CAR‐DC combined with CAR‐T therapy for relapsed/refractory acute myeloid leukaemia: Research progress and future perspectives

Introduction: Our clinical program in chimeric antigen receptor T (CART) cell therapy has focused on targeting EGFR oncoproteins for glioblastoma (GBM). We have completed a phase I trial using CART cells directed to the EGFRvIII antigen and showed in situ uptake, proliferation, functional activation and antigen editing by CART cells in GBM tissue. This first clinical trial demonstrated two barriers to clinical efficacy. First, GBM heterogeneity was striking spatially and temporally, including the number of variants in EGFR. Second, CART cell therapy was associated with an adaptive anti-GBM response illustrated by an initial wave of CART activation followed by dramatic immunosuppression. These two features indicate that we need strategies to overcome GBM heterogeneity and the immunosuppressive tumor microenvironment (TME). Methods: We have used multiple CART variants, targeting both IL13Rα2 and EGFRvIII, in combination with selected immune checkpoint blockade inhibitors, to explore possible additive effects in both in vitro and in vivo GBM model systems on limiting the immunosuppressive GBM TME. Independently, we have developed a portfolio of CARTs with different binding specificities that target other homo- and heterodimers of EGFR oncoproteins, to address GBM heterogeneity. Results: Combination studies of CART cells with immune checkpoint blockade inhibitors revealed a non-homogenous response. Different CART structures and targets showed increased tumor killing activity with specific immune checkpoint blockade inhibition. Specifically, IL13Rα2-targeting CART cells had the best effect when paired with anti-CTLA4 treatment, while EGFRvIII-targeting CART cells had the largest effect when paired with anti-PD-1 treatment. In parallel to our combination therapy work, we have expanded our repertoire of CART constructs for targeting EGFR oncogenic alleles with extracellular domain mutations by utilizing antibody phage display technology. EGFR-targeting antibody sequences were incorporated into viral constructs and transduced into T cells. We have shown variable killing with these novel CART constructs in GSC/PDX models of GBM heterogeneity. Conclusion: Our initial experience with CART cells in GBM suggested that although a single intravenous infusion results in CART cell bioactivity in the brain, overcoming the adaptive changes in the local TME and addressing antigen heterogeneity may improve the clinical efficacy of CART-directed strategies. Our combination work showed that different CART constructs cooperate with immune checkpoint blockade inhibition differentially. This selective pairing suggests that a more personalized immunophenotypic assessment may result in higher efficacy therapeutic combinations. In addition, development of a broad portfolio of both selective and promiscuous CART constructs targeting EGFR in its various forms gives us the potential to cover a larger percentage of both the GBM tumor volume and regionally-specific tumor cells within a single GBM. Citation Format: Zev A. Binder, Yibo Yin, Radhika Thokala, Donald M. O'Rourke. Combinatorial platform for CART cell therapy for glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-340.

This is a title in sentence case:early intrathecal dexamethasone and methotrexate as an effective approach for immune effector cell-associated neurotoxicity syndrome after CAR-T cell therapies

The treatment of B cell malignancies has dramatically changed with the introduction of immunotherapy, especially chimeric antigen receptor T (CAR-T) cell therapy. However, only limited efficacy is observed in acute...

Human C-type lectin-like molecule 1 (CLL-1) represents a promising therapeutic target for Chimeric antigen receptor T (CAR-T) cells therapy in the treatment of acute myeloid leukemia (AML). In this study, we aimed to evaluate the efficacy and safety profile of donor-derived CLL-1 CAR-T cells in AML patients who experienced relapsed post-transplantation. 14 AML patients who experienced relapse following allogeneic HSCT were enrolled in our clinical trial. However, 2 patients withdrew from the study due to rapid disease progression. 12 participants received donor-derived CLL-1 CAR-T cells and were categorized into 3 groups based on the dosage of infused CAR-T cells dose (Group A:0.5×106/kg, Group B:1×106/kg, Group C:1.5×106/kg). And scRNA-seq was used to reveal CLL-1 CAR-T cells dynamics in a CAR-T cells infusion products and PBMCs at the peak of expansion for patient 4. CLL-1 CAR-T cells were well tolerated by all 12 patients. Cytokine Release Syndrome (CRS) was observed in all patients, with 5 patients experiencing grade ≥3. 3 patients developed cytokine release syndrome-associated encephalopathy (CRES), and 1 patient had a grade 4 severity level. All patients demonstrated a reduction in tumor burden, while 7 patients (58.33%) achieved MRD-CR and 2 patients (16.67%) reached MRD+CR. CAR-T cells expansion was detectable in all 12 patients, with the median time of peak expansion was 9 days (range: 7-11 days). In patient 4, compared to the pre-reinfusion state, CD4+ cells at the peak of expansion showed upregulation of cell killing-related genes and memory T cell-related genes (P < 0.01). The CLL-1 CAR-T cells therapy derived from allogeneic donors demonstrates both safety and efficacy in the management of relapsed AML following allogeneic HSCT. And adjusting the ratio of CD4+ CAR-T cells and CD8+ CAR-T cells prior to infusion may help mitigate CAR-T cell-related side effects. https://www.chictr.org.cn/, identifier ChiCTR2000041054.

Developing “Off-the-Shelf” CLL1 CAR-DNT Therapeutics for the R/R Acute Myeloid Leukemia

Promoter usage regulating the surface density of CAR molecules may modulate the kinetics of CAR-T cells in vivo

Secondary Myeloid Neoplasms after CD19 CAR-T Cell Therapy: Real-Life Multicenter Data from the Clonhema Study

Autologous chimeric antigen receptor T (CAR-T) cell therapy has demonstrated efficacy in the treatment of acute myeloid leukemia (AML). Nevertheless, the intrinsic characteristics of autologous therapy, such as extended manufacturing timelines and patient-specific limitations, contribute to delays in treatment availability. More critically, relapse due to antigen escape following single-targeted CAR-T therapy constitutes a significant clinical obstacle. To address the dual challenges of delayed treatment accessibility and antigen escape relapse, this study proposes the development of universal tandem CAR-T cells. These cells, engineered to target CD123 and B7-H3 through clustered regularly interspaced short palindromic repeats (CRISPR) gene editing technology, represent an innovative therapeutic strategy for AML. In this study, an immune phage display nanobody library was developed for the purpose of screening CD123-specific nanobodies. The CRISPR/CRISPR-associated protein 9 (CRISPR/Cas9) gene editing system was utilized to disrupt the TRAC and B2M genes present in T cells, resulting in the generation of universal CD123/B7-H3 bispecific universal CAR-T (UCAR-T) cells. The efficacy of these dual-specific UCAR-T cells in combating tumors was subsequently assessed through in vitro and in vivo experiments. Through four rounds of panning against CD123 from an immunized camelid VHH library, we identified 21 antigen-specific nanobodies. Tandem bispecific UCAR-T engineered with these binders demonstrated CAR transduction efficiencies ranging from 82% to 87%. In vitro functional profiling revealed a significantly enhanced cytotoxicity of bispecific UCAR-Ts against CD123+/B7-H3+ AML cell lines when compared to single-target constructs, while effectively regulating the secretion of effector cytokines (IL-2, IFN-γ, TNF-α). In AML xenograft models, treatment with bispecific UCAR-T notably inhibited tumor progression, extended the survival of tumor-bearing mice with recurrence-free persistence throughout the observation period, and did not result in significant body weight loss or cytokine release syndrome. The findings of the study address the issue of tumor antigen evasion in the treatment of AML, circumvent certain constraints associated with autologous CAR-T cell therapy, and offer novel insights and strategies for managing AML.

BackgroundImmunotherapy, such as chimeric antigen receptor T (CAR-T) cells targeting CD33 or CD123, has been well developed over the past decade for the treatment of acute myeloid leukemia (AML). However, the inability to sustain tumor-free survival and the possibility of relapse due to antigen loss have raised concerns. A dual targeting of CD33 and CD123 is needed for better outcomes.MethodsBased on our previously constructed CD33 and CD123 monovalent CAR-T, Loop33 × 123 and Loop123 × 33 CAR-T were constructed with molecular cloning techniques. All CAR-T cells were generated by lentivirus transduction of T cells from healthy donors. Phenotype detection was evaluated on day 7 concerning activation, exhaustion, and subtype proportions. Coculture killing assays were conducted using various AML cell lines and primary AML cells. Degranulation and cytokine secretion levels were detected by flow cytometry. Cell-derived xenograft models were established using wild-type Molm 13 cell lines, or a mixture of Molm 13-KO33 and Molm 13-KO123 cells as an ideal model of immune escape. By monitoring body weight and survival of tumor-bearing mice, Loop33 × 123 and Loop123 × 33 CAR-T cells were further assessed for their efficacy in vivo.ResultsIn vitro study, our results demonstrated that Loop33 × 123 CAR-T cells could efficiently eliminate AML cell lines and primary AML cells with elevated degranulation and cytokine secretion levels. Compared with our previously constructed monovalent CD33 or CD123 CAR-T cells, Loop33 × 123 CAR-T cells showed superior advantages in an immune escape model. In vivo studies further confirmed that Loop33 × 123 CAR-T cells could effectively prolong the survival of mice without significant toxicity. However, Loop123 × 33 CAR-T cells failed to show the same effects. Furthermore, Loop33 × 123 CAR-T cells efficiently circumvented potential immune escape, a challenge where monovalent CAR-T cells failed.ConclusionsLoop33 × 123 CAR-T targeting CD33 and CD123 could efficiently eliminate AML cells and prolong survival of tumor-bearing mice, while addressing the issue of immune escape.

Chimeric antigen receptor T (CAR-T) cell therapies have transformed the treatment of relapsed/refractory (R/R) B-cell malignancies and multiple myeloma by redirecting activated T cells to CD19- or BCMA-expressing tumor cells. However, this approach has yet to be approved for acute myeloid leukemia (AML), the most common acute leukemia in adults and the elderly. Simultaneously, CAR-T cell therapies continue to face significant challenges in the treatment of solid tumors. The primary challenge in developing CAR-T cell therapies for AML is the absence of an ideal target antigen that is both effective and safe, as AML cells share most surface antigens with healthy hematopoietic stem and progenitor cells (HSPCs). Simultaneously targeting antigen expression on both AML cells and HSPCs may result in life-threatening on-target/off-tumor toxicities such as prolonged myeloablation. In addition, the immunosuppressive nature of the AML tumor microenvironment has a detrimental effect on the immune response. This review begins with a comprehensive overview of CAR-T cell therapy for cancer, covering the structure of CAR-T cells and the history of their clinical application. It then explores the current landscape of CAR-T cell therapy in both hematologic malignancies and solid tumors. Finally, the review delves into the specific challenges of applying CAR-T cell therapy to AML, highlights ongoing global clinical trials, and outlines potential future directions for developing effective CAR-T cell-based treatments for relapsed/refractory AML.

DAP10 integration in CAR-T cells enhances the killing of heterogeneous tumors by harnessing endogenous NKG2D

Glioblastoma (GBM) is the most common malignant primary brain cancer in adults. It is always resistant to existing treatments, including surgical resection, postoperative radiotherapy, and chemotherapy, which leads to a dismal prognosis and a high relapse rate. Therefore, novel curative therapies are urgently needed for GBM. Chimeric antigen receptor T (CAR-T) cell therapy has significantly improved life expectancy for hematological malignancies patients, and thus it increases the interest in applying CAR-T cell therapy for solid tumors. In the recently published research, it is indicated that there are numerous obstacles to achieve clinical benefits for solid tumors, especially for GBM, because of GBM anatomical characteristics (the blood–brain barrier and suppressive tumor microenvironment) and the tumor heterogeneity. CAR-T cells are difficult to penetrate blood–brain barrier, and immunosuppressive tumor microenvironment (TME), which induces CAR-T cell exhaustion, impairs CAR-T cell therapy response. Moreover, under the pressure of CAR-T cell therapy, the tumor heterogeneity and tumor plasticity drive tumor evolution and therapy resistance, such as antigen escape. Nonetheless, scientists strive for strategies to overcome these hurdles, including novel CAR-T cell designs and regional delivery. For instance, the structure of multi-antigen-targeted CAR-T cells can enrich CAR-T accumulation in tumor TME and eliminate abundant tumor cells to avoid tumor antigen heterogeneity. Additionally, paired with an immune modifier and one or more stimulating domains, different generation of innovations in the structure and manufacturing of CAR-T cells have improved efficacy and persistence. While single CAR-T cell therapy receives limited clinical survival benefit. Compared with single CAR-T cell therapy, the combination therapies have supplemented the treatment paradigm. Combinatorial treatment methods consolidate the CAR-T cells efficacy by regulating the tumor microenvironment, optimizing the CAR structure, targeting the CAR-T cells to the tumor cells, reversing the tumor-immune escape mechanisms, and represent a promising avenue against GBM, based on multiple impressive research. Moreover, exciting results are also reported to be realized through combining effective therapies with CAR-T cells in preclinical and clinical trials samples, have aroused inspiration to explore the antitumor function of combination therapies. In summary, this study aims to summarize the limitation of CAR-T cell therapies and introduces novel strategies to enhance CAR-T cell function as well as prospect the potential of the therapeutic combination.

Prophylactic Tocilizumab in Patients with Relapsed Refractory Lymphoma Treated with Anti CD19 Chimeric Antigen Receptor T-Cell Therapy

Leukemia Stem Cells Are Characterized By CLEC12A Expression and Chemotherapy Refractoriness That Can be Overcome By Targeting with Chimeric Antigen Receptor T Cells

A Multi-Omic Single-Cell Landscape of Cytokine Release Syndrome in Multiple Myeloma Patients after Anti-BCMA CAR-T Cell Therapy