Endothelial, pericyte and tumor cell expression in glioblastoma identifies fibroblast activation protein (FAP) as an excellent target for immunotherapy

Lisa M. Ebert, Wenbo Yu, Tessa Gargett, John Toubia, Paris M. Kollis, Melinda N. Tea, Brenton W. Ebert, Cedric Bardy, Mark van den Hurk, Claudine S. Bonder, Jim Manavis, Kathleen S. Ensbey, Mariana Oksdath Mansilla, Kaitlin G. Scheer, Sally L. Perrin, Rebecca J. Ormsby, Santosh Poonnoose, Barbara Koszyca, Stuart M. Pitson, Bryan W. DayGuillermo A. Gomez, Michael P. Brown

Research output: Contribution to journalArticlepeer-review

Abstract

Objectives: Targeted immunotherapies such as chimeric antigen receptor (CAR)-T cells are emerging as attractive treatment options for glioblastoma, but rely on identification of a suitable tumor antigen. We validated a new target antigen for glioblastoma, fibroblast activation protein (FAP), by undertaking a detailed expression study of human samples. Methods: Glioblastoma and normal tissues were assessed using immunostaining, supported by analyses of published transcriptomic datasets. Short-term cultures of glioma neural stem (GNS) cells were compared to cultures of healthy astrocytes and neurons using flow cytometry. Glioblastoma tissues were dissociated and analysed by high-parameter flow cytometry and single-cell transcriptomics (scRNAseq). Results: Compared to normal brain, FAP was overexpressed at the gene and protein level in a large percentage of glioblastoma tissues, with highest levels of expression associated with poorer prognosis. FAP was also overexpressed in several paediatric brain cancers. FAP was commonly expressed by cultured GNS cells but absent from normal neurons and astrocytes. Within glioblastoma tissues, the strongest expression of FAP was around blood vessels. In fact, almost every tumor vessel was highlighted by FAP expression, whereas normal tissue vessels and cultured endothelial cells (ECs) lacked expression. Single-cell analyses of dissociated tumors facilitated a detailed characterisation of the main cellular components of the glioblastoma microenvironment and revealed that vessel-localised FAP is because of expression on both ECs and pericytes. Conclusion: Fibroblast activation protein is expressed by multiple cell types within glioblastoma, highlighting it as an ideal immunotherapy antigen to target destruction of both tumor cells and their supporting vascular network.

Original languageEnglish
Article numbere1191
Number of pages24
JournalClinical and Translational Immunology
Volume9
Issue number10
DOIs
Publication statusPublished - 14 Oct 2020

Bibliographical note

Funding Information:
This work was supported by the Health Services Charitable Gifts Board (Adelaide) and a SA Cancer Council Beat Cancer Project Hospital Research Support Package (MB); the Neurosurgical Research Foundation (LE, MB, TG, MT, SP, GG); generous donations from the Sid Faithfull Family (KE, BD); a Senior Research Fellowship (1156693) from the National Health and Medical Research Council of Australia (SP); Flinders Foundation (CB); Cure Brain Cancer Foundation (CB, MB, GG, BD); Perpetual Impact Philanthropy (CB); the Brain Foundation (CB); Rebecca L. Cooper Foundation (CB); the Cancer Council SA Beat Cancer Project (to MPB and GAG); the Australian Research Council (FT160100366 to GAG); and a Netherlands Organisation for Scientific Research Rubicon Fellowship 019.163LW.032 (MvdH). SLP is supported by an Australian Government Research Training Program Scholarship. The authors would like to thank Samantha Escarbe and Nathalie Nataren for excellent technical assistance, Erica Yeo and Natansh Modi for assisting with the development of FAP immunostaining and Mark DeNichilo for HUVEC culture. We also acknowledge the support and generosity of the patients and medical and technical staff, including from SA Pathology and the SA Neurological Tumour Bank (supported by Flinders University, Flinders Foundation and The Neurosurgical Research Foundation), who made collection of tissue specimens possible.

Publisher Copyright:
© 2020 The Authors. Clinical & Translational Immunology published by John Wiley & Sons Australia, Ltd on behalf of Australian and New Zealand Society for Immunology, Inc.

Copyright:
Copyright 2020 Elsevier B.V., All rights reserved.

Keywords

  • blood vessels
  • fibroblast activation protein
  • glioblastoma
  • immunotherapy
  • scRNAseq
  • target antigen

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