Chimeric antigen receptor (CAR)-engineered immune cell therapy represents an important advance in cancer treatments. However, the complex ex vivo cell manufacturing process and stringent patient selection criteria curtail its widespread use. In vivo CAR engineering is emerging as a promising off-the-shelf therapy, providing advantages such as streamlined production, elimination of patient-specific manufacturing, reduced costs and simplified logistics. A large set of preclinical findings has inspired further investigation into treatments for hard-to-treat diseases such as solid tumours and has facilitated the development of advanced products to enhance in vivo CAR engineering efficacy, the persistence of the cellular therapeutic and safety. In this Review, we summarize current in vivo CAR engineering strategies, including nanoparticle-based and viral delivery systems as well as bioinstructive implantable scaffolds, and discuss their advantages and disadvantages. Additionally, we provide a systematic comparison between in vivo and conventional ex vivo CAR engineering methods and address the challenges and future prospects of in vivo CAR engineering.