Identification, validation and biological characterisation of novel glioblastoma tumour microenvironment subtypes: implications for precision immunotherapy

K. White; K. Connor; M. Meylan; A. Bougoüin; M. Salvucci; F. Bielle; A. C. O'Farrell; K. Sweeney; L. Weng; G. Bergers; P. Dicker; D. M. Ashley; E. S. Lipp; J. T. Low; J. Zhao; P. Wen; R. Prins; M. Verreault; A. Idbaih; A. Biswas; J. H.M. Prehn; D. Lambrechts; I. Arijs; F. Lodi; G. Dilcan; M. Lamfers; S. Leenstra; F. Fabro; I. Ntafoulis; J. M. Kros; J. Cryan; F. Brett; E. Quissac; A. Beausang; S. MacNally; P. O'Halloran; J. Clerkin; O. Bacon; A. Kremer; R. T. Chi Yen; F. S. Varn; R. G.W. Verhaak; C. Sautès-Fridman; W. H. Fridman; A. T. Byrne

doi:10.1016/j.annonc.2022.11.008

Identification, validation and biological characterisation of novel glioblastoma tumour microenvironment subtypes: implications for precision immunotherapy

K. White
, K. Connor
, M. Meylan
, A. Bougoüin
, M. Salvucci
, F. Bielle
, A. C. O'Farrell
, K. Sweeney
, L. Weng
, G. Bergers
, P. Dicker
, D. M. Ashley
, E. S. Lipp
, J. T. Low
, J. Zhao
, P. Wen
, R. Prins
, M. Verreault
, A. Idbaih
, A. Biswas

J. H.M. Prehn, D. Lambrechts, I. Arijs, F. Lodi, G. Dilcan, M. Lamfers, S. Leenstra, F. Fabro, I. Ntafoulis, J. M. Kros, J. Cryan, F. Brett, E. Quissac, A. Beausang, S. MacNally, P. O'Halloran, J. Clerkin, O. Bacon, A. Kremer, R. T. Chi Yen, F. S. Varn, R. G.W. Verhaak, C. Sautès-Fridman, W. H. Fridman, A. T. Byrne^*

^*Corresponding author for this work

Royal College of Surgeons in Ireland
Sorbonne Université
Hôpital La Pitié-Salpêtrière
KU Leuven
Duke University
Dana-Farber Cancer Institute
Flanders Institute for Biotechnology
Information Technology for Translational Medicine (ITTM) S.A
Jackson Laboratory
Columbia University
University of California
Information Technology for Translational Medicine (ITTM)
VU University Medical Center
The Jackson Laboratory for Genomic Medicine

Research output: Contribution to journal › Article › Academic › peer-review

104 Citations (Scopus)

183 Downloads (Pure)

Abstract

Background: New precision medicine therapies are urgently required for glioblastoma (GBM). However, to date, efforts to subtype patients based on molecular profiles have failed to direct treatment strategies. We hypothesised that interrogation of the GBM tumour microenvironment (TME) and identification of novel TME-specific subtypes could inform new precision immunotherapy treatment strategies. Materials and methods: A refined and validated microenvironment cell population (MCP) counter method was applied to >800 GBM patient tumours (GBM-MCP-counter). Specifically, partition around medoids (PAM) clustering of GBM-MCP-counter scores in the GLIOTRAIN discovery cohort identified three novel patient clusters, uniquely characterised by TME composition, functional orientation markers and immune checkpoint proteins. Validation was carried out in three independent GBM-RNA-seq datasets. Neoantigen, mutational and gene ontology analysis identified mutations and uniquely altered pathways across subtypes. The longitudinal Glioma Longitudinal AnalySiS (GLASS) cohort and three immunotherapy clinical trial cohorts [treatment with neoadjuvant/adjuvant anti-programmed cell death protein 1 (PD-1) or PSVRIPO] were further interrogated to assess subtype alterations between primary and recurrent tumours and to assess the utility of TME classifiers as immunotherapy biomarkers. Results: TME^High tumours (30%) displayed elevated lymphocyte, myeloid cell immune checkpoint, programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated protein 4 transcripts. TME^High/mesenchymal+ patients featured tertiary lymphoid structures. TME^Med (46%) tumours were enriched for endothelial cell gene expression profiles and displayed heterogeneous immune populations. TME^Low (24%) tumours were manifest as an ‘immune-desert’ group. TME subtype transitions upon recurrence were identified in the longitudinal GLASS cohort. Assessment of GBM immunotherapy trial datasets revealed that TME^High patients receiving neoadjuvant anti-PD-1 had significantly increased overall survival (P = 0.04). Moreover, TME^High patients treated with adjuvant anti-PD-1 or oncolytic virus (PVSRIPO) showed a trend towards improved survival. Conclusions: We have established a novel TME-based classification system for application in intracranial malignancies. TME subtypes represent canonical ‘termini a quo’ (starting points) to support an improved precision immunotherapy treatment approach.

Original language	English
Pages (from-to)	300-314
Number of pages	15
Journal	Annals of Oncology
Volume	34
Issue number	3
DOIs	https://doi.org/10.1016/j.annonc.2022.11.008
Publication status	Published - Mar 2023

Bibliographical note

Funding Information:
This project has received funding from the European Union’s Horizon 2020 and 2021 research and innovation programme under the Marie Skłodowska-Curie ITN initiatives [grant number #766069], GLIOTRAIN ( http://www.gliotrain.eu ) and ‘GLIORESOLVE’ projects [grant number #101073386]. Additional support was received by donation from the Paolo Iacovelli memorial endowment. We are also grateful for the financial support from the Beaumont Hospital Foundation , and from Brain Tumour Ireland to establish the Beaumont Hospital Brain Tumour Biorepository. The authors further acknowledge financial contribution towards staff salaries from RCSI and Champions Oncology (Baltimore, USA).

Funding Information:
The work presented in this paper is dedicated to the memory of Mr Paolo Iacovelli. We would like to thank J. Heffernan, L. M. Houlihan and J. Mythen (Beaumont Hospital Neuropathology Department) for their help in establishing the GLIOTRAIN biobank in Dublin. The authors gratefully acknowledge all patients who kindly donated tumour tissue and clinical/genomic data used in this study. This project has received funding from the European Union's Horizon 2020 and 2021 research and innovation programme under the Marie Skłodowska-Curie ITN initiatives [grant number #766069], GLIOTRAIN (http://www.gliotrain.eu) and ‘GLIORESOLVE’ projects [grant number #101073386]. Additional support was received by donation from the Paolo Iacovelli memorial endowment. We are also grateful for the financial support from the Beaumont Hospital Foundation, and from Brain Tumour Ireland to establish the Beaumont Hospital Brain Tumour Biorepository. The authors further acknowledge financial contribution towards staff salaries from RCSI and Champions Oncology (Baltimore, USA). The authors have declared no conflicts of interest.

Publisher Copyright:
© 2022 The Authors

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Identification, validation and biological characterisation of novel glioblastoma tumour microenvironment subtypesFinal published version, 0.99 MBLicence: CC BY-NC-ND

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White, K., Connor, K., Meylan, M., Bougoüin, A., Salvucci, M., Bielle, F., O'Farrell, A. C., Sweeney, K., Weng, L., Bergers, G., Dicker, P., Ashley, D. M., Lipp, E. S., Low, J. T., Zhao, J., Wen, P., Prins, R., Verreault, M., Idbaih, A., ... Byrne, A. T. (2023). Identification, validation and biological characterisation of novel glioblastoma tumour microenvironment subtypes: implications for precision immunotherapy. Annals of Oncology, 34(3), 300-314. https://doi.org/10.1016/j.annonc.2022.11.008

@article{b94eade709054b67aedcc8514f107f19,

title = "Identification, validation and biological characterisation of novel glioblastoma tumour microenvironment subtypes: implications for precision immunotherapy",

abstract = "Background: New precision medicine therapies are urgently required for glioblastoma (GBM). However, to date, efforts to subtype patients based on molecular profiles have failed to direct treatment strategies. We hypothesised that interrogation of the GBM tumour microenvironment (TME) and identification of novel TME-specific subtypes could inform new precision immunotherapy treatment strategies. Materials and methods: A refined and validated microenvironment cell population (MCP) counter method was applied to >800 GBM patient tumours (GBM-MCP-counter). Specifically, partition around medoids (PAM) clustering of GBM-MCP-counter scores in the GLIOTRAIN discovery cohort identified three novel patient clusters, uniquely characterised by TME composition, functional orientation markers and immune checkpoint proteins. Validation was carried out in three independent GBM-RNA-seq datasets. Neoantigen, mutational and gene ontology analysis identified mutations and uniquely altered pathways across subtypes. The longitudinal Glioma Longitudinal AnalySiS (GLASS) cohort and three immunotherapy clinical trial cohorts [treatment with neoadjuvant/adjuvant anti-programmed cell death protein 1 (PD-1) or PSVRIPO] were further interrogated to assess subtype alterations between primary and recurrent tumours and to assess the utility of TME classifiers as immunotherapy biomarkers. Results: TMEHigh tumours (30\%) displayed elevated lymphocyte, myeloid cell immune checkpoint, programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated protein 4 transcripts. TMEHigh/mesenchymal+ patients featured tertiary lymphoid structures. TMEMed (46\%) tumours were enriched for endothelial cell gene expression profiles and displayed heterogeneous immune populations. TMELow (24\%) tumours were manifest as an {\textquoteleft}immune-desert{\textquoteright} group. TME subtype transitions upon recurrence were identified in the longitudinal GLASS cohort. Assessment of GBM immunotherapy trial datasets revealed that TMEHigh patients receiving neoadjuvant anti-PD-1 had significantly increased overall survival (P = 0.04). Moreover, TMEHigh patients treated with adjuvant anti-PD-1 or oncolytic virus (PVSRIPO) showed a trend towards improved survival. Conclusions: We have established a novel TME-based classification system for application in intracranial malignancies. TME subtypes represent canonical {\textquoteleft}termini a quo{\textquoteright} (starting points) to support an improved precision immunotherapy treatment approach.",

author = "K. White and K. Connor and M. Meylan and A. Bougo{\"u}in and M. Salvucci and F. Bielle and O'Farrell, \{A. C.\} and K. Sweeney and L. Weng and G. Bergers and P. Dicker and Ashley, \{D. M.\} and Lipp, \{E. S.\} and Low, \{J. T.\} and J. Zhao and P. Wen and R. Prins and M. Verreault and A. Idbaih and A. Biswas and Prehn, \{J. H.M.\} and D. Lambrechts and I. Arijs and F. Lodi and G. Dilcan and M. Lamfers and S. Leenstra and F. Fabro and I. Ntafoulis and Kros, \{J. M.\} and J. Cryan and F. Brett and E. Quissac and A. Beausang and S. MacNally and P. O'Halloran and J. Clerkin and O. Bacon and A. Kremer and \{Chi Yen\}, \{R. T.\} and Varn, \{F. S.\} and Verhaak, \{R. G.W.\} and C. Saut{\`e}s-Fridman and Fridman, \{W. H.\} and Byrne, \{A. T.\}",

note = "Funding Information: This project has received funding from the European Union{\textquoteright}s Horizon 2020 and 2021 research and innovation programme under the Marie Sk{\l}odowska-Curie ITN initiatives [grant number \#766069], GLIOTRAIN ( http://www.gliotrain.eu ) and {\textquoteleft}GLIORESOLVE{\textquoteright} projects [grant number \#101073386]. Additional support was received by donation from the Paolo Iacovelli memorial endowment. We are also grateful for the financial support from the Beaumont Hospital Foundation , and from Brain Tumour Ireland to establish the Beaumont Hospital Brain Tumour Biorepository. The authors further acknowledge financial contribution towards staff salaries from RCSI and Champions Oncology (Baltimore, USA). Funding Information: The work presented in this paper is dedicated to the memory of Mr Paolo Iacovelli. We would like to thank J. Heffernan, L. M. Houlihan and J. Mythen (Beaumont Hospital Neuropathology Department) for their help in establishing the GLIOTRAIN biobank in Dublin. The authors gratefully acknowledge all patients who kindly donated tumour tissue and clinical/genomic data used in this study. This project has received funding from the European Union's Horizon 2020 and 2021 research and innovation programme under the Marie Sk{\l}odowska-Curie ITN initiatives [grant number \#766069], GLIOTRAIN (http://www.gliotrain.eu) and {\textquoteleft}GLIORESOLVE{\textquoteright} projects [grant number \#101073386]. Additional support was received by donation from the Paolo Iacovelli memorial endowment. We are also grateful for the financial support from the Beaumont Hospital Foundation, and from Brain Tumour Ireland to establish the Beaumont Hospital Brain Tumour Biorepository. The authors further acknowledge financial contribution towards staff salaries from RCSI and Champions Oncology (Baltimore, USA). The authors have declared no conflicts of interest. Publisher Copyright: {\textcopyright} 2022 The Authors",

year = "2023",

month = mar,

doi = "10.1016/j.annonc.2022.11.008",

language = "English",

volume = "34",

pages = "300--314",

journal = "Annals of Oncology",

issn = "0923-7534",

publisher = "Elsevier Ltd.",

number = "3",

}

White, K, Connor, K, Meylan, M, Bougoüin, A, Salvucci, M, Bielle, F, O'Farrell, AC, Sweeney, K, Weng, L, Bergers, G, Dicker, P, Ashley, DM, Lipp, ES, Low, JT, Zhao, J, Wen, P, Prins, R, Verreault, M, Idbaih, A, Biswas, A, Prehn, JHM, Lambrechts, D, Arijs, I, Lodi, F, Dilcan, G, Lamfers, M , Leenstra, S , Fabro, F, Ntafoulis, I, Kros, JM, Cryan, J, Brett, F, Quissac, E, Beausang, A, MacNally, S, O'Halloran, P, Clerkin, J, Bacon, O, Kremer, A, Chi Yen, RT, Varn, FS, Verhaak, RGW, Sautès-Fridman, C, Fridman, WH & Byrne, AT 2023, 'Identification, validation and biological characterisation of novel glioblastoma tumour microenvironment subtypes: implications for precision immunotherapy', Annals of Oncology, vol. 34, no. 3, pp. 300-314. https://doi.org/10.1016/j.annonc.2022.11.008

TY - JOUR

T1 - Identification, validation and biological characterisation of novel glioblastoma tumour microenvironment subtypes

T2 - implications for precision immunotherapy

AU - White, K.

AU - Connor, K.

AU - Meylan, M.

AU - Bougoüin, A.

AU - Salvucci, M.

AU - Bielle, F.

AU - O'Farrell, A. C.

AU - Sweeney, K.

AU - Weng, L.

AU - Bergers, G.

AU - Dicker, P.

AU - Ashley, D. M.

AU - Lipp, E. S.

AU - Low, J. T.

AU - Zhao, J.

AU - Wen, P.

AU - Prins, R.

AU - Verreault, M.

AU - Idbaih, A.

AU - Biswas, A.

AU - Prehn, J. H.M.

AU - Lambrechts, D.

AU - Arijs, I.

AU - Lodi, F.

AU - Dilcan, G.

AU - Lamfers, M.

AU - Leenstra, S.

AU - Fabro, F.

AU - Ntafoulis, I.

AU - Kros, J. M.

AU - Cryan, J.

AU - Brett, F.

AU - Quissac, E.

AU - Beausang, A.

AU - MacNally, S.

AU - O'Halloran, P.

AU - Clerkin, J.

AU - Bacon, O.

AU - Kremer, A.

AU - Chi Yen, R. T.

AU - Varn, F. S.

AU - Verhaak, R. G.W.

AU - Sautès-Fridman, C.

AU - Fridman, W. H.

AU - Byrne, A. T.

N1 - Funding Information: This project has received funding from the European Union’s Horizon 2020 and 2021 research and innovation programme under the Marie Skłodowska-Curie ITN initiatives [grant number #766069], GLIOTRAIN ( http://www.gliotrain.eu ) and ‘GLIORESOLVE’ projects [grant number #101073386]. Additional support was received by donation from the Paolo Iacovelli memorial endowment. We are also grateful for the financial support from the Beaumont Hospital Foundation , and from Brain Tumour Ireland to establish the Beaumont Hospital Brain Tumour Biorepository. The authors further acknowledge financial contribution towards staff salaries from RCSI and Champions Oncology (Baltimore, USA). Funding Information: The work presented in this paper is dedicated to the memory of Mr Paolo Iacovelli. We would like to thank J. Heffernan, L. M. Houlihan and J. Mythen (Beaumont Hospital Neuropathology Department) for their help in establishing the GLIOTRAIN biobank in Dublin. The authors gratefully acknowledge all patients who kindly donated tumour tissue and clinical/genomic data used in this study. This project has received funding from the European Union's Horizon 2020 and 2021 research and innovation programme under the Marie Skłodowska-Curie ITN initiatives [grant number #766069], GLIOTRAIN (http://www.gliotrain.eu) and ‘GLIORESOLVE’ projects [grant number #101073386]. Additional support was received by donation from the Paolo Iacovelli memorial endowment. We are also grateful for the financial support from the Beaumont Hospital Foundation, and from Brain Tumour Ireland to establish the Beaumont Hospital Brain Tumour Biorepository. The authors further acknowledge financial contribution towards staff salaries from RCSI and Champions Oncology (Baltimore, USA). The authors have declared no conflicts of interest. Publisher Copyright: © 2022 The Authors

PY - 2023/3

Y1 - 2023/3

N2 - Background: New precision medicine therapies are urgently required for glioblastoma (GBM). However, to date, efforts to subtype patients based on molecular profiles have failed to direct treatment strategies. We hypothesised that interrogation of the GBM tumour microenvironment (TME) and identification of novel TME-specific subtypes could inform new precision immunotherapy treatment strategies. Materials and methods: A refined and validated microenvironment cell population (MCP) counter method was applied to >800 GBM patient tumours (GBM-MCP-counter). Specifically, partition around medoids (PAM) clustering of GBM-MCP-counter scores in the GLIOTRAIN discovery cohort identified three novel patient clusters, uniquely characterised by TME composition, functional orientation markers and immune checkpoint proteins. Validation was carried out in three independent GBM-RNA-seq datasets. Neoantigen, mutational and gene ontology analysis identified mutations and uniquely altered pathways across subtypes. The longitudinal Glioma Longitudinal AnalySiS (GLASS) cohort and three immunotherapy clinical trial cohorts [treatment with neoadjuvant/adjuvant anti-programmed cell death protein 1 (PD-1) or PSVRIPO] were further interrogated to assess subtype alterations between primary and recurrent tumours and to assess the utility of TME classifiers as immunotherapy biomarkers. Results: TMEHigh tumours (30%) displayed elevated lymphocyte, myeloid cell immune checkpoint, programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated protein 4 transcripts. TMEHigh/mesenchymal+ patients featured tertiary lymphoid structures. TMEMed (46%) tumours were enriched for endothelial cell gene expression profiles and displayed heterogeneous immune populations. TMELow (24%) tumours were manifest as an ‘immune-desert’ group. TME subtype transitions upon recurrence were identified in the longitudinal GLASS cohort. Assessment of GBM immunotherapy trial datasets revealed that TMEHigh patients receiving neoadjuvant anti-PD-1 had significantly increased overall survival (P = 0.04). Moreover, TMEHigh patients treated with adjuvant anti-PD-1 or oncolytic virus (PVSRIPO) showed a trend towards improved survival. Conclusions: We have established a novel TME-based classification system for application in intracranial malignancies. TME subtypes represent canonical ‘termini a quo’ (starting points) to support an improved precision immunotherapy treatment approach.

AB - Background: New precision medicine therapies are urgently required for glioblastoma (GBM). However, to date, efforts to subtype patients based on molecular profiles have failed to direct treatment strategies. We hypothesised that interrogation of the GBM tumour microenvironment (TME) and identification of novel TME-specific subtypes could inform new precision immunotherapy treatment strategies. Materials and methods: A refined and validated microenvironment cell population (MCP) counter method was applied to >800 GBM patient tumours (GBM-MCP-counter). Specifically, partition around medoids (PAM) clustering of GBM-MCP-counter scores in the GLIOTRAIN discovery cohort identified three novel patient clusters, uniquely characterised by TME composition, functional orientation markers and immune checkpoint proteins. Validation was carried out in three independent GBM-RNA-seq datasets. Neoantigen, mutational and gene ontology analysis identified mutations and uniquely altered pathways across subtypes. The longitudinal Glioma Longitudinal AnalySiS (GLASS) cohort and three immunotherapy clinical trial cohorts [treatment with neoadjuvant/adjuvant anti-programmed cell death protein 1 (PD-1) or PSVRIPO] were further interrogated to assess subtype alterations between primary and recurrent tumours and to assess the utility of TME classifiers as immunotherapy biomarkers. Results: TMEHigh tumours (30%) displayed elevated lymphocyte, myeloid cell immune checkpoint, programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated protein 4 transcripts. TMEHigh/mesenchymal+ patients featured tertiary lymphoid structures. TMEMed (46%) tumours were enriched for endothelial cell gene expression profiles and displayed heterogeneous immune populations. TMELow (24%) tumours were manifest as an ‘immune-desert’ group. TME subtype transitions upon recurrence were identified in the longitudinal GLASS cohort. Assessment of GBM immunotherapy trial datasets revealed that TMEHigh patients receiving neoadjuvant anti-PD-1 had significantly increased overall survival (P = 0.04). Moreover, TMEHigh patients treated with adjuvant anti-PD-1 or oncolytic virus (PVSRIPO) showed a trend towards improved survival. Conclusions: We have established a novel TME-based classification system for application in intracranial malignancies. TME subtypes represent canonical ‘termini a quo’ (starting points) to support an improved precision immunotherapy treatment approach.

UR - https://www.scopus.com/pages/publications/85147361098

U2 - 10.1016/j.annonc.2022.11.008

DO - 10.1016/j.annonc.2022.11.008

M3 - Article

C2 - 36494005

AN - SCOPUS:85147361098

SN - 0923-7534

VL - 34

SP - 300

EP - 314

JO - Annals of Oncology

JF - Annals of Oncology

IS - 3

ER -

Identification, validation and biological characterisation of novel glioblastoma tumour microenvironment subtypes: implications for precision immunotherapy

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