery route.


Assuntos
Ésteres , Polietilenoglicóis , Aminas , Técnicas de Transferência de Genes , Tamanho da Partícula , Transfecção
20.
Gene ; 788: 145664, 2021 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-33887371

RESUMO

The angiogenic gene therapy is an attractive approach for the treatment of ischemic muscle diseases, including peripheral arterial disease and ischemic heart diseases. Although a variety of gene transfer methods have been developed, the efficiency of gene transfer is still limited. We have been developing the needleless high-energy bioinjector device, Pyro-drive Jet Injector (PJI), based on pyrotechnics using a combination of ignition powder and gunpowder, however, the utility of PJI in gene transfer into muscle tissues remains unclear. pcDNA3.1 plasmid containing Flag was injected to the thigh muscles of C57BL/6J mice using PJI or needle, as a control. Histological analysis demonstrated that the protein expression of Flag was observed in a wider range in PJI group than in needle group. To assess the validity of PJI for gene therapy, pcDNA3.1-human fibroblast growth factor 2 (FGF2), which has angiogenic activity and tissue protective properties, was injected into the ischemic thigh muscles with PJI or needle. ELISA assay revealed that the protein expression of FGF2 was increased in the thigh muscle tissues by PJI-mediated gene delivery. Significantly, histological analyses revealed that muscle fiber cross-sectional area and the number of endothelial marker CD31 (+) cells was increased in ischemic hind-limb tissues of the PJI-FGF2 group but not in those of needle-FGF2 group. To expand the applicability of the PJI-mediated gene transfer, pcDNA3.1-venus plasmid was injected into murine hearts with PJI or needle. PJI method was successful in gene transfer into murine hearts, especially into cardiomyocytes, with high efficiency when compared to needle method. Collectively, the non-needle, non-liposomal and non-viral gene transfer by PJI could be a novel therapeutic approach for muscle diseases.


Assuntos
Fator 2 de Crescimento de Fibroblastos/administração & dosagem , Técnicas de Transferência de Genes/instrumentação , Músculo Esquelético/metabolismo , Miocárdio/metabolismo , Animais , Linhagem Celular , Fator 2 de Crescimento de Fibroblastos/genética , Fator 2 de Crescimento de Fibroblastos/metabolismo , Terapia Genética/instrumentação , Vetores Genéticos/administração & dosagem , Vetores Genéticos/farmacologia , Membro Posterior , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Doenças Musculares/genética , Doenças Musculares/metabolismo , Doenças Musculares/terapia , Plasmídeos/genética
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