RIG-I-dependent tumor-intrinsic type I interferon signaling restricts growth in breast cancer 3D culture

Liu, K.; Mangiante, L.; Levin-Konigsberg, R.; Sotomayor-Vivas, C.; Yang, W.; Spees, K.; Ma, Z.; Caswell-Jin, J. L.; Curtis, C. N.; Bassik, M. C.

doi:10.1101/2025.08.10.669545

bioRxiv · 10.1101/2025.08.10.669545

RIG-I-dependent tumor-intrinsic type I interferon signaling restricts growth in breast cancer 3D culture

Liu, K.·Mangiante, L.·Levin-Konigsberg, R.·Sotomayor-Vivas, C.·Yang, W.·Spees, K.·Ma, Z.·Caswell-Jin, J. L.·Curtis, C. N.·Bassik, M. C.

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Abstract

Discovery of key growth drivers that can be targeted for therapy is a central goal in cancer research. While high-throughput CRISPR screens have revolutionized our ability to identify gene dependencies in cancer, most large-scale screens are conducted in two-dimensional (2D) culture systems that fail to recapitulate tumor organization and behavior. To uncover architecture-dependent vulnerabilities in breast cancer, we performed parallel CRISPR interference (CRISPRi) screens in 2D and three-dimensional (3D) cultures of MCF7 cells, an estrogen receptor-positive (ER+) breast cancer model representative of a high risk of relapse, luminal subtype. Knockdown of IFNAR2 and TYK2 conferred a growth advantage in 3D cultures, implicating type I interferon signaling as a tumor-intrinsic suppressor of proliferation in 3D spheroids. Transcriptomic and functional analyses demonstrated that type I IFN signaling is endogenously activated in 3D spheroids via RIG-I-mediated sensing of cytosolic double-stranded RNA, leading to TBK1 activation and induction of interferon-stimulated genes (ISGs). This tumor-intrinsic IFN response slowed proliferation in 3D culture, independent of exogenous stimuli or the presence of immune cells. Analysis of bulk, single-cell, and spatial transcriptomic datasets from breast cancer patients revealed that a subset of tumors exhibit elevated IFN signaling in cancer cells, including in immune-depleted tumor cores, consistent with a tumor-intrinsic IFN signature. Our findings uncover an IFN-mediated growth-suppressive program shaped by 3D tumor architecture, and contribute towards a better understanding of the role of tumor-intrinsic IFN activity.

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Liu, K., Mangiante, L., Levin-Konigsberg, R., Sotomayor-Vivas, C., Yang, W., Spees, K., Ma, Z., Caswell-Jin, J. L., Curtis, C. N., Bassik, M. C.. 2025-08-12. RIG-I-dependent tumor-intrinsic type I interferon signaling restricts growth in breast cancer 3D culture. https://doi.org/10.1101/2025.08.10.669545

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