Engineered amphiphilic peptides enable delivery of proteins and CRISPR-associated nucleases to airway epithelia

Krishnamurthy, Sateesh; Wohlford‐Lenane, Christine L.; Kandimalla, Suhas; Sartre, Gilles; Meyerholz, David K.; Théberge, Vanessa; Hallée, Stéphanie; Duperré, Anne-Marie; Del’Guidice, Thomas; Lepetit-Stoffaes, Jean-Pascal; Barbeau, Xavier; Guay, David R.P.; McCray, Paul B.

doi:10.1038/s41467-019-12922-y

Cited by 101 publications

(102 citation statements)

References 76 publications

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“…All formalin-fixed paraffin-embedded tissues were sectioned (~4 µm) and hydrated through a series of xylene and alcohol baths to water. Immunohistochemical techniques were used for the following markers: angiotensin-converting enzyme 2 (ACE2) [30] , surfactant protein C (SP-C) [31] , and mucin 5B (MUC5B) [32] . For more specifics about the reagents please see Supplemental Table 3 .…”

Section: Methodsmentioning

confidence: 99%

Heterogeneous expression of the SARS-Coronavirus-2 receptor ACE2 in the human respiratory tract

Ortiz¹,

Thurman²,

Pezzulo³

et al. 2020

EBioMedicine

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170

146

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Background Zoonotically transmitted coronaviruses are responsible for three disease outbreaks since 2002, including the current COVID-19 pandemic, caused by SARS-CoV-2. Its efficient transmission and range of disease severity raise questions regarding the contributions of virus-receptor interactions. ACE2 is a host ectopeptidase and the receptor for SARS-CoV-2. Numerous reports describe ACE2 mRNA abundance and tissue distribution; however, mRNA abundance is not always representative of protein levels. Currently, there is limited data evaluating ACE2 protein and its correlation with other SARS-CoV-2 susceptibility factors. Materials and methods We systematically examined the human upper and lower respiratory tract using single-cell RNA sequencing and immunohistochemistry to determine receptor expression and evaluated its association with risk factors for severe COVID-19. Findings Our results reveal that ACE2 protein is highest within regions of the sinonasal cavity and pulmonary alveoli, sites of presumptive viral transmission and severe disease development, respectively. In the lung parenchyma, ACE2 protein was found on the apical surface of a small subset of alveolar type II cells and colocalized with TMPRSS2, a cofactor for SARS-CoV2 entry. ACE2 protein was not increased by pulmonary risk factors for severe COVID-19. Additionally, ACE2 protein was not reduced in children, a demographic with a lower incidence of severe COVID-19. Interpretation These results offer new insights into ACE2 protein localization in the human respiratory tract and its relationship with susceptibility factors to COVID-19.

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Section: Methodsmentioning

confidence: 99%

Heterogeneous expression of the SARS-Coronavirus-2 receptor ACE2 in the human respiratory tract

Ortiz¹,

Thurman²,

Pezzulo³

et al. 2020

EBioMedicine

Self Cite

170

146

View full text Add to dashboard Cite

show abstract

“…This was achieved through a peptide combining a cationic CPP for complexing and a targeting integrin-specific arginylglycylaspartic acid (iRGD) peptide that can interact with αvβ3 integrins found on cancer cells [ 110 ]. The iRGD peptide is a tumor homing motif routinely used for cancer targeted delivery of drugs, biologics and nanoparticles [ 126 ]. Using amphiphilic peptides, another study established the delivery of CRISPR/Cas9 and Cas12 RNPs to airway epithelial cells in mice [ 112 ].…”

Section: The Evolution Of Peptide Delivery Systems For Crispr-cas mentioning

confidence: 99%

“…Importantly, cell-surface receptors have been shown to engage in sequence-specific uptake of CPPs. For example, neuronal nicotinic acetylcholine receptors which are known to bind rabies virus glyocoprotein 29 (RVG29) peptide [ 131 , 153 ], cancer-expressed integrins that bind iRGD peptide [ 110 , 126 ], and prohibitins on adipocytes that bind ATS [ 96 , 154 ].…”

Section: Mechanism Of Cpp-uptake and Releasementioning

confidence: 99%

Tissue-Specific Delivery of CRISPR Therapeutics: Strategies and Mechanisms of Non-Viral Vectors

Shalaby

Aouida

El‐Agnaf

2020

IJMS

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The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) genome editing system has been the focus of intense research in the last decade due to its superior ability to desirably target and edit DNA sequences. The applicability of the CRISPR-Cas system to in vivo genome editing has acquired substantial credit for a future in vivo gene-based therapeutic. Challenges such as targeting the wrong tissue, undesirable genetic mutations, or immunogenic responses, need to be tackled before CRISPR-Cas systems can be translated for clinical use. Hence, there is an evident gap in the field for a strategy to enhance the specificity of delivery of CRISPR-Cas gene editing systems for in vivo applications. Current approaches using viral vectors do not address these main challenges and, therefore, strategies to develop non-viral delivery systems are being explored. Peptide-based systems represent an attractive approach to developing gene-based therapeutics due to their specificity of targeting, scale-up potential, lack of an immunogenic response and resistance to proteolysis. In this review, we discuss the most recent efforts towards novel non-viral delivery systems, focusing on strategies and mechanisms of peptide-based delivery systems, that can specifically deliver CRISPR components to different cell types for therapeutic and research purposes.

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“…Delivery of coding DNA sequences can be performed by both viral (including adeno-associated virus and adenovirus) and non-viral methods; packaging and delivery of mRNA/RNA and RNPs are usually performed by non-viral methods [ 16 ]. Nanotechnological methods mostly rely on the use of liposomes and cationic lipids [ 44 , 45 , 46 ], amphiphilic peptides [ 47 ], DNA nanoclews [ 48 , 49 ], gold nanoparticles [ 50 , 51 , 52 ], and graphene-based nanosheets [ 53 ].…”

Section: Introductionmentioning

confidence: 99%

Gene Editing by Extracellular Vesicles

Kostyushev

Kostyusheva

Brezgin

et al. 2020

IJMS

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CRISPR/Cas technologies have advanced dramatically in recent years. Many different systems with new properties have been characterized and a plethora of hybrid CRISPR/Cas systems able to modify the epigenome, regulate transcription, and correct mutations in DNA and RNA have been devised. However, practical application of CRISPR/Cas systems is severely limited by the lack of effective delivery tools. In this review, recent advances in developing vehicles for the delivery of CRISPR/Cas in the form of ribonucleoprotein complexes are outlined. Most importantly, we emphasize the use of extracellular vesicles (EVs) for CRISPR/Cas delivery and describe their unique properties: biocompatibility, safety, capacity for rational design, and ability to cross biological barriers. Available molecular tools that enable loading of desired protein and/or RNA cargo into the vesicles in a controllable manner and shape the surface of EVs for targeted delivery into specific tissues (e.g., using targeting ligands, peptides, or nanobodies) are discussed. Opportunities for both endogenous (intracellular production of CRISPR/Cas) and exogenous (post-production) loading of EVs are presented.

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Engineered amphiphilic peptides enable delivery of proteins and CRISPR-associated nucleases to airway epithelia

Cited by 101 publications

References 76 publications

Heterogeneous expression of the SARS-Coronavirus-2 receptor ACE2 in the human respiratory tract

Heterogeneous expression of the SARS-Coronavirus-2 receptor ACE2 in the human respiratory tract

Tissue-Specific Delivery of CRISPR Therapeutics: Strategies and Mechanisms of Non-Viral Vectors

Gene Editing by Extracellular Vesicles

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