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Background: The interleukin-1 receptor accessory protein (IL1RAP) is highly expressed on acute myeloid leukemia (AML) bulk blasts and leukemic stem cells (LSCs), but not on normal hematopoietic stem cells (HSCs), providing an opportunity to target and eliminate the disease, while sparing normal hematopoiesis. Herein, we report the activity of BIF002, a novel anti-IL1RAP/CD3 T cell engager (TCE) in AML. Methods: Antibodies to IL1RAP were isolated from CD138+ B cells collected from the immunized mice by optoelectric positioning and single cell sequencing. Individual mouse monoclonal antibodies (mAbs) were produced and characterized, from which we generated BIF002, an anti-human IL1RAP/CD3 TCE using Fab arm exchange. Mutations in human IgG1 Fc were introduced to reduce FcγR binding. The antileukemic activity of BIF002 was characterized in vitro and in vivo using multiple cell lines and patient derived AML samples. Results: IL1RAP was found to be highly expressed on most human AML cell lines and primary blasts, including CD34+ LSC-enriched subpopulation from patients with both de novo and relapsed/refractory (R/R) leukemia, but not on normal HSCs. In co-culture of T cells from healthy donors and IL1RAPhigh AML cell lines and primary blasts, BIF002 induced dose- and effector-to-target (E:T) ratio-dependent T cell activation and leukemic cell lysis at subnanomolar concentrations. BIF002 administered intravenously along with human T cells led to depletion of leukemic cells, and significantly prolonged survival of IL1RAPhigh MOLM13 or AML patient-derived xenografts with no off-target side effects, compared to controls. Of note, BiF002 effectively redirects T cells to eliminate LSCs, as evidenced by the absence of disease initiation in secondary recipients of bone marrow (BM) from BIF002+T cells-treated donors (median survival not reached; all survived > 200 days) compared with recipients of BM from vehicle- (median survival: 26 days; p = 0.0004) or isotype control antibody+T cells-treated donors (26 days; p = 0.0002). Conclusions: The novel anti-IL1RAP/CD3 TCE, BIF002, eradicates LSCs and significantly prolongs survival of AML xenografts, representing a promising, novel treatment for AML. [ABSTRACT FROM AUTHOR]

The adaptive T cell immune response requires cellular plasticity to generate distinct subsets with diverse functional and migratory capacities. Studies of CAR T cells have primarily focused on a limited number of phenotypic markers in blood, representing an incomplete view of CAR T cell complexity. Here, we adapted mass cytometry to simultaneously analyze trafficking and functional proteins expression in CD19 CAR T cells across patients' tissues, including leukapheresis T cells, CAR product, CAR T cells in peripheral blood, bone marrow, and cerebrospinal fluid post infusion and correlate them with phenotypes. This approach revealed spatiotemporal plasticity of CAR T cells. Patients' CAR product revealed upregulation in many trafficking and activation molecules compared to leukapheresis T cells as baseline. Including statistically significant upregulation in CD4 and CD8 integrin-β7, CD4 granzyme B, and CD11a as well as CD8 CD25 and CD95. Moreover, patients' tissues showed spatiotemporal alteration in trafficking, activation, maturation, and exhaustion features, with a distinct signature in the central nervous system niche. Compared to peripheral blood samples, cerebrospinal fluid samples were statistically significant enriched in CD4 and CD8 trafficking and memory phenotype proteins integrin β7, CCR7, CXCR4, and CD8 CD69. Our data provide a potential framework to remodel CAR T cells and enhance immunotherapy efficacy. [ABSTRACT FROM AUTHOR]

AbstractReactive pleocytosis in the CSF has been observed with blinatumomab but has not been well-described. We performed a retrospective study of 88 patients who received intrathecal chemotherapy (IT) while on blinatumomab with CSF analyzed to determine if pleocytosis had an impact efficacy and safety. Blinatumomab was used for relapsed/refractory 62.5%, MRD-positive 31.8%, and consolidation in MRD-negative 5.7%. The incidence of pleocytosis in CSF was 51% and was more frequent after day 15 (55.8% vs. 18.2%, p = 0.025). Pleocytosis did not impact CR, clearance of MRD positivity, PFS and OS rates. Lower incidence of non-CNS extramedullary relapse was seen (3.7% vs. 30.8%, p = 0.011) with pleocytosis in CSF. Analysis of CSF by flow cytometry showed median CD4:CD8 ratio of 1.34. In conclusion, CSF pleocytosis is prevalent with blinatumomab but only demonstrated lower rates of non-CNS extramedullary relapse but no impact on CNS relapse or neurotoxicity. [ABSTRACT FROM AUTHOR]

Immune checkpoint blockade has emerged as a potent new tool in the war on cancer. However, only a subset of cancer patients benefit from this therapeutic modality, sparking a search for combination therapies to increase the fraction of responding patients. We argue here that inhibition of protein phosphatase 2A (PP2A) is a promising approach to increase responses to immune checkpoint blockade and other therapies that rely on the presence of tumor‐reactive T cells. Inhibition of PP2A increases neoantigen expression on tumor cells, activates the cGAS/STING pathway, suppresses regulatory T cells, and increases cytotoxic T cell activation. In preclinical models, inhibition of PP2A synergizes with immune checkpoint blockade and emerging evidence indicates that patients who have tumors with mutations in PP2A respond better to immune checkpoint blockade. Therefore, inhibition of PP2A activity may be an effective way to sensitize cancer cells to immune checkpoint blockade and cell‐based therapies using tumor‐reactive T cells. [ABSTRACT FROM AUTHOR]

High-grade (WHO grades III-IV) glioma remains one of the most lethal human cancers. Adoptive transfer of tumor-targeting chimeric antigen receptor (CAR)-redirected T cells for high-grade glioma has revealed promising indications of anti-tumor activity, but objective clinical responses remain elusive for most patients. A significant challenge to effective immunotherapy is the highly heterogeneous structure of these tumors, including large variations in the magnitudes and distributions of target antigen expression, observed both within individual tumors and between patients. To obtain a more detailed understanding of immunotherapy target antigens within patient tumors, we immunochemically mapped at single cell resolution three clinically-relevant targets, IL13Rα2, HER2 and EGFR, on tumor samples drawn from a 43-patient cohort. We observed that within individual tumor samples, expression of these antigens was neither random nor uniform, but rather that they mapped into local neighborhoods – phenotypically similar cells within regions of cellular tumor – reflecting not well understood properties of tumor cells and their milieu. Notably, tumor cell neighborhoods of high antigen expression were not arranged independently within regions. For example, in cellular tumor regions, neighborhoods of high IL13Rα2 and HER2 expression appeared to be reciprocal to those of EGFR, while in areas of pseudopalisading necrosis, expression of IL13Rα2 and HER2, but not EGFR, appeared to reflect the radial organization of tumor cells around hypoxic cores. Other structural features affecting expression of immunotherapy target antigens remain to be elucidated. This structured but heterogeneous organization of antigen expression in high grade glioma is highly permissive for antigen escape, and combinatorial antigen targeting is a commonly suggested potential mitigating strategy. Deeper understanding of antigen expression within and between patient tumors will enhance optimization of combination immunotherapies, the most immediate clinical application of the observations presented here being the importance of including (wild-type) EGFR as a target antigen. [ABSTRACT FROM AUTHOR]