Foltz, Jennifer A.

Abstract Purpose: Standard care for high-grade gliomas (HGG) involves maximal surgical resection followed by radiation and temozolomide. Postoperative adjuvant therapy frequently causes lymphopenia, which is associated with poor prognosis. Interleukin 7 (IL7) is essential for lymphocyte development, homeostasis, and survival. NT-I7 (efineptakin alfa), a long-acting recombinant IL7, reverses lymphopenia and improves survival in murine glioma models. However, the safety, maximum tolerated dose (MTD), and impact of NT-I7 on immune cells in patients with HGG remain unknown. Patients and Methods: We conducted a phase I trial (NCT03687957) examining the MTD and effect of NT-I7 on lymphocytes in patients with newly diagnosed HGG. The primary endpoint was dose-limiting toxicity; secondary endpoints included absolute lymphocyte count (ALC) changes over time, overall response, progression-free survival, and overall survival. Exploratory endpoints included immune profiling at different time points using single-cell RNA sequencing (scRNA-seq) in a subset of patients. Results: NT-I7 was well tolerated with a MTD of 720 µg/kg. Moreover, NT-I7 significantly increased ALCs for more than 12 weeks in duration. Early elevations in CD4+, CD8+ T cells and NK cells further coincided with increased TNF and CXCL9 cytokine levels. Comprehensive immune profiling of peripheral blood T cells revealed selective clonotype expansion within CD8+, but not CD4+, T cells following NT-I7 administration. Finally, a subset of our patients with MGMT promoter–unmethylated glioblastoma, which are typically associated with a poorer prognosis, demonstrated promising clinical responses. Conclusions: NT-I7 has the potential to maintain and increase lymphocyte counts in patients with HGG and warrants further investigation, particularly in combination with immune-based therapies.

Abstract Lymphopenia following chemoradiation for high grade gliomas (HGG) is common and is associated with decreased survival. NT-I7 (efineptakin alfa) is a long-acting analog of the lymphocyte pro-survival cytokine, interleukin-7. Our Phase I dose-escalation study of 19 patients with HGG demonstrated early elevations in CD4+ and CD8+ T cells in the first 4 weeks after administration of NT-I7. In this study, we explored the diversity and potential functionality of the NT-I7-expanded T cell population. A subset of 4 subjects with glioblastoma multiforme (GBM) treated at 720 µg/kg of NT-I7 during adjuvant temozolomide in our study (NCT03687957) underwent Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-seq) of peripheral blood. CITE-seq with TCR clonotype sequencing (10x Genomics 5’) permitted simultaneous tracking of the T cell phenotype at the protein level, individual T cell clones, and gene expression dynamics between 4 timepoints: pre-treatment, and post-treatment weeks 1, 2, and 13. Here, we report the results of differential expression analyses between timepoints and patient-specific differences in T cell clonotype expansions. Both CD4+ and CD8+ T cells in our study patients had increased expression of T cell activation markers LTB, CISH, and CD74. CD8+ T cell subsets demonstrated additional elevations in genes important in cytotoxic function (granzymes (GZMA, GZMB) and perforin (PRF1)), MHC Class II expression (HLA-DRA, HLA-DRB1), and AP-1 transcription factor subunits (JUND, JUN, JUNB, FOS, FOSB). After excluding known viral antigen clonotypes, 2 of 4 patients demonstrated TCR clonotype expansion. Notably, nearly all the expanded TCR clonotypes were found within CD8+ T cells, but not CD4+ T cells. NT-I7 promotes activation, cytotoxic function, and selective clonotype expansion of CD8+ T cells in patients with HGG. This, combined with its role in increased CD4+ T cell counts, suggests NT-I7 may be a powerful tool to restore immune function after treatment-associated lymphopenia in HGG.