127 Preclinical evaluation of NKX019, a CD19-targeting CAR NK Cell

Abstract

BackgroundNatural killer (NK) cells are highly effective and fast-acting cytolytic cells capable of eradicating target cells with limited adverse effects such as cytokine release syndrome (CRS) or graft-versus-host disease. Chimeric antigen receptors (CARs)-engineered NK cells have been recently used against leukemia with encouraging clinical outcomes.1 The surface antigen CD19, expressed by B-lymphoblasts, represents an ideal CAR target against B cell acute lymphoblastic leukemia (B-ALL). We developed a highly potent CD19 -directed CAR NK cell therapy, NKX019, with an extended in vivo half-life aimed at killing CD19-expressing target.MethodsNK cells isolated from healthy PBMCs were expanded in the presence of NKSTIM cells, IL-2, IL-12, IL-18 and transduced with both a CD19-targeted CAR construct and a membrane-bound form of IL-15 (mbIL-15). Control (non-engineered) NK cells were produced in parallel. Cytotoxic activity of NKX019 against CD19+ B-ALL cell line (REH), pre-B ALL cell line (Nalm-6), allogeneic PBMCs was assessed using Incucyte® or flow cytometry. NSG mice bearing either Nalm-6.fluc (Nalm6) or REH.fluc (REH) tumor received different concentrations of NKX019 or control NK cells. In-life analysis of tumor-bearing and naïve NSG mice include: 1) bioluminescence imaging, 2) clinical observations, 3) serum cytokines and 4) CAR+ NK cell persistency.ResultsNKX019 showed enhanced cytolytic activity against REH and Nalm-6 tumor cells compared to control NK cells and CAR19+ T cells. The superiority of NKX019 over CAR19+ T cells was more pronounced at the earlier time point (24 hours) with near identical calculated EC50 observed at 72 hours for both cell types. Increased cytolytic activity of NKX019 was limited to CD19+ cells in bulk PBMCs. Consistent with our in vitro observations, NKX019 controlled Nalm-6 and REH tumor growth in doses as low as 2 × 106 cells/kg for up to 30 days with no apparent increase in cytokines commonly associated with CRS. Increased Nalm-6 tumor growth coincided with an apparent decrease in measurable NKX019 in the periphery. In tumor-naïve NSG mice, NKX019 was detectable in the blood for up to 9 weeks post-infusion consistent with its extended half-life.ConclusionsNKX019 expresses mbIL-15 and is produced in the presence of IL-12 and IL-18, resulting in enhanced in vitro expansion and longer in vivo half-life than non-engineered NK cells. NKX019 also exhibited advantages compared to CAR19+ T cells including faster cytotoxic kinetics and limited production of cytokines associated with CRS. A first-in-human trial of NKX019 in B cell malignancies is planned for 2021.Ethics ApprovalThe animal procedures described in this abstract were conducted in accordance with Explora BioLabs Animal Care and Use Protocol approved by Explora BioLabs Institutional Animal Care and Use Committee.ReferenceLiu, et al. 2020 NEJM

Are we ready for personalizedCAR‐Ttherapy?

“…20 The results provided by Daher et al 20 showed that the antitumor activity of armored inter- and pre-B ALL cell line (Nalm-6). 21 These findings were then implemented in a phase 1 study in patients with R/R non-Hodgkin lymphoma (NHL) or ALL. 24 A major challenge associated with the implementation of CAR-NK is their poor resistance to cryopreservation and subsequent thawing.…”

“…Genetic editing of macrophages with CAR (CAR‐Ms) resulted in sustained polarization to a pro‐inflammatory M1 phenotype, promotion of antitumor T‐cell activity and antigen‐specific phagocytosis 18 . Numerous studies have focused on natural killer (NK) cells 19–23 . Auto CAR‐T cells based on the NK group 2D (NKG2D) receptor—CYAD‐01—were administered to patients with R/R acute myeloid leukemia (AML), myelodysplastic syndromes (MDS), or multiple myeloma (MM).…”

“…Obtained from PB anti‐CD19 CAR‐NK cells—NKX019—developed by Nkarta were enriched with a membrane‐bound form of IL‐15 (mbIL‐15). In a mouse model, NKX019 showed high cytotoxicity, with simultaneous limited release of cytokines, potentially related to cytokine release syndrome (CRS), against CD19+ B‐cell acute lymphoblastic lymphoma (B‐ALL) (REH) and pre‐B ALL cell line (Nalm‐6) 21 . These findings were then implemented in a phase 1 study in patients with R/R non‐Hodgkin lymphoma (NHL) or ALL 24 .…”

CD19-Targeted Immunotherapies for Diffuse Large B-Cell Lymphoma

“…The purpose of this phase 1 study is to identify the optimal treatment dose with NKX019 product of Nkarta Therapeutics (https://ash.confex.com/ash/ 2021/webprogram/Paper146602.html). NKX019 expresses a CD19-targeted CAR, OX40 costimulatory domain, CD3z signaling moiety, and a membrane-bound form of IL-15 (mbIL-15) (78). Equipping CAR-NK cells with on-board cytokines, such as IL-15, lays the foundations for new therapeutic options aimed at improving clinical efficacy by enhancing both persistence and cytotoxicity against tumor cells (79).…”

902 NKX019, an off-the-shelf CD19 CAR-NK cell, mediates improved anti-tumor activity and persistence in combination with CD20-directed therapeutic mAbs

“…NKX019 exhibits more rapid cytotoxic kinetics than CD19directed CAR T cells, and lower production of cytokines associated with cytokine release syndrome (CRS). 1 The safety and clinical activity of NKX019 are currently being evaluated in a phase I [GM1] clinical study [NCT05020678]. Recent studies 2,3 have shown that combining NK cell therapies with monoclonal antibodies (mAbs) may improve targeted NK cell activation and overcome some of the disadvantages associated with the stand-alone use of therapeutic mAbs.…”