Despite impressive progress, more than 50% of patients treated with CD19-targeting chimeric antigen receptor T cells (CAR19) experience progressive disease. Ten of 16 patients with large B cell lymphoma (LBCL) with progressive disease after CAR19 treatment had absent or low CD19. Lower surface CD19 density pretreatment was associated with progressive disease. To prevent relapse with CD19− or CD19lo disease, we tested a bispecific CAR targeting CD19 and/or CD22 (CD19-22.BB.z-CAR) in a phase I clinical trial (NCT03233854) of adults with relapsed/refractory B cell acute lymphoblastic leukemia (B-ALL) and LBCL. The primary end points were manufacturing feasibility and safety with a secondary efficacy end point. Primary end points were met; 97% of products met protocol-specified dose and no dose-limiting toxicities occurred during dose escalation. In B-ALL (n = 17), 100% of patients responded with 88% minimal residual disease-negative complete remission (CR); in LBCL (n = 21), 62% of patients responded with 29% CR. Relapses were CD19−/lo in 50% (5 out of 10) of patients with B-ALL and 29% (4 out of 14) of patients with LBCL but were not associated with CD22−/lo disease. CD19/22-CAR products demonstrated reduced cytokine production when stimulated with CD22 versus CD19. Our results further implicate antigen loss as a major cause of CAR T cell resistance, highlight the challenge of engineering multi-specific CAR T cells with equivalent potency across targets and identify cytokine production as an important quality indicator for CAR T cell potency.

HLA-haploidentical hematopoietic cell transplantation (Haplo-HCT) using posttransplantation cyclophosphamide (PT-Cy) has improved donor availability. However, a matched sibling donor (MSD) is still considered the optimal donor. Using the Center for International Blood and Marrow Transplant Research database, we compared outcomes after Haplo-HCT vs MSD in patients with acute myeloid leukemia (AML) in first complete remission (CR1). Data from 1205 adult CR1 AML patients (2008-2015) were analyzed. A total of 336 patients underwent PT-Cy–based Haplo-HCT and 869 underwent MSD using calcineurin inhibitor–based graft-versus-host disease (GVHD) prophylaxis. The Haplo-HCT group included more reduced-intensity conditioning (65% vs 30%) and bone marrow grafts (62% vs 7%), consistent with current practice. In multivariable analysis, Haplo-HCT and MSD groups were not different with regard to overall survival (P = .15), leukemia-free survival (P = .50), nonrelapse mortality (P = .16), relapse (P = .90), or grade II-IV acute GVHD (P = .98). However, the Haplo-HCT group had a significantly lower rate of chronic GVHD (hazard ratio, 0.38; 95% confidence interval, 0.30-0.48; P < .001). Results of subgroup analyses by conditioning intensity and graft source suggested that the reduced incidence of chronic GVHD in Haplo-HCT is not limited to a specific graft source or conditioning intensity. Center effect and minimal residual disease–donor type interaction were not predictors of outcome. Our results indicate a lower rate of chronic GVHD after PT-Cy–based Haplo-HCT vs MSD using calcineurin inhibitor–based GVHD prophylaxis, but similar other outcomes, in patients with AML in CR1. Haplo-HCT is a viable alternative to MSD in these patients.

Bortezomib (V), lenalidomide (R), cyclophosphamide (C) and dexamethasone (D) are components of the most commonly used modern doublet (RD, VD) or triplet (VRD, CVD) initial induction regimens prior to autologous hematopoietic cell transplantation (AHCT) for multiple myeloma (MM) in the US. In this study we evaluated 693 patients receiving “upfront” AHCT after initial induction therapy with modern doublet or triplet regimens using data reported to the Center for International Blood and Marrow Transplant Research from 2008–2013. Analysis was limited to those receiving a single AHCT after one line of induction therapy within 12 months from treatment initiation for MM. In multivariate analysis, progression-free (PFS) and overall survival were similar irrespective of induction regimen. However high risk cytogenetics and non-receipt of post-transplant maintenance/consolidation therapy were associated with higher risk of relapse. Patients receiving post-transplant therapy had significantly improved 3-year PFS vs. no post-transplant therapy (55% vs. 39%, p=0.0001). This benefit was most evident in patients not achieving at least CR post-AHCT (p=0.005). In patients receiving upfront AHCT, the choice of induction regimen (doublet or triplet therapies) appears to be of lower impact than use of post-transplant therapy.

PURPOSE Although the majority of patients with relapsed or refractory large B-cell lymphoma respond to axicabtagene ciloleucel (axi-cel), only a minority of patients have durable remissions. This prospective multicenter study explored the prognostic value of circulating tumor DNA (ctDNA) before and after standard-of-care axi-cel for predicting patient outcomes. METHODS Lymphoma-specific variable, diversity, and joining gene segments (VDJ) clonotype ctDNA sequences were frequently monitored via next-generation sequencing from the time of starting lymphodepleting chemotherapy until progression or 1 year after axi-cel infusion. We assessed the prognostic value of ctDNA to predict outcomes and axi-cel–related toxicity. RESULTS A tumor clonotype was successfully detected in 69 of 72 (96%) enrolled patients. Higher pretreatment ctDNA concentrations were associated with progression after axi-cel infusion and developing cytokine release syndrome and/or immune effector cell–associated neurotoxicity syndrome. Twenty-three of 33 (70%) durably responding patients versus 4 of 31 (13%) progressing patients demonstrated nondetectable ctDNA 1 week after axi-cel infusion ( P < .0001). At day 28, patients with detectable ctDNA compared with those with undetectable ctDNA had a median progression-free survival and OS of 3 months versus not reached ( P < .0001) and 19 months versus not reached ( P = .0080), respectively. In patients with a radiographic partial response or stable disease on day 28, 1 of 10 patients with concurrently undetectable ctDNA relapsed; by contrast, 15 of 17 patients with concurrently detectable ctDNA relapsed ( P = .0001). ctDNA was detected at or before radiographic relapse in 29 of 30 (94%) patients. All durably responding patients had undetectable ctDNA at or before 3 months after axi-cel infusion. CONCLUSION Noninvasive ctDNA assessments can risk stratify and predict outcomes of patients undergoing axi-cel for the treatment of large B-cell lymphoma. These results provide a rationale for designing ctDNA-based risk-adaptive chimeric antigen receptor T-cell clinical trials.

Eukaryotic initiation factor 2A is a single polypeptide that acts to negatively regulate IRES-mediated translation during normal cellular conditions. We have found that eIF2A (encoded by YGR054w) abundance is reduced at both the mRNA and protein level during 6% ethanol stress (or 37°C heat shock) under conditions that mimic the diauxic shift in the yeast Saccharomyces cerevisiae. Furthermore, eIF2A protein is posttranslationally modified during ethanol stress. Unlike ethanol and heat shock stress, H2O2 and sorbitol treatment induce the loss of eIF2A mRNA, but not protein and without protein modification. To investigate the mechanism of eIF2A function we employed immunoprecipitation-mass spectrometry and identified an interaction between eIF2A and eEF1A. The interaction between eIF2A and eEF1A increases during ethanol stress, which correlates with an increase in IRES-mediated translation from the URE2 IRES element. These data suggest that eIF2A acts as a switch to regulate IRES-mediated translation, and eEF1A may be an important mediator of translational activation during ethanol stress.

The outcomes of patients with DLBCL and primary treatment failure (PTF) in the rituximab era are unclear. We analyzed 331 patients with PTF, defined as primary progression while on upfront chemoimmunotherapy (PP), residual disease at the end of upfront therapy (RD) or relapse <6 months from end of therapy (early relapse; ER). Median age was 58 years and response to salvage was 41.7%. Two-year OS was 18.5% in PP, 30.6% in RD and 45.5% in ER. The presence of PP, intermediate-high/high NCCN-IPI at time of PTF or MYC translocation predicted 2-year OS of 13.6% constituting ultra-high risk (UHR) features. Among the 132 patients who underwent autologous hematopoietic cell transplantation, 2-year OS was 74.3%, 59.6% and 10.7% for patients with 0,1 and 2–3 UHR features respectively. Patients with PTF and UHR features should be prioritized for clinical trials with newer agents and innovative cellular therapy.

Classic Hodgkin lymphoma (cHL) patients with relapsed or refractory disease may benefit from allogeneic hematopoietic cell transplantation (allo-HCT), but many lack a matched sibling donor (MSD). Herein, we compare outcomes of 2 reduced-intensity conditioning (RIC) HCT platforms in cHL: T cellÀreplete related donor haploidentical (haplo) HCT

Autologous CD19 directed CAR T-cell therapy has response rates of >70% in adult acute lymphoblastic leukemia (ALL) and >40% in adult diffuse large B cell lymphoma (DLBCL). Large trials (ZUMA-1/JULIET/TRANSCEND) have highlighted that many patients fail to achieve durable responses. Several groups have reported on the phenomenon of CD19 immune escape as a cause (Grupp et al, NEJM 2013, Neelapu et al, NEJM 2017) and the NIH Pediatric Oncology Branch has shown CD22 as an alternative target (Fry et al, Nat Med. 2018). We developed a bi-specific CAR construct targeting CD19 & CD22 with intracellular signaling domains incorporating 4-1BB and CD3ζ (CD19/CD22.BB.z) to overcome CD19 immune escape. Here, we present our Phase I experience with this bi-specific CAR in adults. This is a single institution phase I dose escalation study enrolling patients Age ≥ 18 years with relapsed/refractory B-ALL or DLBCL after standard therapies. Primary aim is to determine feasibility of manufacturing the bi-specific CAR and safety at three dose levels (1 x 106 CAR T cells/kg, 3 x 106 CAR T cells/kg, 1 x 107 CAR T cells/kg). Clinical response was evaluated as a secondary endpoint utilizing standard response criteria for ALL and DLBCL. All patients underwent lymphodepletion with cyclophosphamide (500mg/m2 daily x3 doses) and fludarabine (30mg/m2 daily x 3 doses) followed by CAR infusion two days later. Patients were assessed at pre-defined time-points (Day 28, Month 3, 6, 9, 12 then every 6-12 months) with correlative assessments including immunophenotyping, single cell RNAseq, CAR qtPCR, serum and single cell cytokine analysis. Seven adult patients (5 DLBCL, 2 ALL), aged 35 - 75 years have been enrolled and 6 treated, all at dose Level 1 [Table 1]. The first 3 patients received freshly harvested cells and the remaining received cryopreserved cells (1 patient treated twice received initial fresh then cryopreserved product). None received systemic bridging therapy before CAR T infusion. Six patients developed reversible cytokine release syndrome (CRS,4 with Grade 1, 2 with Grade 2), onset between Day 1 to 13, and 2 patients received tocilizumab & systemic steroids. Three patients developed neurotoxicity (1 with grade 2, Day 8-11 and the others grade 1) with grade 2 neurotoxicity managed with steroids. Four patients required growth factor support beyond Day 28 and all treated patients show persistent B-cell aplasia. Two patients achieved CR: an ALL patient with disease in bone marrow/blood/CNS was MRD negative at day 28 & 60; a 75yo DLBCL patient achieved PR at day 28 and CR at month 3. Three others have ongoing PR and one died of progressive disease after initial PR at Day 28. A patient with PD at Day 28 subsequently treated with radiation and 2-months of revlimid/rituximab, now has no detectable disease 6 months post CAR-T. One patient with initial 6-month PR received a second infusion due to PD, did not develop CRS or CRES with 2nd infusion and has SD at Day 28 Notably, the patient experienced a lack of CAR-T expansion with the second infusion, raising the possibility of an immunogenic response to the CAR-T cell infusion. Flow analysis of all patients' peripheral blood showed CAR expansion peaked at median Day 13 (range Day 10-20) and CARs remained detectable [Figure 1]. Multi-parametric CyTOF phenotyping of the CAR19-22 products showed significant numbers of transduced CAR-T memory stem cells (phenotype: CD3+CD8+CD45RA+CD127+CD27+CCR7+). Single cell cytokine secretion analysis (Isoplexis,Rossi et al Blood 2018) revealed high polyfunctional strength index (PSI) in both CD4+ and CD8+ cell subsets in each patient's pre-infusion CAR product that reflected phenotypic expansion in patients. Additional correlative studies, including cytokine analysis, qtPCR based CAR quantification and CyTOF phenotypic analysis of the CAR-T cells will be presented. This first adult phase I trial of bi-specific CAR targeting CD19 & CD22 shows low toxicity with promising efficacy including achievement of CR in adult DLBCL and ALL patients. We have escalated dose to 3x 106 CAR T cells/kg and an expansion study of 60 patients will follow. CAR-T cells expanded within the first 20 days and continue to be detectable through 6 months. Disclosures Muffly: Shire Pharmaceuticals: Research Funding; Adaptive Biotechnologies: Research Funding. Miklos:Janssen: Consultancy, Research Funding; Genentech: Research Funding; Pharmacyclics - Abbot: Consultancy, Research Funding; Kite - Gilead: Consultancy, Research Funding; Adaptive Biotechnologies: Consultancy, Research Funding; Novartis: Consultancy, Research Funding.