Your browser does not allow saving data (e.g. private mode): bookmarks will be lost when the tab closes.
Search PubMed
Keywords, author, journal, years — results from PubMed, abstracts with Russian translation, bookmarks in your browser.
Found: 26 179
Page 884 of 1000-
Somatostatin receptor-targeted polymeric nanoplatform for efficient CRISPR/Cas9 gene editing to enhance synergistic hepatocellular carcinoma therapy.
The CRISPR/Cas9 system-based gene therapy can fundamentally address the issues of cancer occurrence, development, progression, and metastasis. However, the lack of targeting and effectiveness hinders gene therapy from entering clinical application. Herein, a somatostatin receptor-targeted polymeric nanoplatform is developed for the delivery of a PD-L1-targeted CRISPR/Cas9 system and synergistic treatment of hepatocellular carcinoma. This nanoplatform can effectively incorporate the CRISPR/Cas9 system and the chemotherapeutic drug paclitaxel to simultaneously address the biological safety and packaging capacity issues of viral vectors. After the octreotide-modified polymer (LNA-PEG-OCT) guided the nanoparticle into hepatoma carcinoma cells, the nanoparticle protected the CRISPR/Cas9 ribonucleoprotein complex (RNP) and achieved lysosomal escape. Then, the RNP reached the target gene (PD-L1) under the guidance of the single guide RNA (sgRNA) in the RNP. The PD-L1 gene editing efficiency reached up to 55.8% for HepG2 cells in vitro and 46.0% for tumor tissues in vivo, leading to effective suppression of PD-L1 protein expression. Substantial inhibition of hepatocellular carcinoma cell proliferation and further 79.45% growth repression against subcutaneous xenograft tumors were achieved. Overall, this somatostatin receptor-targeted polymeric nanoplatform system not only provides a promising nanocarrier for CRISPR/Cas9 system delivery, but also expands the potential of combining gene editing and chemotherapy.
-
CRISPR-Cas9 Ribonucleoprotein-Mediated Genomic Editing in Primary Innate Immune Cells.
CRISPR-Cas9 genome engineering can be used to functionally investigate the complex mechanisms of immune system regulation. Decades of work have aimed to genetically reprogram innate immunity, but current approaches are inefficient or nonspecific, limiting their use. Here, we detail an optimized strategy for non-viral CRISPR-Cas9 ribonucleoprotein (cRNP) genomic editing of primary innate lymphocytes (ILCs) and myeloid lineage cells, resulting in high-efficiency editing of target gene expression from a single electroporation. For complete details on the use and execution of this protocol, please refer to Riggan et al. (2020).
-
TIGR on the loose: A dual-guide RNA system for DNA targeting.
A recent study1 unveils tandem interspaced guide RNAs (TIGRs) that simultaneously engage both strands of target DNA and direct Tas protein activity. It offers insights into the evolution of RNA-guided proteins and introduces a promising tool for genome editing.
-
AcrIIA5 Inhibits a Broad Range of Cas9 Orthologs by Preventing DNA Target Cleavage.
CRISPR-Cas9 is an adaptive immune system for prokaryotes to defend against invasive genetic elements such as phages and has been used as a powerful tool for genome editing and modulation. To overcome CRISPR immunity, phages encode anti-CRISPR proteins (Acrs) to inhibit Cas9, providing an efficient "off-switch" tool for Cas9-based applications. Here, we characterized AcrIIA5, which is a Cas9 inhibitor discovered in a virulent phage of Streptococcus thermophilus. We found that AcrIIA5 is a potent and broad-spectrum inhibitor of CRISPR-Cas9, which can inhibit diverse Cas9 orthologs of type II-A, type II-B, and type II-C. AcrIIA5 inhibits Cas9 by preventing DNA target cleavage, but DNA target binding of Cas9 is unaffected. Importantly, it can affect the activity of the RuvC nuclease domain of Cas9 independent of the HNH nuclease domain. Our work expands the diversity of the inhibitory mechanisms used by Acrs and provides the guidance for developing controlling tools in Cas9-based applications.
-
CRISPR/Cas-Mediated Genome Editing for Developing Herbicide Tolerant Rice: A Step-by-Step Protocol.
Weed management in rice cultivation has predominantly relied on acetolactate synthase (ALS) and acetyl-CoA carboxylase (ACCase) inhibiting herbicides, particularly against Echinochloa spp. (watergrass). However, these herbicides carry a high risk of resistance evolution, as evidenced by the numerous resistant biotypes reported worldwide. The emergence of herbicide resistance necessitates innovative and sustainable weed control strategies. Genome editing, particularly through the CRISPR/Cas system, provides a precise and efficient platform for introducing targeted genetic modifications to develop herbicide-tolerant (HT) rice cultivars. In this protocol, we present a step-by-step approach for generating bispyribac sodium-tolerant rice using the CRISPR/Cas-mediated editing of the ALS gene. The method encompasses guide RNA design, vector construction, transformation, selection of edited plants, and molecular confirmation of targeted mutations. This approach offers a robust framework for producing HT rice lines, potentially reducing reliance on conventional herbicide regimes and mitigating the risk of resistance development in weed populations.
-
A quantitative assay for the efficiency of RNA-guided genome editing in plants.
RNA-guided endonucleases originating from the bacterial CRISPR/Cas system are a versatile tool for targeted gene editing. To determine the functional relevance of a gene of interest, deletion of the entire open reading frame (ORF) by two independent double-strand breaks (DSBs) is particularly attractive. This strategy greatly benefits from high editing efficiency, which is strongly influenced by the Cas endonuclease version used. We developed two reporter switch-on assays, for quantitative comparison and optimization of Cas constructs. The assays are based on four components: (i) A reporter gene, the mRNA of which carries a hairpin (HP) loop targeted by (ii) the endoribonuclease Csy4. Cleavage of the mRNA at the HP loop by Csy4 abolishes the translation of the reporter. Csy4 was used as the target for full deletion. (iii) A Cas system targeting sites flanking the Csy4 ORF with a 20-bp spacer either side to preferentially detect full-deletion events. Loss of functional Csy4 would lead to reporter gene expression, allowing indirect quantification of Cas-mediated deletion events. (iv) A reference gene for normalization. We tested these assays on Nicotiana benthamiana leaves and Lotus japonicus calli induced on hypocotyl sections, using Firefly luciferase and mCitrine as reporter genes and Renilla luciferase and hygromycin phosphotransferase II as reference genes, respectively. We observed a >90% correlation between reporter expression and full Csy4 deletion events, demonstrating the validity of these assays. The principle of using the Csy4-HP module as Cas target should be applicable to other editing goals including single DSBs in all organisms.
-
Roadmap and Considerations for Genome Editing in a Non-Model Organism: Genetic Variations and Off-Target Profiling.
The CRISPR/Cas genome editing approach in non-model organisms poses challenges that remain to be resolved. Here, we demonstrated a generalized roadmap for a de novo genome annotation approach applied to the non-model organism Macrobrachium rosenbergii. We also addressed the typical genome editing challenges arising from genetic variations, such as a high frequency of single nucleotide polymorphisms, differences in sex chromosomes, and repetitive sequences that can lead to off-target events. For the genome editing of M. rosenbergii, our laboratory recently adapted the CRISPR/Cas genome editing approach to embryos and the embryonic primary cell culture. In this continuation study, an annotation pipeline was trained to predict the gene models by leveraging the available genomic, transcriptomic, and proteomic data, and enabling accurate gene prediction and guide design for knock-outs. A next-generation sequencing analysis demonstrated a high frequency of genetic variations in genes on both autosomal and sex chromosomes, which have been shown to affect the accuracy of editing analyses. To enable future applications based on the CRISPR/Cas tool in non-model organisms, we also verified the reliability of editing efficiency and tracked off-target frequencies. Despite the lack of comprehensive information on non-model organisms, this study provides an example of the feasibility of selecting and editing specific genes with a high degree of certainty.
-
VERVE-101: a promising CRISPR-based gene editing therapy that reduces LDL-C and PCSK9 levels in HeFH patients.
-
Generation of the Fip1l1-Pdgfra fusion gene using CRISPR/Cas genome editing.
-
Advancements in Gene Therapy for Non-Small Cell Lung Cancer: Current Approaches and Future Prospects.
Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death worldwide, characterized by late diagnosis and resistance to conventional therapies. Gene therapy has emerged as a promising alternative for NSCLC therapy, especially for patients with advanced disease who have exhausted conventional treatments. This article delved into the current developments in gene therapy for NSCLC, including gene replacement and tumor suppressor gene therapy, gene silencing, CRISPR/Cas9 gene editing, and immune modulation with CAR-T cell therapy. In addition, the challenges and future prospects of gene-based therapies for NSCLC were discussed.
PubMed returns only the first 10,000 results — refine the query to see the rest.