Potent antitumor efficacy of anti-GD2 CAR T cells in H3-K27M+ diffuse midline gliomas

Abstract

Diffuse intrinsic pontine glioma (DIPG) and other diffuse midline gliomas (DMGs) with mutated histone H3 K27M (H3-K27M)1,2,3,4,5 are aggressive and universally fatal pediatric brain cancers6. Chimeric antigen receptor (CAR)-expressing T cells have mediated impressive clinical activity in B cell malignancies7,8,9,10, and recent results suggest benefit in central nervous system malignancies11,12,13. Here, we report that patient-derived H3-K27M-mutant glioma cell cultures exhibit uniform, high expression of the disialoganglioside GD2. Anti-GD2 CAR T cells incorporating a 4-1BBz costimulatory domain14 demonstrated robust antigen-dependent cytokine generation and killing of DMG cells in vitro. In five independent patient-derived H3-K27M+ DMG orthotopic xenograft models, systemic administration of GD2-targeted CAR T cells cleared engrafted tumors except for a small number of residual GD2lo glioma cells. To date, GD2-targeted CAR T cells have been well tolerated in clinical trials15,16,17. Although GD2-targeted CAR T cell administration was tolerated in the majority of mice bearing orthotopic xenografts, peritumoral neuroinflammation during the acute phase of antitumor activity resulted in hydrocephalus that was lethal in a fraction of animals. Given the precarious neuroanatomical location of midline gliomas, careful monitoring and aggressive neurointensive care management will be required for human translation. With a cautious multidisciplinary clinical approach, GD2-targeted CAR T cell therapy for H3-K27M+ diffuse gliomas of pons, thalamus and spinal cord could prove transformative for these lethal childhood cancers.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: GD2 is a target for immunotherapy in DIPG.
Fig. 2: GD2-CAR T cells mediate a potent and lasting antitumor response in DIPG orthotopic xenografts.
Fig. 3: GD2-CAR T cell therapy improves survival in mice with DIPG orthotopic xenografts.
Fig. 4: GD2 CAR T cell therapy effectively clears other midline H3-K27M-mutant pediatric DMGs but is associated with toxicity in thalamic xenografts.

References

  1. 1.

    Paugh, B. S. et al. Novel oncogenic PDGFRA mutations in pediatric high-grade gliomas. Cancer Res. 73, 6219–6229 (2013).

    CAS  Article  Google Scholar 

  2. 2.

    Wu, G. et al. Somatic histone H3 alterations in pediatric diffuse intrinsic pontine gliomas and non-brainstem glioblastomas. Nat. Genet. 44, 251–253 (2012).

    CAS  Article  Google Scholar 

  3. 3.

    Buczkowicz, P. et al. Genomic analysis of diffuse intrinsic pontine gliomas identifies three molecular subgroups and recurrent activating ACVR1 mutations. Nat. Genet. 46, 451–456 (2014).

    CAS  Article  Google Scholar 

  4. 4.

    Taylor, K. R. et al. Recurrent activating ACVR1 mutations in diffuse intrinsic pontine glioma. Nat. Genet. 46, 457–461 (2014).

    CAS  Article  Google Scholar 

  5. 5.

    Wu, G. et al. The genomic landscape of diffuse intrinsic pontine glioma and pediatric non-brainstem high-grade glioma. Nat. Genet. 46, 444–450 (2014).

    CAS  Article  Google Scholar 

  6. 6.

    Jones, C. et al. Pediatric high-grade glioma: biologically and clinically in need of new thinking. Neuro Oncol. 19, 153–161 (2016).

    PubMed Central  Google Scholar 

  7. 7.

    Lee, D. W. et al. T cells expressing CD19 chimeric antigen receptors for acute lymphoblastic leukaemia in children and young adults: a phase 1 dose-escalation trial. Lancet 385, 517–528 (2015).

    CAS  Article  Google Scholar 

  8. 8.

    Davila, M. L. et al. Efficacy and toxicity management of 19-28z CAR T cell therapy in B cell acute lymphoblastic leukemia. Sci. Transl. Med. 6, 224ra25 (2014).

    Article  Google Scholar 

  9. 9.

    Maude, S. L. et al. Chimeric antigen receptor T cells for sustained remissions in leukemia. N. Engl. J. Med. 371, 1507–1517 (2014).

    Article  Google Scholar 

  10. 10.

    Gardner, R. A. et al. Intent-to-treat leukemia remission by CD19 CAR T cells of defined formulation and dose in children and young adults. Blood 129, 3322–3331 (2017).

    CAS  Article  Google Scholar 

  11. 11.

    Brown, C. E. et al. Regression of glioblastoma after chimeric antigen receptor T-cell therapy. N. Engl. J. Med. 375, 2561–2569 (2016).

    CAS  Article  Google Scholar 

  12. 12.

    Ahmed, N. et al. HER2-specific chimeric antigen receptor-modified virus-specific T cells for progressive glioblastoma: a phase 1 dose-escalation Trial. JAMA Oncol. 3, 1094–1101 (2017).

    Article  Google Scholar 

  13. 13.

    O’Rourke, D.M. et al. A single dose of peripherally infused EGFRvIII-directed CAR T cells mediates antigen loss and induces adaptive resistance in patients with recurrent glioblastoma. Sci. Transl. Med. 9, eaaa0984 (2017).

    Article  Google Scholar 

  14. 14.

    Long, A. H. et al. 4-1BB costimulation ameliorates T cell exhaustion induced by tonic signaling of chimeric antigen receptors. Nat. Med. 21, 581–590 (2015).

    CAS  Article  Google Scholar 

  15. 15.

    Pule, M. A. et al. Virus-specific T cells engineered to coexpress tumor-specific receptors: persistence and antitumor activity in individuals with neuroblastoma. Nat. Med. 14, 1264–1270 (2008).

    CAS  Article  Google Scholar 

  16. 16.

    Louis, C. U. et al. Antitumor activity and long-term fate of chimeric antigen receptor-positive T cells in patients with neuroblastoma. Blood 118, 6050–6056 (2011).

    CAS  Article  Google Scholar 

  17. 17.

    Heczey, A. et al. CAR T cells administered in combination with lymphodepletion and PD-1 inhibition to patients with neuroblastoma. Mol. Ther. 25, 2214–2224 (2017).

    CAS  Article  Google Scholar 

  18. 18.

    Majzner, R. G., Heitzeneder, S. & Mackall, C. L. Harnessing the immunotherapy revolution for the treatment of childhood cancers. Cancer Cell 31, 476–485 (2017).

    CAS  Article  Google Scholar 

  19. 19.

    Khuong-Quang, D. A. et al. K27M mutation in histone H3.3 defines clinically and biologically distinct subgroups of pediatric diffuse intrinsic pontine gliomas. Acta Neuropathol. 124, 439–447 (2012).

    CAS  Article  Google Scholar 

  20. 20.

    Schwartzentruber, J. et al. Driver mutations in histone H3.3 and chromatin remodelling genes in paediatric glioblastoma. Nature 482, 226–231 (2012).

    CAS  Article  Google Scholar 

  21. 21.

    Thomas, S., Straathof, K., Himoudi, N., Anderson, J. & Pule, M. An optimized GD2-targeting retroviral cassette for more potent and safer cellular therapy of neuroblastoma and other cancers. PLoS One 11, e0152196 (2016).

    Article  Google Scholar 

  22. 22.

    Long, A. H. et al. Reduction of MDSCs with all-trans retinoic acid improves CAR therapy efficacy for sarcomas. Cancer Immunol. Res. 4, 869–880 (2016).

    CAS  Article  Google Scholar 

  23. 23.

    Yu, A. L. et al. Anti-GD2 antibody with GM-CSF, interleukin-2, and isotretinoin for neuroblastoma. N. Engl. J. Med. 363, 1324–1334 (2010).

    CAS  Article  Google Scholar 

  24. 24.

    Perez Horta, Z., Goldberg, J. L. & Sondel, P. M. Anti-GD2 mAbs and next-generation mAb-based agents for cancer therapy. Immunotherapy 8, 1097–1117 (2016).

    Article  Google Scholar 

  25. 25.

    Hong, J. J. et al. Successful treatment of melanoma brain metastases with adoptive cell therapy. Clin. Cancer Res. 16, 4892–4898 (2010).

    CAS  Article  Google Scholar 

  26. 26.

    Suzuki, K. The pattern of mammalian brain gangliosides. II. Evaluation of the extraction procedures, postmortem changes and the effect of formalin preservation. J. Neurochem. 12, 629–638 (1965).

    CAS  Article  Google Scholar 

  27. 27.

    Kramer, K. et al. Compartmental intrathecal radioimmunotherapy: results for treatment for metastatic CNS neuroblastoma. J. Neurooncol. 97, 409–418 (2010).

    Article  Google Scholar 

  28. 28.

    Kramer, K. et al. A phase II study of radioimmunotherapy with intraventricular 131 I-3F8 for medulloblastoma. Pediatr. Blood Cancer 65, e26754 (2018).

    Article  Google Scholar 

  29. 29.

    Monje, M. et al. Hedgehog-responsive candidate cell of origin for diffuse intrinsic pontine glioma. Proc. Natl Acad. Sci. USA 108, 4453–4458 (2011).

    CAS  Article  Google Scholar 

  30. 30.

    Qin, E. Y. et al. Neural precursor-derived pleiotrophin mediates subventricular zone invasion by glioma. Cell 170, 845–859 (2017).

    CAS  Article  Google Scholar 

  31. 31.

    Grasso, C. S. et al. Functionally defined therapeutic targets in diffuse intrinsic pontine glioma. Nat. Med. 21, 555–559 (2015).

    CAS  Article  Google Scholar 

  32. 32.

    Fry, T. J. et al. CD22-targeted CAR T cells induce remission in B-ALL that is naive or resistant to CD19-targeted CAR immunotherapy. Nat. Med. 24, 20–28 (2018).

    CAS  Article  Google Scholar 

  33. 33.

    Walker, A. J. et al. Tumor antigen and receptor densities regulate efficacy of a chimeric antigen receptor targeting anaplastic lymphoma kinase. Mol. Ther. 25, 2189–2201 (2017).

    CAS  Article  Google Scholar 

  34. 34.

    Ali, N. et al. Xenogeneic graft-versus-host-disease in NOD-scid IL-2Rγnull mice display a T-effector memory phenotype. PLoS One 7, e44219 (2012).

    CAS  Article  Google Scholar 

  35. 35.

    Nagaraja, S. et al. Transcriptional dependencies in diffuse intrinsic pontine glioma. Cancer Cell 31, 635–652 (2017).

    CAS  Article  Google Scholar 

  36. 36.

    Louveau, A. et al. Structural and functional features of central nervous system lymphatic vessels. Nature 523, 337–341 (2015).

    CAS  Article  Google Scholar 

  37. 37.

    Louis, D. N. et al. The 2016 World Health Organization classification of tumors of the central nervous system: a summary. Acta Neuropathol. 131, 803–820 (2016).

    Article  Google Scholar 

  38. 38.

    Wolchok, J. D. et al. Guidelines for the evaluation of immune therapy activity in solid tumors: immune-related response criteria. Clin. Cancer Res. 15, 7412–7420 (2009).

    CAS  Article  Google Scholar 

  39. 39.

    Gust, J. et al. Endothelial activation and blood–brain barrier disruption in neurotoxicity after adoptive immunotherapy with CD19 CAR-T cells. Cancer Discov. 7, 1404–1419 (2017).

    CAS  Article  Google Scholar 

  40. 40.

    Lin, G. L. & Monje, M. A protocol for rapid post-mortem cell culture of diffuse intrinsic pontine glioma (DIPG). J. Vis. Exp. 7, e55360 (2017).

    Google Scholar 

  41. 41.

    Lynn, R. C. et al. Targeting of folate receptor β on acute myeloid leukemia blasts with chimeric antigen receptor-expressing T cells. Blood 125, 3466–3476 (2015).

    CAS  Article  Google Scholar 

  42. 42.

    Haso, W. et al. Anti-CD22-chimeric antigen receptors targeting B-cell precursor acute lymphoblastic leukemia. Blood 121, 1165–1174 (2013).

    CAS  Article  Google Scholar 

  43. 43.

    Sen, G., Chakraborty, M., Foon, K. A., Reisfeld, R. A. & Bhattacharya-Chatterjee, M. B. Induction of IgG antibodies by an anti-idiotype antibody mimicking disialoganglioside GD2. J. Immunother. 21, 75–83 (1998).

    CAS  Article  Google Scholar 

  44. 44.

    Jena, B. et al. Chimeric antigen receptor (CAR)-specific monoclonal antibody to detect CD19-specific T cells in clinical trials. PLoS One 8, e57838 (2013).

    CAS  Article  Google Scholar 

  45. 45.

    Hendel, A. et al. Chemically modified guide RNAs enhance CRISPR-Cas genome editing in human primary cells. Nat. Biotechnol. 33, 985–989 (2015).

    CAS  Article  Google Scholar 

  46. 46.

    Chu, V. T. et al. Increasing the efficiency of homology-directed repair for CRISPR–Cas9-induced precise gene editing in mammalian cells. Nat. Biotechnol. 33, 543–548 (2015).

    CAS  Article  Google Scholar 

  47. 47.

    Brinkman, E. K., Chen, T., Amendola, M. & van Steensel, B. Easy quantitative assessment of genome editing by sequence trace decomposition. Nucleic Acids Res. 42, e168 (2014).

    Article  Google Scholar 

Download references

Acknowledgements

We thank the following for generously providing cell cultures: A. Moore (University of Queensland) and C. Jones (Institute of Cancer Research) for QCTB R059, R. Seeger (Children's Hospital Los Angeles) for CHLA136, 255, C. Khanna (National Cancer Institute) for MG63-3 and L. Helman (Children's Hospital Los Angeles) for EW8 and TC32.

This work was supported by a Stand Up To Cancer–St. Baldrick’s–National Cancer Institute Pediatric Dream Team Translational Cancer Research Grant (C.L.M.). Stand Up To Cancer is a program of the Entertainment Industry Foundation administered by the American Association for Cancer Research. C.L.M is a member of the Parker Institute for Cancer Immunotherapy, which supports the Stanford University Cancer Immunotherapy Program. The authors gratefully acknowledge support from the National Institute of Neurological Disorders and Stroke (F31NS098554 to C.W.M. and R01NS092597 to M.M.), Abbie’s Army Foundation (M.M.), Unravel Pediatric Cancer (M.M.), Maiy’s Miracle Foundation (E.P.A.), Stella S. Jones Foundation (M.M.), McKenna Claire Foundation (M.M.), Alex’s Lemonade Stand Foundation (M.M.), Izzy's Infantry Foundation (M.M.), The Cure Starts Now Foundation and DIPG Collaborative (M.M.), Lyla Nsouli Foundation (M.M.), Declan Gloster Memorial Fund (M.M.), N8 Foundation (M.M.), Fly a Kite Foundation (M.M.), Liwei Wang Research Fund (M.M.), Virginia and D.K. Ludwig Fund for Cancer Research (M.M. and C.L.M.), Sam Jeffers Foundation (M.M.), Michael Mosier DEFEAT DIPG Foundation (M.M.), ChadTough Foundation (M.M.), Reller Family Research Fund, Child Health Research Institute at Stanford and SPARK program (M.M.) and the Anne T. and Robert M. Bass Endowed Faculty Scholarship in Pediatric Cancer and Blood Diseases (M.M.).

Author information

Affiliations

Authors

Contributions

C.W.M. and E.P.A. performed the antibody array screening. C.W.M. and M.M. identified GD2 as a target in H3-K27M+ DMGs. C.W.M. and S.S. performed immunohistochemistry and immunofluorescence microscopy on primary and xenograft tissue. L.L. and R.G.M. designed CAR constructs. R.G.M. and M.K. prepared CAR T cells for in vivo experiments. C.W.M. and P.J.W. conducted in vivo experiments. R.G.M., M.K. and S.P.R. performed in vitro T cell experiments and flow cytometry. E.H. contributed VUMC-DIPG10 and data on ganglioside synthesis pathway expression. H.V. performed neuropathological review of brain tissue. S.H. performed CRISPR–Cas9-mediated gene editing. C.W.M., R.G.M., H.V., M.M. and C.L.M. contributed to data analysis and interpretation. C.W.M., M.M., R.G.M. and C.L.M wrote the manuscript. C.W.M. and R.G.M. made the figures. M.M. and C.L.M. supervised all aspects of the work.

Corresponding authors

Correspondence to Michelle Monje or Crystal L. Mackall.

Ethics declarations

Competing interests

C.L.M, M.M., R.G.M. and C.W.M. are inventors on a patent application for GD2-directed CAR use for H3-K27M DMG.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–10 and Supplementary Tables 2 and 3

Reporting Summary

Supplementary Table 1

Surface Panel Flow Cytometry Screening Data

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Mount, C.W., Majzner, R.G., Sundaresh, S. et al. Potent antitumor efficacy of anti-GD2 CAR T cells in H3-K27M+ diffuse midline gliomas. Nat Med 24, 572–579 (2018). https://doi.org/10.1038/s41591-018-0006-x

Download citation

Further reading

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing