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Regular and Young Investigator Award Abstracts
Cellular Therapies
323 Development of nuclear envelope protein-targeting CAR T cells for treatment of hyperproliferative MYC-driven group 3 medulloblastoma
  1. Yujin Suk1,
  2. Martin Rossotti2,
  3. Jorge Ibanez3,
  4. Iqra Chaudhry1,
  5. Erika Apel1,
  6. Minomi Subapanditha1,
  7. Petar Miletic1,
  8. Stefan Custers1,
  9. Laura Escudero1,
  10. Alberto Delaidelli4,
  11. Yiyun Chen5,
  12. Takuma Nakashima6,
  13. Yuxi Xiao7,
  14. Jason Moffat7,
  15. Hiromichi Suzuki6,
  16. Poul Sorensen8,
  17. Kevin Henry2,
  18. Elena Sotillo9,
  19. Chitra Venugopal1,
  20. Crystal L Mackall10,
  21. Giedre Krenciute3 and
  22. Sheila K Singh1
  1. 1McMaster University, Hamilton, ON, Canada
  2. 2National Research Council Canada, Ottawa, ON, Canada
  3. 3St. Jude Children’s Research Hospital, Memphis, TN, USA
  4. 4University of British Columbia, Vancouver, BC, Canada
  5. 5Stanford Center for Cancer Cell Therapy, Palo Alto, CA, USA
  6. 6National Cancer Center Research Institute (Japan), Tokyo, Japan
  7. 7University of Toronto, Toronto, ON, Canada
  8. 8BC Cancer Research Centre, Vancouver, BC, Canada
  9. 9Stanford University School of Medicine, Stanford, CA, USA
  10. 10Stanford University, Palo Alto, CA, USA

  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.

Abstract

Background Medulloblastoma (MB) is the most common, malignant pediatric brain tumour comprised of four distinct molecular subgroups (WNT, SHH, Group 3, and Group 4). A subset of Group 3 MB tumors harbor focal amplifications of the MYC oncogene (MYC-G3MB) and are particularly prone to tumour recurrence and leptomeningeal spread due to their highly proliferative nature linked to its embryonic stem cell-of-origin and programing. Therefore, there is an urgent need for the development of therapeutic modalities that are safe and have potent antitumour efficacy against recurrent MB.

Methods We discovered that LBR and TMPO, while present in all cells at the NE, is aberrantly presented to the cell surface in MYC-G3MB cell lines. Flow cytometry analysis identified highly abundant cell surface expression in all MYC-G3MB cell lines with low to no expression in human neural stem cells and other MB subgroups. Transcriptomic profiling of identifies significant NE protein enrichment in MB compared to normal tissue and with high expression during fetal development that dampens postnatally. Immunohistochemistry analysis of LBR on MB patient samples identified that positive LBR staining correlated with a significantly worse prognosis and was also identified to be enriched in recurrent tissue compared to their matched primary samples.

Results Mechanistically, high resolution microscopy of endogenously tagged HALO-LBR and mNeon-TMPO reveals cell surface presentation to be linked to ER or ER-like vesicles that are trafficked to the cell surface. Transcriptomic analysis of LBR cell surface positive and negative cells identify that cell surface positivity is significantly linked to cell division and mitotic processes. LBR mislocalization is enriched following exposure to chemotherapy and radiation in vitro and in vivo and may act as a marker for the brain tumor initiating cells that seed recurrence.

We then generated single domain antibodies (sdAb) to develop LBR and TMPO-specific CAR T cells. Validation using LBR KO MYC-G3MB cells show LBR sdAbs to have on-target binding and comparable affinity and avidity to commercial antibodies. Preliminary in vitro cytotoxicity assays of LBR-CAR T cell therapy reveal potent antitumour efficacy and CAR T cell activation following co-culture with of MYC-G3MB cells.

Conclusions There is an undeniable paucity of recurrent tumor-specific targets for MB patients stemming from the lack of neoantigens and genomic aberrations in pediatric tumors. The preclinical development and validation of novel immunotherapeutic targets such as LBR for treatment of therapy-resistant, hyperproliferative MYC-G3MB provides a novel option for patients who otherwise face palliation (figure 1).

Abstract 323 Figure 1
Abstract 323 Figure 1

Graphical abstract

http://creativecommons.org/licenses/by-nc/4.0/

This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See http://creativecommons.org/licenses/by-nc/4.0/.

https://doi.org/10.1136/jitc-2024-SITC2024.0323

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