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2062 Background: GBM is the most common malignant brain tumor with a median survival of 15 months despite surgery and radio-chemotherapy. The most important mechanism of TMZ resistance is the O6-methylguanine-DNA methyltransferase (MGMT) gene which repairs temozolamide-induced DNA methylation. The MGMT inhibitor O6-benzylguanine (BG) demonstrated efficacy in depleting MGMT and maximizing tumor response in early phase clinical trials. However, MGMT expression is also low in hematopoietic cells, so this approach led to unacceptable bone marrow toxicity and thus has been abandoned. We hypothesized that chemoprotection of hematopoietic HPC with an MGMT mutant (MGMT-P140K) characterized by normal methyltransferase activity, coupled with low affinity for BG would maximize anti-tumor response while enabling patients to tolerate TMZ & BG dose escalation with minimal toxicity. A phase I trial was performed to test this hypothesis. Methods: 10 adults with newly diagnosed MGMT unmethylated, IDH-1 WT, GBM underwent standard surgery and radiation, followed by transplantation with autologous CD34+ HPC engineered to express MGMT-P140K using a lentiviral vector. We tested tolerance and efficacy of three different paradigms for conditioning bone marrow and re-infusion of HPC. To assess chemo-protection, patients’ blood counts and transgene marking were monitored during and after treatment, as was toxicity, response, and progression-free and overall survival. Results: Treatment was moderately toxic with 3/10 patients suffering grade 3-4 hematologic toxicity; no high grade non-hematologic toxicity was observed . Viral transduction rates ranged from 3-75% and were clearly improved in Arm III utilizing BCNU conditioning and intra-patient dose escalation of TMZ/O6GB. In patients tolerating 3 cycles or more, P140K-MGMT gene markings in peripheral blood and bone marrow cells increased 3-26-fold with only mild (Grade 2-3) mylosuppression consistent with chemo-protection as hypothesized. Median PFS and OS was 22 and 31 months respectfully, and three patients in Arm III are healthy and progression free at 36-39 months. OS exceeded RPA predicted survival by 3.3-fold suggesting clinical benefit. Viral insertion site analysis demonstrate lack of clonal dominance. Conclusions: P140K-MGMT transfected HPC enables TMZ/ BG dose escalation with acceptable toxicity and increased survival in a small cohort of selected patients. A phase II study is ongoing. Clinical trial information: NCT01269424.

2510 Background: Anti-CD19 CAR-T cell therapy is a breakthrough treatment (tx) for patients (pts) with relapsed/refractory (R/R) B-cell non-Hodgkin lymphoma (NHL). Despite impressive outcomes, non-response and relapse with CD19 negative disease remain challenges. Through dual B-cell antigen targeting of CD20 and CD19, with a first-in-human bispecific lentiviral CAR-T cell (LV20.19CAR), we aim to improve response rates while limiting CD19 negative relapse. Methods: Pts were treated on a Phase 1 dose escalation + expansion trial (NCT03019055) to demonstrate safety of a 41BB/CD3z LV20.19CAR T cell for adults with R/R B-cell NHL. Safety was assessed by incidence of dose limiting toxicities (DLTs) within 28 days post-infusion. Starting dose was 2.5 x 10^5 cells/kg with a target dose of 2.5 x 10^6 cells/kg. All pts received fludarabine+cyclophosphamide for lymphodepletion. Results: 11 pts have completed tx to date. 9 pts in dose escalation and 2 pts in expansion phase. Median age was 54 years (46-67) and histology included DLBCL = 5 pts, MCL = 4 pts, and CLL = 2 pts. In dose escalation, 3 pts were treated at 2.5 x 10^5 cells/kg, 3 pts at 7.5 x 10^5 cells/kg, and 3 pts at 2.5 x 10^6 cells/kg with no DLTs. As a result, 2.5 x 10^6 cells/kg was selected for expansion. In terms of safety, 6 pts developed Grade 1-2 cytokine release syndrome (CRS) and 3 pts had Grade 1-2 neurotoxicity (NTX). No patient had grade 3-4 CRS or NTX and none required ICU level care. 4 pts required 1-2 doses of tocilizumab for CRS. The day 28 overall response rate (ORR) for all pts was 82% (6/11 = complete response (CR) and 3/11 = partial response). All CR pts remain in remission, the longest > 1 year. All progressing pts underwent repeat biopsy, and all retained either CD19 or CD20 positivity. Additional pts are being enrolled in the expansion phase and updated data will be presented. Conclusions: Phase 1 results from the LV20.19 CAR T clinical trial demonstrate that infusion of 2.5 x 10^6 cells/kg is safe for further investigation with no DLTs among treated pts. Down-regulation of target antigens was not identified as a mechanism of resistance in progressing pts. With limited toxicity and encouraging ORR, dual targeted LV20.19CAR T cells merits further investigation. Clinical trial information: NCT03019055.

2539 Background: AntiCD19 CAR-T cells have shown encouraging anti-lymphoma activity. Decreasing the time from apheresis to CAR-T infusion can make this therapy available to pts with rapid progression. We present the interim results of a phase I clinical trial using on-site CAR-T manufacture. Methods: Adult pts with r/r CD19+ B cell lymphomas who failed ≥ 2 lines of therapy were enrolled. Autologous T cells were transduced with a lentiviral vector (Lentigen Technology, Inc,LTG1563) encoding an antiCD19 binding motif, CD8 linker and TNFRSF19 transmembrane region, and 4-lBB/CD3z domains. GMP-compliant manufacture was done using CliniMACS Prodigy, in a 12-day culture. Dose levels were 0.5, 1 and 2 x 106 CAR-T cells/kg. Lymphodepletion was done with cyclophosphamide (60mg/kg x 1) and fludarabine (25mg/m2/d x 3). Results: 7 pts (4 women, 3 men) were enrolled. Median age was 60y [range 43-69]. Diagnoses were DLBCL (n = 3) PMBCL, follicular lymphoma (FL), transformed FL, and transformed lymphoplasmacytic lymphoma; with a median of 4 previous treatments. Six pts had symptomatic refractory disease. CAR-T cell product manufacture was successful in all pts. Mean transduction rate was 44% [range 29-57]. CAR-T cell doses were 0.5 x 106/kg (n = 3) and 1 x 106/kg (n = 4). Median apheresis to infusion time was 13 days [range 13–20], 5 products were infused fresh. CAR-T persistence based on vector sequence, peaked in peripheral blood MNCs between days 14-21. Five pts are evaluable for safety. CRS grade 1 - 2 (Lee) occurred in 4 pts; with 3 requiring treatment. Grade 4 CRES (CARTOX-10) occurred in 1 pt, with resolution after corticosteroids; considered a DLT as it lasted more than 72 hours. No treatment-related mortality has occurred. 4/5 evaluable pts have achieved complete response. One pt did not respond and died. After a median follow up 3 months, all responding pts are alive and 1 relapsed 6 mo after treatment. Conclusions: Second generation antiCD19 CAR-T cells with TNFRS19 transmembrane domain have clinical activity against refractory NHL. Short manufacture time achieved by local CAR-T cell manufacture with the CliniMACS Prodigy enables treatment of a very high risk NHL population. Clinical trial information: NCT03434769.

3043 Background: Studies in adoptive T-cell transfer have suggested that persistence of the transduced T-cells is central to the viability of this therapeutic option. The objectives of this phase I...

TPS1616 Background: The administration of radiation therapy (RT) and concomitant and adjuvant temozolomide (TMZ) for the treatment of newly-diagnosed glioblastoma (GBM) is associated with modestly prolonged survival. However, the benefit of concurrent therapy is restricted to a subgroup of patients whose tumor exhibits methylation of the promoter for methylguanine-DNA-methyltransferase (MGMT). Promoter methylation silences the expression of DNA alkyltransferase (AGT), the product of the MGMT gene which functions to repair alkylated DNA. Unfortunately, the majority of GBMs have non-methylated MGMT promoters and do not appear to benefit from adding TMZ to radiotherapy. Two potential strategies for targeting MGMT-induced chemoprotection in gliomas include depletion of tumor MGMT, and dose escalation of TMZ accomplished by minimizing TMZ-induced bone marrow toxicity. In this study, we propose to combine the irreversible MGMT inhibitor O6-benzylguanine (BG) with infusion of autologous hematopoetic stem cells (HSCs) transduced with lentiviral P140K-MGMT, an MGMT mutant which is highly resistant to inactivation by BG, with the intent to reduce or prevent the myelosuppressive effects of TMZ. The primary objective is to assess the safety and feasibility of combining these two strategies in patients with newly-diagnosed GBM. Methods: This is a three cohort, Phase I study for patients with newly resected supratentorial GBM. There will be at least four evaluable patients per cohort. Cohort 1 will receive standard RT and concomitant TMZ prior to infusion of autologous lentiviral P140K-MGMT transduced HSCs, followed by adjuvant TMZ. Cohort 2 will receive an induction dose of TMZ+BG followed by infusion of autologous LV-P140K-MGMT HSCs prior to concomitant RT +TMZ and adjuvant TMZ. Cohort 3 will include intra-patient dose escalation of TMZ in patients who exhibit detectable levels of P140K-MGMT marked cells in vivo, using the design (i.e. cohort 1 or 2) proven to be safest and most effective. The study is open and has accrued the first patient.