Excision of HIV-1 DNA by gene editing: a proof-of-concept in vivo study

Gene Therapy, Год журнала: 2016, Номер 23(8-9), С. 690 - 695

Опубликована: Май 19, 2016

Язык: Английский

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Engineered extracellular vesicles as versatile ribonucleoprotein delivery vehicles for efficient and safe CRISPR genome editing

Journal of Extracellular Vesicles, Год журнала: 2021, Номер 10(5)

Опубликована: Март 1, 2021

Abstract Transient delivery of CRISPR‐based genome editing effectors is important to reduce off‐target effects and immune responses. Recently extracellular vesicles (EVs) have been explored for Cas9 ribonucleoprotein (RNP) delivery. However, lack mechanisms enrich RNPs into EVs limited the efficiency as a RNP vehicle. Here we describe mechanism actively EVs. We used specific interaction between RNA aptamer aptamer‐binding protein (ABP) inserted com single guide (sgRNA), fused com‐binding ABP Com both termini tetraspan CD63 that abundant in exosomes. found Com/com enriched adenine base editor (ABE) EVs, via forming three‐component complex including CD63‐Com fusion protein, com‐modified sgRNA or ABE. The are efficient transiently expressed. system capable delivering targeting multiple loci multiplex editing. In addition, from different species can be together. EV‐delivered active vivo. data show interactions utilized improved safety.

Язык: Английский

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CRISPR/Cas gene therapy

Journal of Cellular Physiology, Год журнала: 2020, Номер 236(4), С. 2459 - 2481

Опубликована: Сен. 22, 2020

Abstract Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated enzyme (Cas) is a naturally occurring genome editing tool adopted from the prokaryotic adaptive immune defense system. Currently, CRISPR/Cas9‐based has been becoming one of most promising tools for treating human genetic diseases, including cardiovascular neuro‐disorders, and cancers. As quick modification CRISPR/Cas9 system, delivery gene therapy extensively studied in preclinic clinic treatments. CRISPR/Cas also robust to create animal models studying disorders, particularly diseases associated with point mutations. However, significant challenges remain before technology can be routinely employed different which include toxicity response treated cells component, highly throughput method, potential off‐target impact. The effect major concerns therapy, more research should focused on limiting this impact by designing high specific gRNAs using specificity Cas enzymes. Modifying method not only targets tissue/cell but potentially limits

Язык: Английский

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Advances in CRISPR therapeutics

Nature Reviews Nephrology, Год журнала: 2022, Номер 19(1), С. 9 - 22

Опубликована: Окт. 24, 2022

Язык: Английский

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Towards application of CRISPR-Cas12a in the design of modern viral DNA detection tools (Review)

Journal of Nanobiotechnology, Год журнала: 2022, Номер 20(1)

Опубликована: Янв. 21, 2022

Abstract Early detection of viral pathogens by DNA-sensors in clinical samples, contaminated foods, soil or water can dramatically improve outcomes and reduce the socioeconomic impact diseases such as COVID-19. Clustered regularly interspaced short palindromic repeat (CRISPR) its associated protein Cas12a (previously known CRISPR-Cpf1) technology is an innovative new-generation genomic engineering tool, also ‘genetic scissors’, that has demonstrated accuracy recently been effectively applied appropriate (E-CRISPR) DNA-sensor to detect nucleic acid interest. The CRISPR-Cas12a from Prevotella Francisella 1 are guided a CRISPR RNA (gRNA). unique simultaneous cis- trans- DNA cleavage after target sequence recognition at PAM site, sticky-end (5–7 bp) employment, ssDNA/dsDNA hybrid strategies manipulate attractive nature CRISPR–Cas12a reviewed. based on for rapid, robust, sensitive, inexpensive, selective virus without additional sample purification, amplification, fluorescent-agent- and/or quencher-labeling relevant becoming increasingly important industrial medical applications. In addition, system shows great potential field E-CRISPR-based bioassay research technologies. Therefore, we highlighting insights this direction. Graphical

Язык: Английский

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CRISPR editing of CCR5 and HIV-1 facilitates viral elimination in antiretroviral drug-suppressed virus-infected humanized mice

Proceedings of the National Academy of Sciences, Год журнала: 2023, Номер 120(19)

Опубликована: Май 1, 2023

Treatment of HIV-1 ADA -infected CD34+ NSG-humanized mice with long-acting ester prodrugs cabotegravir, lamivudine, and abacavir in combination native rilpivirine was followed by dual CRISPR-Cas9 C-C chemokine receptor type five (CCR5) proviral DNA gene editing. This led to sequential viral suppression, restoration absolute human CD4 + T cell numbers, then elimination replication-competent virus 58% infected mice. Dual CRISPR therapies enabled the excision integrated cells contained within live animals. Highly sensitive nucleic acid nested droplet digital PCR, RNAscope, outgrowth assays affirmed elimination. not detected blood, spleen, lung, kidney, liver, gut, bone marrow, brain virus-free Progeny from adoptively transferred CRISPR-treated neither nor recovered. Residual fragments were easily seen untreated viral-rebounded No evidence off-target toxicities recorded any treated Importantly, therapy demonstrated statistically significant improvements cure percentages compared single treatments. Taken together, these observations underscore a pivotal role combinatorial editing achieving infection.

Язык: Английский

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Exploring the potential of genome editing CRISPR-Cas9 technology

Gene, Год журнала: 2016, Номер 599, С. 1 - 18

Опубликована: Ноя. 9, 2016

Язык: Английский

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Brain Macrophages in Simian Immunodeficiency Virus-Infected, Antiretroviral-Suppressed Macaques: a Functional Latent Reservoir

mBio, Год журнала: 2017, Номер 8(4)

Опубликована: Авг. 16, 2017

A human immunodeficiency virus (HIV) infection cure requires an understanding of the cellular and anatomical sites harboring that contribute to viral rebound upon treatment interruption. Despite antiretroviral therapy (ART), HIV-associated neurocognitive disorders (HAND) are reported in HIV-infected individuals on ART. Biomarkers for macrophage activation neuronal damage cerebrospinal fluid (CSF) demonstrate continued effects HIV brain suggest central nervous system (CNS) may serve as a reservoir. Using simian (SIV)/macaque model encephalitis AIDS, we evaluated whether infected cells persist despite Eight SIV-infected pig-tailed macaques were virally suppressed with ART, plasma CSF viremia levels analyzed longitudinally. To assess persisted macrophages (BrMΦ) these macaques, used quantitative outgrowth assay (MΦ-QVOA), PCR, situ hybridization (ISH) measure frequency RNA DNA brain. Viral tissue was undetectable, although detected all animals. The MΦ-QVOA demonstrated majority animals contained latently BrMΦ. We also showed produced MΦ-QVOAs replication competent, suggesting BrMΦ capable reestablishing productive This report provides first confirmation presence replication-competent SIV ART-suppressed suggests highly debated issue latency macrophages, at least brain, has been addressed treated ART.IMPORTANCE Resting CD4+ T currently only fit definition latent However, recent evidence HIV/SIV-infected Markers observed suppressive ART regimens, CNS infection. controversy exists represent source In this study, reservoir brains ART-treated using our established quantitate genomes. Our results support idea existence other reservoirs addition resting underscore importance developing strategies eradicate HIV.

Язык: Английский

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Various Aspects of a Gene Editing System—CRISPR–Cas9

International Journal of Molecular Sciences, Год журнала: 2020, Номер 21(24), С. 9604 - 9604

Опубликована: Дек. 16, 2020

The discovery of clustered, regularly interspaced short palindromic repeats (CRISPR) and their cooperation with CRISPR-associated (Cas) genes is one the greatest advances century has marked application as a powerful genome engineering tool. CRISPR–Cas system was discovered part adaptive immune in bacteria archaea to defend from plasmids phages. CRISPR been found be an advanced alternative zinc-finger nucleases (ZFN) transcription activator-like effector (TALEN) for gene editing regulation, CRISPR–Cas9 protein remains same various targets just guide RNA sequence needs altered redirect site-specific cleavage. Due its high efficiency precision, Cas9 derived type II have applications many fields science. Although allows easy number benefits, we should not ignore important ethical biosafety issues. Moreover, any tool that great potential offers significant capabilities carries level risk being used non-legal purposes. In this review, present brief history mechanism system. We also describe on technology regulation editing; treatment cancer other diseases; limitations concerns use CRISPR–Cas9.

Язык: Английский

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A combinational CRISPR/Cas9 gene-editing approach can halt HIV replication and prevent viral escape

Scientific Reports, Год журнала: 2017, Номер 7(1)

Опубликована: Фев. 8, 2017

Abstract HIV presents one of the highest evolutionary rates ever detected and combination antiretroviral therapy is needed to overcome plasticity virus population control viral replication. Conventional treatments lack ability clear latent reservoir, which remains major obstacle towards a cure. Novel strategies, such as CRISPR/Cas9 gRNA-based genome-editing, can permanently disrupt genome. However, genome-editing may accelerate escape, questioning feasibility approach. Here, we demonstrate that targeting single loci, only partially inhibits replication facilitates rapid escape at target site. A combinatorial approach two strong gRNAs different regions genome completely abrogate prevent escape. Our data shows accelerating effect gene-editing on be provide future alternative for HIV-infection.

Язык: Английский

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