Dex and IL-7 enhance persistence of CAR T cells to promote long-term remission in mouse models of multiple myeloma

Abstract

Abstract Introduction: Chimeric antigen receptor (CAR) T cells have had phenomenal response rates in hematological malignancies, including >80% complete response in multiple myeloma (MM). However, ~50% of MM patients who responded to CAR T cells will relapse within the first year, which is attributed to poor CAR T cell persistence. An emerging method to enhance persistence is the leveraging of interleukin 7 receptor (IL-7R) signaling. Although poised to be influential, this method may be limited by the highly variable IL-7R expression on CAR T cells; ~30-90% and ~0-50% IL-7R from healthy donors and patients, respectively. One method to overcome this limitation is to overexpress constitutively active IL-7R, yet the potential for oncogenesis limits its clinical viability. We previously found that dexamethasone (dex) administered ex vivo during manufacturing orin vivo during therapyupregulates IL7R alpha expression on CAR T cells, can increase persistence and overall efficacy of CAR T cells when combined with exogenous IL-7 and importantly, does not inhibit CAR T cell expansion or activity. One major benefit of using dex to upregulate IL-7R versus engineering overexpressing constitutively active IL-7R is the reversibility upon removal of dex, which allows this strategy to increase efficacy with low oncogenesis potential and thus a better safety profile for patients. While lymphodepletion increases serum IL-7, the level of increase is highly patient-dependent and is not a reliable method for the CAR T cell and dex strategy. NeoImmuneTech (NIT) developed a long-acting IL-7 fusion protein (NT-I7; rhIL-7-hyFc) that has been shown to increase persistence of CAR T cells preclinically and clinically. We partnered with NIT to test our combination strategy of dex, CAR T cells, and NT-I7 in primary and extramedullary MM. We hypothesize that the dex-mediated upregulation of IL-7R alpha in CAR T cells combined with NT-I7 will enhance anti-tumor efficacy against MM. Methods: We generated patient-derived BCMA-targeted CAR T cells by activating T cells isolated from MM patients with CD3/CD28 beads, transduced with BCMA CAR lentivirus, and expanded the resulting CAR T cells for 16 days (d). To test our combination strategy, we established two challenging xenograft mouse models of primary and extramedullary MM. For the primary model, we engrafted 2 million (M) MM.1R cells, a dex resistant MM cell line expressing GFP and firefly luciferase, by intratibial injection. After 7d, we injected tumor-bearing mice with a suboptimal dose of CAR T cells (0.1M) intravenously. On the same day as CAR T cells, we injected a single dose of NT-I7 (10mg/kg) subcutaneously and dex (10mg/kg) intraperitoneally, which we administered every 48 hours for 3 total doses. For the more challenging extramedullary model, we engrafted 10M MM.1R cells by subcutaneous injection in the flank. After 14d, allowing for high tumor burden, we injected tumor-bearing mice with a suboptimal dose of CAR T cells (0.2M) intravenously. On the same day as CAR T cell infusion, we administered NT-I7 and dex as in the primary model. For both models, we followed anti-tumor activity by live mouse imaging and collected blood and bone marrow at euthanasia to analyze long-term persistence, comparing treatment group to mice treated with CAR T cells alone or with single agent, dex or NT-I7. Results: For the primary model, CAR T cells alone were unable to control tumor growth, while CAR T cells with either dex or NT-I7 alone and triple combination achieved remission by d21. While mice treated with both CAR and either single agent relapsed by d35, mice treated with CAR T cells, dex, and NT-I7 achieved deep remission, leading to 100% survival for >100 days without relapse. For the extramedullary model, only the combination of CAR T cells with dex and NT-I7 was able to control tumor growth, with observable antitumor efficacy by d14 and remission by d35. At euthanasia, we observed a higher number of CAR T cells in the blood and bone marrow of mice that received all three agents than mice treated with CAR T cells and dex or NT-I7 alone or CAR T cells alone, which suggest the potential of this combination to enhance CAR T cell expansion and persistence even at severely suboptimal doses.Conclusions: Combining CAR T cells with dex and NT-I7 significantly enhanced antitumor efficacy of CAR T cell therapy for primary and extramedullary, leading to deep remission in both models.