Author Correction: Immune-orthogonal orthologues of AAV capsids and of Cas9 circumvent the immune response to the administration of gene therapy

Moreno, Ana M.; Palmer, Nathan; Alemán, Fernando; Chen, Genghao; Pla, Andrew; Jiang, Ning; Chew, Wei Leong; Law, Mansun; Mali, Prashant

doi:10.1038/s41551-019-0456-6

Cited by 7 publications

(5 citation statements)

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“…We conjectured that the prolonged expression via icRNAs could facilitate genome and especially epigenome targeting. However, this same feature of persistence could also aggravate immune responses in therapeutic settings as CRISPR systems are derived from prokaryotes (32)(33)(34). Thus, to enable compatibility between persistence of expression and immunogenicity, we sought first to develop a methodology to screen progressively deimmunized SpCas9 proteins by combinatorially mutating particularly immunogenic epitopes (35).…”

Section: Resultsmentioning

confidence: 99%

Extensive in vitro and in vivo protein translation via in situ circularized RNAs

Kumar

Palmer

Miyasaki

et al. 2022

Preprint

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RNAs are a powerful therapeutic class. However their inherent transience impacts their activity both as an interacting moiety as well as a template. Circularization of RNA has been demonstrated as a means to improve persistence, however simple and scalable approaches to achieve this are lacking. Utilizing autocatalytic RNA circularization, here we engineer in situ circularized RNAs (icRNAs). This approach enables icRNA delivery as simple linear RNA that is circularized upon delivery into the cell, thus making them compatible with routine synthesis, purification, and delivery formulations. We confirmed extensive protein translation from icRNAs both in vitro and in vivo and explored their utility in three contexts: first, we delivered the SARS-CoV-2 Omicron spike protein in vivo as icRNAs and showed corresponding induction of humoral immune responses; second, we demonstrated robust genome targeting via zinc finger nucleases delivered as icRNAs; and third, to enable compatibility between persistence of expression and immunogenicity, we developed a novel long range multiplexed (LORAX) protein engineering methodology to screen progressively deimmunized Cas9 proteins, and demonstrated efficient genome and epigenome targeting via their delivery as icRNAs. We anticipate this highly simple and scalable icRNA methodology could have broad utility in basic science and therapeutic applications.

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Section: Resultsmentioning

confidence: 99%

Extensive in vitro and in vivo protein translation via in situ circularized RNAs

Kumar

Palmer

Miyasaki

et al. 2022

Preprint

Self Cite

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“…Importantly, we show that strategies commonly used to minimize cellular immune responses (e.g., tissue-specific promoter, prednisolone immune suppression) are insufficient to bypass Cas9 immunity. Comprehensive approaches, including but not limited to the use of an immune-attenuated/immune-privileged Cas9 gene expression cassette, nonviral vector-mediated transient Cas9 gene expression, immune tolerance induction, more potent immune suppression, and pre-screening for Cas9 immunity, might be necessary in order to translate AAV CRISPR therapy to DMD patients 10 , 13 , 31 .…”

Section: Resultsmentioning

confidence: 99%

Cas9-specific immune responses compromise local and systemic AAV CRISPR therapy in multiple dystrophic canine models

et al. 2021

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Adeno-associated virus (AAV)-mediated CRISPR-Cas9 editing holds promise to treat many diseases. The immune response to bacterial-derived Cas9 has been speculated as a hurdle for AAV-CRISPR therapy. However, immunological consequences of AAV-mediated Cas9 expression have thus far not been thoroughly investigated in large mammals. We evaluate Cas9-specific immune responses in canine models of Duchenne muscular dystrophy (DMD) following intramuscular and intravenous AAV-CRISPR therapy. Treatment results initially in robust dystrophin restoration in affected dogs but also induces muscle inflammation, and Cas9-specific humoral and cytotoxic T-lymphocyte (CTL) responses that are not prevented by the muscle-specific promoter and transient prednisolone immune suppression. In normal dogs, AAV-mediated Cas9 expression induces similar, though milder, immune responses. In contrast, other therapeutic (micro-dystrophin and SERCA2a) and reporter (alkaline phosphatase, AP) vectors result in persistent expression without inducing muscle inflammation. Our results suggest Cas9 immunity may represent a critical barrier for AAV-CRISPR therapy in large mammals.

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“…To address these concerns, researchers have explored different Cas9 orthologs and AAV serotypes based on sequence similarities and predicted binding strength to major histocompatibility complex (MHC) class I and class II molecules. While no ortholog or serotype has been found to completely evade immune recognition, Moreno et al identified three Cas9 orthologs, namely, SpCas9, SaCas9, and Campylobacter jejuni Cas9 (CjCas9), that showed robust editing efficiency and tolerated repeated administration with reduced immunogenic toxicity in mice immunized against AAV and Cas9 ( Moreno et al, 2019 ). However, pre-existing immunity against SpCas9 and SaCas9 in humans leaves CjCas9 as the only current option for repeated gene therapy in this cohort of patients.…”

Section: Tackling Limitations: Recent Advancements In Crispr/cas Systemmentioning

confidence: 99%

Revolutionizing in vivo therapy with CRISPR/Cas genome editing: breakthroughs, opportunities and challenges

Macarrón Palacios,

Korus,

Wilkens

et al. 2024

Front. Genome Ed.

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Genome editing using the CRISPR/Cas system has revolutionized the field of genetic engineering, offering unprecedented opportunities for therapeutic applications in vivo. Despite the numerous ongoing clinical trials focusing on ex vivo genome editing, recent studies emphasize the therapeutic promise of in vivo gene editing using CRISPR/Cas technology. However, it is worth noting that the complete attainment of the inherent capabilities of in vivo therapy in humans is yet to be accomplished. Before the full realization of in vivo therapeutic potential, it is crucial to achieve enhanced specificity in selectively targeting defective cells while minimizing harm to healthy cells. This review examines emerging studies, focusing on CRISPR/Cas-based pre-clinical and clinical trials for innovative therapeutic approaches for a wide range of diseases. Furthermore, we emphasize targeting cancer-specific sequences target in genes associated with tumors, shedding light on the diverse strategies employed in cancer treatment. We highlight the various challenges associated with in vivo CRISPR/Cas-based cancer therapy and explore their prospective clinical translatability and the strategies employed to overcome these obstacles.

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Author Correction: Immune-orthogonal orthologues of AAV capsids and of Cas9 circumvent the immune response to the administration of gene therapy

Cited by 7 publications

References 0 publications

Extensive in vitro and in vivo protein translation via in situ circularized RNAs

Extensive in vitro and in vivo protein translation via in situ circularized RNAs

Cas9-specific immune responses compromise local and systemic AAV CRISPR therapy in multiple dystrophic canine models

Revolutionizing in vivo therapy with CRISPR/Cas genome editing: breakthroughs, opportunities and challenges

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