Tumor immune microenvironment reconstitution in patient-derived organoids enables therapy modeling for NSCLC

  • Cell Rep Methods. 2026 Jun 15;6(6):101339. doi: 10.1016/j.crmeth.2026.101339.
Enrique Podaza  1 ,  Jared Capuano  2 ,  Hui-Hsuan Kuo  2 ,  Majd Al Assaad  3 ,  Geoffrey Markowitz  4 ,  M Victoria Revuelta  5 ,  John Nguyen  2 ,  Adriana Irizarry  2 ,  Hiranmayi Ravichandran  2 ,  Sarah Ackermann  2 ,  Troy Kane  2 ,  Jyothi Manohar  2 ,  Alyssa Duren-Lubanski  2 ,  Michael Sigouros  2 ,  Jenna Moyer  2 ,  Bhavneet Bhinder  6 ,  Pooja Chandra  6 ,  Murtaza Malbari  4 ,  Karsten Boehnke  7 ,  Juan Miguel Mosquera  3 ,  Vivek Mittal  4 ,  Andrea Sboner  8 ,  Hamza Gokozan  9 ,  Nasser Altorki  4 ,  Olivier Elemento  10 ,  M Laura Martin  11
Affiliations
  • 1. Caryl and Israel Englander Institute for Precision Medicine, Weill Cornell Medicine, New York, NY 10021, USA; Department of Physiology and Biophysics, Weill Cornell Medicine, New York, NY 10021, USA.
  • 2. Caryl and Israel Englander Institute for Precision Medicine, Weill Cornell Medicine, New York, NY 10021, USA.
  • 3. Caryl and Israel Englander Institute for Precision Medicine, Weill Cornell Medicine, New York, NY 10021, USA; Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY 10021, USA.
  • 4. Cardiothoracic Surgery, Weill Cornell Medical College, New York, NY 10021, USA.
  • 5. Department of Hematology and Oncology, Weill Cornell Medical College, New York, NY 10021, USA.
  • 6. Caryl and Israel Englander Institute for Precision Medicine, Weill Cornell Medicine, New York, NY 10021, USA; Institute for Computational Biomedicine, Weill Cornell Medicine, New York, NY 10021, USA.
  • 7. Eli Lilly and Company, Lilly Oncology, Discovery Technologies, New York, NY 10016, USA.
  • 8. Caryl and Israel Englander Institute for Precision Medicine, Weill Cornell Medicine, New York, NY 10021, USA; Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY 10021, USA; Institute for Computational Biomedicine, Weill Cornell Medicine, New York, NY 10021, USA.
  • 9. Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY 10021, USA.
  • 10. Caryl and Israel Englander Institute for Precision Medicine, Weill Cornell Medicine, New York, NY 10021, USA; Department of Physiology and Biophysics, Weill Cornell Medicine, New York, NY 10021, USA; Institute for Computational Biomedicine, Weill Cornell Medicine, New York, NY 10021, USA.
  • 11. Caryl and Israel Englander Institute for Precision Medicine, Weill Cornell Medicine, New York, NY 10021, USA. Electronic address: [email protected].
PMID: 42134319 DOI: 10.1016/j.crmeth.2026.101339
Abstract

Non-Small Cell Lung Cancer (NSCLC) remains a leading cause of cancer-related mortality. Despite various therapeutic options, treatment resistance is common, underscoring the need for effective combination therapies and reliable pre-clinical models for patient-specific evaluation. Here, we describe strategies for reconstituting tumor immune microenvironment (TIME) components within patient-derived tumor Organoid (PDTO) cultures. We established a tumor processing pipeline that enables concurrent expansion of tumor-infiltrating lymphocytes (TILs) and PDTO generation from the same resection. We optimized scalable assays to assess IFN-γ secretion and T cell cytotoxicity with immune checkpoint inhibitors (alone or in combination) and targeted inhibitors, capturing inter-patient heterogeneity and intra-patient variations between TILs and peripheral blood mononuclear cells (PBMCs). Additionally, we developed methods for differentiating PDTO-specific tumor-associated Macrophages (TAMs) and established PDTO-TAM co-culture systems to evaluate TAM effects on PDTO growth and chemotherapy sensitivity. All approaches are scalable to high-throughput levels, highlighting the value of TIME-PDTO co-cultures for therapeutic modeling and precision medicine.

Keywords
CP: cancer biology; CP: immunology; T cells; co-cultures; immunotherapy; tumor microenviroment; tumor organoids; tumor-infiltrating lymphocytes.
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