AAV vectors with enhancer-controlled ITR promoters

Yanjiang Zheng; Luzi Yang; Kaiyu Zhou; Qirun Wang; Tiange Li; Jiahao Wu; Jiangwei Qin; Fei Gao; Pingzhu Zhou; Tian Lu; Yangpo Cao; Yimin Hua; Yuxuan Guo; Yifei Li

doi:10.64898/2025.12.03.692098

BioengineeringbioRxivDec 5, 2025

Preprint

AAV vectors with enhancer-controlled ITR promoters

Yanjiang Zheng, Luzi Yang iD, Kaiyu Zhou iD, Qirun Wang iD, Tiange Li iD, Jiahao Wu iD, Jiangwei Qin, Fei Gao iD, Pingzhu Zhou iD, Tian Lu iD, Yangpo Cao iD, Yimin Hua iD, Yuxuan Guo iD, Yifei Li iD

Abstract

Adeno-associated virus (AAV) is limited by its packaging capacity and the unwanted promoter activity of inverted terminal repeats (ITRs). Here we utilized mini-enhancers to regulate ITR promoters and drive AAV cargo expression in the absence of the canonical promoters, which not only solved the ITR issue but also released more payload for the cargo. As an example, this new design successfully enabled robust, tissue-specific SpCas9 gene editing via a single AAV, which otherwise requires a dual-AAV system due to the large size of SpCas9.

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