Lymphotoxin-driven cancer cell eradication by tumoricidal CD8+ TIL

  • bioRxiv. 2025 Nov 20:2025.11.19.689204. doi: 10.1101/2025.11.19.689204.
Hongyan Xie  1  2  3 ,  Aiping Jiang  1  2  3 ,  Aonkon Dey  1  2  3  4 ,  Joseph W Dean  5 ,  Jonathan J Perera  3 ,  Neal P Smith  1  2  3  4 ,  Alex C Y Chen  1  2  3  4 ,  Seth Anderson  1  3 ,  Angelina M Cicerchia  1 ,  Yi Sun  1 ,  William A Michaud  6 ,  Maria Florentin  1 ,  Jacy Fang  1  3 ,  Or-Yam Revach  1  2  3 ,  Tatyana Sharova  6 ,  Aleigha Lawless  6 ,  Katherine H Xu  1 ,  Yuhui Song  1 ,  Bidish Patel  1 ,  Jonathan D Stevens  7 ,  William J Lane  7 ,  Derin B Keskin  2  3  8  9  10  11 ,  Sonia Cohen  1  2  6 ,  Donald P Lawrence  2  12 ,  Ryan J Sullivan  2  12 ,  Keith T Flaherty  2  12 ,  Genevieve M Boland  1  2  6 ,  Linda T Nieman  1  2 ,  Moshe Sade-Feldman  1  2  3 ,  Nir Hacohen  1  2  3 ,  Debattama R Sen  1  2  3 ,  Catherine Wu  2  9 ,  Brian Gastman  5 ,  Rongsu Qi  5 ,  Hequn Yin  5 ,  Alexandra-Chloé Villani  1  2  3  4 ,  Robert T Manguso  1  2  3 ,  Russell W Jenkins  1  2  3  12  13
Affiliations
  • 1. Mass General Cancer Center, Krantz Family Center for Cancer Research, Department of Medicine, Massachusetts General Hospital, Boston, MA, USA.
  • 2. Harvard Medical School, Boston, MA, USA.
  • 3. Broad Institute of MIT and Harvard, Cambridge, MA, USA.
  • 4. Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, MA, USA.
  • 5. Iovance Biotherapeutics, Inc., San Carlos, CA, USA.
  • 6. Division of Gastrointestinal and Oncologic Surgery, Department of Surgery, Massachusetts General Hospital, Boston, MA, USA.
  • 7. Department of Pathology, Brigham and Women's Hospital, Boston, MA 02215, USA.
  • 8. Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA.
  • 9. Translational Immunogenomics Laboratory, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
  • 10. Department of Computer Science, Metropolitan College, Boston University, Boston, Massachusetts, USA.
  • 11. Section for Bioinformatics, Department of Health Technology, Technical University of Denmark, Lyngby, Denmark.
  • 12. Mass General Cancer Center, Department of Medicine, Massachusetts General Hospital, Boston, MA, USA.
  • 13. Present address: Hollings Cancer Center, Medical University of South Carolina, Charleston, South Carolina, USA.
PMID: 41332750 DOI: 10.1101/2025.11.19.689204
Abstract

Tumor-infiltrating lymphocyte (TIL) therapy is FDA-approved for patients with treatment-resistant advanced melanoma, but the TIL subpopulations critical for tumor eradication remains incompletely understood. Using patient-derived TIL-melanoma co-cultures, we identified and characterized a novel subset of CD8+ TIL, capable of class I HLA-independent cancer Cell Lysis. The Lymphotoxin β Receptor (LTβR) and interferon (IFN) sensing pathways were nominated as key determinants of TIL-mediated Cancer cell killing from a whole-genome, loss-of-function CRISPR screen. Validation studies confirmed that dual LTβR and IFN sensing is necessary and sufficient for cancer Cell Lysis, and that expanded CD8+ TIL express high lymphotoxin β (LTB) and upregulate lymphotoxin α (LTA) upon coculture with Cancer cells. Leveraging paired scRNA-seq and scTCR-seq data, we confirmed that enrichment of LTB + CD8 + T cells is associated with clinical response to TIL, and that LTB + CD8 + TIL are expanded from putative neoantigen-reactive, LTB lo CD8+ T cells in resected tumors.

Keywords
cell death; interferon; lymphotoxin; melanoma; tumor-infiltrating lymphocytes.
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