Mature tertiary lymphoid structures predict immune checkpoint inhibitor efficacy in solid tumors independently of pd-l1 expression

ABSTRACT Only a minority of patients derive long-term clinical benefit from anti-programmed cell death protein 1 (anti-PD-1) or anti-programmed death-ligand 1 (anti-PD-L1) monoclonal

antibodies. The presence of tertiary lymphoid structures (TLSs) has been associated with improved survival in several tumor types. Here, using a large-scale retrospective analysis of three

independent cohorts of patients with cancer who were treated with anti-PD-1 or anti-PD-L1 antibodies, we show that the presence of mature TLSs was associated with improved objective response

rates, progression-free survival and overall survival, independent of PD-L1 expression status and CD8+ T cell density. These results pave the way for using TLS detection to select patients

who are more likely to benefit from immune checkpoint blockade. Access through your institution Buy or subscribe This is a preview of subscription content, access via your institution ACCESS

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Read our FAQs * Contact customer support SIMILAR CONTENT BEING VIEWED BY OTHERS THE PD-1 EXPRESSION BALANCE BETWEEN EFFECTOR AND REGULATORY T CELLS PREDICTS THE CLINICAL EFFICACY OF PD-1

BLOCKADE THERAPIES Article 31 August 2020 PERIPHERAL CD4 MEMORY T CELLS PREDICT THE EFFICACY OF IMMUNE CHECKPOINT INHIBITOR THERAPY IN PATIENTS WITH NON-SMALL CELL LUNG CANCER Article Open

access 04 July 2023 TERTIARY LYMPHOID STRUCTURES AND B CELLS DETERMINE CLINICALLY RELEVANT T CELL PHENOTYPES IN OVARIAN CANCER Article Open access 21 March 2024 DATA AVAILABILITY The

datasets that support the findings of this study are not publicly available due to information that could compromise research participant consent. According to French/European regulations,

any re-use of the data must be approved by the ethics committee. Each request for access to the dataset (including the images) will be granted after a request sent to the corresponding

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Download references ACKNOWLEDGEMENTS This study was funded by AstraZeneca. AUTHOR INFORMATION Author notes * These authors contributed equally: Lucile Vanhersecke, Maxime Brunet, Alban

Bessede, Wolf H. Fridman, François Le Loarer, Antoine Italiano. AUTHORS AND AFFILIATIONS * Department of Pathology, Institut Bergonié, Bordeaux, France Lucile Vanhersecke, Isabelle

Soubeyran, Valérie Velasco & François Le Loarer * Faculty of Medicine, University of Bordeaux, Bordeaux, France Lucile Vanhersecke, Maxime Brunet, Mathieu Larroquette, François Le Loarer

 & Antoine Italiano * Department of Medicine, Institut Bergonié, Bordeaux, France Maxime Brunet, Sophie Cousin, Mathieu Larroquette, Maud Toulmonde, Guilhem Roubaud, Simon Pernot, 

Mathilde Cabart, François Chomy, Corentin Lefevre, Kevin Bourcier & Antoine Italiano * Explicyte Immuno-Oncology, Bordeaux, France Jean-Philippe Guégan, Christophe Rey, Félicie Courgeon 

& Alban Bessede * Centre de Recherche des Cordeliers, INSERM, Sorbonne Université, Equipe Labellisée Ligue Nationale Contre le Cancer, USPC Université de Paris, Paris, France Antoine

Bougouin, Ilenia Giglioli, Catherine Sautès-Fridman & Wolf H. Fridman * Department of Oncology, Clinique Marzet, Pau, France Sylvestre Le Moulec * Department of Medicine, Gustave Roussy,

Villejuif, France Benjamin Besse, Yohann Loriot, Aurélien Marabelle, Jean-Charles Soria & Antoine Italiano * Department of Radiology, Institut Bergonié, Bordeaux, France Michèle Kind *

AstraZeneca, Rahway, NJ, USA Ezoglin Oflazoglu & Ariel Savina * Clinical Research and Clinical Epidemiology Unit (ISO 9001 Certified), Institut Bergonié, Comprehensive Cancer Centre,

Bordeaux, France Carine Bellera & Casimir Sofeu Authors * Lucile Vanhersecke View author publications You can also search for this author inPubMed Google Scholar * Maxime Brunet View

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Italiano View author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS A.I., A. Bessede, W.H.F. and F.L.L. conceived of and designed the study. L.V. and

F.L.L. performed the histological analyses. S.C., S.L.M., I.S., M.T., G.R., S.P., M.C., F.C., C.L., K.B., M.K., F.C., B.B., Y.L., A.M., J.-C.S. and A.I. provided study materials or treated

patients. All authors collected and assembled data. A.I., A. Bessede, L.V. and F.L.L. developed the tables and figures. A.I., A. Bessede, M.B., F.L.L., W.H.F. and C.S.-F. conducted the

literature search and wrote the manuscript. All authors were involved in the critical review of the manuscript and approved the final version. CORRESPONDING AUTHOR Correspondence to Antoine

Italiano. ETHICS DECLARATIONS COMPETING INTERESTS A. Bessede, J.-P.G. and C.R. are employees of ImmuSmol/Explicyte. E.O. and A.S. are employees of AstraZeneca. A.I. received research grants

from AstraZeneca, Bayer, Bristol Myers Squibb, Chugai, Merck, MSD, PharmaMar, Novartis and Roche and received personal fees from Epizyme, Bayer, Lilly, Roche and SpringWorks. B.B. received

grants from AstraZeneca, Pfizer, Eli Lilly, Onxeo, Bristol Myers Squibb, Inivata, AbbVie, Amgen, Blueprint Medicines, Celgene, GlaxoSmithKline, Ignyta, Ipsen, Merck KGaA, MSD Oncology,

Nektar, PharmaMar, Sanofi, Spectrum Pharmaceuticals, Takeda, Tiziana Therapeutics, Cristal Therapeutics, Daiichi Sankyo, Janssen Oncology, OSE Immunotherapeutics, BeiGene, Boehringer

Ingelheim, Genentech, Servier and Tolero Pharmaceuticals. Y.L. received grants from Janssen, Sanofi and MSD, personal fees from Janssen, Sanofi, Astellas, Roche, AstraZeneca, Bristol Myers

Squibb, Seattle Genetics, MSD, Clovis, Incyte and Pfizer and non-financial support from Astellas, Roche, AstraZeneca, Bristol Myers Squibb, Seattle Genetics and MSD. A.M. received research

grants from Merus, Bristol Myers Squibb, Boehringer Ingelheim, Transgene and MSD and personal fees from Bristol Myers Squibb, AstraZeneca, MedImmune, Oncovir and Merieux. J.-C.S. received

consultancy fees from AstraZeneca, Astex, Clovis, GlaxoSmithKline, GamaMabs, Lilly, MSD, Mission Therapeutics, Merus, Pfizer, PharmaMar, Pierre Fabre, Roche/Genentech, Sanofi, Servier,

Symphogen and Takeda. L.V., M.B., S.C., S.L.M., M.L., I.S., M.T., G.R., S.P., M.C., F.C., C.L., K.B., M.K., I.G., C.S.-F., V.V., F.C., W.H.F. and F.L.L. declare no competing interests.

ADDITIONAL INFORMATION PEER REVIEW INFORMATION _Nature Cancer_ thanks the anonymous reviewers for their contribution to the peer review of this work. PUBLISHER’S NOTE Springer Nature remains

neutral with regard to jurisdictional claims in published maps and institutional affiliations. EXTENDED DATA EXTENDED DATA FIG. 1 ASSESSMENT OF THE PRESENCE OF TLS AND THEIR MATURATION

STAGE IN TUMORS. A: This is a TLS-positive primary pancreatic adenocarcinoma associated with a T CD8 + lymphocyte density of 154/mm² and negative for PD-L1. The TLS are delineated with the

black lines on the HES slide, highlighting their vicinity to tumor cells. Scale bar indicates 300 µm in size. Representative of 540 tumors analyzed (Discovery cohort n = 328, validation

cohort A n = 131, validation cohort B n = 81). B: This panel shows representative examples of immature and mature TLS observed in tumor samples. Upper panel: This mature TLS is detected in a

primary pancreatic adenocarcinoma associated with a T CD8 + lymphocyte density of 154/mm² and negative for PD-L1. Mature TLS are defined by the presence of a network of CD23-positive

dendritic cells on immunofluorescence (note that in this case, the presence of a germinal center visible on the HES was already diagnostic of a mature TLS). Lower panel: The picture shows an

immature TLS detected in a primary adenocarcinoma of the lung concomitantly showing a high T CD8 + lymphocyte infiltrate of 372/mm² and a PD-L1 TPS score of 1%. The tumor was only

associated with immature TLS displaying no germinal center and no network of CD23-positive dendritic cells on immunofluorescence. Representative of 540 tumors analyzed (Discovery cohort n = 

328, validation cohort A n = 131, validation cohort B n = 81). The pictures from the left to right column correspond to 1) Hematoxylin Eosin Saffron (HES) staining, 2) Double

immunohistochemistry staining of CD3/CD20 (CD3 in brown, CD20 in purple), 3) Double immunohistochemistry staining of CD8/PD-L1 (CD8 in brown, PD-L1 in purple), 4) Multiplex

immunofluorescence assay of CD4 (blue), CD8 (yellow), CD20 (orange), CD21 (green) and CD23 (pink). The scale bars on the HES images indicate 50 µm and 100 µm for the upper and lower panels,

respectively. Black cropped arrows highlight the tumor cells in the samples. EXTENDED DATA FIG. 2 PREDICTIVE VALUE OF TLS STATUS ACCORDING TO CPS PD-L1 SCORES. Objective response rates (OR:

objective response, SD: stable disease; PD: progressive disease; chi squared (χ²) test) and Kaplan-Meier curves of progression-free survival (log-rank test) and overall survival (log-rank

test) of 328 cancer patients treated with anti-PD1/PD-L1 antagonists according to CPS PD-L1 scores (a: CPS PD-L1 < 1, N = 223 patients; b: CPS PD-L1 ≥ 1, N = 105 patients) and TLS status

(red curve: mature-TLS positive tumors; blue line: mature-TLS negative tumors). Source data EXTENDED DATA FIG. 3 PREDICTIVE VALUE OF TLS STATUS ACCORDING TO T CD8 CELL DENSITY. Objective

Response rates (OR: objective response, SD: stable disease, PD: progressive disease; chi squared (χ²) test) and Kaplan-Meier curves of progression-free (log-rank test) and overall survival

(log-rank test) of 328 cancer patients treated with anti-PD1/PD-L1 antagonists according to T CD8 + cell density (A: low density, N = 165 patients; B: high density, N = 163 patients) and TLS

status (red curve: mature-TLS positive tumors; blue line: mature-TLS negative tumors). Source data EXTENDED DATA FIG. 4 OUTCOME OF CANCER PATIENTS (NON-SMALL CELL LUNG CANCER EXCLUDED)

ACCORDING TO TLS STATUS. A) Objective Response rates (OR: objective response, SD: stable disease, PD: progressive disease; chi squared (χ²) test) and Kaplan-Meier curves (log-rank test) of

progression-free (B) and overall survival (C) of 201 cancer patients (all tumor types except non-small cell lung cancer) treated with anti-PD1/PD-L1 antagonists according to TLS status (red

curve: mature-TLS positive tumors; blue line: mature-TLS negative tumors). Source data EXTENDED DATA FIG. 5 COMPARISON OF THE TLS SCREENING WITH PATHOLOGY AND IMMUNOHISTOCHEMISTRY PATHOLOGY

VERSUS THE SCREENING WITH IMMUNOFLUORESCENCE (METHOD BY PETITPREZ ET AL). First line: conspicuous mature TLS with CD23 + follicular dendritic cells network. Second line: mature TLS defined

by isolated CD23 + cell displaying dendritic morphology. Pictures from the left to right column correspond to 1) Hematoxylin Eosin Saffron (HES) staining, 2) Multiplex immunofluorescence

assay of CD23 (pink) and CD20 (orange), 3) Double immunohistochemistry staining of CD23/CD20 (CD23 in brown, CD20 in red). The scale bars on the HES images indicate 50 µm in size.

Representative of 540 tumors analyzed (Discovery cohort n = 328, validation cohort A n = 131, validation cohort B n = 81). SUPPLEMENTARY INFORMATION SUPPLEMENTARY INFORMATION Supplementary

Tables 1 and 2. REPORTING SUMMARY SUPPLEMENTARY TABLES Supplementary Tables 3–5. SOURCE DATA SOURCE DATA FIG. 1 Statistical source data. SOURCE DATA FIG. 2 Statistical source data. SOURCE

DATA EXTENDED DATA FIG. 2 Statistical source data. SOURCE DATA EXTENDED DATA FIG. 3 Statistical source data. SOURCE DATA EXTENDED DATA FIG. 4 Statistical source data. RIGHTS AND PERMISSIONS

Reprints and permissions ABOUT THIS ARTICLE CITE THIS ARTICLE Vanhersecke, L., Brunet, M., Guégan, JP. _et al._ Mature tertiary lymphoid structures predict immune checkpoint inhibitor

efficacy in solid tumors independently of PD-L1 expression. _Nat Cancer_ 2, 794–802 (2021). https://doi.org/10.1038/s43018-021-00232-6 Download citation * Received: 16 January 2021 *

Accepted: 08 June 2021 * Published: 12 August 2021 * Issue Date: August 2021 * DOI: https://doi.org/10.1038/s43018-021-00232-6 SHARE THIS ARTICLE Anyone you share the following link with

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