JOURNAL OF CONTROLLED RELEASE
·
2025
·
IF 11.5
Tong Wang,Tianyi Chen,Dazhao Li,Xiaoxing Hang,Shuangshuang Zhang,Han Yi,Tianyue Jiang,Dawei Ding,Xuenong Zhang
Cancer stem cells (CSCs) play an important role in the development of triple-negative breast cancer (TNBC), including metastasis, invasion, tumorigenicity, and drug resistance. Moreover, non-CSCs can spontaneously transform into CSCs in special tumor microenvironments, thereby leading to poor prognosis or even failed treatments. Therefore, reversing CSCs into normal tumor cells in a sustained-acting manner is a promising strategy. It has been reported that down-regulation of FBXO44 protein expression inhibits tumor cell stemness. Moreover, CRISPR/Cas9 technology, a well-known precise gene editing tool, was adopted to permanently block FBXO44 within the genome upon its successful implementation. Given this, a core-shell nanoparticle (NP) consisting of amphiphilic polymer core and crosslinked-hyaluronic acid shell (nDOX-PL/pFBXO44 NPs) is developed in this work to concurrently deliver FBXO44-targeted CRISPR/Cas9 plasmids (pFBXO44) and doxorubicin (DOX) for combinational CSC reprogramming and chemotherapy of TNBC, which exhibits tumor cell targeting, endosomal escape, and reduction responsiveness to release DOX and plasmids in the cytoplasma. CRISPR/Cas9-mediated downregulation of FBXO44 expression could convert CSC into normal tumor cells, and effectively inhibit tumor growth without obvious side effects in vivo after combining with chemotherapy. In summary, we developed an intelligent system to co-deliver genetic and hydrophobic drugs, achieving effective cancer stemness reversal and synergistic suppression of contractable TNBC.
研究领域:
肿瘤学
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基因编辑
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纳米医学
·
药理学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
JOURNAL OF COLLOID AND INTERFACE SCIENCE
·
2025
·
IF 9.7
Shi-Cheng Tian,Xun-Huan Song,Ke-Ke Feng,Cheng-Lei Li,Yi-Fan Tu,Yong-Shan Hu,Jing-Wei Shao
Photodynamic therapy (PDT) has arisen as a promising method due to its spatiotemporal precision and minimal invasiveness. It encounters significant obstacles in solid tumors due to hypoxia-induced therapeutic resistance and the self-protective mechanisms of cancer cells facilitated by MutT homolog 1 (MTH1), an enzyme involved in oxidative damage repair. Herein, we fabricate a tumor-microenvironment responsive CRISPR nanoplatform based on hollow mesoporous manganese dioxide (H-MnO 2 ) for PDT. This platform utilizes H-MnO 2 to produce oxygen (O 2 ) through the decomposition of hydrogen peroxide (H 2 O 2 ) in TME, thereby mitigating hypoxia and enhancing reactive oxygen species (ROS) generation. The high concentration of glutathione (GSH) and hyaluronidase (HAase) in TME induces the release of CRISPR/Cas9 ribonucleoproteins (RNP) to target the MTH1 gene, thereby impairs oxidative damage repair pathways and amplifys ROS-mediated cytotoxicity. The released Mn 2+ ions function as immunomodulatory agents, activate innate immune responses via stimulating STING signal pathway. In vitro , IHMRH NPs markedly increased intracellular O 2 levels, ROS production, lipid peroxidation and DNA damage, leading to tumor cell death, immune activation, and effective gene editing. In vivo , the nanoplatform suppressed tumor growth, diminished MTH1 gene expression, stimulated dendritic cell (DC) maturation through immunogenic cell death (ICD). This multimodal nanosystem may amplifies oxidative stress, collaborates with innate and adaptive immune activation to surpass the constraints of traditional PDT. The research presents a novel framework for cancer combination therapy by systematically integrating nanotechnology with precision gene editing.
研究领域:
生物医学工程
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肿瘤学
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基因治疗
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免疫学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF PHARMACEUTICS
·
2024
·
IF 5.3
Qian Sun,Hongqian Zhang,Feng Ding,Xue Gao,Zongwei Zhu,Chuanxu Yang
CRISPR-Cas genome editing technology holds great promise for wide-ranging biomedical applications. However, the development of efficient delivery system for CRISPR-Cas components remains challenging. Herein, we synthesized a series of ionizable lipids by conjugation of alkyl-acrylate to different amine molecules and further assembled ionizable lipid nanoparticles (iLNPs) for co-delivery of Cas9 mRNA and sgRNA. Among all the iLNP candidates, 1A14-iLNP with lipids containing spermine as amine head, demonstrated the highest cellular uptake, endosomal escape and mRNA expression in vitro. Co-delivery of Cas9 mRNA and sgRNA targeting EGFP by 1A14-iLNP achieved the highest EGFP knockout efficiency up to 70% in HeLa-EGFP cells. In addition, 1A14-iLNP displayed passive liver-targeting delivery of Cas9 mRNA in vivo with good biocompatibility. Moreover, we developed a simple method of lyophilization-mediated reverse transfection of CRISPR-Cas9 components for efficient genome editing. Therefore, the developed 1A14-iLNP and the lyophilization formulation, represent a potent solution for CRISPR-Cas9 delivery, which might broaden the future of biomedical applications of both mRNA and CRISPR-based therapies.
研究领域:
分子生物学
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基因治疗
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纳米医学
✓ 已验证
DOI
PubMed
高影响因子
npj Precision Oncology
·
2025
·Vol. 9
·期 1
·1-21
·
IF 8
Li Xiaoli,Wang Jiashuo,Guo Jiaxing,Zhang Ming
Cisplatin resistance is a major challenge in laryngeal cancer treatment. Tumor necrosis factor-alpha-induced protein 2 (TNFAIP2) has been implicated in chemoresistance, though its role in laryngeal cancer remains unclear. We identified TNFAIP2 as a key gene associated with cisplatin resistance and developed a second near-infrared (NIR-II) light-responsive CRISPR-Cas9 nanosystem (APC@RBCs) to target it. In cisplatin-resistant Tu177/CDDP cells, light-activated knockout of TNFAIP2 significantly increased cisplatin sensitivity (IC50 reduced from 12.55 to 4.37 µg/mL). Mechanistic studies revealed that TNFAIP2 modulates cisplatin resistance via the NRF2 pathway, affecting oxidative stress response and epithelial-mesenchymal transition. TNFAIP2 deletion also suppressed cancer cell migration and invasion. Our results establish TNFAIP2 as a critical mediator of chemoresistance and demonstrate the potential of APC@RBCs as a precise, non-invasive gene therapy platform. This strategy offers a promising approach to overcoming cisplatin resistance and improving treatment outcomes in laryngeal cancer patients.
研究领域:
肿瘤学
·
分子生物学
·
药理学
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纳米技术
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基因工程
✓ 已验证
产品关联
DOI
PubMed
高影响因子
CANCER RESEARCH
·
2025
·
IF 16.6
Shang Yan,Li Shuaihu,Xu Tian,Sun Yihua,Li Fei
The CRISPR/Cas9 lentiviral system has emerged as a powerful tool for gene knockout in cancer immunology research. However, the persistent expression of exogenous elements, such as Cas9 and resistance markers, often triggers excessive tumor immune rejection. This can lead to prolonged experimental timelines, increased data variability, biased outcomes, and even experimental failures. To address this challenge, several strategies have been investigated that offer partial solutions, but they have failed so far to comprehensively resolve the issue. In response, we developed the VL-AdCre system, a strategy that enables efficient excision of exogenous expression elements following gene knockout. The VL-AdCre system effectively reduced tumor immune rejection in allograft models, streamlined experimental workflows, and improved the reliability of research outcomes. Overall, this optimized CRISPR/Cas9 lentiviral gene knockout system offers a robust and practical solution for studying gene functions in vivo and advancing immunotherapeutic strategies.
研究领域:
免疫学
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肿瘤学
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分子生物学
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基因编辑
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转化医学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
NUCLEIC ACIDS RESEARCH
·
2020
·Vol. 48
·期 20
·11773-11784
·
IF 11.502
Lin Jiao,Liu Yan,Lai Peidong,Ye Huixia,Xu Liang
A variety of nanodevices developed for nucleic acid computation provide great opportunities to construct versatile synthetic circuits for manipulation of gene expressions. In our study, by employing a two-hairpin mediated nucleic acid strand displacement as a processing joint for conditional guide RNA, we aim to build artificial connections between naturally occurring RNA expressions through programmable CRISPR/Cas9 function. This two-hairpin joint possesses a sequence-switching machinery, in which a random trigger strand can be processed to release an unconstrained sequence-independent strand and consequently activate the self-inhibitory guide RNA for conditional gene regulation. This intermediate processor was characterized by the fluorescence reporter system and applied for regulation of the CRISPR/Cas9 binding activity. Using plasmids to generate this sequence-switching machinery in situ, we achieved the autonomous genetic regulation of endogenous RNA expressions controlled by other unrelated endogenous RNAs in both E. coli and human cells. Unlike previously reported strand-displacement genetic circuits, this advanced nucleic acid nanomachine provides a novel approach that can establish regulatory connections between naturally occurring endogenous RNAs. In addition to CRISPR systems, we anticipate this two-hairpin machine can serve as a general processing joint for wide applications in the development of other RNA-based genetic circuits.
研究领域:
合成生物学
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分子遗传学
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基因工程
✓ 已验证
DOI
PubMed
高影响因子
Nanoscale
·
2022
·Vol. 14
·期 39
·14645-14660
·
IF 8.307
Li-Min Zhai,Yan Zhao,Rui-Lin Xiao,Shi-Quan Zhang,Bao-Hua Tian,Xin-Xin Li,Rong Zhang,Ri-Sheng Ma,Hai-Xia Liang
Nuclear targeted delivery has great potential in improving the efficiency of non-viral carrier mediated genome editing. However, direct and efficient delivery of CRISPR/Cas9 plasmid into the nucleus remains a challenge. In this study, a nuclear targeted gene delivery platform based on fluorescent carbon quantum dots (CQDs) was developed. Polyethylenimine (PEI) and polyethylene glycol (PEG) synergistically passivated the surface of CQDs, providing an excitation-independent green-emitting fluorescent CQDs–PEI–PEG conjugate (CQDs–PP) with an ultra-small size and positive surface charge. Here we show that CQDs–PP could bind CRISPR/Cas9 plasmid to form a nano-complex by electrostatic attraction, which can bypass lysosomes and enter the nucleus by passive diffusion, and thereby improve the transfection efficiency. Also, CQDs–PP could deliver CRISPR/Cas9 plasmid into HeLa cells, resulting in the insertion/deletion mutation of the target EFHD1 gene. More importantly, CQDs–PP exhibited a considerably higher gene editing efficiency as well as comparable or lower cytotoxicity relative to Lipo2000 and PEI-passivated CQDs–PEI (CQDs–P). Thus, the nuclear-targeted CQDs–PP is expected to constitute an efficient CRISPR/Cas9 delivery carrier in vitro with imaging-trackable ability.
研究领域:
遗传学
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分子生物学
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生物医学工程
·
纳米技术
✓ 已验证
产品关联
DOI
PubMed
高影响因子
MOLECULAR AND CELLULAR BIOCHEMISTRY
·
2023
·
IF 4.3
Lu Junnan,Tong Ying,Dong Rui,Yang Yijun,Hu Wen,Zhang Minghong,Liu Quan,Zhao Siting,Adams John H.,Qin Li,Chen Xiaoping
CRISPR/Cas9 technology applied to Plasmodium falciparum offers the potential to greatly improve gene editing, but such expectations including large DNA fragment knock-ins and sequential gene editing have remained unfulfilled. Here, we achieved a major advance in addressing this challenge, especially for creating large DNA fragment knock-ins and sequential editing, by modifying our suicide-rescue-based system that has already been demonstrated to be highly efficient for conventional gene editing. This improved approach was confirmed to mediate efficient knock-ins of DNA fragments up to 6.3 kb, to produce “marker-free” genetically engineered parasites and to show potential for sequential gene editing. This represents an important advancement in establishing platforms for large-scale genome editing, which might gain a better understanding of gene function for the most lethal cause of malaria and contribute to adjusting synthetic biology strategies to live parasite malaria vaccine development. Site-directed knock-in of large DNA fragments is highly efficient using suicide-rescue-based CRISPR/Cas9 system, and sequential gene insertion is feasible but further confirmation is still needed.
研究领域:
寄生虫学
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分子生物学
·
基因组编辑
✓ 已验证
DOI
PubMed
高影响因子
BIOTECHNOLOGY LETTERS
·
2023
·Vol. 45
·期 3
·401-410
·
IF 2.7
Tian Li,Yan Bingyu,Huo Dandan,Sun Wenhui,Cui Sufang,Li Xiaojing,Zhang Xiangmei,Dong Huijun
Objectives To develop a modified CRISPR/Cas9 system with the β-glucuronidase (GusA) reporter and a dual sgRNA cassette for Nonomuraea gerenzanensis ( N. gerenzanensis ). Results With the aid of a visual GusA reporter, the complicated and tedious process of cloning and gene identification could be abandoned entirely in the genetic editing of N. gerenzanensis . Moreover, introducing a dual sgRNA cassette into the CRISPR/Cas9 system significantly improved gene deletion efficiency compared to the single sgRNA element. Furthermore, the length of the homologous flanking sequences set to the lowest value of 500 bp in this system could still reach the relatively higher conjugation transfer frequency. Conclusions The enhanced CRISPR/Cas9 system could efficiently perform genetic manipulation on the rare actinomycete N. gerenzanensis .
研究领域:
微生物学
·
分子生物学
·
遗传工程
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Synthetic and Systems Biotechnology
·
2019
·
IF 0
Jingjun Mo,Shuwen Wang,Wan Zhang,Chunyu Li,Zixin Deng,Lixin Zhang,Xudong Qu
Actinobacteria able to produce varieties of bioactive natural products have been long appreciated by the field of drug discovery and development. Recently, a few of CRISPR/Cas9 systems bearing different types of replicons (pSG5 and pIJ101) were developed to efficiently edit their genomes. Despite wide application in gene editing, their utility in editing challenging DNA regions e.g. high sequence identity has not been compared. In this study, we confirmed that the widely used temperature-sensitive pSG5 replicon is indeed not suitable for editing modular polyketide synthase (PKS) genes due to causing unpredicted gene recombination. This problem can be addressed by replacing the pSG5 with the segregationally unstable pIJ101 replicon. By introducing a counter-selection marker CodA, convenient cloning sites in the single guide RNAs (sgRNAs) and homologous template scaffolds, we developed a new CRISPR-Cas9 system pMWCas9. This system was successfully used to delete/replace erythromycin PKS and other biosynthetic genes in Saccharopolyspora erythraea and Streptomyces sp. AL2110. By swapping the promoters of antB and antC with ermE and kasOp , we achieved a deacyl-antimycin hyper producer which produces a 9-fold higher yield than the original Streptomyces sp. AL2110 strain. Our results provide a robust and useful Cas9 tool for genetic studies in Actinobacteria .
研究领域:
遗传学
·
微生物学
·
生物技术
·
药物发现
✓ 已验证
DOI
PubMed
Biology-Basel
·
2021
·Vol. 10
·期 10
·1038
·
IF 5.079
Haodong Cui,Min Jiang,Wenhua Zhou,Ming Gao,Rui He,Yifan Huang,Paul K. Chu,Xue-Feng Yu
Simple SummaryCRISPR/Cas9 system as a potential gene editing platform has been widely applied in biological engineering and disease therapies. To achieve precise gene targeting, active CRISPR/Cas9 components must be efficiently transported to targeted cells. As a simple and effective strategy, Cold Atmospheric Plasma (CAP) treatment has been demonstrated for the transmembrane delivery of various exogenous materials. In comparison with carrier-dependent delivery methods, this carrier-free platform provides a promising alternative to circumvent the obstacles of biosafety and complicated preparation processes. In this work, a CAP-based CRISPR/Cas9 carrier-free delivery platform has been established and corresponding mechanism related to efficient transportation has been explored. Briefly, the efficient production of bioactive species in culture media after CAP treatment alters cell membrane potential and permeability, which facilitates cytosolic delivery of active CRISPR/Cas9 components via passive diffusion and ATP-dependent endocytosis pathways, resulting in efficient genome editing and gene silencing. This carrier-free strategy using CAP-based transportation may also be extended to other active biomolecules in drug delivery and gene therapy.AbstractA carrier-free CRISPR/Cas9 ribonucleoprotein delivery strategy for genome editing mediated by a cold atmospheric plasma (CAP) is described. The CAP is promising in many biomedical applications due to efficient production of bioactive ionized species. The MCF-7 cancer cells after CAP exposure exhibit increased extracellular reactive oxygen and nitrogen species (RONS) and altered membrane potential and permeability. Hence, transmembrane transport of Ca2+into the cells increases and accelerates ATP hydrolysis, resulting in enhanced ATP-dependent endocytosis. Afterwards, the increased Ca2+and ATP contents promote the release of cargo into cytoplasm due to the enhanced endosomal escape. The increased membrane permeability also facilitates passive diffusion of foreign species across the membrane into the cytosol. After CAP exposure, the MCF-7 cells incubated with Cas9 ribonucleoprotein (Cas9-sgRNA complex, Cas9sg) with a size of about 15 nm show 88.9% uptake efficiency and 65.9% nuclear import efficiency via passive diffusion and ATP-dependent endocytosis pathways. The efficient transportation of active Cas9sg after the CAP treatment leads to 21.7% and 30.2% indel efficiencies in HEK293T and MCF-7 cells, respectively. This CAP-mediated transportation process provides a simple and robust alternative for the delivery of active CRISPR/Cas9 ribonucleoprotein. Additionally, the technique can be extended to other macro-biomolecules and nanomaterials to cater to different biomedical applications.Keywords:cold atmospheric plasma;CRISPR/Cas9;carrier-free transport;genome editing
研究领域:
生物医学工程
·
遗传学
·
分子生物学
·
治疗学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
ANALYST
·
2023
·Vol. 148
·期 10
·2308-2315
·
IF 4.2
Ming Zhou,Xin Li,Herui Wen,Bin Huang,Jiali Ren,Jialin Zhang
The simple and efficient detection of nucleic acids is important in the diagnosis of tuberculosis (TB) caused by Mycobacterium tuberculosis (M. tuberculosis). However, base mismatch will lead to false positive and false negative nucleic acid test, which seriously interferes with the accuracy of the final results. Herein, we demonstrated a CRISPR/Cas-9-mediated fluorescent strategy utilizing fluorescence resonance energy transfer (FRET) for the detection of bacteria. High-variable region of M. tuberculosis 16S rDNA fragment was used as the target, and CRISPR/Cas9 was used as the recognition element. The binding of the P1 probe of upconversion nanoparticles (UCNPs) @SiO2-P1 and the P2 probe of Fe3O4@Au-P2 caused the fluorescence quenching of UCNPs. In the presence of the target, the P2 probe hybridized with the target to form double-stranded DNA (dsDNA), which was recognized and cleaved by CRISPR/Cas9, resulting in the breaking of the P1–P2 duplex linkage. UCNPs moved away from Fe3O4@Au under a magnetic field, and the fluorescence signal was restored; bacteria were detected under the excitation of a 980 nm laser source. Using the CRISPR/Cas-9-mediated system, the sensor could distinguish single-base mismatches in 10 bases from the protospacer adjacent motif (PAM) region. The limit of detection (LOD) was 20 CFU mL−1 and the detection time was 2 h. It developed a new way of accurate nucleic acid detection for disease diagnosis.
研究领域:
生物技术
·
医学诊断
·
纳米技术
·
分子生物学
✓ 已验证
DOI
PubMed
高影响因子
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