We generated a novel CD19CAR (CAT) with a lower affinity than FMC63, the binder utilised in many clinical studies. CAT CAR T cells showed increased proliferation/cytotoxicity in vitro and enhanced proliferative capacity and anti-tumor activity than FMC63 CAR T cells in a xenograft model. In a clinical study (CARPALL, NCT02443831), 12/14 patients with relapsed/refractory pediatric BALL obtained molecular remission after CAT CAR T cell therapy. CAR T cell expansion compared favourably with published data on other CD19CARs and persistence was demonstrated in 11 of 14 patients at last follow-up. Toxicity was low with no severe cytokine release syndrome. At a median follow up of 14 months, 5/14 patients (37%) remain in molecular CR with circulating CAR T cells.

SummaryAPS1/APECED patients are defined by defects in the autoimmune regulator (AIRE) that mediates central T cell tolerance to many self-antigens. AIRE deficiency also affects B cell tolerance, but this is incompletely understood. Here we show that most APS1/APECED patients displayed B cell autoreactivity toward unique sets of approximately 100 self-proteins. Thereby, autoantibodies from 81 patients collectively detected many thousands of human proteins. The loss of B cell tolerance seemingly occurred during antibody affinity maturation, an obligatorily T cell-dependent step. Consistent with this, many APS1/APECED patients harbored extremely high-affinity, neutralizing autoantibodies, particularly against specific cytokines. Such antibodies were biologically active in vitro and in vivo, and those neutralizing type I interferons (IFNs) showed a striking inverse correlation with type I diabetes, not shown by other anti-cytokine antibodies. Thus, naturally occurring human autoantibodies may actively limit disease and be of therapeutic utility.

Mature T cell cancers are typically aggressive, treatment resistant and associated with poor prognosis. Clinical application of immunotherapeutic approaches has been limited by a lack of target antigens that discriminate malignant from healthy (normal) T cells. Unlike B cell depletion, pan-T cell aplasia is prohibitively toxic. We report a new targeting strategy based on the mutually exclusive expression of T cell receptor β-chain constant domains 1 and 2 (TRBC1 and TRBC2). We identify an antibody with unique TRBC1 specificity and use it to demonstrate that normal and virus-specific T cell populations contain both TRBC1 and TRBC2 compartments, whereas malignancies are restricted to only one. As proof of concept for anti-TRBC immunotherapy, we developed anti-TRBC1 chimeric antigen receptor (CAR) T cells, which recognized and killed normal and malignant TRBC1, but not TRBC2, T cells in vitro and in a disseminated mouse model of leukemia. Unlike nonselective approaches targeting the entire T cell population, TRBC-targeted immunotherapy could eradicate a T cell malignancy while preserving sufficient normal T cells to maintain cellular immunity.

B-cell maturation antigen (BCMA) is a promising therapeutic target for multiple myeloma (MM), but expression is variable, and early reports of BCMA targeting chimeric antigen receptors (CARs) suggest antigen downregulation at relapse. Dual-antigen targeting increases targetable tumor antigens and reduces the risk of antigen-negative disease escape. "A proliferation-inducing ligand" (APRIL) is a natural high-affinity ligand for BCMA and transmembrane activator and calcium-modulator and cyclophilin ligand (TACI). We quantified surface tumor expression of BCMA and TACI on primary MM cells (n = 50). All cases tested expressed BCMA, and 39 (78%) of them also expressed TACI. We engineered a third-generation APRIL-based CAR (ACAR), which killed targets expressing either BCMA or TACI ( < .01 and < .05, respectively, cf. control, effector-to-target [E:T] ratio 16:1). We confirmed cytolysis at antigen levels similar to those on primary MM, at low E:T ratios (56.2% ± 3.9% killing of MM.1s at 48 h, E:T ratio 1:32; < .01) and of primary MM cells (72.9% ± 12.2% killing at 3 days, E:T ratio 1:1; < .05, n = 5). Demonstrating tumor control in the absence of BCMA, we maintained cytolysis of primary tumor expressing both BCMA and TACI in the presence of a BCMA-targeting antibody. Furthermore, using an intramedullary myeloma model, ACAR T cells caused regression of an established tumor within 2 days. Finally, in an in vivo model of tumor escape, there was complete ACAR-mediated tumor clearance of BCMATACI and BCMATACI cells, and a single-chain variable fragment CAR targeting BCMA alone resulted in outgrowth of a BCMA-negative tumor. These results support the clinical potential of this approach.

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