{
  "abstract": "Background Neoadjuvant Chemoimmunotherapy (ChIO) administered prior to surgery has significantly improved the prognosis of patients with locally advanced non-small cell lung cancer (NSCLC), increasing the proportion of individuals achieving complete pathological responses (CPR, i.e. 0% of viable tumour cells) and survival. 1–3Tertiary lymphoid structures (TLSs) are ectopic lymphoid aggregates that resemble the architecture and immunological functions of secondary lymphoid organs. TLSs typically arise in areas of chronic inflammation with persistent antigen exposure, such as the tumour microenvironment of solid malignancies, where their presence has been associated with favourable clinical outcomes.4–8 However, TLS induction, as well as their specific role in mediating responses to neoadjuvant ChIO in NSCLC, remain mainly unexplored.Methods To investigate TLS induction and TLS functionality association with response, we analysed pre- and post-treatment FFPE tumour samples from NSCLC patients treated with perioperative nivolumab plus chemotherapy in the NADIM trial ( NCT03081689). Bulk RNA-seq (n=15 patients, OIRRA, Thermofisher) and spatial transcriptomics (ST) (n=12, Visium Spatial for FFPE, 10x Genomics) were performed. TLS-related regions—mature TLS, non-mature TLS, dispersed B-cells, and low B-cell area—were defined based on H&E staining and gene expression data. Upregulated post-treatment genes (DESeq2) were spatially mapped using ST data and MCP-counter. Differential expression and cell composition analyses (Scran, GSEA, SpaCET) were conducted between CPR and NCPR tumours across spatial regions to characterize response-associated immune features.Results Neoadjuvant ChIO increased the expression of 15 genes, several related to B-cells and TLS induction (JCHAIN, CXCR4, MS4A1...). These genes were statistically enriched in TLS regions, suggesting a TLS induction process during neoadjuvant ChIO ( figure 1).Regarding CPR and NCPR differences in ST areas of interest (figure 2), CPR mTLSs showed 214 overexpressed genes and multiple enriched pathways, pointing towards enhanced major histocompatibility complex (MHC) antigen processing and presentation, and humoral responses.Furthermore, CPR mTLSs display higher plasma cell content (p<0.001), whilst the NCPR mTLSs have more naïve B cells (p<0.001). Higher CPR mTLSs functionality is also supported by increased T follicular helper (Tfh) levels (p=0.002), as well as larger macrophage infiltrate (p<0.001). Additionally, CPR low B-cell area has higher T CD8 and natural killer presence (p<0.001) than the NCPR (figure 2).Conclusions NSCLC neoadjuvant ChIO antitumour response is primarily orchestrated through tertiary lymphoid structures, which serve as pivotal immunological hubs coordinating local adaptive immune responses through increased antigen presentation, humoral response and cytotoxicity within the tumour microenvironment.Trial Registration NADIM trial ( NCT03081689)References Provencio M, Calvo V, Romero A, Spicer JD, Cruz-Bermúdez A. Treatment Sequencing in Resectable Lung Cancer: The Good and the Bad of Adjuvant Versus Neoadjuvant Therapy. American Society of Clinical Oncology Educational Book 2022;(42):711-728. doi:10.1200/edbk_358995Provencio M, Nadal E, Insa A, et al. Neoadjuvant chemotherapy and nivolumab in resectable non-small-cell lung cancer (NADIM): an open-label, multicentre, single-arm, phase 2 trial. Lancet Oncol. 2020;21(11):1413-1422. doi:10.1016/S1470-2045(20)30453-8Provencio M, Nadal E, González-Larriba JL, et al. Perioperative nivolumab and chemotherapy in stage III non-small-cell lung cancer. New England Journal of Medicine 2023;389(6):504-513. doi:10.1056/NEJMoa2215530Fridman WH, Meylan M, Pupier G, Calvez A, Hernandez I, Sautès-Fridman C. Tertiary lymphoid structures and B cells: an intratumoral immunity cycle. Immunity. 2023;56(10):2254-2269. doi:10.1016/j.immuni.2023.08.009Teillaud JL, Houel A, Panouillot M, Riffard C, Dieu-Nosjean MC. Tertiary lymphoid structures in anticancer immunity. Nat Rev Cancer 2024;24(9):629-646. doi:10.1038/s41568-024-00728-0Schumacher TN, Thommen DS. Tertiary lymphoid structures in cancer. Science (1979). 2022;375(6576). doi:10.1126/science.abf9419Flippot R, Teixeira M, Rey-Cardenas M, et al. B cells and the coordination of immune checkpoint inhibitor response in patients with solid tumors. J Immunother Cancer 2024;12(4). doi:10.1136/jitc-2023-008636Laumont CM, Banville AC, Gilardi M, Hollern DP, Nelson BH. Tumour-infiltrating B cells: immunological mechanisms, clinical impact and therapeutic opportunities. Nat Rev Cancer. 2022;22(7):414-430. doi:10.1038/s41568-022-00466-1Ethics Approval Informed consent for the collection of research samples and study protocol was approved by the clinical research ethics committee of Hospital Puerta de Hierro (ref. 20.16) and the Spanish Lung Cancer Group (SLCG) Board (ref. N/A) in accordance with the International Conference on Harmonization Guidelines on Good Clinical Practice and the Declaration of Helsinki. Participants gave informed consent to participate in the study before taking part.Consent Informed consent for the collection of research samples and study protocol was approved by the clinical research ethics committee of Hospital Puerta de Hierro (ref. 20.16) and the Spanish Lung Cancer Group (SLCG) Board (ref. N/A) in accordance with the International Conference on Harmonization Guidelines on Good Clinical Practice and the Declaration of Helsinki. Participants gave informed consent to participate in the study before taking part.Abstract 734 Figure 1Abstract 734 Figure 2",
  "authors": [
    {
      "affiliations": [
        "Fundación Investigación Biomédica Hospital Puerta de Hierro Majadahonda, Madrid, Spain"
      ],
      "name": "María de Los Ángeles Gil González"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Marta Molina-Alejandre"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Belén Sierra-Rodero"
    },
    {
      "affiliations": [
        "Institut Català d’Oncologia (ICO), Bellvitge campus, L’Hospitalet de Llobregat, Barcelona, Spain"
      ],
      "name": "Ernest Nadal"
    },
    {
      "affiliations": [
        "Institut Català d’Oncologia (ICO), Bellvitge campus, L’Hospitalet de Llobregat, Barcelona, Spain"
      ],
      "name": "Ramon Palmero"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Ana Collazo-Lorduy"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Virginia Calvo"
    },
    {
      "affiliations": [
        "Hospital Álvaro Cunqueiro, Vigo, Spain"
      ],
      "name": "Martín Lázaro"
    },
    {
      "affiliations": [
        "Hospital General Universitario Dr. Balmis, Alicante, Spain"
      ],
      "name": "Bartomeu Massuti"
    },
    {
      "affiliations": [
        "Hospital Clínic Universitari, Valencia, Spain"
      ],
      "name": "Amelia Insa"
    },
    {
      "affiliations": [
        "Hospital Universitario Fundación Jiménez Díaz, Madrid, Spain"
      ],
      "name": "Manuel Dómine"
    },
    {
      "affiliations": [
        "Hospital Universitario Reina Sofía, Córdoba, Spain"
      ],
      "name": "Isidoro Barneto"
    },
    {
      "affiliations": [
        "Hospital Insular Gran Canaria, Las Palmas, Spain"
      ],
      "name": "Delvys Rodriguez-Abreu"
    },
    {
      "affiliations": [
        "Hospital Universitario La Paz, Madrid, Spain"
      ],
      "name": "Javier de Castro"
    },
    {
      "affiliations": [
        "Hospital Teresa Herrera, A Coruña, Spain"
      ],
      "name": "Maria Rosario Garcia Campelo"
    },
    {
      "affiliations": [
        "Hospital Universitario de Valencia, Valencia, Spain"
      ],
      "name": "Ana Blasco"
    },
    {
      "affiliations": [
        "Hospital Universitario Virgen del Rocío, Sevilla, Spain"
      ],
      "name": "Reyes Bernabé"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Rafael Muñoz-Viana"
    },
    {
      "affiliations": [
        "Unidad de Microscopía Óptica Avanzada, Instituto de Salud Carlos III, Madrid, Spain"
      ],
      "name": "Diego Megías"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Javier Martín-López"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Juan Manuel Gutierrez-Escobedo"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Cristina Martínez-Toledo"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Alberto Cruz-Bermúdez"
    },
    {
      "affiliations": [
        "Hospital Universitario Puerta de Hierro-Majadahonda, Madrid, Spain"
      ],
      "name": "Mariano Provencio"
    }
  ],
  "title": "734 Tertiary lymphoid structures as hallmarks of response to neoadjuvant chemoimmunotherapy in locally advanced non-small cell lung cancer NADIM trial",
  "uid": "c7d7a22d-abd4-5797-86ca-6a1d20ea5e6c"
}