ALBERT

Description: CD19-targeted chimeric antigen receptor-engineered (CD19 CAR) T cell therapy has shown significant efficacy for relapsed or refractory (R/R) B-cell malignancies. Yet CD19 CAR T cells fail to induce durable responses in most patients. Second infusions of CD19 CAR T cells (CART2) have been considered as a possible approach to improve outcomes. We analyzed data from 44 patients with R/R B-cell malignancies (ALL, n=14; CLL, n=9; NHL, n=21) who received CART2 on a phase 1/2 trial at our institution. Despite a CART2 dose increase in 82% of patients, we observed a low incidence of severe toxicity after CART2 (grade ≥3 CRS, 9%; grade ≥3 neurotoxicity, 11%). After CART2, CR was achieved in 22% of CLL, 19% of NHL, and 21% of ALL patients. The median durations of response after CART2 in CLL, NHL, and ALL patients were 33, 6, and 4 months, respectively. Addition of fludarabine to cyclophosphamide-based lymphodepletion before CART1 and an increase in the CART2 dose compared to CART1 were independently associated with higher overall response rates and longer progression-free survival after CART2. We observed durable CAR T-cell persistence after CART2 in patients who received Cy-Flu lymphodepletion before CART1 and a higher CART2 compared to CART1 cell dose. The identification of two modifiable pre-treatment factors independently associated with better outcomes after CART2 suggests strategies to improve in vivo CAR T-cell kinetics and responses after repeat CAR T-cell infusions, and has implications for the design of trials of novel CAR T-cell products after failure of prior CAR T-cell immunotherapies.

Description: Background CD19-targeted chimeric antigen receptor-engineered (CD19 CAR)-T cell immunotherapy has shown promising efficacy in patients with relapsed or refractory (R/R) B-cell malignancies. The potential benefits of repeat infusions of CD19 CAR-T cells are unknown, and the factors associated with response, CAR-T cell in vivo expansion, and progression-free survival (PFS) after repeat infusion of CD19 CAR-T cells have not been investigated. Methods We analyzed the outcomes of patients with R/R B-cell malignancies after a second infusion of CD19 CAR-T cells (CART2) on a phase 1/2 trial (NCT01865617) at our institution. Responses after CAR-T cell therapy were evaluated around day 28 after infusion and defined according to the 2018 NCCN guidelines for acute lymphoblastic leukemia (ALL), 2018 iwCLL for chronic lymphocytic leukemia (CLL), and the Lugano criteria for non-Hodgkin lymphoma (NHL). Logistic, Cox and linear regression were used for multivariable analyses of response, progression-free survival and peak CD8+ CAR-T in blood, respectively. Bayesian model averaging was performed for variable selection. Results Forty-four patients evaluable for response (ALL, n=14; CLL, n=11; NHL, n=19) were included in this study. The median age at the time of CART2 was 58 (range, 23-73). Patients were heavily pre-treated (median prior therapies, 6; range, 2-13), and 16 patients (36%) had bulky (≥ 5cm) nodal or extramedullary disease. The median time from the first CAR-T infusion (CART1) to CART2 was 70 days (range, 28-712). Twenty-eight patients (64%) had received a CART1 dose ≥ 2x106 CAR-T cells/kg. Fifteen patients (32%) had not responded to CART1, 22 (50%) relapsed or progressed after having initially responded (complete response [CR], n=15; partial response [PR], n=7) to CART1; 7 (16%) received CART2 in PR after CART1. All characteristics are shown in the Table. We observed responses in all disease types, including 3 of 14 ALL patients (21%; all CR/CRi), 4 of 11 CLL patients (36%; CR/CRi, n=3; partial response [PR], n=1), and 9 of 19 NHL patients (47%; CR, n=2; PR, n=7). After a median follow-up of 43 months (range, 16-66) in alive and responding patients, the estimated 4-year PFS probability in responders was 23% (95% CI, 9-59%). The 4-year overall survival probability in responders was 36% (95% CI 19-71%) compared to 24% (95% CI, 12-47) in non-responders. Multivariable logistic regression modeling identified predictors of response after CART2: CART1 lymphodepletion (high-intensity cyclophosphamide and fludarabine [CyFlu] vs no CyFlu, OR=12.19, 95% CI, 1.10-1689.85, p=0.04), and peak of in vivo CAR-T cell expansion after CART2 (OR=2.31 per log10 CD8+ CAR-T cell/µL increase, 95% CI, 1.17-5.29, p=0.01). In a multivariable Cox model, a higher peak of CD8+ CAR-T cells after CART2 (HR=0.47 per log10 CD8+ CAR-T cell/µL increase, 95%CI, 0.33-0.68, p CART1 cell dose was associated with longer PFS (HR=0.36, 95% CI, 0.16-0.86, p=0.02). This suggested that CD8+ CAR-T cell peak after CART2 and factors increasing CART2 peak (e.g. prevention of immune rejection or increase in the infused cell dose) are key elements associated with outcomes of CART2. Hence, we looked at factors associated with higher CD8+ CART2 peak. In multivariable linear regression, CART1 CyFlu predicted a higher peak of CD8+ CAR-T cells after CART2 (high-intensity CyFlu vs no CyFlu, p

Description: Introduction Lymphodepletion chemotherapy followed by infusion of T cells engineered to express a CD19-specific chimeric antigen receptor (CAR) has shown remarkable efficacy in patients (pts) with relapsed/refractory (R/R) CD19+ B-cell malignancies, with high response rates reported in non-Hodgkin lymphoma (NHL). Durable responses have been observed in a subset of pts, but the factors associated with these long-term remissions have not been identified. We studied adults with R/R CD19+ B-cell NHL treated with cyclophosphamide and fludarabine lymphodepletion followed by infusion of 2 x 106 CD19 CAR-T cells/kg, and identified factors before and after CAR-T cell infusion that are associated with progression-free survival (PFS). Methods We conducted a phase 1/2 open-label clinical trial (NCT01865617) with the primary objective of evaluating the feasibility and safety of infusing a defined composition of CD4+ and CD8+ CD19 CAR-T cells after lymphodepletion chemotherapy in pts with R/R CD19+ B-cell malignancies. Best responses are reported according to the Lugano criteria (Cheson, JCO 2014). PFS was defined as the time from CAR-T cell infusion until disease progression or death, without censoring for new therapy. Logistic regression and penalized Cox regression multivariable modeling using elastic net were performed for analysis of response and PFS, respectively. Results Characteristics of the 57 pts in the study are shown in Table 1. One patient with incomplete response assessment was excluded. For the 56 remaining pts, the best overall response rate (ORR) without additional therapy was 57% (95% confidence interval [CI], 43-70%), with 48% achieving complete remission (CR; 95% CI, 35-62%). Most pts with partial response (PR) or stable disease (SD) after initial restaging at 4 weeks after CAR-T cell infusion received new therapy (11 of 15, 73%). All pts with PR/SD on initial restaging who did not receive additional therapy after CAR-T cells (n = 4) subsequently achieved CR. The duration of persistence of CAR-T cells was longer in pts who did not receive new therapy (15.7 vs. 5.3 months; P = .06). Eight of 9 pts with indolent histology achieved CR (89%; 95% CI, 51-99%). For the 47 pts with aggressive NHL, the best ORR was 51% (95% CI, 36-66%), with 40% (95% CI, 27-56%) achieving CR. Among aggressive NHL subtypes, pts with DLBCL (n = 28) had best ORR and CR rates of 50% (95% CI, 33-67%) and 43% (95% CI, 25-63%), respectively. In pts with aggressive lymphoma, multivariable analysis showed that the probability of achieving CR was independently associated with a lower pre-lymphodepletion serum LDH concentration (P = .003) and greater increase in serum MCP-1 concentration from a pre-lymphodepletion timepoint to immediately before CAR-T cell infusion (P = .01). Analysis of pts with all histologic subtypes showed that those achieving CR had better PFS and overall survival (OS) compared to those who did not achieve CR (median PFS: CR, not reached; non-CR, 1.35 month; Figure 1). In pts achieving CR, after a median follow-up of 20.2 months (range 2.5-32.4 months), the 24-month probabilities of PFS and OS were 59% (95% CI, 41-84%) and 79% (95% CI, 64-97%), respectively. No pts with indolent NHL who achieved CR (n = 8) have relapsed with a median follow-up of 14.5 months (range, 10.7-30.1 months). For pts with aggressive lymphoma who achieved CR, after a median follow-up of 26.9 months (range, 2.5-32.4 months), the median PFS was 20.0 months (95% CI, 9.2-not reached), and 24-month probabilities of PFS and OS were 46% (95% CI, 28-76%) and 72% (95% CI, 54-96%), respectively. In aggressive NHL, multivariable analysis suggested that, in addition to being associated with the probability of achieving CR, serum LDH and MCP-1 concentration also impacted the probability of longer PFS. The model found that lower pre-lymphodepletion serum LDH (P = .0004) and higher serum MCP-1 peak after CAR-T cell infusion (P = .05), along with higher serum IL-7 (P = .02) and lower serum IL-18 (P = .02) concentrations before lymphodepletion were independently associated with better PFS. Similar findings were obtained after multivariable analysis was performed only in those who had achieved CR. Conclusion CR after CD19 CAR-T cell therapy appears to be a strong predictor of PFS in adult pts with B-cell NHL. Identification of additional factors associated with better PFS might guide future management strategies for pts achieving CR after CD19 CAR-T cell therapy. Disclosures Hirayama: DAVA Oncology: Honoraria. Hay:DAVA Oncology: Honoraria. Li:Juno Therapeutics: Employment, Equity Ownership. Lynch:Incyte: Research Funding; Johnson Graffe Keay Moniz and Wick LLP: Consultancy; Juno Therapeutics: Research Funding; Rhizen Pharmaceuticals: Research Funding; Takeda: Research Funding. Till:Mustang Bio: Patents & Royalties, Research Funding. Kiem:Homology Medicine: Consultancy; Rocket Pharmaceuticals: Consultancy; Magenta: Consultancy. Ramos:Seattle Genetics: Employment, Equity Ownership. Shadman:Gilead Sciences: Research Funding; Genentech: Consultancy; Pharmacyclics: Research Funding; Celgene: Research Funding; Mustang Biopharma: Research Funding; Genentech: Research Funding; TG Therapeutics: Research Funding; Acerta Pharma: Research Funding; AbbVie: Consultancy; Verastem: Consultancy; Beigene: Research Funding; AstraZeneca: Consultancy; Qilu Puget Sound Biotherapeutics: Consultancy. Cassaday:Jazz Pharmaceuticals: Consultancy; Amgen: Consultancy, Research Funding; Kite Pharma: Research Funding; Adaptive Biotechnologies: Consultancy; Merck: Research Funding; Pfizer: Consultancy, Research Funding; Seattle Genetics: Other: Spouse Employment, Research Funding; Incyte: Research Funding. Acharya:Juno Therapeutics: Research Funding; Teva: Honoraria. Riddell:Cell Medica: Membership on an entity's Board of Directors or advisory committees; Juno Therapeutics: Equity Ownership, Patents & Royalties, Research Funding; NOHLA: Consultancy; Adaptive Biotechnologies: Consultancy. Maloney:GlaxoSmithKline: Research Funding; Juno Therapeutics: Research Funding; Roche/Genentech: Honoraria; Seattle Genetics: Honoraria; Janssen Scientific Affairs: Honoraria. Turtle:Bluebird Bio: Consultancy; Juno Therapeutics / Celgene: Consultancy, Patents & Royalties, Research Funding; Nektar Therapeutics: Consultancy, Research Funding; Eureka Therapeutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Precision Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Caribou Biosciences: Consultancy; Gilead: Consultancy; Adaptive Biotechnologies: Consultancy; Aptevo: Consultancy.

Description: Background We reported durable responses to CD19-specific chimeric antigen receptor-modified T-cell therapy (JCAR014) in relapsed/refractory (R/R) chronic lymphocytic leukemia (CLL) patients (pts) after prior failure of ibrutinib (Turtle, JCO 2017; NCT01865617). In those pts, ibrutinib was not administered during CAR-T cell immunotherapy. Continuation of ibrutinib through leukapheresis, lymphodepletion and CAR-T cell therapy may prevent tumor progression after ibrutinib withdrawal, mobilize tumor into the blood, improve CAR-T cell function, and decrease cytokine release syndrome (CRS). Methods We conducted a phase 1/2 study of CD19 CAR-T cell immunotherapy in R/R CLL pts and established a regimen of cyclophosphamide and fludarabine (Cy/Flu) lymphodepletion followed by JCAR014 at 2 x 106 CAR-T cells/kg (Turtle, JCO 2017). We then compared outcomes of these pts (No-ibr cohort) with a subsequent cohort that received Cy/Flu with 2 x 106/kg JCAR014 CAR-T cells with concurrent ibrutinib (420 mg/d) from at least 2 weeks prior to leukapheresis until at least 3 months after JCAR014 infusion (Ibr cohort). Dose reduction was permitted for toxicity. CRS was graded by consensus criteria (Lee, Blood 2014) and neurotoxicity and other adverse events were graded by CTCAE v4.03. Response was evaluated according to 2008 IWCLL criteria. Results Seventeen and 19 pts were treated in the Ibr and No-ibr cohorts, respectively. Pt characteristics were comparable (Table 1). Progression on ibrutinib was noted in 16 (94%) and 18 pts (95%) in the Ibr and No-ibr cohorts, respectively, and prior ibrutinib intolerance was reported in 1 pt in each cohort. The time to intolerance or failure of ibrutinib prior to treatment with JCAR014 was longer, and the pre-leukapheresis LDH was lower in the Ibr compared to the No-ibr cohort. The median follow-up in responders was 98 and 764 days in the Ibr and No-ibr cohorts, respectively. Administration of ibrutinib with Cy/Flu and JCAR014 was well tolerated in most pts; ibrutinib was reduced or discontinued in 6 pts (35%) at a median of 21 days after JCAR014 infusion. In the Ibr cohort, 1 pt with grade 2 CRS developed fatal presumed cardiac arrhythmia and 1 pt developed a subdural hematoma in the setting of trauma and thrombocytopenia. No differences in the incidences of grade ≥3 cytopenias were observed. Concurrent ibrutinib administration did not appear to affect the frequency or severity of neurotoxicity. Although the proportions of pts with grade ≥1 CRS were similar between cohorts (76% vs 89%, P = 0.39), the severity of CRS (grade ≥3 CRS: Ibr, 0%; No-Ibr, 26%; P = 0.05) and serum peak IL-8 (P = 0.04), IL-15 (P = 0.003) and MCP-1 (P = 0.004) concentrations were lower in the Ibr cohort. However, we found comparable CD8+ (P = 0.29) and higher CD4+ (P = 0.06) CAR-T cell counts in blood in the Ibr cohort. Sixteen pts (94%) and 18 pts (95%) in the Ibr and No-ibr cohorts, respectively, have completed response assessment. We observed a higher proportion of responders (complete and partial remission) by IWCLL criteria in the Ibr compared to the No-ibr cohort (88% vs 56%, respectively, P = 0.06). Ten of 12 pts (83%) with lymph node disease before treatment with Cy/Flu and JCAR014 in the Ibr cohort achieved CR or PR by IWCLL imaging criteria, compared to 10/17 pts (59%) in the No-ibr cohort (P = 0.23). The proportion of pts with pretreatment bone marrow (BM) disease who had no disease by flow cytometry after CAR-T cell immunotherapy was similar in the Ibr compared to the No-ibr cohort (75% vs 65%, P = 0.71). However, among pts with no disease by BM flow cytometry after CAR-T cell immunotherapy, a higher proportion of pts in the Ibr cohort had no malignant IGH sequences at 4 weeks (83% vs 60%, respectively, P = 0.35). We performed univariate logistic regression analysis for response by IWCLL criteria and variables with P 〈 0.10 were considered for stepwise multivariable analysis (Table 2). In the multivariable analysis, the Ibr cohort and a lower pre-treatment SUVmax on PET imaging were each associated with a higher probability of response by IWCLL criteria (Ibr cohort, OR = 14.02, 95%CI [0.52-379.61], P = 0.05; SUVmax, OR = 1.31 per SUV unit decrease, 95%CI [1.05-1.67], P 〈 0.001). Conclusion Administration of ibrutinib from 2 weeks before leukapheresis until 3 months after JCAR014 was well tolerated in most pts. This approach might decrease the incidence of severe CRS and improve responses in pts with R/R CLL. Disclosures Hirayama: DAVA Oncology: Honoraria. Hay:DAVA Oncology: Honoraria. Li:Juno Therapeutics: Employment, Equity Ownership. Lymp:Juno Therapeutics: Employment, Equity Ownership. Till:Mustang Bio: Patents & Royalties, Research Funding. Kiem:Magenta: Consultancy; Homology Medicine: Consultancy; Rocket Pharmaceuticals: Consultancy. Shadman:TG Therapeutics: Research Funding; Celgene: Research Funding; Gilead: Research Funding; Qilu Puget Sound Biotherapeutics: Consultancy; AstraZeneca: Consultancy; Verastem: Consultancy; Beigene: Research Funding; Mustang: Research Funding; Genentech: Consultancy, Research Funding; Pharmacyclics: Research Funding; Acerta: Research Funding; Abbvie: Consultancy. Cassaday:Merck: Research Funding; Pfizer: Consultancy, Research Funding; Amgen: Consultancy, Research Funding; Seattle Genetics: Other: Spouse Employment, Research Funding; Incyte: Research Funding; Kite Pharma: Research Funding; Adaptive Biotechnologies: Consultancy; Jazz Pharmaceuticals: Consultancy. Acharya:Juno Therapeutics: Research Funding; Teva: Honoraria. Riddell:Juno Therapeutics: Equity Ownership, Patents & Royalties, Research Funding; Adaptive Biotechnologies: Consultancy; NOHLA: Consultancy; Cell Medica: Membership on an entity's Board of Directors or advisory committees. Maloney:GlaxoSmithKline: Research Funding; Juno Therapeutics: Research Funding; Seattle Genetics: Honoraria; Roche/Genentech: Honoraria; Janssen Scientific Affairs: Honoraria. Turtle:Nektar Therapeutics: Consultancy, Research Funding; Juno Therapeutics / Celgene: Consultancy, Patents & Royalties, Research Funding; Aptevo: Consultancy; Precision Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Caribou Biosciences: Consultancy; Adaptive Biotechnologies: Consultancy; Eureka Therapeutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Bluebird Bio: Consultancy; Gilead: Consultancy.

Description: INTRODUCTION: When introduced into polyclonal T cells, chimeric antigen receptors (CAR) redirect specificity of the engineered T cells to an antigen recognized by the CAR. We conducted a phase I/II clinical trial of treatment of relapsed and refractory CD19-positive B cell malignancies using a defined formulation of CD4+ and CD8+ CD19-specific CAR-T cells (NCT01865617). Little is known about the transcriptional heterogeneity of CAR-T cells in the infused product and their clonal kinetics after adoptive transfer. METHODS: To understand the factors that impact clonal CAR-T cell behavior in vivo, we performed TCRBV sequencing and single cell transcriptional profiling (10X Genomics) on CD8+ CAR-T cells isolated from infused products and the blood of treated patients. TCRBV sequencing was performed on 0.8 to 1.5 million cells from the infused product and 700-65,000 CAR-T cells from blood after CAR-T infusion. For single-cell RNA sequencing (scRNAseq), we obtained paired 5' gene expression and V(D)J data from individual CAR-T cells isolated from infused products and from the blood at the peak of in vivo expansion, after contraction, and at a late time point. RESULTS: High-throughput sequencing of the TCRBV genes revealed that CAR-T cells were polyclonal in the infused products, and during in vivo expansion and contraction, and at late times (≥ 3 months) after adoptive transfer. We evaluated the diversity of the TCRBV repertoire using the Shannon entropy index, and found that clonal diversity was highest in the infused product and declined at later time points after adoptive transfer. Loss of diversity after adoptive transfer was due to both expansion of higher frequency CAR-T cell clones and loss of low-frequency clones. We identified distinct CAR-T cell clones in the infused product and in blood at multiple time points after infusion that exhibited different kinetics of expansion and contraction. To examine the transcriptional programs that regulate the fate of CAR-T cells after infusion, we performed scRNAseq on CD8+ CAR-T cells, and found transcriptional heterogeneity in the infused products, which declined in CD8+ CAR-T cells isolated from patient blood after adoptive transfer. Gene set enrichment analysis showed that the infused products expressed higher levels of genes associated with hypoxia, glycolysis, and proliferation, and lower levels of genes associated with cytotoxicity compared to CAR-T cells isolated after adoptive transfer. In the infused product, genes associated with cytotoxicity were expressed at higher levels in CAR-T cells harboring clonotypes that were subsequently represented at relatively higher levels in vivo after adoptive transfer. CONCLUSIONS: There is transcriptional heterogeneity in the infused product and distinct CAR-T cell clones exhibit different kinetics of expansion and contraction after infusion. A better understanding of the kinetics of clonal expansion of CAR-T cells after adoptive transfer may provide insight into strategies to improve CAR-T cell immunotherapy. Disclosures Turtle: Nektar Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Precision Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Eureka Therapeutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Caribou Biosciences: Membership on an entity's Board of Directors or advisory committees; Juno/Celgene: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding.

Description: Background CD19 CAR T immunotherapy has been successful in achieving durable remissions in some patients with relapsed/refractory B cell lymphomas, but disease progression and loss of CAR T cell persistence remains problematic. Interleukin 15 (IL-15) is known to support T cell proliferation and survival, and therefore may enhance CAR T cell efficacy, however, utilizing native IL-15 is challenging due to its short half-life and poor tolerability in the clinical setting. NKTR-255 is a polymer-conjugated IL-15 that retains binding affinity to IL15Rα and exhibits reduced clearance, providing sustained pharmacodynamic responses. We investigated the effects of NKTR-255 on human CD19 CAR T cells both in vitro and in an in vivo xenogeneic B cell lymphoma model and found improved survival of lymphoma bearing mice receiving NKTR-255 and CAR T cells compared to CAR T cells alone. Here, we extend upon these findings to further characterize CAR T cells in vivo and examine potential mechanisms underlying improved anti-tumor efficacy. Methods CD19 CAR T cells incorporating 4-1BB co-stimulation were generated from CD8 and CD4 T cells isolated from healthy donors. For in vitro studies, CAR T cells were incubated with NKTR-255 or native IL-15 with and without CD19 antigen. STAT5 phosphorylation, CAR T cell phenotype and CFSE dilution were assessed by flow cytometry and cytokine production by Luminex. For in vivo studies, NSG mice received 5x105 Raji lymphoma cells IV on day (D)-7 and a subtherapeutic dose (0.8x106) of CAR T cells (1:1 CD4:CD8) on D0. To determine optimal start date of NKTR-255, mice were treated weekly starting on D-1, 7, or 14 post CAR T cell infusion. Tumors were assessed by bioluminescence imaging. Tumor-free mice were re-challenged with Raji cells. For necropsy studies mice received NKTR-255 every 7 days following CAR T cell infusion and were euthanized at various timepoints post CAR T cell infusion. Results Treatment of CD8 and CD4 CAR T cells in vitro with NKTR-255 resulted in dose dependent STAT5 phosphorylation and antigen independent proliferation. Co-culture of CD8 CAR T cells with CD19 positive targets and NKTR-255 led to enhanced proliferation, expansion and TNFα and IFNγ production, particularly at lower effector to target ratios. Further studies showed that treatment of CD8 CAR T cells with NKTR-255 led to decreased expression of activated caspase 3 and increased expression of bcl-2. In Raji lymphoma bearing NSG mice, administration of NKTR-255 in combination with CAR T cells increased peak CAR T cell numbers, Ki-67 expression and persistence in the bone marrow compared to mice receiving CAR T cells alone. There was a higher percentage of EMRA like (CD45RA+CCR7-) CD4 and CD8 CAR T cells in NKTR-255 treated mice compared to mice treated with CAR T cells alone and persistent CAR T cells in mice treated with NKTR-255 were able to reject re-challenge of Raji tumor cells. Additionally, starting NKTR-255 on D7 post T cell infusion resulted in superior tumor control and survival compared to starting NKTR-255 on D-1 or D14. Conclusion Administration of NKTR-255 in combination with CD19 CAR T cells leads to improved anti-tumor efficacy making NKTR-255 an attractive candidate for enhancing CAR T cell therapy in the clinic. Disclosures Chou: Nektar Therapeutics: Other: Travel grant. Fraessle:Technical University of Munich: Patents & Royalties. Busch:Juno Therapeutics/Celgene: Consultancy, Equity Ownership, Research Funding; Kite Pharma: Equity Ownership; Technical University of Munich: Patents & Royalties. Miyazaki:Nektar Therapeutics: Employment, Equity Ownership. Marcondes:Nektar Therapeutics: Employment, Equity Ownership. Riddell:Juno Therapeutics: Equity Ownership, Patents & Royalties, Research Funding; Adaptive Biotechnologies: Consultancy; Lyell Immunopharma: Equity Ownership, Patents & Royalties, Research Funding. Turtle:Allogene: Other: Ad hoc advisory board member; Novartis: Other: Ad hoc advisory board member; Humanigen: Other: Ad hoc advisory board member; Nektar Therapeutics: Other: Ad hoc advisory board member, Research Funding; Caribou Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; T-CURX: Membership on an entity's Board of Directors or advisory committees; Juno Therapeutics: Patents & Royalties: Co-inventor with staff from Juno Therapeutics; pending, Research Funding; Precision Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Eureka Therapeutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Kite/Gilead: Other: Ad hoc advisory board member.

Description: Patients with follicular lymphoma (FL) with early relapse after initial chemoimmunotherapy, refractory disease, or histologic transformation (tFL) have limited progression-free and overall survival. We report efficacy and long-term follow-up of 21 patients with relapsed/refractory (R/R) FL (n = 8) and tFL (n = 13) treated on a phase 1/2 clinical trial with cyclophosphamide and fludarabine lymphodepletion followed by infusion of 2 × 106 CD19-directed chimeric antigen receptor–modified T (CAR-T) cells per kilogram. The complete remission (CR) rates by the Lugano criteria were 88% and 46% for patients with FL and tFL, respectively. All patients with FL who achieved CR remained in remission at a median follow-up of 24 months. The median duration of response for patients with tFL was 10.2 months at a median follow-up of 38 months. Cytokine release syndrome occurred in 50% and 39%, and neurotoxicity in 50% and 23% of patients with FL and tFL, respectively, with no severe adverse events (grade ≥3). No significant differences in CAR-T cell in vivo expansion/persistence were observed between FL and tFL patients. CD19 CAR-T cell immunotherapy is highly effective in adults with clinically aggressive R/R FL with or without transformation, with durable remission in a high proportion of FL patients. This trial was registered at clinicaltrials.gov as #NCT01865617.

Description: Factors associated with durable remission after CD19 chimeric antigen receptor (CAR)-modified T-cell immunotherapy for aggressive B-cell non-Hodgkin lymphoma (NHL) have not been identified. We report multivariable analyses of factors affecting response and progression-free survival (PFS) in patients with aggressive NHL treated with cyclophosphamide and fludarabine lymphodepletion followed by 2 × 106 CD19-directed CAR T cells/kg. The best overall response rate was 51%, with 40% of patients achieving complete remission. The median PFS of patients with aggressive NHL who achieved complete remission was 20.0 months (median follow-up, 26.9 months). Multivariable analysis of clinical and treatment characteristics, serum biomarkers, and CAR T-cell manufacturing and pharmacokinetic data showed that a lower pre-lymphodepletion serum lactate dehydrogenase (LDH) level and a favorable cytokine profile, defined as serum day 0 monocyte chemoattractant protein-1 (MCP-1) and peak interleukin-7 (IL-7) concentrations above the median, were associated with better PFS. MCP-1 and IL-7 concentrations increased after lymphodepletion, and higher intensity of cyclophosphamide and fludarabine lymphodepletion was associated with higher probability of a favorable cytokine profile. PFS was superior in patients who received high-intensity lymphodepletion and achieved a favorable cytokine profile compared with those who received the same intensity of lymphodepletion without achieving a favorable cytokine profile. Even in high-risk patients with pre-lymphodepletion serum LDH levels above normal, a favorable cytokine profile after lymphodepletion was associated with a low risk of a PFS event. Strategies to augment the cytokine response to lymphodepletion could be tested in future studies of CD19 CAR T-cell immunotherapy for aggressive B-cell NHL. This trial was registered at www.clinicaltrials.gov as #NCT01865617.

Description: Background We previously reported high response rates and durable remissions in patients (pts) with relapsed/refractory (R/R) B-cell acute lymphoblastic leukemia (ALL; Turtle, JCI 2016) and non-Hodgkin lymphoma (NHL; Turtle, Sci Transl Med 2016) treated with CD19-specific chimeric antigen receptor T (CD19 CAR-T) cells. In a subset of pts, we identified CD8+ T cell responses to epitopes in the murine CD19-binding single chain variable fragment (scFv) of the CAR that could limit CAR-T cell persistence and responses to subsequent infusions. In an effort to reduce the potential for immune CAR-T cell rejection, the murine CD19-binding scFv of the CAR was replaced with a fully human scFv linked to 4-1BB and CD3z signaling domains (JCAR021; Sommermeyer, Leukemia 2017). Here we report the initial clinical results of immunotherapy with JCAR021. Methods We initiated a phase I trial investigating lymphodepletion with cyclophosphamide 300 mg/m2/d and fludarabine 30 mg/m2/d for 3 days (Cy/Flu) followed by infusion of JCAR021 in pts with R/R ALL and aggressive NHL (NCT03103971). Pts were enrolled into 1 of 3 cohorts: high marrow burden ALL (HMB; 〉 5% blasts in bone marrow [BM] before lymphodepletion); low marrow burden ALL (LMB; ≤ 5% blasts in BM before lymphodepletion); and NHL. The starting dose was 7x104 JCAR021 cells/kg for the HMB ALL cohort, and 7x105 JCAR021 cells/kg in both the LMB ALL and NHL cohorts. Dose escalation/de-escalation follows a modified toxicity probability interval algorithm (Guo, Contemp Clin Trials 2017). Responses in the NHL cohort and in the HMB/LMB ALL cohorts were determined by the Lugano criteria (Cheson, JCO 2014) and the 2018 NCCN guidelines, respectively. Cytokine release syndrome (CRS) was graded according to consensus criteria (Lee, Blood 2014) and neurotoxicity was graded according to CTCAE v4.03. Results Pt characteristics are detailed in Table 1. As of June 15, 2018, 9 pts were enrolled on the trial. Two pts did not receive JCAR021: one pt was excluded after aggressive NHL was reclassified as indolent after pathology review and one pt had no detectable disease upon pre-treatment restaging. The 7 pts who received JCAR021 had a median age of 63 years (range: 29 - 69). Both pts in the LMB ALL cohort had bulky extramedullary disease (〉 5 cm diameter). One patient (LMB ALL cohort) had failed two allogeneic transplants and one patient (HMB ALL cohort) had failed an allogeneic transplant prior to treatment with JCAR021. Four of 4 pts in the NHL cohort and 2 of 2 pts in the LMB ALL cohort received 7x105 JCAR021 cells/kg. The pt treated in the HMB ALL cohort received 7x104 JCAR021 cells/kg. No pt in any cohort developed grade ≥ 3 CRS. All ALL pts developed grade 2 CRS. No pts with NHL developed CRS; one pt in the NHL cohort who had CNS disease prior to CAR-T cell immunotherapy developed grade 3 neurotoxicity in the absence of CRS. We did not observe other neurologic events. No other grade ≥ 3 non-hematopoietic organ toxicity was observed and all 7 treated pts have completed response evaluation. Four weeks after infusion of a low dose of JCAR021, both patients in the LMB ALL cohort had undetectable marrow disease by high resolution flow cytometry and regression of bulky extramedullary disease (1 complete response [CR] and 1 partial response [PR] by PET-CT). One pt treated with a low dose (7x104 cells/kg) of JCAR021 in the HMB ALL cohort did not achieve CR (decrease in BM blasts from 79.8% to 29.5%) but CNS disease was cleared by flow cytometry. In the NHL cohort, we observed objective responses in 2 of 4 patients (1 CR, 1 PR). JCAR021 was detected in blood by flow cytometry and/or quantitative PCR for up to 112 days after infusion. Conclusion JCAR021 appears to have a favorable toxicity profile in R/R ALL and NHL pts. JCAR021 cells expanded in vivo and have persisted in all pts. We observed responses at very low doses of CAR-T cells in ALL pts with bulky disease. This trial continues to enroll to define optimal dosing and determine the safety and efficacy of JCAR021. Disclosures Hirayama: DAVA Oncology: Honoraria. Hay:DAVA Oncology: Honoraria. Till:Mustang Bio: Patents & Royalties, Research Funding. Kiem:Homology Medicine: Consultancy; Magenta: Consultancy; Rocket Pharmaceuticals: Consultancy. Shadman:TG Therapeutics: Research Funding; Mustang: Research Funding; Gilead: Research Funding; Pharmacyclics: Research Funding; AstraZeneca: Consultancy; Qilu Puget Sound Biotherapeutics: Consultancy; Acerta: Research Funding; Abbvie: Consultancy; Verastem: Consultancy; Genentech: Consultancy, Research Funding; Beigene: Research Funding; Celgene: Research Funding. Cassaday:Amgen: Consultancy, Research Funding; Seattle Genetics: Other: Spouse Employment, Research Funding; Adaptive Biotechnologies: Consultancy; Incyte: Research Funding; Pfizer: Consultancy, Research Funding; Merck: Research Funding; Kite Pharma: Research Funding; Jazz Pharmaceuticals: Consultancy. Acharya:Teva: Honoraria; Juno Therapeutics: Research Funding. Riddell:NOHLA: Consultancy; Adaptive Biotechnologies: Consultancy; Cell Medica: Membership on an entity's Board of Directors or advisory committees; Juno Therapeutics: Equity Ownership, Patents & Royalties, Research Funding. Maloney:Juno Therapeutics: Research Funding; Seattle Genetics: Honoraria; Janssen Scientific Affairs: Honoraria; GlaxoSmithKline: Research Funding; Roche/Genentech: Honoraria. Turtle:Aptevo: Consultancy; Nektar Therapeutics: Consultancy, Research Funding; Caribou Biosciences: Consultancy; Gilead: Consultancy; Juno Therapeutics / Celgene: Consultancy, Patents & Royalties, Research Funding; Bluebird Bio: Consultancy; Eureka Therapeutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Adaptive Biotechnologies: Consultancy; Precision Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees.

Description: Introduction Autologous T cells engineered to express a CD19-specific chimeric antigen receptor (CAR) have shown high overall response rates (ORR) in otherwise treatment-refractory CD19+ B-cell non-Hodgkin lymphoma (NHL); however, not all patients (pts) achieve complete remission (CR). PD-L1 expression on tumor cells and/or other tissues could impair the function of PD-1+ CAR-T cells and the efficacy of CD19 CAR-T cell immunotherapy. PD-1 pathway blockade may enhance the function and antitumor activity of CD19 CAR-T cells. Here we report preliminary data from a phase 1 dose-finding study (NCT02706405) of the safety and feasibility of combination therapy with JCAR014 CD19-specific 4-1BB-costimulated CAR-T cells and escalating doses of durvalumab, an anti-PD-L1 monoclonal antibody, in adults with relapsed/refractory aggressive B-cell NHL. Methods Pts are treated in one of two groups. All pts receive lymphodepletion chemotherapy with cyclophosphamide and fludarabine followed by infusion of JCAR014. Pts in group 1 receive the first infusion of durvalumab (225 mg, 750 mg, or 1500 mg) 21-28 days after treatment with JCAR014. Pts in group 2 receive the first dose of durvalumab (7.5 mg, 22.5 mg, 75 mg, 225 mg, 750 mg, or 1500 mg) 1 day prior to JCAR014 infusion. Up to 10 doses of durvalumab are administered after JCAR014 at the highest identified safe dose at 4-week intervals until toxicity or disease progression. We evaluated the safety, tolerability, and efficacy of the combination therapy and the pharmacokinetic profile of JCAR014 after infusion. Adverse events were graded using the Common Terminology Criteria for Adverse Events (CTCAE) v4.03, with the exception of cytokine release syndrome (CRS), which was graded according to consensus criteria (Lee, Blood 2014). Positron emission tomography/computed tomography was performed approximately 1, 2, 4, 6, 9, and 12 months after JCAR014 infusion and the best anti-tumor response was reported according to the Lugano criteria (Cheson, JCO 2014). Results Patient characteristics are shown in Table 1. Fifteen pts have been treated, including 6 in group 1 who received post-JCAR014 durvalumab doses of 225 mg (n = 3) and 750 mg (n = 3), and 9 in group 2 who received pre-JCAR014 durvalumab doses of 7.5 mg (n = 1), 22.5 mg (n = 1), 75 mg (n = 3), or 225 mg (n = 4). Durvalumab dose escalation is ongoing. JCAR014 manufacturing was successful for all pts. All pts received 2 x 106 JCAR014 CAR-T cells/kg, except the first 2 pts treated on the study who received 7 x 105 CAR-T cells/kg. Of the 13 pts who received JCAR014 at 2 x 106 CAR-T cells/kg, 5 pts (38%) developed CRS (2 grade 1, 2 grade 2, and 1 grade 4) and one (8%) developed grade 1 neurotoxicity. CRS and/or neurotoxicity occurred within 4 weeks of JCAR014 infusion, and were not observed when durvalumab was administered after JCAR014. With the exception of B cell aplasia, no autoimmune adverse events were observed. Twelve of 13 pts who received 2 x 106 CAR-T cells/kg were evaluable for response. One patient, who had grade 4 CRS and biopsy evidence of extensive CAR-T cell infiltration into persistent sites of disease, elected to receive hospice care and died on day 32 after JCAR014 infusion without full response evaluation. The overall response rate was 50% (5 CR, 42%; 1 PR, 8%). Of the 5 pts who achieved CR, 3 were in CR at the first restaging after JCAR014 and 2 subsequently converted to CR after the first post-JCAR014 durvalumab infusion. Only one patient who achieved CR has relapsed (median follow-up 10.6 months, range 3.7-11.8). Continued stable disease or evidence of regression was seen in 4 of 6 (67%) initially non-responding pts who continued durvalumab therapy (median 5 doses, range 1-6). CAR-T cell counts expanded in the peripheral blood within 14 days of JCAR014 infusion in all pts. Higher peak and day 28 CAR-T cell copy numbers in blood by qPCR were observed in responding pts. CAR-T cells were detected for a median of 5.1 months (range, 1.7 to 9.1 months) in responding pts. In vivo re-accumulation of CAR-T cells after the first post-JCAR014 durvalumab dose was observed in the blood of two patients in group 2. Conclusion The combination of JCAR014 with durvalumab for the treatment of adult pts with aggressive B-cell NHL appears safe; however, dose escalation is ongoing. Complete responses were observed both at initial restaging after JCAR014 infusion, and also subsequently in pts continuing durvalumab therapy after initially failing to achieve CR. Disclosures Hirayama: DAVA Oncology: Honoraria. Hay:DAVA Oncology: Honoraria. Till:Mustang Bio: Patents & Royalties, Research Funding. Kiem:Homology Medicine: Consultancy; Magenta: Consultancy; Rocket Pharmaceuticals: Consultancy. Shadman:Verastem: Consultancy; Beigene: Research Funding; Mustang Biopharma: Research Funding; Gilead Sciences: Research Funding; TG Therapeutics: Research Funding; AbbVie: Consultancy; Genentech: Research Funding; Pharmacyclics: Research Funding; Celgene: Research Funding; Qilu Puget Sound Biotherapeutics: Consultancy; Genentech: Consultancy; AstraZeneca: Consultancy; Acerta Pharma: Research Funding. Cassaday:Jazz Pharmaceuticals: Consultancy; Amgen: Consultancy, Research Funding; Merck: Research Funding; Seattle Genetics: Other: Spouse Employment, Research Funding; Pfizer: Consultancy, Research Funding; Adaptive Biotechnologies: Consultancy; Kite Pharma: Research Funding; Incyte: Research Funding. Acharya:Juno Therapeutics: Research Funding; Teva: Honoraria. Riddell:Cell Medica: Membership on an entity's Board of Directors or advisory committees; Juno Therapeutics: Equity Ownership, Patents & Royalties, Research Funding; Adaptive Biotechnologies: Consultancy; NOHLA: Consultancy. Maloney:Roche/Genentech: Honoraria; Juno Therapeutics: Research Funding; Janssen Scientific Affairs: Honoraria; GlaxoSmithKline: Research Funding; Seattle Genetics: Honoraria. Turtle:Precision Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Adaptive Biotechnologies: Consultancy; Bluebird Bio: Consultancy; Gilead: Consultancy; Nektar Therapeutics: Consultancy, Research Funding; Eureka Therapeutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Juno Therapeutics / Celgene: Consultancy, Patents & Royalties, Research Funding; Caribou Biosciences: Consultancy; Aptevo: Consultancy.