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Rational engineering of minimally immunogenic nucleases for gene therapy

ABSTRACT: Genome editing using CRISPR-Cas systems is a promising avenue for the treatment of genetic diseases. However, cellular and humoral immunogenicity of genome editing tools, which originate from bacteria, complicates their clinical use. Here we report reduced immunogenicity (Red)(i)-variants of two clinically-relevant nucleases, SaCas9 and AsCas12a. Through MHC-associated peptide proteomics (MAPPs) analysis, we identified putative immunogenic epitopes on each nuclease. Then, we used computational modeling to rationally design these proteins to evade the immune response. SaCas9 and AsCas12a Redi variants were substantially less recognized by adaptive immune components, including reduced binding affinity to MHC molecules and attenuated generation of cytotoxic T cell responses, while maintaining wild-type levels of activity and specificity. In vivo editing of PCSK9 with SaCas9.Redi.1 was comparable in efficiency to wild-type SaCas9, but significantly reduced undesired immune responses. This demonstrates the utility of this approach in engineering proteins to evade immune detection.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Mammary Epithelial Cell Line

DISEASE(S): Breast Cancer

SUBMITTER: Rumya Raghavan

LAB HEAD: Feng Zhang

PROVIDER: PXD054579 | Pride | 2024-11-07

REPOSITORIES: Pride

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			Action	DRS
	FR_BRIN1351_PX601_AB.msf	Msf
	FR_BRIN1351_PX601_AB_1.mgf	Mgf
	FR_BRIN1351_PX601_AB_1.raw	Raw
	FR_BRIN1351_PX601_AB_2.mgf	Mgf
	FR_BRIN1351_PX601_AB_2.raw	Raw

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