Project description:To evaluate whether the findings of our pre-clinical study hold true in patients treated with CAR19/IL-15 NK cells, we isolated and profiled CAR-NK cells and B cells from peripheral blood collected at multiple timepoints from two patients diagnosed with CD19+ relapse/refractory hematological malignancies and treated with CAR19/IL-15 NK cells (11). Each patient received one infusion of CAR-NK cells
Project description:CEACAM5-specific CAR NK-92 cells were engineered to target CEACAM5-expressing lung tumor cells. To study their transcriptional response during tumor engagement, parental NK-92 cells, CEACAM5-CAR NK-92 cells, hydrocortisone-treated CAR NK-92 cells, NR3C1-knockout (cortisol-resistant) CAR NK-92 cells, and hydrocortisone-treated NR3C1-knockout CAR NK-92 cells were co-cultured with CEACAM5⁺ A549 lung cancer cells for 16 hours. Following co-culture, NK-92–derived effector cells were isolated by flow cytometry and processed for bulk RNA sequencing. This dataset captures transcriptional programs associated with CAR activation, hydrocortisone exposure, and glucocorticoid receptor deficiency in NK-92–based effector cells responding to CEACAM5⁺ tumor targets.
Project description:Steroids enriched within the lung tumor microenvironment suppress the cytotoxic activity of tumor-infiltrating NK cells and intensify hypoxic stress. To model these conditions in vitro, CEACAM5-specific CAR-NK cells and NR3C1-knockout CAR-NK cells were exposed to hydrocortisone (1 µM), hypoxia-mimicking CoCl₂ (100 µM), or their combination. NK cells were treated for 24 hours, followed by a 6-hour co-culture with A549 lung cancer cells to activate NK-cell effector programs. After co-culture, NK cells were isolated to high purity and subjected to bulk RNA sequencing to characterize transcriptional changes driven by steroid signalling, hypoxia, and glucocorticoid receptor deficiency. This dataset includes multiple treatment groups: untreated CAR-NK cells, hypoxia-treated CAR-NK cells, combined hypoxia + hydrocortisone–treated CAR-NK cells, and corresponding NR3C1-knockout CAR-NK cell groups under identical conditions.
Project description:The purpose of this experiment to examine how the gene expression profile and differential gene expression in NK cells, TIGITKO NK cells, TIGITKOGD2.CAR NK cells or GD2.CAR NK cells changes after 72 hours of neuroblastoma tumor exposure.
Project description:Chimeric antigen receptor (CAR) cell therapy has revolutionized the treatment of hematological malignancies; however, its efficacy in solid tumors remains limited due to dense stromal barriers and the immunosuppressive tumor microenvironment (TME), which hinder immune cell infiltration and persistence. To overcome these challenges, we propose an innovative strategy utilizing induced pluripotent stem cell (iPSC)-derived CAR-NK extracellular vesicles (CAR-iNEV), engineered from iPSC-differentiated NK cells. Unlike conventional CAR-NK cell therapies that rely on immortalized NK92 cell lines, our approach leverages iPSC-derived NK cells, reducing tumorigenic risk and enhancing CAR transduction efficiency. Additionally, we incorporate nanobody-based CAR constructs in place of traditional single-chain variable fragments (scFv), significantly improving CAR stability and expression efficiency. Beyond direct cytotoxicity, CAR-iNEV offers superior therapeutic advantages, including reduced systemic toxicity and enhanced drug delivery capabilities compared to CAR-iNK cells. In vivo studies demonstrate that CAR-iNEV exhibits exceptional safety and potent antitumor activity across multiple solid tumor xenograft and patient-derived xenograft (PDX) models in humanized mice. Mechanistically, CAR-iNEV not only directly eliminates tumor cells but also reprograms the TME by activating macrophage-derived nitric oxide synthase (NOS2), amplifying host antitumor immunity. These findings establish CAR-iNEV as a promising platform for solid tumor immunotherapy, particularly in relapsed and metastatic settings, providing a novel and effective strategy that bridges direct tumor targeting with immune microenvironment remodeling. This study lays the foundation for future clinical translation, advancing the next generation of cell-free CAR-based immunotherapies.
Project description:The purpose of this experiment was two-fold. The first was to examine how the gene expression profile changes over time in C7R-CAR NK cells. C7R is a constitutively active IL-7 receptor that provides persistent activation of STAT5. The second was to examine the differential gene expression in C7R CAR NK cells or CAR NK treated with exogenous IL-15 (exIL15) after 2 weeks of stimulation.
Project description:This is a preclinical study investigating the role of CREM transcription factor in CAR NK cells. We found thata CREM is a regulatory checkpoint induced by CAR CD3z signaling and IL-15 stimulation that leads to suppression of NK cell function. CREM KO enhances the anti-tumor efficacy of CAR NK cells.
Project description:Chronic stimulation with Acute Myeloid Leukemia can induce dysfunction in NK and CAR-NK cells. We evaluated transcriptomic differences on the day 10 of chronic antigen stimulation in NK cells secreting IL15 compared to non-secreting NK cells
Project description:Chimeric Antigen Receptor (CAR)-cell-based therapies have shown remarkable achievements in cancer treatment, especially for hematological malignancies. In CAR-T cells, the costimulatory[P1] domain is a key factor that significantly affects their therapeutic success. However, the effects of different costimulatory domains on the activation and performance of CAR-natural killer (CAR-NK) cells have not been sufficiently studied. We designed novel CAR constructs incorporating NK-specific costimulatory domains, such as 2B4 (CD244) and DAP12, to enhance CAR NK functionality. We evaluated their efficacy using in vitro cytotoxicity assays, cytokine profiling, and in vivo tumor models. We further investigated dasatinib, a tyrosine kinase inhibitor, as a pharmacological modulator of CAR NK cell activity. Short-term dasatinib exposure temporarily and reversibly suppressed CAR-NK activity yet enhanced function upon withdrawal. Accordingly, 2B4-DAP12 CAR-NK cells demonstrated superior tumor control and survival compared to conventional 4-1BBζ CAR-NK cells in vivo following dasatinib pretreatment. Our findings highlight that NK-optimized co-stimulation (2B4-DAP12) with reversible dasatinib pharmacological control synergistically enhanced CAR-NK cytotoxic function, providing an innovative strategy for next-generation cellular therapies.
Project description:Chimeric antigen receptor (CAR)-NK therapy has emerged as a highly promising alternative to CAR-T cell therapy, due to its remarkable efficacy and safety in hematological tumors. However, the efficacy of CAR-NK cells currently remains limited in solid tumors. Here, we successfully developed CISH locus-specific integrated CAR-NK cells using a non-viral mini-circular single-stranded DNA (mcssDNA)-based CRISPR/Cas9 targeted genome editing (mcssDNA/CRISPR/Cas9) technology via a single-step electroporation procedure. The developed CISH-knockout (CISH-KO) CAR-NK cells exhibited the enhanced proliferation and cytotoxicity against hepatocellular carcinoma (HCC) compared with conventional lentivirus-transduced CAR-NK (LV-CAR-NK) cells. Single-cell RNA sequencing analysis revealed an important subset of adaptive NK cells that were highly enriched in CISH-KO CAR-NK cells compared to LV-CAR-NK cells and showed the up-regulated JAK/STAT, energy metabolism and activating receptor signaling. Furthermore, non-viral CISH locus-specific integrated IL-15-armored CAR-NK cells were generated and achieved persistent tumor regression and a significant survival advantage in a lung metastatic mouse model of HCC. Collectively, our results demonstrate that non-viral CISH locus-specific integrated CAR-NK cells can be generated by a simplified, single-step manufacturing procedure and achieve potent efficacy against HCC, thus providing an innovative technology for CAR-NK cell therapy against solid tumors.