Rewiring endogenous genes in CAR T cells for tumour-restricted payload delivery PMID: 40604285

ADC Targets in Oncology

2026-04-10

Rewiring endogenous genes in CAR T cells for tumour-restricted payload delivery

Nature. 2025 Aug;644(8075):241-251. doi: 10.1038/s41586-025-09212-7.

Amanda X Y Chen ^# ¹ ² , Kah Min Yap ^# ³ ⁴ , Joelle S Kim ⁵ ⁶ , Kevin Sek ⁵ ⁶ , Yu-Kuan Huang ⁵ ⁶ , Phoebe A Dunbar ⁵ ⁶ , Volker Wiebking ⁷ , Jesse D Armitage ⁸ ⁹ , Isabelle Munoz ⁵ ⁶ , Kirsten L Todd ⁵ ⁶ , Emily B Derrick ⁵ ⁶ , Dat Nguyen ⁵ ⁶ , Junming Tong ⁵ ⁶ , Cheok Weng Chan ⁵ ⁶ , Thang X Hoang ⁵ ⁶ , Katherine M Audsley ⁵ ⁶ , Marit J van Elsas ⁵ ⁶ , Jim Middelburg ⁵ ⁶ , Joel N Lee ⁵ ⁶ , Maria N de Menezes ⁵ ⁶ , Thomas J Cole ⁵ ⁶ , Jasmine Li ⁵ ⁶ , Christina Scheffler ⁵ ⁶ , Andrew M Scott ¹⁰ ¹¹ , Laura K Mackay ¹² , Jason Waithman ⁸ ⁹ , Jane Oliaro ⁵ ⁶ , Simon J Harrison ⁶ ¹³ , Ian A Parish ⁵ ⁶ , Junyun Lai ⁵ ⁶ , Matthew H Porteus ⁷ , Imran G House ⁵ ⁶ , Phillip K Darcy ¹⁴ ¹⁵ ¹⁶ , Paul A Beavis ¹⁷ ¹⁸

Affiliations

1. Cancer Immunology Program, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia. [email protected].
2. Sir Peter MacCallum Department of Oncology, The University of Melbourne, Parkville, Victoria, Australia. [email protected].
3. Cancer Immunology Program, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia. [email protected].
4. Sir Peter MacCallum Department of Oncology, The University of Melbourne, Parkville, Victoria, Australia. [email protected].
5. Cancer Immunology Program, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.
6. Sir Peter MacCallum Department of Oncology, The University of Melbourne, Parkville, Victoria, Australia.
7. Department of Pediatrics, Stanford University, Stanford, CA, USA.
8. School of Biomedical Sciences, University of Western Australia, Perth, Western Australia, Australia.
9. Leukaemia Translational Research Laboratory, Telethon Kids Cancer Centre, Telethon Kids Institute, Perth, Western Australia, Australia.
10. Olivia Newton-John Cancer Research Institute and School of Cancer Medicine, La Trobe University, Melbourne, Victoria, Australia.
11. Faculty of Medicine, The University of Melbourne, Parkville, Victoria, Australia.
12. Department of Microbiology and Immunology at the Peter Doherty Institute for Infection and Immunity, The University of Melbourne, Parkville, Victoria, Australia.
13. Clinical Haematology and Centre of Excellence for Cellular Immunotherapies, Peter MacCallum Cancer Centre and Royal Melbourne Hospital, Melbourne, Victoria, Australia.
14. Cancer Immunology Program, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia. [email protected].
15. Sir Peter MacCallum Department of Oncology, The University of Melbourne, Parkville, Victoria, Australia. [email protected].
16. Department of Immunology, Monash University, Clayton, Australia. [email protected].
17. Cancer Immunology Program, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia. [email protected].
18. Sir Peter MacCallum Department of Oncology, The University of Melbourne, Parkville, Victoria, Australia. [email protected].
# Contributed equally.

Abstract

The efficacy of chimeric antigen receptor (CAR) T cell therapy in solid tumours is limited by immunosuppression and antigen heterogeneity^1-3. To overcome these barriers, 'armoured' CAR T cells, which secrete proinflammatory cytokines, have been developed⁴. However, their clinical application has been limited because of toxicity related to peripheral expression of the armouring transgene⁵. Here, we have developed a CRISPR knock-in strategy that leverages the regulatory mechanisms of endogenous genes to drive transgene expression in a tumour-localized manner. By screening endogenous genes with tumour-restricted expression, we have identified the NR4A2 and RGS16 promoters as promising candidates to support the delivery of cytokines such as IL-12 and IL-2 directly to the tumour site, leading to enhanced antitumour efficacy and long-term survival of mice in both syngeneic and xenogeneic models. This effect was concomitant with improved CAR T cell polyfunctionality, activation of endogenous antitumour immunity and a favourable safety profile, and was applicable in CAR T cells from patients.