A Non-Human Primate Model That Recapitulates B Cell-directed Chimeric Antigen Receptor (CAR) T cell-mediated Cytokine Release Syndrome (CRS) and Neurologic Toxicity
Agne Taraseviciute1, Leslie S. Kean1, Michael C. Jensen1
1Seattle Children’s Research Institute, Seattle, WA
CD19 Chimeric Antigen Receptor (CAR) T cell therapy has revolutionized the treatment for pediatric and adult patients with relapsed and refractory acute lymphoblastic leukemia (ALL). In the PLAT-02 trial at Seattle Children’s Hospital, treatment with CD19 CARs induced complete remissions in >90% of patients with refractory B-cell ALL. Although extremely successful, CD19 CAR T cell therapy is associated with significant and potentially life-threatening side effects, including cytokine release syndrome (CRS) and neurotoxicity, with symptoms ranging from fevers and hypotension to headaches, tremors, seizures and fatal cerebral edema. Our current understanding of CRS and neurotoxicity has been significantly limited by the lack of animal models that faithfully recapitulate these symptoms.
We created a non-human primate (NHP) model of B-cell-directed CAR T cell therapy targeting CD20 in Macaca mulatta, given the close similarity of rhesus macaque and human immune systems. Rhesus macaques (n=3) were treated with 30-40mg/kg cyclophosphamide followed 3-6 days later by an infusion of CAR T cells at a dose of 1x107 transduced cells/kg. Recipient animals were monitored for clinical signs and symptoms of CRS as well as neurotoxicity, and data were collected longitudinally to determine CAR T cell expansion and persistence, B cell aplasia, as well as clinical labs of CRS and cytokine levels. Infusion of autologous CD20 CAR T cells in 3 animals resulted in significant expansion of the CAR T cells with persistence for as long as 43 days post-infusion, which corresponded to concurrent B cell aplasia. CD20 CAR T cell recipients also developed clinical signs and symptoms of CRS as well as neurotoxicity which was manifested by decreased activity, extremity tremors and gait abnormalities beginning between days 5 to 7 following CAR T cell infusion, with the onset of clinical symptoms coinciding with maximum CAR T cell expansion and activation. These neurologic symptoms were responsive to treatment with the anti-epileptic medication levetiracetam. The clinical syndrome was accompanied by elevations in CRP, Ferritin, LDH and serum cytokines, including IL-6, IL-8 and ITAC, recapitulating data from patient clinical trials using CD19 CAR T cells. A drastic expansion of CD20 CAR T cells on day 7 following infusion was also observed in the CSF in the animals, and coincided with the onset of neurotoxicity. Strikingly, we also detected CD20 CAR T cells in multiple regions of the brain via flow cytometry, including the frontal, parietal, occipital lobes and cerebellum, and demonstrated an increased number of infiltrating T cells by immunofluorescence in the brains of animals treated with CD20 CAR T cells when compared to healthy controls.
These data demonstrate the successful establishment of a large animal model of B-cell directed CAR T cell therapy that recapitulates the most significant and potentially life threatening toxicities of CAR T cell therapy, including CRS and neurotoxicity. This model will be instrumental in elucidating the mechanisms driving clinical toxicities and will allow for pre-clinical evaluation of potentially novel and targeted therapies designed to prevent or abort such toxicities.