JOURNAL OF CONTROLLED RELEASE
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2025
·
IF 11.5
Enze Li,Liewei Wen,Caiyun Yin,Nan Wang,Suleixin Yang,Wenzheng Feng,Meiwan Chen
Cuproptosis as a copper-dependent cell death modality driven by pathological aggregation of lipoylated proteins and proteotoxic stress resulting from destabilization of iron-sulfur (Fe S) cluster proteins, represents a promising therapeutic strategy for cancer. However, the therapeutic efficacy of cuproptosis can be compromised by the intrinsic compensatory mechanisms within cancers, particularly low intracellular copper ion concentration and protective autophagy, which facilitates cellular adaptation and survival under stress. To overcome this limitation, a self-amplifying cuproptosis nanoregulator (SHK-Cu/ TK PF/pATG5@HA, abbreviated as SC/TpA@HA) for CD44-targeted delivery is developed that integrate shikonin‑copper (SHK-Cu) coordination complexes with CRISPR/Cas9 plasmids targeting ATG5, condensed by fluorinated polyethyleneimine ( TK PF)-condensed to enhance cancer therapy. Briefly, Cu + is released from the dissociated SHK-Cu complex upon intracellular GSH activation induces dihydrolipoamide S -acetyltransferase (DLAT) oligomerization and reduces Fe S cluster proteins, triggering tricarboxylic acid (TCA) cycle collapse and irreversible mitochondrial damage. Concurrently, CRISPR/Cas9-mediated ATG5 knockout prevents autophagosome formation, creating an autophagic flux trap that accumulates copper-damaged mitochondria. Notably, such mitochondrial dysfunction as induced by copper overload combined with impaired cellular damage clearance from autophagy blockade elevates cuproptosis. In addition, the immunogenic cell death through cuproptosis in cancer cells, as validated by the exposure of calreticulin and the extracellular release of HMGB1, triggers a potent anti-tumor immune response. This response is enhanced through autophagy inhibition, as ATG5 deletion blocks the downstream signaling of copper-activated Unc-51-like autophagy activating kinase 1/2 (ULK1/2), ultimately amplifying cytotoxic T lymphocyte infiltration. Therefore, this dual intervention through copper overload and autophagy blockade potentiates both cuproptosis and anti-tumor immune effect, representing an innovative strategy of cuproptosis treatment.
研究领域:
肿瘤学
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分子生物学
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纳米医学
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免疫学
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细胞死亡机制
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基因编辑
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代谢调控
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转化医学
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高影响因子
Neuroscience Bulletin
·
2021
·Vol. 37
·期 9
·1271-1288
·
IF 5.203
Li Hao,Wu Shihao,Ma Xia,Li Xiao,Cheng Tianlin,Chen Zhifang,Wu Jing,Lv Longbao,Li Ling,Xu Liqi,Wang Wenchao,Hu Yingzhou,Jiang Haisong,Yin Yong,Qiu Zilong,Hu Xintian
Whether direct manipulation of Parkinson’s disease (PD) risk genes in the adult monkey brain can elicit a Parkinsonian phenotype remains an unsolved issue. Here, we used an adeno-associated virus serotype 9 (AAV9)-delivered CRISPR/Cas9 system to directly co-edit PINK1 and DJ-1 genes in the substantia nigras (SNs) of two monkey groups: an old group and a middle-aged group. After the operation, the old group exhibited all the classic PD symptoms, including bradykinesia, tremor, and postural instability, accompanied by key pathological hallmarks of PD, such as severe nigral dopaminergic neuron loss (>64%) and evident α-synuclein pathology in the gene-edited SN. In contrast, the phenotype of their middle-aged counterparts, which also showed clear PD symptoms and pathological hallmarks, were less severe. In addition to the higher final total PD scores and more severe pathological changes, the old group were also more susceptible to gene editing by showing a faster process of PD progression. These results suggested that both genetic and aging factors played important roles in the development of PD in the monkeys. Taken together, this system can effectively develop a large number of genetically-edited PD monkeys in a short time (6–10 months), and thus provides a practical transgenic monkey model for future PD studies.
研究领域:
神经科学
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分子生物学
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遗传学
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衰老研究
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疾病模型
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高影响因子
FOOD CHEMISTRY
·
2026
·
IF 10.4
Yipeng Zhang,Shijia Wu,Yisheng Chen
A portable biosensor for the peanut allergen Ara h 1 was developed by integrating a polydopamine-functionalized pipette tip, CRISPR/Cas12a signal amplification, and a personal glucose meter. Ara h 1 binding induced partial displacement of a cDNA activator, which activated Cas12a trans-cleavage and released glucose oxidase from GOx–DNA conjugates. The released enzyme catalyzed glucose oxidation, producing a quantitative decrease in glucose concentration. Under optimized conditions, the assay showed a linear range of 50–1000 ng/mL and a solution-based limit of detection of 22.19 ng/mL. Post-extraction matrix-spike tests in commercial peanut-biscuit and peanut-beverage extracts yielded recoveries of 101.58–104.68% with relative standard deviations below 3.44%. These results demonstrate the analytical feasibility and preliminary matrix compatibility of the platform, although validation using pre-extraction fortified, incurred, and naturally contaminated foods is still required.
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Journal of Biological Engineering
·
2026
·Vol. 20
·期 1
·29
·
IF 6.5
Ye Changchuan,Zhang Yan,Zhang Jie,Shi Menglei,Nie Feixue,Liu Qinghua
Background Vitamin K 2 (VK 2 ), as a derivative of the menaquinone family, plays an important role in the prevention of osteoporosis and cardiovascular calcification. The realization of the industrialization of VK 2 and the reduction of its production cost have become the focus of attention. Results In this work, an E. coli strain with high VK 2 accumulation was constructed through rational metabolic engineering and stepwise improvement based on regulatory metabolic information and CRISPR/Cas9-mediated gene knockout. We first constructed a recombinant E. coli strain BW-T7/MU to produce menaquinol-8 (MKH 2 -8, a reduced form of VK 2 ) by overexpressing menA and ubiE genes, which encoding the rate-limiting enzymes of the menaquinol pathway. After 24 h and 48 h of fermentation, this strain BW-T7/MU reach a titer of 303 mg/L and 232 mg/L. Secondly, we overexpressed different related genes wrbA (oxidative stress mitigation), qorB (reduction of quinones) and menF (conversion of chorismate to isochorismate), respectively. Among these recombinant strains, the strain BW-T7/MUW (overexpressing menA , ubiE and wrbA genes) reached the highest titer of VK 2 after 48 h of fermentation. The optimization of the medium led to an increase in the accumulation of VK 2 . Subsequently, the rational metabolic engineering of gene knockout further increased the titer of VK 2 . The recombinant strain ΔB/MUW was selected as the dominant strain for further optimization, with a high VK 2 titer of 724 mg/L. A final attempt is to overexpress ispB gene to increased flux of isoprenoid side chain synthesis, resulting in strain ΔB/MUWI with a titer of 859 mg/L in a shake flask and 1360 mg/L in a 5 L fermenter after 48 h cultivation. Conclusions The stepwise engineering strategy raised the VK 2 titer from the initial 303 mg/L to 859 mg/L through rational pathway modification and systematic gene expression. Further optimization in batch fermentation increased the VK 2 titer to 1360 mg/L, which highlights the strong engineering impact of our strategy.
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高影响因子
BIOSENSORS & BIOELECTRONICS
·
2026
·
IF 11.8
Yan Wang,Boya Xu,Zehan Zeng,Haixin Wang,Lin Shi,Wanting Liu,Yanyan Chen,Xuexian Deng,Jun Chen,Jin-Xiang Chen
Extracellular vesicle (EV)-based liquid biopsy holds great promise for glioma diagnosis, but its clinical translation remains hindered by inefficient isolation of disease-relevant EV subpopulations and insufficiently integrated signal validation. Herein, we develop a platform that integrates a bifunctional nanozyme system (phosphatase-like MOF and peroxidase-like Pt) with a CHA-CRISPR/Cas12a cascade, achieving selective isolation and ultrasensitive detection of EGFR-positive glioma-derived EVs. Defective UiO-66-NH 2 loaded with Pt nanoparticles and functionalized with EGFR aptamers (UiO@Pt@Apt) is immobilized on Hook strand-modified glass 96-well plates via Apt-Hook hybridization. Upon introduction of EGFR-positive EVs, they specifically bind to aptamers on UiO@Pt@Apt and induce the release of UiO@Pt@Apt-EV complexes through perturbation and destabilization of the Apt-Hook interface. After EV lysis, EV-derived miRNA-21 activates the CHA-CRISPR/Cas12a cascade to generate a fluorescent signal, while the nucleotide fragments produced by Cas12a trans-cleavage are proposed to be hydrolyzed by phosphatase-like defective UiO-66-NH 2 to generate PO 4 3− . The PO 4 3− may contribute to Pt-mediated TMB oxidation, thereby supporting colorimetric signal amplification. The platform achieved a colorimetric EV detection limit of 427 particles/μL after coupling with the CHA-CRISPR system, representing a 6.9-fold improvement over the UiO@Pt@Apt system alone, and enabled a miRNA-21 detection limit of 87.0 fM. In plasma samples from 25 glioma patients and 20 healthy donors, the combined readout achieved an AUC of 0.968, showing numerically better discriminatory performance than either single readout, although the improvement was not statistically significant by DeLong analysis. This work provides a promising strategy for EV-based glioma liquid biopsy and offers a potentially adaptable framework for cascade-coupled dual-signal biosensing.
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2025
·
IF 7.7
Zhengfang Liang,Shuwei Xiao,Junhong Li,Guoli Zuo,Jihua Wei,Yuanxun Gong,Qianli Tang,Kai Zhang,Xianjiu Liao
Apurinic/apyrimidinic endonuclease 1 (APE1), a key enzyme in the base excision repair pathway, is frequently overexpressed in multiple cancers and has emerged as an important biomarker for early diagnosis and therapeutic response evaluation. In this work, a highly sensitive electrochemiluminescence (ECL) biosensor was developed for APE1 detection by integrating a PAM-regulated CRISPR-Cas12a activation mechanism with a multifunctional nanocomposite-modified electrode. A Ti₃C₂/AuNPs/CsPbBr₃ ternary nanocomposite was fabricated through ultrasonic assembly to provide enhanced conductivity, water stability, and probe-loading capacity. Meanwhile, a DNA duplex containing an AP site within the PAM region was engineered to regulate Cas12a activation through APE1-mediated cleavage and polymerase-assisted repair, thereby minimizing background interference and enabling highly specific signal generation. Ferrocene-labeled DNA tetrahedrons (TDNs) were further introduced as a spatially ordered probe scaffold to improve hybridization efficiency and facilitate Cas12a trans-cleavage–mediated signal release. This work presents three key innovations: (1) Construction of a Ti₃C₂/AuNPs/CsPbBr₃ nanocomposite that simultaneously enhances ECL stability, conductivity, and electron-transfer efficiency, addressing the inherent water-instability of perovskite emitters. (2) Coupling of a PAM-regulated APE1–KF repair mechanism with Cas12a activation to achieve highly specific AP-site recognition and selective signal generation. (3) Introduction of a TDN-based probe architecture that provides rigid spatial orientation and high-density probe loading, markedly improving hybridization efficiency and Cas12a trans-cleavage performance. Benefiting from these synergistic innovations, the biosensor achieves a detection limit of 6.28 × 10⁻⁸ U μL⁻¹ and a wide linear range (1 × 10⁻⁷ to 1 × 10⁻ 1 U μL⁻¹). The platform also demonstrates excellent specificity, reproducibility, and reliable performance in diluted serum, highlighting its promise for clinical and biochemical applications.
研究领域:
生物传感器
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纳米技术
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癌症诊断
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电分析化学
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分子生物学
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CRISPR技术
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高影响因子
Journal of Future Foods
·
2026
·
IF 7.2
Xiaobo Zhang,Zekun Long,Wei Wang,Yafeng Jiang,Qiuyue Zheng,Jijuan Cao
Aflatoxin B1 (AFB1), the most toxic mycotoxin among aflatoxin family, which presents a risk to food safety and human health at a trace level. Herein, the self-enhanced electrochemiluminescence (ECL) emitters of Au nanoclusters-anchored g-C 3 N 4 nanosheet (Au NCs@g-C 3 N 4 ) were synthesized, in which the Au NCs as a co-reaction accelerator enhanced the ECL stability and intensity. Integrating with CRISPR/Cas12a system, the Au NCs@g-C 3 N 4 behaved as sensing substrate to construct a “off-on” switch-type ECL sensor for AFB1 detection based on ECL resonance energy transfer (ECL-RET). The presence of targets AFB1 can specifically bind its aptamer to trigger the release of the activator. Using dual aptamer-regulated activation of CRISPR-Cas12a, AFB1 can be quantified via the ECL sensor, displaying a wide detection range from 0.2 pg/mL-100 ng/mL and a detection limit of 0.08 pg/mL. Furthermore, good practicability was verified in real samples assay, providing an efficient tool for the early warning of AFB1 pollution.
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高影响因子
MICROCHEMICAL JOURNAL
·
2026
·
IF 5.2
Chunlei Yu,Keyun Ren,Xuezhen Xu,Jinmiao Ma,Falan Li,Yanyan Zhang,Qingqing Yang
Most applications of CRISPR-Cas12a technology for mycotoxin detection rely on aptamers as recognition elements. However, although antibodies exhibit high affinity and specificity, they require converting immune signals into nucleic acid signals and cannot directly activate the CRISPR-Cas12a system, thereby limiting their utility for highly sensitive small-molecule target detection. In this work, we conjugated CRISPR-Cas12a activator DNA (aDNA) to antibodies via strain-promoted azide-alkyne cycloaddition (SPAAC) to achieve dual functionality: transducing small-molecule target signals into nucleic acid signals and partially immobilizing the Cas12a protein. Magnetic spherical nucleic acids (MSNAs) served as trans-cleavage substrates for the CRISPR-Cas12a system. Terminal deoxynucleotidyl transferase (TdT) was used to extend polyA sequences by adding dATP to the 3’-OH ends of numerous cleaved DNA fragments. This process, combined with high-concentration NaCl-induced aggregation and color change of gold nanoparticles (AuNPs), established a cascade signal amplification system. We developed a novel ultrasensitive immunosensor, termed Cas12a-TdT Cascade Amplification for Antigen-Toxin Immunoassay (CTCA-ATI), for mycotoxin quantification. The limits of detection for aflatoxin B1 and deoxynivalenol (DON) were 2.63 × 10 −5 ng/mL and 6.23 × 10 −4 ng/mL, respectively, with linear ranges of 1 × 10 −4 –10 ng/mL for aflatoxin B1 and 1 × 10 −3 –10 ng/mL for DON. This study provides new insights into the development of cascade-amplified biosensors using CRISPR-Cas12a and demonstrates substantial potential for integrating antibodies with CRISPR-Cas technology to detect other critical targets.
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高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2023
·
IF 6.1
Zhong Qiang Jia,Su Gui Zhang,Ying Wang,Jun Heng Pan,Fei Fan Liu,En Ling Zhan,Eman Atef Fouad,Ya Li Fu,Qi Rui Pan,Chun Qing Zhao
In insect ionotropic γ-aminobutyric acid receptor (iGABAR) subunits, only resistance to dieldrin (RDL) can be individually and functionally expressed in vitro. In lepidopteran, two to three RDL subtypes are identified; however, their physiological roles have not been distinguished in vivo. In this study, SlRdl1 and SlRdl2 of S. litura were individually knocked out using CRISPR/Cas9, respectively. The mortality and larval and pupal duration of KOSlRdl1 and KOSlRdl2 were increased. The flight time and distance were increased by 43.30%–80.66% and 58.96%–198.22%, respectively, in KOSlRdl1. The GABA-induced current was significantly decreased by 53.57%–74.28% and 46.91%–63.34% in the ventral nerve cord, and the GABA titer was significantly reduced by 17.65%–28.05% and 19.85%–42.46% in KOSlRdl1 and KOSlRdl2, respectively. In conclusion, SlRdl1 and SlRdl2 are necessary for the transmission of GABA-induced neural signals; however, only SlRdl1 could regulate the flight capability of S. litura. Our results provided a new avenue to study lepidopteran iGABARs.
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高影响因子
BIOSENSORS & BIOELECTRONICS
·
2021
·
IF 10.618
Kai Zhang,Zhenqiang Fan,Bo Yao,Yuedi Ding,Jing Zhao,Minhao Xie,Jianbin Pan
Sialic acid-binding immunoglobulin (Ig)-like lectins (Siglecs) is a type I transmembrane receptor on the cell surface. Siglec-5, as one of the Siglecs family, play an important role as an inhibitory receptor for leukocytes in the human body. The development of novel siglec-5 assays can help to study the pathogenesis of related diseases as well as to develop novel therapeutic drugs. We use catalytic hairpin assembly (CHA) amplification strategy combined with CRISPR-Cas12a′s side-cutting feature to build a 2D ultra-thin Ti 3 C 2 T x (MXene) based electrochemiluminescence (ECL) biosensor for the detection of Siglec-5. By using this ECL biosensor, the cleavage of CRISPR-Cas12a is reasonably combined with CHA-mediated isothermal amplification, thereby realizing the sensitive amplification assay Siglec-5 with 20.22 fM sensitivity. By introducing pairs of sites that are not in the same double-stranded DNA into the DNA duplex, the hybridization sequence of CRISPR-Cas12a complements the targeting mechanism to enhance indirect Siglec-5 amplification assay. Also, the double-strand DNA (dsDNA) design based on CRISPR-Cas12a amplification allows the same CRISPR RNA (crRNA, also known as guide RNA (gRNA)) to detect the output of DNA duplexes from different intermediate DNAs, which provides a common way for biomarker detection based on the conversion of protein analytes to intermediate DNA strategy. This work extends the application scope of CRISPR-Cas12a to the construction of ECL biosensors, evaluates the role of lectins, which can be used for the biochemical research and clinical diagnosis of protein markers. This is the first investigative work exploring the Trans-Cleavage activity of CRISPR-Cas12a for Mxene-based ECL biosensor establishment to the best of our knowledge.
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高影响因子
Small
·
2020
·Vol. 16
·期 52
·2004879
·
IF 11.459
Liang Zhao,Yingli Luo,Qiaoyi Huang,Ziyang Cao,Xianzhu Yang
Blocking immune checkpoint pathways with an antibody or small interfering RNA (siRNA) has become a promising method to reactivate antitumor responses for cancer treatment. However, both blockade strategies achieve only temporary inhibition of these immune checkpoints. Herein, a photoswitched CRISPR/Cas9 system for genomic disruption of the PD-L1 gene is developed to achieve permanent blockade of the PD-1/PD-L1 pathway; this system is constructed by using a photoactivated self-degradable polyethyleneimine derivative and the plasmid pX330/sgPD-L1 (expression of the Cas9 protein and single-guide RNA targeting PD-L1). Under light irradiation, this photoswitched CRISPR/Cas9 system efficiently genetically disrupts the PD-L1 gene in not only bulk cancer cells but also cancer stem-like cells. As a result, the photoswitched CRISPR/Cas9 system significantly increases the infiltration of CD8 + T cells into tumor tissue, leading to effective activation of a T cell-mediated antitumor response against cancer cells and cancer stem-like cells. This study provides an alternative strategy to block the PD-1/PD-L1 pathway for efficacious immune checkpoint therapy.
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2021
·
IF 7.46
Kai-Ren Zhao,Li Wang,Peng-Fei Liu,Xiao-Min Hang,Hui-Yi Wang,Shu-Ying Ye,Zhi-Jun Liu,Guo-Xi Liang
Herein, we report a “on-off” signal-switchable electrochemiluminescence (ECL) biosensor for simple, sensitive and quantitative detection of two kinds of virus genes, human immunodeficiency virus (HIV) and human papilloma virus (HPV-16) DNAs, by the integration of spherical nucleic acid (SNA) with CRISPR/Cas12a. The concept behind this approach is “on” signal based on a sandwich type assay after ECL luminophores of carbon dots (CDs) labeled SNA incubated with the biosensor in the presence of HIV DNA; and “off” signal upon the biosensor incubation with assistant DNA (HIV DNA sequence), CDs-labeled SNA and Cas12a/crRNA/HPV-16 DNA ternary complex relying on the collaterally cleavage of Cas12a. The cross-interference between HIV and HPV-16 DNAs can be avoided by adding Cas12a effector to the system or not. Using this strategy, HIV and HPV-16 DNAs are successfully quantitatively detection with detection limits of 30 fM and 0.32 pM (S/N = 3), respectively. Importantly, this system does not rely on multiplex signal reporting molecules, thus simplifying the detection system and avoiding the likelihood of cross interferences presented in the assay. Furthermore, multiplex detection of HIV/HPV-16 DNAs in human serum samples are successfully achieved within 2 h without additional nucleic acid amplification step. As demonstrated, the proposed biosensor has the potential for multiplex detection in clinical diagnosis.
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