GFAP and NfL increase during neurotoxicity from high baseline levels in pediatric CD19-CAR T-cell patients.

There is a need for biomarkers to predict and measure the severity of immune effector cell-associated neurotoxicity syndrome (ICANS). Glial fibrillary acidic protein (GFAP) and neurofilament light chain (NfL) are well-validated biomarkers of astroglial and neuronal injury, respectively. We hypothesized that pretreatment GFAP and NfL levels can predict the risk of subsequent ICANS and that increases in GFAP and NfL levels during treatment reflect ICANS severity. We measured cerebrospinal fluid GFAP (cGFAP) and NfL (cNfL) along with serum NfL (sNfL) levels at pretreatment and day 7 to 10 after chimeric antigen receptor (CAR) T-cell infusion in 3 pediatric cohorts treated with CD19- or CD19/CD22-directed CAR T cells. cGFAP and cNfL levels increased during grade ≥1 ICANS in patients treated with CD19-directed CAR T cells but not in those who received CD19/CD22-directed CAR T cells. The sNfL levels did not increase during ICANS. Prelymphodepletion cGFAP, cNfL, and sNfL levels were not predictive of subsequent ICANS. Elevated baseline cGFAP levels were associated with a history of transplantation. Patients with prior central nervous system (CNS) radiation had higher cNfL levels, and elevated baseline sNfL levels were associated with a history of peripheral neuropathy. Thus, cGFAP and cNfL may be useful biomarkers for measuring the severity of CNS injury during ICANS in children. Elevated baseline levels of cGFAP, cNfL, and sNfL likely reflect the cumulative injury to the central and peripheral nervous systems from prior treatment. However, levels of any of the 3 biomarkers before CAR T-cell infusion did not predict the risk of ICANS.

EEG Correlates of Delirium in Children and Young Adults With CD19-Directed CAR T Cell Treatment-Related Neurotoxicity.

INTRODUCTION: EEG patterns in chimeric antigen receptor T cell treatment-associated neurotoxicity (immune effector cell-associated neurotoxicity syndrome) have not yet been systematically studied. We tested the hypothesis that EEG background abnormalities in immune effector cell-associated neurotoxicity syndrome correlate with clinical signs of neurotoxicity. In addition, we describe ictal and interictal EEG patterns to better understand the natural history of immune effector cell-associated neurotoxicity syndrome-associated seizures. METHODS: EEGs were obtained in 19 of 100 subjects in a prospective cohort study of children and young adults undergoing CD19-directed chimeric antigen receptor T cell therapy. We classified the EEG background on a severity scale of 0 to 5 during 30-minute epochs. EEG grades were compared with neurotoxicity scored by Common Terminology Criteria for Adverse Events and Cornell Assessment of Pediatric Delirium scores. Descriptive analysis was conducted for ictal and interictal EEG abnormalities. RESULTS: EEG background abnormality scores correlated well with Common Terminology Criteria for Adverse Events neurotoxicity scores (P = 0.0022) and Cornell Assessment of Pediatric Delirium scores (P = 0.0085). EEG was better able to differentiate the severity of coma patterns compared with the clinical scores. The EEG captured electroclinical seizures in 4 of 19 subjects, 3 of whom had additional electrographic-only seizures. Seizures most often arose from posterior head regions. Interictal epileptiform discharges were focal, multifocal, or lateralized periodic discharges. No seizures or interictal epileptiform abnormalities were seen in subjects without previous clinical seizures. CONCLUSIONS: Continuous EEG monitoring is high yield for seizure detection in high-risk chimeric antigen receptor T cell patients, and electrographic-only seizures are common. Increasing severity of EEG background abnormalities correlates with increasing neurotoxicity grade.

Knock-in of the Wt1 R394W mutation causes MDS and cooperates with Flt3/ITD to drive aggressive myeloid neoplasms in mice.

Wilms tumor 1 (WT1) is a zinc finger transcriptional regulator, and has been implicated as both a tumor suppressor and oncogene in various malignancies. Mutations in the DNA-binding domain of the WT1 gene are described in 10-15% of normal-karyotype AML (NK-AML) in pediatric and adult patients. Similar WT1 mutations have been reported in adult patients with myelodysplastic syndrome (MDS). WT1 mutations have been independently associated with treatment failure and poor prognosis in NK-AML. Internal tandem duplication (ITD) mutations of FMS-like tyrosine kinase 3 (FLT3) commonly co-occur with WT1-mutant AML, suggesting a cooperative role in leukemogenesis. The functional role of WT1 mutations in hematologic malignancies appears to be complex and is not yet fully elucidated. Here, we describe the hematologic phenotype of a knock-in mouse model of a Wt1 mutation (R394W), described in cases of human leukemia. We show that Wt1 +/R394W mice develop MDS which becomes 100% penetrant in a transplant model, exhibit an aberrant expansion of myeloid progenitor cells, and demonstrate enhanced self-renewal of hematopoietic progenitor cells in vitro. We crossbred Wt1 +/R394W mice with knock-in Flt3 +/ITD mice, and show that mice with both mutations (Flt3 +/ITD/Wt1 +/R394W) develop a transplantable MDS/MPN, with more aggressive features compared to either single mutant mouse model.

The Evolution and Future of CAR T Cells for B-Cell Acute Lymphoblastic Leukemia.

Several CAR T designs with CD19 specificity have been associated with consistent responses in clinical trials with complete remission (CR) rates ranging from 70-90%. Relevant challenges remain to be addressed, such as production time, early loss of CAR T cells, relapse due to loss of the target antigen, and prevention of severe cytokine release syndrome and neurotoxicity. This review describes constructs, clinical trial results, side effects, and future direction of CAR T-cell therapy in B-ALL.

POL5551, a novel and potent CXCR4 antagonist, enhances sensitivity to chemotherapy in pediatric ALL.

The importance of the cell surface receptor CXCR4 and the chemokine stromal cell-derived factor-1 (SDF-1/CXCL12) is well-established in normal and malignant hematopoiesis. The Protein Epitope Mimetic POL5551 is a novel and potent antagonist of CXCR4. POL5551 efficiently mobilizes hematopoietic stem and progenitor cells, but its effects in acute lymphoblastic leukemia (ALL) have not been reported. Here, we demonstrate that POL5551 is a potent antagonist of CXCR4 in pre-B and T cell ALL cell lines and pediatric ALL primary samples. POL5551 has activity at nanomolar concentrations in decreasing CXCR4 antibody binding, blocking SDF-1α-mediated phosphorylation of ERK1/2, inhibiting SDF-1α-induced chemotaxis, and reversing stromal-mediated protection from chemotherapy. POL5551 is significantly more effective at inhibiting CXCR4 antibody binding than the FDA-approved CXCR4 inhibitor plerixafor in ALL cell lines and primary samples. We also show that treatment with POL5551 in vitro and cytarabine +/- POL5551 in vivo modulates surface expression of adhesion molecules, findings that may guide the optimal clinical use of POL5551. Finally, we demonstrate that POL5551 increases sensitivity to cytarabine in a xenograft model of a high-risk pediatric ALL, infant MLL-rearranged (MLL-R) ALL. Therefore, disruption of the CXCR4/SDF-1 axis with POL5551 may improve outcomes in children with high-risk ALL.

Novel agents for the treatment of childhood acute leukemia.

Together, acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML) make up approximately one-third of all pediatric cancer diagnoses. Despite remarkable improvement in the treatment outcomes of these diseases over the past several decades, the prognosis for certain high-risk groups of leukemia and for relapsed disease remains poor. However, recent insights into different types of 'driver' lesions of leukemogenesis, such as the aberrant activation of signaling pathways and various epigenetic modifications, have led to the discovery of novel agents that specifically target the mechanism of transformation. In parallel, emerging approaches in cancer immunotherapy have led to newer therapies that can exploit and harness cytotoxic immunity directed against malignant cells. This review details the rationale and implementation of recent and specifically targeted therapies in acute pediatric leukemia. Topics covered include the inhibition of critical cell signaling pathways [BCR-ABL, FMS-like tyrosine kinase 3 (FLT3), mammalian target of rapamycin (mTOR), and Janus-associated kinase (JAK)], proteasome inhibition, inhibition of epigenetic regulators of gene expression [DNA methyltransferase (DNMT) inhibitors, histone deacetylase (HDAC) inhibitors, and disruptor of telomeric signaling-1 (DOT1L) inhibitors], monoclonal antibodies and immunoconjugated toxins, bispecific T-cell engaging (BiTE) antibodies, and chimeric antigen receptor-modified (CAR) T cells.

Plerixafor as a chemosensitizing agent in pediatric acute lymphoblastic leukemia: efficacy and potential mechanisms of resistance to CXCR4 inhibition.

In spite of advances in the treatment of pediatric acute lymphoblastic leukemia (ALL), a significant number of children with ALL are not cured of their disease. We and others have shown that signaling from the bone marrow microenvironment confers therapeutic resistance, and that the interaction between CXCR4 and stromal cell-derived factor-1 (SDF-1 or CXCL12) is a key mediator of this effect. We demonstrate that ALL cells that upregulate surface CXCR4 in response to chemotherapy treatment are protected from chemotherapy-induced apoptosis when co-cultured with bone marrow stroma. Treatment with the CXCR4 inhibitor plerixafor diminishes stromal protection and confers chemosensitivity. Using xenograft models of high-risk pediatric ALL, plerixafor plus chemotherapy induces significantly decreased leukemic burden, compared to chemotherapy alone. Further, treatment with plerixafor and chemotherapy influences surface expression of CXCR4, VLA-4, and CXCR7 in surviving ALL blasts. Finally, prolonged exposure of ALL blasts to plerixafor leads to a persistent increase in surface CXCR4 expression, along with modulation of surface expression of additional adhesion molecules, and enhanced SDF-1α-induced chemotaxis, findings that may have implications for therapeutic resistance. Our results suggest that while CXCR4 inhibition may prove useful in ALL, further study is needed to understand the full effects of targeting the leukemic microenvironment.

The Biology and Targeting of FLT3 in Pediatric Leukemia.

Despite remarkable improvement in treatment outcomes in pediatric leukemia over the past several decades, the prognosis for high-risk groups of acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL), as well as for relapsed leukemia, remains poor. Intensification of chemotherapy regimens for those at highest risk has improved success rates, but at the cost of significantly increased morbidity and long-term adverse effects. With the success of imatinib in Philadelphia-chromosome-positive leukemia and all-trans retinoic acid in acute promyelocytic leukemia, the quest to find additional molecularly targeted therapies has generated much excitement over recent years. Another such possible target in pediatric acute leukemia is FMS-like tyrosine kinase 3 (FLT3). FLT3 aberrations are among the most frequently identified transforming events in AML, and have significant clinical implications in both high-risk pediatric AML and in certain high-risk groups of pediatric ALL. Therefore, the successful targeting of FLT3 has tremendous potential to improve outcomes in these subsets of patients. This article will give an overview of the molecular function and signaling of the FLT3 receptor, as well as its pathogenic role in leukemia. We review the discovery of targeting FLT3, discuss currently available FLT3 inhibitors in pediatric leukemia and results of clinical trials to date, and finally, consider the future promise and challenges of FLT3 inhibitor therapy.