Equity Holder In Publicly-traded Company Research Articles

Targeted Degradation of the Wiz Transcription Factor for Gamma Globin De-Repression

Disclosures Sickle cell disease (SCD) is a life-threatening genetic condition arising from a point mutation in the β-globin gene that causes hemoglobin polymerization and vaso-occlusive sickling of erythrocytes. Hereditary persistence or pharmacologic induction of gamma-globin (HBG1 or HBG2) expression leads to increases in fetal hemoglobin (HbF) that biophysically oppose hemoglobin polymerization. Indeed, the standard-of-care, hydroxyurea (HU), increases expression of HbF in the erythroblast, and in patients the magnitude of response (9-24% HbF) correlates with a reduction in pain crises and acute complications. However, the use of HU is limited by variable response, myelosuppression, GI side effects and carcinogenicity risk. New targeted therapeutic approaches to induce HbF are therefore being intently pursued, leveraging the increasingly detailed functional characterization of the globin locus. The recent discovery of BCL11A as a repressor of HBG has prompted the widespread innovation of genetically engineered hematopoietic stem cells that suppress BCL11A expression or function. While a major advance in SCD therapeutics, the complexity and clinical implementation of stem cell transplantation may limit access and impact, especially in developing countries where SCD burden is greatest. We therefore undertook to discover a small-molecule HbF inducer, focusing on transcription factors that repress gamma-globin expression. Although transcription factors as a target class have historically been considered "undruggable," recent mechanistic characterization of phthalimide medicines as serendipitous, cereblon (CRBN)-dependent degraders of IKZF1 and IKZF3 by our organization and others, suggested the plausible degradation of new zinc-finger containing transcription factors by this emerging class of molecular glues. Therefore, we elaborated a large library of CRBN-biased ligands for study in target-directed and phenotypic drug discovery campaigns. Here we report for the first time the phenotypic discovery and structure-based optimization of two molecular glue degraders of the transcription factor, WIZ, which lead to pronounced de-repression of HbF. Using a chemical biology approach, the first molecule, dWIZ-1, was identified by phenotypic screening of a CRBN-biased chemical library for compounds that induce HbF without deleterious effect on erythroid cell proliferation and differentiation. Subsequent target deconvolution by global proteomics revealed WIZ as the target. WIZ knockout using two independent sgRNA elevated the proportion of γ-globin mRNA, HbF protein levels and HbF + cells (Figure 1) in vitro in primary human erythroblasts, validating WIZ as a previously unrecognized repressor of HbF. Optimization of dWIZ-1 led to a second tool compound, dWIZ-2, which was suitable to assess WIZ degradation and HbF induction in vivo due to its superior pharmacokinetic properties. dWIZ-2 treatment increased the proportion of γ-globin mRNA, HbF + cells and HbF protein levels in vitro in sickle cell disease patient-derived erythroblasts (Figure 2). Because many IMiDs lack activity in rodents, we used a humanized mouse model to measure the impact of dWIZ-2 on WIZ degradation and HbF expression in human erythroblasts in vivo. NBSGW mice humanized with CD34 + hematopoietic stem and progenitor cells from healthy human donors were orally dosed once daily with dWIZ-2 for 21 days. dWIZ-2 treatment resulted in robust dose-dependent WIZ degradation and an increase in total HbF and the proportion of HbF + human erythroblasts in the bone marrow. To further study the kinetics of HbF induction in the peripheral blood, and assess the effects of dWIZ-2 on steady-state hematopoiesis, we orally dosed naïve healthy cynomolgus monkeys with vehicle or 30 mg/kg dWIZ-2, once daily for 28 days. We detected elevated levels of γ-globin mRNA in the blood on day 28, reaching up to 37% of β-like globins. Additionally, on day 28, we observed up to 95% HbF + reticulocytes.dWIZ-2 was well-tolerated based on the absence of notable clinical observations during the study and consistent body weight for all animals. There were no dWIZ-2-related changes in hematology, coagulation or clinical chemistry measurements, specifically including reticulocytes, hematocrit and neutrophil counts. These findings support the development of an oral WIZ degrader as a globally accessible therapeutic approach for SCD. Ting: Novartis: Current Employment, Current equity holder in publicly-traded company; Light Horse Therapeutics: Current equity holder in private company. Borikar:Novartis: Current Employment, Current equity holder in publicly-traded company. Kerrigan:Novartis: Current Employment, Current equity holder in publicly-traded company. Thomsen:Novartis: Current Employment, Current equity holder in publicly-traded company. Aghania:Novartis: Current Employment. Hinman:Novartis: Current Employment, Current equity holder in publicly-traded company. Pizzato:Novartis: Current Employment. Reyes:Novartis: Current Employment, Current equity holder in publicly-traded company. Fodor:Novartis: Current Employment, Current equity holder in publicly-traded company. Wu:Novartis: Current Employment, Current equity holder in publicly-traded company. Belew:Notch Therapeutics: Current Employment, Current equity holder in private company. Mao:Novartis: Current Employment, Current equity holder in publicly-traded company. Wang:Novartis: Current Employment. Chitnis:Novartis: Current Employment, Current equity holder in publicly-traded company. Ashcroft:Novartis: Current Employment, Current equity holder in publicly-traded company. Niu:Novartis: Current equity holder in publicly-traded company; Arvinas: Current Employment, Current equity holder in publicly-traded company. Hachey:Novartis: Current Employment. Cobb:Novartis: Current Employment, Current equity holder in publicly-traded company. Savage:Novartis: Current Employment, Current equity holder in publicly-traded company. Burke:Novartis: Current Employment, Current equity holder in publicly-traded company; Arvinas: Current equity holder in publicly-traded company. Paulk:Novartis: Current Employment. Dovala:Novartis: Current Employment, Current equity holder in publicly-traded company; Twist Biosciences: Current equity holder in publicly-traded company; Vicinitas Therapeutics: Current equity holder in private company. Lin:Novartis: Current Employment, Current equity holder in publicly-traded company. Clifton:Novartis: Current Employment, Current equity holder in publicly-traded company; Nurix Therapeutics: Current equity holder in publicly-traded company. Ornelas:Novartis: Current Employment. Ware:Novartis: Current Employment, Current equity holder in publicly-traded company. Sanchez:Novartis: Current Employment, Current equity holder in publicly-traded company; Merck: Current equity holder in publicly-traded company. Terranova:Novartis: Current Employment, Current equity holder in publicly-traded company. Knehr:Novartis: Current Employment. Altorfer:Novartis: Current Employment, Current equity holder in publicly-traded company. Barnes:Novartis: Current Employment. Taraszka:Novartis: Current Employment, Current equity holder in publicly-traded company. Lubicka:Novartis: Current Employment, Current equity holder in publicly-traded company. Manning:Novartis: Current Employment, Current equity holder in publicly-traded company. Miranda:Novartis: Current Employment, Current equity holder in publicly-traded company. Beckwith:Novartis: Current equity holder in publicly-traded company; Neomorph: Current Employment, Current equity holder in private company. Solomon:Novartis: Current Employment, Current equity holder in publicly-traded company. Forrester:Novartis: Current Employment, Current equity holder in publicly-traded company. Hollingworth:Novartis: Current Employment, Current equity holder in publicly-traded company. Dales:Novartis: Ended employment in the past 24 months; Odyssey Therapeutics: Current Employment, Current equity holder in publicly-traded company. Patterson:Novartis: Current Employment, Current equity holder in publicly-traded company. Bouwmeester:Novartis: Current Employment, Current equity holder in publicly-traded company. Wagner:Novartis: Current Employment, Current equity holder in publicly-traded company. Dranoff:Novartis: Current Employment, Current equity holder in publicly-traded company. Stevenson:Novartis: Current Employment, Current equity holder in publicly-traded company. Bradner:Novartis: Current Employment, Current equity holder in publicly-traded company. Figure 1

Safety and Efficacy of Axatilimab at 3 Different Doses in Patients with Chronic Graft-Versus-Host Disease (AGAVE-201)

Disclosures Background: Chronic graft-versus-host disease (cGVHD) is an immune-mediated complication of allogeneic hematopoietic cell transplant (alloHCT) that affects multiple organs with inflammatory and fibrotic pathology, leading to significant patient burden and mortality. Colony-stimulating factor 1 receptor (CSF-1R)-dependent monocytes and macrophages play a key role in cGVHD inflammation and fibrosis. Axatilimab (SNDX-6352) is an investigational, high-affinity anti-CSF-1R monoclonal antibody that targets monocytes and macrophages. We previously demonstrated the biological and clinical activity of axatilimab with organ-specific responses and symptom improvement in a phase 1/2 study (NCT03604692) in patients with cGVHD. Methods: This pivotal phase 2, open-label, randomized, multicenter study evaluated axatilimab at 3 different doses in alloHCT patients with recurrent or refractory cGVHD (AGAVE-201; NCT04710576). Eligible patients were randomized 1:1:1 to receive intravenous (IV) axatilimab at 0.3 mg/kg every 2 weeks (Q2W), 1 mg/kg Q2W, or 3 mg/kg every 4 weeks (Q4W). Randomization was stratified by severity of cGVHD and prior use of ibrutinib, ruxolitinib, or belumosudil. Concomitant use of corticosteroids, calcineurin inhibitors, or mammalian target of rapamycin inhibitors (sirolimus or everolimus) was allowed. Axatilimab treatment could be continued as long as there was clinical benefit as assessed by the investigator. The primary efficacy endpoint was overall response rate (ORR) in the first 6 cycles (24 weeks) as defined by the NIH 2014 consensus criteria; the efficacy boundary of the ORR is based on the lower bound of the 95% CI exceeding 30%. The key secondary endpoint was the proportion of patients reporting a clinically significant reduction of symptoms, as measured by the modified Lee Symptom Scale (mLSS) score with a threshold of ≥7 points. Safety endpoints included frequency and severity of treatment-related adverse events (TRAEs) and treatment-emergent adverse events (TEAEs). Results: A total of 241 patients were enrolled across 121 study sites and 239 (99.2%) patients were treated with axatilimab at the data cutoff of 7 April 2023. Patients were heavily pretreated with a median of 4 prior lines of therapy, including ruxolitinib (74%), belumosudil (23%), and ibrutinib (31%). Demographics and disease characteristics were balanced among the 3 dose cohorts (Table 1). ORR (95% CI) was 74% (63, 83) with 0.3 mg/kg Q2W, 67% (55, 77) with 1 mg/kg Q2W, and 50% (39, 61) with 3 mg/kg Q4W (Table 1). Median duration of response (DOR) has not been reached in any of the cohorts, with 60%, 60% and 53% of patients maintaining response at 12 months in the 0.3 mg/kg Q2W, 1 mg/kg Q2W, and 3 mg/kg Q4W dose cohorts, respectively. Clinical benefit, as measured by reduction in mLSS score, was reported in 55%, 54%, and 36% of patients in the 0.3 mg/kg Q2W, 1 mg/kg Q2W, and 3 mg/kg Q4W cohorts, respectively, in the first 6 cycles. Drug discontinuation owing to TEAEs occurred in 6% of patients with 0.3 mg/kg Q2W, 22% with 1 mg/kg Q2W, and 18% with 3 mg/kg Q4W. The most common TEAEs are summarized in Table 2. Fatal TEAEs occurred in 1.3%, 8.6%, and 7.6% of patients in the 0.3 mg/kg Q2W, 1 mg/kg Q2W, and 3 mg/kg Q4W dose cohorts, respectively. The frequency of TRAEs and grade ≥ 3 TRAEs were dose dependent, consistent with CSF-1R inhibition-mediated macrophage clearance. Most infections were mild, with 3 reported cytomegalovirus infections, including reactivations, in the higher dose cohorts. Conclusions: The AGAVE-201 pivotal trial met its primary endpoint for all doses studied. Axatilimabtreatment of refractory cGVHD in heavily pretreated patients resulted in robust clinical activity and durable responses in all 3 dose cohorts, with the highest ORR and least toxicity at the 0.3 mg/kg Q2W dose. Adverse events consisted primarily of transient laboratory abnormalities and other on-target effects known to be associated with CSF-1R block. Wolff: Novartis: Honoraria, Research Funding; Mallickrodt: Honoraria; Incyte: Honoraria; Sanofi: Honoraria; Behring: Honoraria; Takeda: Honoraria; Gilead: Honoraria. Cutler:Sanofi: Consultancy; Ruth L. Kirschstein Postdoctoral Individual National Research Service Award: Research Funding; Allovir: Other: Data Safety Monitoring Board (DSMB); Pluristem Therapeutics: Other: DSMB; InhibRx: Consultancy; Astellas: Consultancy; Rigel: Consultancy; Oxford Immune Algorithmics: Membership on an entity’s Board of Directors or advisory committees; Cimeio: Membership on an entity’s Board of Directors or advisory committees. Lee:Amgen, AstraZeneca, Incyte, Kadmon, Pfizer, Syndax: Research Funding; Novartis: Other: Steering Committee member; Janssen: Other: Study medication provider; Equillium, Kadmon, Mallinckrodt: Consultancy. Pusic:Incyte: Honoraria; Syndax: Honoraria. White:Novartis: Honoraria. Hamadani:Kite, a Gilead Company: Consultancy, Speakers Bureau; AstraZeneca: Speakers Bureau; Caribou: Consultancy; BeiGene: Speakers Bureau; Gamida Cell: Consultancy; Incyte: Consultancy; Genmab: Consultancy; Kadmon: Consultancy; Legend Biotech: Consultancy; MorphoSys: Consultancy; Novartis: Consultancy; SeaGen: Consultancy; Astra Zeneca: Speakers Bureau; BeiGene: Speakers Bureau; Sanofi Genzyme: Speakers Bureau; Astellas: Research Funding; Spectrum Pharmaceuticals: Research Funding; Takeda: Research Funding; Genentech: Honoraria; Myeloid Therapeutics: Honoraria; Bristol Myers Squibb: Consultancy; Genmab: Consultancy; CRISPR: Consultancy; Omeros: Consultancy; Abbvie: Consultancy; ADC therapeutics: Consultancy, Honoraria, Research Funding, Speakers Bureau. Salhotra:Gilead: Research Funding; Kura Oncology: Research Funding; OrcaBio: Research Funding; Jazz Pharma: Research Funding; Rigel Pharma: Research Funding; Sobi: Membership on an entity’s Board of Directors or advisory committees; Sanofi: Speakers Bureau; BMS: Research Funding. Choe:Opna: Other: Receipt of equipment, materials, drugs to institution, Research Funding; Incyte: Consultancy, Membership on an entity’s Board of Directors or advisory committees, Other: Receipt of equipment, materials, drugs to institution; MJH Life Sciences: Honoraria; Actinium Pharmaceuticals: Other: Support for attending meetings and/or travel; NIH National Cancer Institute: Research Funding. Kwon:Kite-Gilead: Consultancy, Speakers Bureau; Jazz: Speakers Bureau; Pfizer: Speakers Bureau. Bermúdez:Janssen: Consultancy, Speakers Bureau; Novartis: Consultancy, Speakers Bureau; Pfitzer: Consultancy, Speakers Bureau. Radojcic:Syndax: Current Employment, Current equity holder in publicly-traded company. O’Toole:Syndax: Current Employment, Current equity holder in publicly-traded company. Tian:Incyte: Current Employment, Current holder of stock options in a privately-held company. Ordentlich:Syndax: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties: Patents assigned to Syndax Pharmaceuticals. DeFilipp:Incyte: Consultancy, Honoraria, Research Funding; Regimmune: Research Funding; Taiho Oncology: Research Funding; Sanofi: Consultancy; MorphoSys: Consultancy, Honoraria; Inhibrx: Consultancy; PharmaBiome AG: Consultancy; Ono Pharmaceuticals: Consultancy. Kitko:Horizon: Membership on an entity’s Board of Directors or advisory committees. Figure 1

The POLARIX Study: Polatuzumab Vedotin with Rituximab, Cyclophosphamide, Doxorubicin, and Prednisone (pola-R-CHP) Versus Rituximab, Cyclophosphamide, Doxorubicin, Vincristine and Prednisone (R-CHOP) Therapy in Patients with Previously Untreated Diffuse Large B-Cell Lymphoma

Background: The current standard of care for patients with newly diagnosed diffuse large B-cell lymphoma (DLBCL) is R CHOP; however, approximately 40% of patients are not cured. The CD79b-targeting antibody-drug conjugate, polatuzumab vedotin, is approved in relapsed/refractory DLBCL in combination with bendamustine and rituximab, and has also demonstrated promising first line activity and safety when combined with rituximab, cyclophosphamide, doxorubicin, and prednisone (pola-R-CHP) in a Phase Ib/II study (Tilly, et al. Lancet Oncol 2019). Thus, in the Phase III POLARIX study (NCT03274492) we compared pola-R-CHP with R-CHOP in patients with previously untreated DLBCL.Methods: In this double-blind, placebo-controlled, international study, patients with previously untreated DLBCL and an International Prognostic Index (IPI) of 2-5 were randomized 1:1 to receive six cycles of pola-R-CHP (with a vincristine placebo) or R-CHOP (with a polatuzumab vedotin placebo); all patients also received two additional cycles of rituximab. Patients received polatuzumab vedotin 1.8mg/kg or vincristine 1.4mg/m² administered on Day 1, plus intravenous rituximab 375mg/m2, cyclophosphamide 750mg/m², doxorubicin 50mg/m², and placebo on Day 1, and oral prednisone 100mg once daily on Days 1-5. The primary endpoint was investigator-assessed progression-free survival (PFS). Secondary endpoints included investigator-assessed event-free survival (EFS), independent review committee-assessed complete response (CR) rate at the end of treatment by positron emission tomography-computed tomography (PET-CT), disease-free survival (DFS), overall survival (OS), and safety.Results: Overall, 879 patients were randomized, 440 to pola-R-CHP and 439 to R-CHOP. Median age was 65 (range 19-80) years, and the majority of patients had IPI 3-5 (62.0%). At the data cut-off of June 28, 2021, and after a median follow-up of 28.2 months, PFS was superior with pola-R-CHP vs R CHOP (hazard ratio [HR] 0.73; 95% confidence interval [CI]: 0.57-0.95; P<0.02). The 2-year PFS rate was 76.7% (95% CI: 72.7-80.8) with pola-R-CHP vs 70.2% (95% CI: 65.8-74.6) with R-CHOP. EFS favored pola-R-CHP compared with R-CHOP (HR 0.75; 95% CI: 0.58-0.96; P=0.02). The end-of-treatment PET-CT CR rate was not significantly different with pola-R-CHP vs R-CHOP (78.0% vs 74.0%; P=0.16); however, DFS suggested responses were more durable with pola-R-CHP than with R-CHOP (HR 0.70; 95% CI: 0.50-0.98). There was no difference in OS between treatment arms (HR 0.94; 95% CI: 0.65-1.37; P=0.75). At the time of data cut-off, 99 (23%) and 133 (30%) patients in the pola-R-CHP and R-CHOP arms, respectively, had received at least one subsequent anti-lymphoma therapy. Fewer patients in the pola-R-CHP than the R-CHOP arm received subsequent anti-lymphoma treatments (radiotherapy, 9.3% vs 13.0%; stem cell transplantation, 3.9% vs 7.1%; chimeric antigen receptor T-cell therapy, 2.0% vs 3.6%). The safety profile was comparable for pola-R-CHP vs R-CHOP, including rates of grade 3-4 adverse events (AEs; 57.7% vs 57.5%), serious AEs (34.0% vs 30.6%), grade 5 AEs (3.0% vs 2.3%), and AEs leading to dose reduction (9.2% vs 13.0%), respectively. The frequency and severity of peripheral neuropathy were similar for pola-R-CHP vs R-CHOP (any grade, 52.9% vs 53.9%; grade 3-4, 1.6% vs 1.1%).Conclusion: The pola-R-CHP combination demonstrated a 27% reduction in the relative risk of disease progression, relapse, or death compared with R-CHOP, with a similar safety profile in the first-line treatment of patients with DLBCL. DisclosuresTilly: Karyopharm: Membership on an entity's Board of Directors or advisory committees; F. Hoffmann-La Roche Ltd: Membership on an entity's Board of Directors or advisory committees, Other: Meeting attendance and travel, Research Funding; AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Janssen-Cilag: Membership on an entity's Board of Directors or advisory committees; Incyte: Membership on an entity's Board of Directors or advisory committees. Morschhauser: AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; AstraZenenca: Membership on an entity's Board of Directors or advisory committees; Epizyme: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees; F. Hoffmann-La Roche Ltd: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Chugai: Honoraria; Genentech, Inc.: Consultancy; Janssen: Honoraria; Genmab: Membership on an entity's Board of Directors or advisory committees; Servier: Consultancy; Incyte: Membership on an entity's Board of Directors or advisory committees. Sehn: Novartis: Consultancy; Genmab: Consultancy; Debiopharm: Consultancy; Teva: Consultancy, Research Funding; Roche/Genentech: Consultancy, Research Funding; AbbVie: Consultancy; Acerta: Consultancy; Amgen: Consultancy; Apobiologix: Consultancy; AstraZeneca: Consultancy; Celgene: Consultancy; Gilead: Consultancy; Incyte: Consultancy; Janssen: Consultancy; Kite: Consultancy; Karyopharm: Consultancy; Lundbeck: Consultancy; Merck: Consultancy; Morphosys: Consultancy; Sandoz: Consultancy; Seattle Genetics: Consultancy; Takeda: Consultancy; TG Therapeutics: Consultancy; Verastem: Consultancy. Friedberg: Bayer: Membership on an entity's Board of Directors or advisory committees; Acerta: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees. Trněný: Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Gilead Sciences: Consultancy, Honoraria; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; F. Hoffmann-La Roche Ltd: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Morphosys: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Portola: Honoraria, Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Honoraria. Sharman: AbbVie: Consultancy; AstraZeneca: Consultancy; BeiGene: Consultancy; Bristol-Myers Squibb: Consultancy; Lilly: Consultancy; Pharmacyclics LLC, an AbbVie Company: Consultancy; TG Therapeutics: Consultancy; Centessa: Current holder of stock options in a privately-held company, Membership on an entity's Board of Directors or advisory committees; Genentech, Inc.: Consultancy; Velos: Current holder of stock options in a privately-held company, Membership on an entity's Board of Directors or advisory committees. Herbaux: Takeda: Honoraria, Research Funding; AbbVie: Honoraria, Research Funding; F. Hoffmann-La Roche Ltd: Honoraria; Janssen: Honoraria. Burke: Verastem: Consultancy; AstraZeneca: Consultancy; Morphosys: Consultancy; Adaptive Biotechnologies: Consultancy; Epizyme: Consultancy; Kura: Consultancy; AbbVie: Consultancy; BeiGene: Consultancy, Speakers Bureau; Kymera: Consultancy; Bristol-Myers Squibb: Consultancy; X4 Pharmaceuticals: Consultancy; Seattle Genetics: Consultancy, Speakers Bureau; Gilead: Consultancy; Genentech, Inc.: Consultancy. Matasar: Memorial Sloan Kettering Cancer Center: Current Employment; Merck Sharp & Dohme: Consultancy, Current holder of individual stocks in a privately-held company, Membership on an entity's Board of Directors or advisory committees; Genentech, Inc.: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel, Research Funding; F. Hoffmann-La Roche Ltd: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel, Research Funding; GlaxoSmithKline: Honoraria, Research Funding; Bayer: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel, Research Funding; IGM Biosciences: Research Funding; Pharmacyclics: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Rocket Medical: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Seattle Genetics: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel, Research Funding; ImmunoVaccine Technologies: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Daiichi Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Juno Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees; Teva: Consultancy, Membership on an entity's Board of Directors or advisory committees; TG Therapeutics: Consultancy, Honoraria. Rai: Chugai Pharmaceutical Co., Ltd: Speakers Bureau; ONO Pharmaceutical Co., Ltd: Speakers Bureau; Janssen Pharmaceutical: Speakers Bureau; Eisai Co., Ltd: Speakers Bureau. Izutsu: AbbVie: Honoraria; Allergan Japan: Honoraria; AstraZeneca: Honoraria, Research Funding; Bayer: Research Funding; BeiGene: Research Funding; Celgene: Honoraria, Research Funding; Chugai: Honoraria, Research Funding; Daiichi Sankyo: Honoraria, Research Funding; Eisai: Honoraria, Research Funding; Fuji Film Toyama Chemical: Honoraria; Genmab: Honoraria, Research Funding; Huya Biosciences: Research Funding; Incyte: Research Funding; Janssen: Honoraria, Research Funding; Kyowa Kirin: Honoraria, Research Funding; MSD: Research Funding; Novartis: Honoraria, Research Funding; Ono Pharmaceutical: Honoraria, Research Funding; Pfizer: Research Funding; Solasia: Research Funding; Symbio: Honoraria; Takeda: Honoraria, Research Funding; Yakult: Research Funding. Mehta-Shah: C4 Therapeutics: Consultancy; Kiowa Hakko Kirin: Consultancy; Karyopharm: Consultancy; Ono Pharmaceuticals: Consultancy; Secura Bio: Consultancy, Research Funding; Daiichi Sankyo: Consultancy, Research Funding; AstraZeneca: Research Funding; Bristol-Myers Squibb: Research Funding; Celgene: Research Funding; Innate Pharmaceuticals: Research Funding; Roche/Genentech: Research Funding; Corvus Pharmaceuticals: Research Funding; Verastem: Research Funding. Oberic: Celgene: Honoraria; F. Hoffmann-La Roche Ltd: Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel; Janssen: Honoraria, Other: Support for attending meetings and/or travel; Gilead: Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel; AbbVie: Other: Support for attending meetings and/or travel; Incyte: Membership on an entity's Board of Directors or advisory committees. Jurczak: Maria Sklodowska-Curie National Research Institute of Oncology: Current Employment; Jagiellonian University: Ended employment in the past 24 months. Greil: Sandoz: Honoraria, Research Funding; Amgen: Honoraria, Other: Travel, Accommodations, Expenses, Research Funding; AstraZeneca: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Daiichi Sankyo: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Sankyo: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses; Merck Sharp & Dohme: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; F. Hoffmann-La Roche Ltd: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding. Pinto: Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel; Gilead Sciences: Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel; MSD: Honoraria; Servier: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy, Honoraria, Other: Support for attending meetings and/or travel; F. Hoffmann-La Roche Ltd: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel, Speakers Bureau. Abrisqueta Costa: Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; F. Hoffmann-La Roche Ltd: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Honoraria, Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees. Hirata: Genentech, Inc.: Current Employment; Genentech/Roche: Current holder of stock options in a privately-held company. Jiang: Genentech, Inc.: Current Employment; F. Hoffmann-La Roche Ltd: Current equity holder in publicly-traded company. Yan: F. Hoffmann-La Roche Ltd: Current Employment, Current equity holder in publicly-traded company. Lee: Genentech, Inc.: Current Employment; F. Hoffmann-La Roche Ltd: Current equity holder in publicly-traded company. Flowers: AbbVie: Consultancy, Research Funding; Bayer: Consultancy, Research Funding; BeiGene: Consultancy; Celgene: Consultancy, Research Funding; Denovo Biopharma: Consultancy; Epizyme: Consultancy; Roche/Genentech: Consultancy, Research Funding; Genmab: Consultancy; Gilead: Consultancy, Research Funding; Karyopharm: Consultancy; Pharmacyclics/Janssen: Consultancy; Seattle Genetics: Consultancy; Spectrum: Consultancy; 4D: Research Funding; Acerta: Research Funding; Adaptimmune: Research Funding; Allogene: Research Funding; Amgen: Research Funding; Cellectis: Research Funding; EMD: Research Funding; Guardant: Research Funding; Iovance: Research Funding; Janssen: Research Funding; Kite: Research Funding; Morphosys: Research Funding; Nektar: Research Funding; Novartis: Research Funding; Pfizer: Research Funding; Sanofi: Research Funding; Takeda: Research Funding; TG Therapeutics: Research Funding; Xencor: Research Funding; Ziopharm: Research Funding; Burroughs Wellcome Fund: Research Funding; Eastern Cooperative Oncology Group: Research Funding; National Cancer Institute: Research Funding; Cancer Prevention and Research Institute of Texas: CPRIT Scholar in Cancer Research: Research Funding; Pharmacyclics: Research Funding. Salles: Bayer: Honoraria; AbbVie: Consultancy, Honoraria; BeiGene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb/Celgene: Consultancy, Honoraria; Debiopharm: Consultancy, Membership on an entity's Board of Directors or advisory committees; Epizyme: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche/Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees; Genmab: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees; Ipsen: Consultancy; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Kite/Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; Loxo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Miltenyi: Consultancy; Morphosys: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Rapt: Consultancy, Membership on an entity's Board of Directors or advisory committees; Regeneron: Consultancy, Honoraria; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; Velosbio: Consultancy, Membership on an entity's Board of Directors or advisory committees; Allogene: Consultancy. OffLabel Disclosure:Polatuzumab vedotin is an antibody-drug conjugate targeting CD79b on malignant B-cells. Polatuzumab vedotin in combination with bendamustine and rituximab (pola-BR) improved complete response rate and overall survival compared with BR alone in patients with relapsed/refractory diffuse large B-cell lymphoma. Pola-BR is approved for the treatment of adult patients with relapsed or refractory diffuse large B-cell lymphoma, not otherwise specified, after at least two prior therapies.

Fitusiran, an Investigational siRNA Therapeutic Targeting Antithrombin for the Treatment of Hemophilia: First Results from a Phase 3 Study to Evaluate Efficacy and Safety in People with Hemophilia a or B without Inhibitors (ATLAS-A/B)

BackgroundHemophilia is a rare bleeding disorder characterized by deficiency of FVIII or FIX resulting in ineffective clot formation due to impaired thrombin generation. Fitusiran is a subcutaneously (SC) administered investigational siRNA therapeutic agent which targets antithrombin to enhance thrombin generation potential and rebalance hemostasis in people with hemophilia (PwH), irrespective of inhibitor status. Here, we present results from a Phase 3 study of the efficacy and safety of fitusiran prophylaxis compared with on-demand (OD) treatment with factor concentrates in PwH A or B without inhibitors (ATLAS-A/B; NCT03417245).MethodsThis Phase 3, multicenter, multinational, randomized, open-label study evaluated the efficacy and safety of fitusiran in males aged ≥12 years with severe hemophilia A or B without inhibitors, previously treated OD. Eligible participants (pts) were randomized 2:1 to receive either once-monthly 80 mg SC fitusiran prophylaxis (fitusiran arm) or OD factor concentrates for treatment of bleeding episodes (OD arm) for a treatment period of 9 months. The primary endpoint was annualized bleeding rate (ABR) in the efficacy period (Day 29 post-first fitusiran dose up to Day 246). Secondary endpoints included annualized spontaneous bleeding rate (AsBR) and annualized joint bleeding rate (AJBR) in the efficacy period and health-related quality of life (HRQoL) in the treatment period, measured by Haem-A-QoL. Safety and tolerability were assessed throughout the study.ResultsOverall, 120 pts were randomized (fitusiran arm n=80; OD arm n=40); 79 pts (98.8%) in the fitusiran arm and 37 pts (92.5%) in the OD arm completed the study. A total of 93 pts had hemophilia A (fitusiran arm n=62, OD arm n=31) and 27 pts had hemophilia B (fitusiran arm n=18, OD arm n=9). Baseline demographics and characteristics were similar in both arms. Median observed ABR was 0.0 (IQR, 0.0 to 3.4) in the fitusiran arm and 21.8 (IQR, 8.4 to 41.0) in the OD arm; 40 pts (50.6%) in the fitusiran arm experienced no bleeds that required treatment with OD factor concentrates. A significant reduction in estimated ABR was achieved in the fitusiran arm vs the OD arm (89.9% reduction [95% CI, 84.1% to 93.6%], p<0.0001; Table). Median observed AsBR was 0.0 (IQR, 0.0 to 1.7) in the fitusiran arm and 16.1 (IQR, 3.4 to 27.6) in the OD arm. A significant reduction in estimated AsBR was achieved in the fitusiran arm vs the OD arm (91.7% reduction [95% CI, 85.9% to 95.1%], p<0.0001; Table). Median observed AJBR was 0.0 (IQR, 0.0 to 3.4) in the fitusiran arm and 15.9 (IQR, 4.2 to 33.5) in the OD arm. There was a significant reduction in estimated AJBR in the fitusiran arm vs the OD arm (90.3% reduction [95% CI, 83.9% to 94.1%], p<0.0001; Table). A significant improvement was also achieved in the transformed total and physical health score in the fitusiran arm vs the OD arm (LS mean difference -7.07 [95% CI, -11.23 to -2.90], p=0.0011; -19.75 [95% CI, -27.00 to -12.50], p<0.0001, respectively).Seventy-nine pts received at least 1 dose of fitusiran and 40 patients were randomized to the OD arm and included in the safety analysis. Overall, 62 pts (78.5%) in the fitusiran arm and 18 pts (45%) in the OD arm experienced ≥1 treatment emergent adverse event (TEAE). A total of 5 treatment emergent serious adverse events (TESAEs) were reported in 5 pts (6.3%) in the fitusiran arm and 9 TESAEs were reported in 5 pts (12.5%) in the OD arm. TESAEs in the fitusiran arm included cholelithiasis (2 pts, 2.5%), cholecystitis, lower respiratory tract infection, and asthma (1 pt each, 1.3%). In the fitusiran arm, 2 pts (2.5%) experienced TEAEs that resulted in fitusiran discontinuation (cholecystitis and increased alanine aminotransferase). No TEAEs of thrombosis and no fatal TEAEs were reported.ConclusionsAll key primary and secondary endpoints were met in this Phase 3 study. Specifically, once-monthly 80 mg SC fitusiran prophylaxis demonstrated a significant reduction in ABR, AsBR and AJBR (all ~90%) in people with severe hemophilia A or B without inhibitors compared with OD treatment. This reduction in bleeding was associated with a meaningful improvement in HRQoL. Reported TESAEs were generally consistent with previously identified risks of fitusiran. With the aim of further enhancing the benefit-risk profile of fitusiran, a revised regimen with reduced dose and frequency is currently being evaluated in ongoing clinical studies. [Display omitted] DisclosuresSrivastava: Roche: Other: Advisory Board, Research Funding; Novo Nordisk: Other: Advisory Board, Research Funding; Sanofi: Other: Advisory Board, Research Funding; Pfizer: Other: Advisory Board, Research Funding; Takeda: Other: Advisory Board, Research Funding; Bayer Healthcare: Other: Grant Review & Awards Committee. Rangarajan: Sanofi: Other: Advisory Board; Pfizer: Other: Advisory Board; Reliance Life Sciences: Consultancy; Takeda: Other: Advisory Board, Conference Support, Speakers Bureau. Kavakli: Pfizer, Bayer, Takeda, Roche, Novo Nordisk: Honoraria; Pfizer, Bayer, Roche, Novo Nordisk, Takeda: Speakers Bureau; Roche, Bayer, Pfizer, Novo Nordisk, Takeda: Membership on an entity's Board of Directors or advisory committees. Klamroth: Bayer, Leo: Research Funding; Bayer, Biotest, Biomarin, BMS, CSL Behring, Daiichi Sankyo, Leo, Novo Nordisk, Octapharma, Pfizer, Roche, Sanofi, SOBI, Takeda: Honoraria; Bayer, Biotest, Biomarin, BMS, CSL Behring, Daiichi Sankyo, Novo Nordisk, Octapharma, Pfizer, Roche, Sanofi, SOBI, Takeda: Speakers Bureau. Kenet: Alnylam: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bayer: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Opko Biologics: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Shire: Research Funding, Speakers Bureau; Novo Nordisk: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Takeda: Consultancy, Honoraria, Research Funding; Nat'l Hemophilia Ctr and Coagulation Unit and Amalia Biron Res Inst of Thromb & Hemost, Sheba Medical Ctr, Tel Hashomer, Israel: Current Employment; BioMarin Pharmaceutical: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; CSL: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Uniquore: Honoraria, Membership on an entity's Board of Directors or advisory committees; BPL: Research Funding. Khoo: NSW Health Pathology: Current Employment; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Novo Nordisk: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Takeda: Speakers Bureau. You: Eulji University Hospital: Current Employment. Malan: PHOENIX Pharma (Pty) Ltd, Qgeberha, South Africa: Current Employment, Research Funding; St Francis Hospital, Qgeberha, South Africa: Membership on an entity's Board of Directors or advisory committees. Frenzel: Pfizer: Research Funding; Roche: Research Funding; SOBI: Research Funding; CSL Behring: Research Funding. Stasyshyn: CSL Behring: Consultancy, Honoraria, Research Funding, Speakers Bureau; Novo Nordisk: Consultancy, Research Funding, Speakers Bureau; Octapharma: Consultancy, Research Funding, Speakers Bureau; Pfizer: Consultancy, Research Funding, Speakers Bureau; Takeda: Consultancy, Research Funding, Speakers Bureau; Grifols: Consultancy, Speakers Bureau; Shire: Consultancy, Honoraria, Research Funding; Institute of Blood Pathology and Transfusion Medicine of National Academy of Medical Sciences of Ukraine: Current Employment; Sanofi: Honoraria, Research Funding; Roche: Speakers Bureau; LFB: Honoraria, Research Funding. Poloskey: Sanofi: Current Employment, Current equity holder in publicly-traded company. Andersson: Sanofi: Current Employment, Current equity holder in publicly-traded company; WEST advisory committee member: Membership on an entity's Board of Directors or advisory committees. Mei: Sanofi: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months, Patents & Royalties. Pipe: Sangamo Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Scientific Advisory Board; ASC Therapeutics: Consultancy, Other: Scientific Advisory Board; Apcintex: Consultancy; Bayer: Consultancy; Biomarin: Consultancy; Catalyst Biosciences: Consultancy; CSL Behring: Consultancy; Freeline: Consultancy; Grifols: Consultancy; HEMA Biologics: Consultancy; Novo Nordisk: Consultancy; Octapharma: Consultancy; Pfizer: Consultancy; Roche/Genentech: Consultancy; Sanofi: Consultancy; Takeda: Consultancy; Spark Therapeutics: Consultancy; uniQure: Consultancy; GeneVentiv: Consultancy, Membership on an entity's Board of Directors or advisory committees; YewSavin: Research Funding; Siemens: Research Funding.

Tisagenlecleucel Vs Standard of Care As Second-Line Therapy of Primary Refractory or Relapsed Aggressive B-Cell Non-Hodgkin Lymphoma: Analysis of the Phase III Belinda Study

Background: Standard of care (SOC) for second-line (2L) therapy (tx) of relapsed/refractory aggressive B-cell non-Hodgkin lymphomas (R/R aNHL) includes platinum-based chemotherapy (PCT) followed by autologous hematopoietic cell transplantation (aHCT) in responders. Outcomes are poor for patients (pts) with R/R aNHL who experience progression during or within 12 months (mo) of 1L tx. Tisagenlecleucel (tisa-cel) is an autologous chimeric antigen receptor (CAR) T-cell tx targeting CD19 approved for pts with large B-cell lymphoma (LBCL; US) and diffuse LBCL (non-US) after ≥2 lines of tx. BELINDA (NCT03570892) is a Phase III, randomized, global (18 countries) study of tisa-cel vs SOC as 2L tx for R/R aNHL.Methods: Adults with confirmed R/R aNHL within 12 mo after 1L chemo-immunotherapy were eligible. All pts underwent leukapheresis for tisa-cel production and were randomized 1:1 to receive tisa-cel (Arm A) or SOC (Arm B) and stratified by 1L duration of response, International Prognostic Index (IPI), and geographic region. Arm A received optional bridging tx (investigator choice of protocol-defined PCT regimens) followed by lymphodepletion (LD; generally, fludarabine 25 mg/m 2/day [d] + cyclophosphamide 250 mg/m 2/d for 3 d) and followed by a single tisa-cel infusion (0.6-6×10 8 CAR-T cells). Arm B received investigator choice of PCT regimen followed by aHCT in responders or a second PCT in nonresponders. Disease assessments were performed at 6 and 12 weeks (wk), then planned every 3 mo for year (y) 1 and every 6 mo for y 2. The primary endpoint was event-free survival (EFS), defined as time from randomization to stable disease (SD) or progressive disease (PD) at or after wk 12 assessment or death at any time. SD/PD at wk 6 was not considered an event on either Arm. Arm A's wk 6 assessment evaluated disease burden before tisa-cel infusion and after bridging if administered. Arm B's wk 6 assessment determined if response was sufficient for aHCT or if a second PCT regimen was needed prior to aHCT.Results: As of May 6, 2021, 322 pts were randomized: 162 to Arm A and 160 to Arm B. In Arms A and B, 33% and 29% were ≥65 y, and 66% and 67% had primary refractory disease, respectively. Baseline characteristics indicated imbalances with more high-grade B-cell lymphomas (24% vs 17%) and IPI ≥2 (65% vs 58%) in Arm A vs B. In Arm A, 47% received ≥2 cycles of bridging PCT, 36% received 1 cycle, and 17% received 0. In Arm A, 96% received tisa-cel; the median dose was 2.9×10 8 cells. In Arm B, 96% received ≥2 PCT cycles and 54% received ≥2 different PCT regimens; 33% received aHCT, including 10% requiring ≥2 different PCT regimens before aHCT. Median time from randomization to aHCT was 92 d (range, 61-158 d). Median follow-up was 10 mo (range, 2.9-23.2 mo). At wk 6 assessment, 26% had PD in Arm A vs 14% in Arm B. Median EFS in Arms A and B was 3 mo (HR 1.07; 95% CI, 0.82-1.40; P=0.69); Arm A pts with PD at wk 6 had shorter EFS. Overall response rate (ORR) at wk 12 in Arm A was 46% vs 43% in Arm B; complete response rate in both arms was 28%. Some pts responded to tisa-cel after SD/PD at wk 12 without additional tx but were counted as an event in EFS analysis. In Arm B, 81 pts (51%) crossed over to receive tisa-cel, 71 without receiving aHCT. Of 72 crossover pts with response assessment, ORR to tisa-cel was 40%. Overall survival was immature at data cutoff. In Arm A, 84% had a grade ≥3 adverse event (AE; vs 90% in Arm B), including grade ≥3 cytokine release syndrome in 5% (CRS; Lee 2014) and grade ≥3 neurologic events (NE) in 3% (CTCAE v5.0), with no grade 5 CRS/NE. Median times to CRS and NE onset were 4 and 5 d, respectively; median time to resolution was 5 and 9 d. Fifty-two (32%) and 45 (28%) pts in Arms A and B died on study, including 42 (26%) and 32 (20%) deaths due to PD, respectively. Ten pts in Arm A and 13 in Arm B died of AEs.Conclusions: Tisa-cel as 2L tx in R/R aNHL pts did not have a higher EFS vs SOC. Possible contributing factors include study design elements, such as optional PCT bridging tx in Arm A with potential delay of tisa-cel infusion until after wk 6 assessment, allowance of a different PCT regimen to reach aHCT in Arm B after inadequate response to first PCT, and imbalances in relevant pt characteristics. Insights from this randomized Phase III study will inform use of cellular tx in the 2L R/R aNHL setting and the design of future CAR-T trials. [Display omitted] DisclosuresBishop: Arcellx, Autolus, Bristol-Myers Squibb, CRISPR, Kite/Gilead, and Novartis: Consultancy, Research Funding; Bristol-Myers Squibb and Kite/Gilead: Other: fees for non-CME/CE services . Dickinson: Novartis: Consultancy, Honoraria, Research Funding, Speakers Bureau; Roche: Consultancy, Honoraria, Other: travel, accommodation, expenses, Research Funding, Speakers Bureau; Gilead Sciences: Consultancy, Honoraria, Speakers Bureau; MSD: Consultancy, Honoraria, Research Funding, Speakers Bureau; Janssen: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria; Takeda: Research Funding; Celgene: Research Funding; Amgen: Honoraria. Purtill: Novartis: Honoraria; Gilead: Honoraria; BMS Celgene: Honoraria. Barba: Amgen, Celgene, Gilead, Incyte, Jazz Pharmaceuticals, MSD, Novartis, Pfizer and Roche, Jazz Phar,aceuticals: Honoraria; Cqrlos III heqlth Institute, aSOCIACION espanola contra el cancer, PERIS: Research Funding. Santoro: Arqule: Consultancy, Speakers Bureau; Sanofi: Consultancy; Takeda: Speakers Bureau; BMS: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Roche: Speakers Bureau; AbbVie: Speakers Bureau; Amgen: Speakers Bureau; Celgene: Speakers Bureau; Servier: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Gilead: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; AstraZeneca: Speakers Bureau; Pfizer: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Eli-Lilly: Speakers Bureau; Sandoz: Speakers Bureau; Eisai: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Speakers Bureau; Bayer: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; MSD: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Hamad: Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Kato: Abbvie: Consultancy, Research Funding; AstraZeneca: Consultancy; Bristol-Myers Squibb: Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Chugai: Consultancy, Honoraria, Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Dainippon-Sumitomo: Honoraria; Eisai: Consultancy, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Kyowa Kirin: Honoraria, Research Funding; MSD: Honoraria; Mundi: Honoraria; Novartis: Consultancy, Research Funding; Ono: Honoraria, Research Funding. Sureda: Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel, Research Funding, Speakers Bureau; BMS/Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings and/or travel, Speakers Bureau; MSD: Consultancy, Honoraria, Speakers Bureau; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Kite, a Gilead Company: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; GSK: Consultancy, Honoraria, Speakers Bureau; Roche: Other: Support for attending meetings and/or travel; Bluebird: Membership on an entity's Board of Directors or advisory committees; Mundipharma: Consultancy. Greil: Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; F. Hoffmann-La Roche Ltd: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; AstraZeneca: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses; Merck Sharp & Dohme: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Gilead: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Daiichi Sankyo: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Sankyo: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses, Research Funding; Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Other: Travel, Accommodations, Expenses, Research Funding; Sandoz: Honoraria, Research Funding. Thieblemont: Gilead Sciences: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses ; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses ; Bristol Myers Squibb/Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses ; Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses , Research Funding; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Kyte: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses ; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees; Abbvie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses ; Cellectis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses ; Hospira: Research Funding; Bayer: Honoraria; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Accommodations, Expenses . Morschhauser: F. Hoffmann-La Roche Ltd: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees; Epizyme: Consultancy, Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; Chugai: Honoraria; Janssen: Honoraria; Genmab: Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Genentech, Inc.: Consultancy; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; AstraZenenca: Membership on an entity's Board of Directors or advisory committees; Servier: Consultancy; Incyte: Membership on an entity's Board of Directors or advisory committees. Janz: Janssen: Honoraria, Other: Meeting Fees; Novartis: Honoraria; Takeda: Honoraria, Other: Travel Accommodations, Expenses, Meeting fees; Roche: Honoraria; Gilead: Other: Travel Accommodations, Expenses, Meeting fees. Flinn: Servier Pharmaceuticals: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Yingli Pharmaceuticals: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Seagen: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Servier Pharmaceuticals: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Unum Therapeutics: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute, Research Funding; Johnson & Johnson: Current equity holder in publicly-traded company; Seattle Genetics: Research Funding; Sarah Cannon Research Institute: Current Employment; Vincerx Pharma: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Hutchison MediPharma: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Century Therapeutics: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Triphase Research & Development Corp.: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Trillium Therapeutics: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Teva: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Rhizen Pharmaceuticals: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Portola Pharmaceuticals: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Pfizer: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Novartis: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Merck: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Loxo: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Karyopharm Therapeutics: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Infinity Pharmaceuticals: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Incyte: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; IGM Biosciences: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Forty Seven: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Forma Therapeutics: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Curis: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Constellation Pharmaceuticals: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Celgene: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Calithera Biosciences: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; ArQule: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Agios: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Acerta Pharma: Other: All research funding payments made to Sarah Cannon Research Institute, Research Funding; Yingli Pharmaceuticals: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Verastem: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Unum Therapeutics: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; TG Therapeutics: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Takeda: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Seagen: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Roche: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Pharmacyclics LLC, an AbbVie Company: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Nurix Therapeutics: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; MorphoSys: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Kite, a Gilead Company: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Juno Therapeutics: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Janssen: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Iksuda Therapeutics: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Great Point Partners: Consultancy, Other: All consultancy payments made to Sarah Cannon Research Institute; Gilead Sciences: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; Genentech: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; BeiGene: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; AstraZeneca: Consultancy, Other: All consultancy and research funding payments made to Sarah Cannon Research Institute, Research Funding; AbbVie: Consultancy, Other: All Consultancy and Research Funding payments made to Sarah Cannon Research Institute, Research Funding. Kwong: Novartis: Consultancy, Honoraria, Research Funding. Kersten: Novartis: Consultancy, Honoraria, Other: Travel support; Takeda: Research Funding; Kite, a Gilead Company: Consultancy, Honoraria, Other: Travel support, Research Funding; Celgene: Research Funding; Roche: Consultancy, Honoraria, Other: Travel support, Research Funding; Miltenyi Biotec: Consultancy, Honoraria, Other: Travel support; BMS/Celgene: Consultancy, Honoraria. Minnema: Janssen: Consultancy, Honoraria; Celgene: Other: Travel expenses; BMS: Consultancy; Kite/Gilead: Consultancy; Alnylam: Consultancy. Holte: Roche: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Nanovector: Membership on an entity's Board of Directors or advisory committees, Other: lectures honorarias; Novartis: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Nordic: Membership on an entity's Board of Directors or advisory committees. Martinez-Lopez: Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees; GSK: Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; Adaptive Biotechnologies: Honoraria, Membership on an entity's Board of Directors or advisory committees. Maziarz: Novartis: Consultancy, Other: Data and Safety Monitoring board, Research Funding; Incyte Corporation: Consultancy, Honoraria; Bristol-Myers, Squibb/Celgene, Intellia, Kite: Honoraria; Allovir: Consultancy, Research Funding; Artiva Therapeutics: Consultancy; CRISPR Therapeutics: Consultancy; Intellia: Honoraria; Omeros: Research Funding; Athersys: Other: Data and Safety Monitoring Board, Patents & Royalties; Vor Pharma: Other: Data and Safety Monitoring Board. McGuirk: Kite/ Gilead: Consultancy, Honoraria, Other: travel accommodations, expense, Kite a Gilead company, Research Funding, Speakers Bureau; Juno Therapeutics: Consultancy, Honoraria, Research Funding; Allovir: Consultancy, Honoraria, Research Funding; Magenta Therapeutics: Consultancy, Honoraria, Research Funding; EcoR1 Capital: Consultancy; Novartis: Research Funding; Fresenius Biotech: Research Funding; Astelllas Pharma: Research Funding; Bellicum Pharmaceuticals: Research Funding; Novartis: Research Funding; Gamida Cell: Research Funding; Pluristem Therapeutics: Research Funding. Bachy: Kite, a Gilead Company: Honoraria; Novartis: Honoraria; Daiishi: Research Funding; Roche: Consultancy; Takeda: Consultancy; Incyte: Consultancy. Dreyling: Amgen: Speakers Bureau; Astra Zeneca: Consultancy, Speakers Bureau; Bayer: Consultancy, Research Funding, Speakers Bureau; Celgene: Consultancy, Research Funding, Speakers Bureau; Gilead Kite: Consultancy, Research Funding, Speakers Bureau; Incyte: Consultancy, Speakers Bureau; Janssen: Consultancy, Research Funding, Speakers Bureau; Novartis: Consultancy, Speakers Bureau; Roche: Consultancy, Research Funding, Speakers Bureau; AbbVie: Research Funding; BeiGene: Consultancy, Speakers Bureau; Genmab: Consultancy; MorphoSys: Consultancy. Harigae: Bristol Myers Squibb: Honoraria; Novartis Pharma: Honoraria, Research Funding; Chugai Pharma: Honoraria; Janssen Pharma: Honoraria; Ono pharma: Honoraria, Other: Subsidies or Donations; Astellas Pharma: Other: Subsidies or Donations; Kyowakirin: Other: Subsidies or Donations. Bond: Kite/Gilead: Honoraria. Andreadis: Merck: Research Funding; BMS: Research Funding; CRISPR Therapeutics: Research Funding; GenMAB: Research Funding; Novartis: Research Funding; Roche: Current equity holder in publicly-traded company, Ended employment in the past 24 months; Epizyme: Honoraria; Incyte: Honoraria; TG Therapeutics: Honoraria; Kite: Honoraria; Karyopharm: Honoraria; Atara: Consultancy, Honoraria. McSweeney: Kite-Gilead: Consultancy, Honoraria, Research Funding, Speakers Bureau; Gamida: Consultancy, Honoraria; TG Theraputics: Consultancy, Honoraria; Autolous Limited: Research Funding; Novartis: Research Funding; NKARTA: Research Funding; Allovir: Research Funding. Newsome: Novartis: Current Employment. Degtyarev: Novartis: Current Employment, Current equity holder in publicly-traded company. Del Corral: Novartis: Current Employment. Andreola: Novartis: Current Employment, Current holder of stock options in a privately-held company. Masood: Novartis: Current Employment, Current holder of stock options in a privately-held company. Schuster: AbbVie, Acerta, Celgene/Juno, DTRM Bio, Genentech, Incyte, Merck, Novartis, Portola, and TG Therapeutics: Research Funding; Acerta, AlloGene, AstraZeneca, BeiGene, Celgene/Juno, Genentech/Roche, LoxoOncology, Novartis, and Tessa Therapeutics: Consultancy; Acerta, AlloGene, AstraZeneca, BeiGene, Celgene, Genentech/Roche, LoxoOncology, Novartis, Nordic Nanovector, Pfizer, and Tessa Therapeutics;: Honoraria; Novartis: Other: Personal fees, Patents & Royalties; AbbVie, Celgene, Novartis, Juno, Nordic Nanovector, and Pfizer: Other: Steering Committee Participation. Jaeger: BMS/Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Norvartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees. Westin: Kite, a Gilead Company: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Bristol Myers Squibb: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; MorphoSys: Consultancy, Research Funding; AstraZeneca: Consultancy, Research Funding; ADC Therapeutics: Consultancy, Research Funding; Umoja: Consultancy; Iksuda Therapeutics: Consultancy; Curis: Research Funding; Morphosys: Research Funding; 47 Inc: Research Funding.

Integrated Genomic Analysis Identifies UBTF Tandem Duplications As a Subtype-Defining Lesion in Pediatric Acute Myeloid Leukemia

Children with acute myeloid leukemia (AML) have a dismal prognosis due to a high relapse rate; however, the molecular basis leading to relapsed pediatric AML has not yet been fully characterized. To define the spectrum of alterations common at relapse, we performed integrated profiling of 136 relapsed pediatric AML cases with RNA sequencing (RNA-seq), whole-genome sequencing, and target-capture sequencing. In addition to well-characterized fusion oncoproteins, such as those involving KMT2A (n=36, 26.5%) or NUP98 (n=18, 13.2%), we also identified somatic mutations in UBTF (upstream binding transcription factor) in 12 of 136 cases (8.8%) of this relapsed cohort.Somatic alterations of the UBTF gene, which encodes a nucleolar protein that is a component of the RNA Pol I pre-initiation complex to ribosomal DNA promoters, have rarely been observed in AML. In our cohort, all alterations can be described as heterozygous in-frame exon 13 tandem duplications (UBTF-TD), either at the 3' end of exon 13 of UBTF or of the entire exon 13 (Fig. A). As we noticed limited detection in our pipeline as a result of complex secondary indels alongside the duplications, we established a soft-clipped read-based screening method to detect UBTF-TD more efficiently. Applying the screening to RNA-seq data of 417 additional pediatric AMLs from previous studies and our clinical service, we identified 15 additional UBTF-TDs, many of which have not been previously reported. At the amino acid level, UBTF-TDs caused amino acid insertions of variable sizes (15-181 amino acids), duplicating a portion of high mobility group domain 4 (HMG4), which includes short leucine-rich sequences.UBTF-TD AMLs commonly occurred in early adolescence (median age: 12.6, range: 2.4-19.6), and 19 of the total 27 cases had either normal karyotype (n=12) or trisomy 8 (n=7). UBTF-TD is mutually exclusive from other recurrent fusion oncoproteins, such as NUP98 and KMT2A rearrangements (Fig. B), but frequently occurred with FLT3-ITD (44.4%) or WT1 mutations (40.7%). The median variant allele fraction (VAF) of the UBTF-TD was 48.0% (range: 9.7-66.7%). In four cases with data at multiple disease time points, the identical UBTF-TDs were present at high allele fractions at all time points, suggesting that UBTF-TD is a clonal alteration. tSNE analysis of the transcriptome dataset showed that UBTF-TD AMLs share a similar expression pattern with NPM1 mutant and NUP98-NSD1 AML subtypes, including NKX2-3 and HOXB cluster genes (Fig. C) . Altogether, these findings suggest that UBTF-TD is a unique subtype of pediatric AML.To address the impact of UBTF-TD expression in primary hematopoietic cells, we introduced UBTF-TD and UBTF wildtype expression vectors into cord blood CD34+ cells via lentiviral transduction. UBTF-TD expression promotes colony-forming activity and cell growth, yielding cells with a persistent blast-like morphology (Fig. D). Further, transcriptional profiling of these cells demonstrated expression of HOXB genes and NKX2-3, similar to UBTF-TD AMLs in patients, indicating that UBTF-TD is sufficient to induce the leukemic phenotype.To investigate the prevalence of UBTF-TDs in larger de novo AML cohorts, we applied the above UBTF-TD screening method to the available de novo AML cohorts of TCGA (n=151, adult), BeatAML (n=220, pediatric and adult), and AAML1031 (n=1035, pediatric). We identified UBTF-TDs in 4.3% (45/1035) of the pediatric AAML1031 cohort, while the alteration is less common (0.9%: 3/329, p=0.002) in the adult AML cohorts (Fig. E). In the AAML1031 cohort, UBTF-TDs remain mutually exclusive with known molecular subtypes of AML and commonly occur with FLT3-ITD (66.7%) and WT1 (40.0%) mutations and either normal karyotype or trisomy 8. The presence of UBTF-TDs in the AAML1031 cohort is associated with a poor outcome (Fig. F, median overall survival, 2.3 years) and MRD positivity; multivariate analysis revealed that UBTF-TD and WT1 are independent risk factors for overall survival within FLT3-ITD+ AMLs.In conclusion, we demonstrate UBTF-TD defines a unique subtype of AMLs that previously lacked a clear oncogenic driver. While independent of subtype-defining oncogenic fusions, UBTF-TD AMLs are associated with FLT3-ITD and WT1 mutations, adolescent age, and poor outcomes. These alterations have been under-recognized by standard bioinformatic approaches yet will be critical for future risk-stratification of pediatric AML. [Display omitted] DisclosuresIacobucci: Amgen: Honoraria; Mission Bio: Honoraria. Miller: Johnson & Johnson's Janssen: Current Employment. Mullighan: Pfizer: Research Funding; Illumina: Membership on an entity's Board of Directors or advisory committees; AbbVie: Research Funding; Amgen: Current equity holder in publicly-traded company.

Conversion of HbS to Hb G-Makassar By Adenine Base Editing Is Compatible with Normal Hemoglobin Function

Conversion of the pathogenic sickle allele to a naturally occurring, non-pathogenic hemoglobin variant, Hb G-Makassar, represents a long-term and durable treatment strategy for sickle cell disease (SCD). Using our engineered adenine base editor, we achieved highly efficient base editing in mobilized sickle trait (HbAS) and non-mobilized homozygous sickle (HbSS) CD34 + cells that led to >70% conversion of sickle allele to Makassar allele in in vitro erythroid differentiated (IVED) cells derived from ex vivo edited CD34 + cells. At this level of editing, >70% bi-allelic Makassar editing could be achieved in HbSS IVED cells, with ~20% of cells being mono-allelically Makassar edited. These mono-allelically edited cells behaved similarly to sickle trait (HbAS) cells, when exposed to hypoxic conditions in vitro. In vivo proof of concept xenotransplantation studies demonstrated that Makassar edited HbAS CD34 + cells achieved long-term, multi-lineage hematopoietic engraftment as well as Makassar globin protein expression in human erythroid glycophorin A + cells in thebone marrow of immunocompromised mice.Although the Makassar variant is naturally occurring in human genetics and present in individuals in Southeast Asia with normal hematologic parameters in both heterozygous and homozygous states, we sought to further characterize Makassar hemoglobin and assess its biophysical and biochemical properties. Recombinant Makassar globin was co-expressed with alpha globin in E. coli and tetramers were purified to homogeneity. Recombinant tetramers were assessed for identity, purity, globin content, and heme content demonstrating comparability to hemoglobin tetramers isolated from primary sources (whole blood). Several characterization methods were employed, to assess size, molecular weight, oligomerization state, tetramer composition, and oxygen binding properties. These studies indicated Makassar globin could properly assemble into hemoglobin tetramers, displaying biochemical properties characteristic of hemoglobins. Furthermore, we assessed polymerization potential using a temperature jump method previously employed for kinetic measurements of sickle-fiber formation and found Makassar hemoglobin did not polymerize in vitro under conditions where sickle hemoglobin (HbS) readily polymerizes, consistent with behavior observed previously by others. Finally, a crystal structure of Hb G-Makassar has been determined at the 2.2 Å resolution and showed high similarity to the HbA (wildtype hemoglobin) structure with a RMSD of 0.385 Å for all the Cα atoms, which indicates that the glutamic acid to alanine (E6A) substitution in beta-hemoglobin does not seem to induce any significant conformational change in hemoglobin structures.Altogether, our biophysical and biochemical characterization shows that the Makassar variant behaves as a functional hemoglobin. By replacing the pathogenic sickle globin with a benign hemoglobin variant with normal function, our base editing approach provides a promising autologous investigational cell therapy for the treatment of SCD. DisclosuresChu: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Ortega: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company. Feliciano: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company. Winton: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company. Xu: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Haupt: Beam Therapeutics: Current Employment, Current holder of stock options in a privately-held company. McDonald: Beam Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Martinez: Beam Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Liquori: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company. Marshall: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company. Lam: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Olins: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Rinaldi: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Rehberger: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Lazarra: Beam Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Decker: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Gantzer: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Bohnuud: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Born: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Barrera: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Yan: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Slaymaker: Beam Therapeutics: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties. Packer: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Smith: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Zambonelli: Beam Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Lee: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Gaudelli: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Hartigan: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Ciaramella: Beam Therapeutics: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company, Membership on an entity's Board of Directors or advisory committees.

Pre-Clinical Development of a Highly Efficient TALEN ®-Based Correction of the β-Globin Gene in Patient-Derived Hematopoietic Stem and Progenitor Cells (HSPCs) to Treat Sickle Cell Disease

Sickle cell disease (SCD) is one of the most common inherited diseases affecting millions of people worldwide. SCD stems from a single point mutation (A>T) in exon 1 of the HBB gene which results in sickle hemoglobin. The only available curative treatment of SCD is allogeneic hematopoietic stem cell transplant, which is only viable for ~20% of SCD patients.Ex-vivo gene therapy approaches have shown to be a promising therapeutic option for patients. Most products currently in the clinic have focused on methods to enhance functional hemoglobin production (e.g., disruption of BCL11A to encourage β-globin gene switching or direct insertion of a functional β-globin gene into patient HSPCs).The Cellectis approach is to directly repair the mutated HBB gene in order to restore HbA production. TALGlobin01 is an autologous HSPC-based gene therapy product designed with a TALEN ® optimized to cleave the sickle HBB gene (TALEN-HBB01) and an AAV based engineering process leading to highly efficient HBB gene correction via endogenous homology directed repair (HDR), while mitigating potential risks of HBB gene knock-out (KO).Use of TALGlobin01 resulted in up to 70% of HDR-mediated HBB gene correction (56% mean frequency) in homozygous sickle (HbSS) patient HSPCs with only 20% of NHEJ-dependent insertion/deletion (indels) events detected. This gene correction process did not affect cell viability, hematopoietic stem/progenitor immunophenotype or differentiation potential of corrected HSPCs. Allelic editing at clonal resolution in single BFU-E colonies showed that up to 72% (with a mean of 50%) of progenitors contained at least one corrected allele, while only 23% were either not corrected or had indels on one allele. Notably, our optimized engineering process led to only 9% of colonies harboring bi-allelic indels events.To evaluate the ability of TALGlobin01 to prevent the sickling phenotype associated with SCD, we performed in vitro differentiation of HbSS patient-edited cells into late-stage erythroid cells and assessed HbA protein production by HPLC. We observed that HbA accounted for up to 60% (with a mean of 49%) of the total Hb with a concomitant decrease of HbS production from 90% to 19%. Interestingly, our gene correction process maintained a balanced α chain/non-α chain ratio, consistent with our genotyping results showing a low frequency of clones harboring bi-allelic indels. More importantly, efficient expression of HbA was translated into a sharp decrease of hypoxia-induced sickling rate of in vitro-generated erythrocytes when comparing unedited to TALGlobin01 edited cells (from 95% to 13%, respectively).When injected in vivo, engineered HSPCs from non-mobilized SCD patients retained the capacity to engraft into immunodeficient NSG mice with levels of allelic correction comparable to the input (~40%) and still detectable at 16-17 weeks post-transplantation.An unbiased genome wide approach (OCA), coupled to target enrichment high-throughput sequencing screening, confirmed the TALEN-HBB01 cleavage activity at only one off-target site located at the HBD locus. TALEN-HBB01 cleavage activity was assessed at this off-target site in patient HSPCs engineered with TALGlobin01 and found to be very low compared to the on-site cleavage activity (50.7% indels at the on-site versus 1.2% at the off-site).Furthermore, we developed a clinical-scale TALGlobin01 manufacturing process that achieved up to 60% of HDR-mediated HBB gene correction and less than 10% of NHEJ-mediated HBB gene inactivation in healthy donor derived HSPCs.These results are the first demonstration that a TALEN-based engineering process could be used to efficiently correct the SCD mutated HBB gene in HbSS patient-derived CD34+ HSPCs. Taken together, the high level of HbA expression and reversion of sickling phenotype, the efficient in vivo long-term engraftment potential of TALGlobin01 edited cells and the low levels of HBB KO or off-target cleavage generated by our gene correction process, warrant the clinical evaluation of TALGlobin01 to treat SCD patients. DisclosuresMoiani: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company. Hong: Cellectis, Inc.: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Letort: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company. Lizot: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company. Chirinos: Cellectis, S.A.: Current Employment. Temburni-Blake: Cellectis, Inc.: Current Employment, Current equity holder in publicly-traded company. Mayer: Cellectis, Inc.: Current Employment, Current equity holder in publicly-traded company. Leduc: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company. Pinard: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company. Foray: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company. Boyne: Cellectis, Inc.: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months, Patents & Royalties. Kazancioglu: Cellectis, Inc.: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Dusseaux: Cellectis, S.A.: Current equity holder in publicly-traded company, Ended employment in the past 24 months. Gouble: Cellectis SA: Current Employment, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Frattini: Celgene/BMS: Current equity holder in publicly-traded company, Ended employment in the past 24 months; Cellectis, Inc.: Current Employment, Current equity holder in publicly-traded company. Brownstein: BMS: Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months, Ended employment in the past 24 months; Cellectis, Inc.: Current Employment, Current equity holder in publicly-traded company. Duclert: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Shiffer-Mannoui: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Juillerat: Cellectis, Inc.: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Duchateau: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Valton: Cellectis, S.A.: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties.

Potent and Orally Bioavailable BCL6 PROTAC TM Degraders Demonstrate Efficacy in Pre-Clinical Models of Diffuse Large B-Cell Lymphoma (DLBCL)

The BCL6 transcription repressor (B-cell lymphoma 6, BCL6) protein has been shown to be a key molecular driver of diffuse large B-cell lymphoma (DLBCL). Somatic mutations of the BCL6 gene include gross- or cryptic-chromosome translocations and point mutations that have been shown to result in the deregulation of BCL6 expression. These BCL6 abnormalities also contribute to a subgroup of high-risk (HR) aggressive double- and triple-hit (DH/TH) lymphomas with very poor outcomes. We have developed highly specific, potent and orally bioavailable BCL6 PROteolysis TArgeting Chimera (PROTAC TM) degraders that demonstrate potent in-vitro and in-vivo efficacy in multiple pre-clinical DLBCL models.Ten of 12 germinal center B-cell (GCB) and two of four activated B-cell (ABC) DLBCL cell lines show significant growth inhibition in-vitro with BCL6 PROTAC TM treatment, demonstrating a critical dependence on BCL6. This array of sensitivity across genetically variable cell lines suggests that BCL6-dependence is not just associated with BCL6-mutated DLBCLs. A more advanced BCL6 PROTAC TM, ARVN-71228, achieves >95% BCL6 D max in-vitro at a DC 50 of <1 nM in the OCI-Ly1 model following 24 hr treatment, degrading BCL6 equally well in the nuclear, chromatin-bound and cytosolic cell fractions. BCL6 degradation is associated with dose-dependent G1 cell cycle arrest and elevated apoptosis that increases over time (24 vs 72 hours). In head-to-head BCL6 degradation and growth inhibition studies using OCI-Ly1, the ARVN-71228 BCL6 PROTAC TM demonstrates superior activity compared to recently published BCL6-targeted degraders/inhibitors and heterobifunctional molecules.Importantly, medicinal chemistry efforts have resulted in the successful development of orally bioavailable BCL6 PROTAC TM degraders for in-vivo dosing. Time-course studies show >95% BCL6 loss within four hours which is maintained at 8-, 16- and 24-hours. Genes repressed by BCL6 such as BLIMP1 and PTPN6 are derepressed and show increased protein levels 24 hours post-dose. ARVN-71228 achieves regressions in the GCB OCI-Ly1 CDX model. Future studies plan to look at rational drug combinations with BCL6 PROTAC TM degraders to find collaborative or synergistic pathways to target, especially in the HR-DLBCL subtypes where there is a high unmet medical need. DisclosuresGough: Arvinas: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Sherman: Arvinas: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. DeCarr: Arvinas: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Eaton: Arvinas: Current Employment, Current equity holder in publicly-traded company. Milanovic: Arvinas: Current Employment, Current equity holder in publicly-traded company. Bookbinder: Arvinas: Current Employment, Current equity holder in publicly-traded company. Pizzano: Arvinas: Current Employment, Current equity holder in publicly-traded company. Altieri: Arvinas: Current Employment, Current equity holder in publicly-traded company. Corradi: Arvinas: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Xiao: Arvinas: Current Employment, Current equity holder in publicly-traded company. Gallego: Arvinas: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Soto: Arvinas: Current Employment, Current equity holder in publicly-traded company. Lingamaneni: Arvinas: Current Employment, Current equity holder in publicly-traded company. Chen: Arvinas: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Zhang: Arvinas: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Wang: Arvinas: Current Employment, Current equity holder in publicly-traded company. Dong: Arvinas: Current Employment, Current equity holder in publicly-traded company. Chirnomas: Arvinas: Current Employment, Current equity holder in publicly-traded company. Berlin: Arvinas: Current Employment, Current equity holder in publicly-traded company. Hornberger: Arvinas: Current Employment, Current equity holder in publicly-traded company. Snyder: Arvinas: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months. Taylor: Arvinas: Current Employment, Current equity holder in publicly-traded company, Divested equity in a private or publicly-traded company in the past 24 months.

Gene Therapy for Fanconi Anemia [Group A]: Interim Results of RP-L102 Clinical Trials

▪Background: Fanconi anemia (FA) is a disorder of defective deoxyribonucleic acid (DNA) repair, progressive bone marrow failure (BMF), and a predisposition to hematologic malignancies and solid tumors. Approximately 60 to 70% of all cases result from a mutation in the Fanconi Anemia Complementation Group A (FANCA) gene (FA-A). 80% of FA patients experience BMF within the first decade of life. Allogeneic hematopoietic stem cell transplant (alloHSCT) is potentially curative for BMF; however, its efficacy is limited by human leukocyte antigen (HLA)-matched sibling donor availability and transplant-related toxicities. Lentiviral mediated gene therapy utilizing autologous FA-A CD34+ enriched hematopoietic stem and progenitor cells (HSPCs) confers a proliferative advantage to gene-corrected HSPCs as demonstrated in preclinical studies and the FANCOLEN-I clinical trial conducted in Madrid, Spain. We report results from ongoing RP-L102 studies using “Process B” manufacturing optimizations including transduction enhancers, commercial grade vector, and modified cell processing.Methods: Patients (pts) with a FANCA gene mutation, age ≥1 year with no HLA-matched sibling donor and at least 30 CD34+ cells/µL in bone marrow (BM) are eligible. Peripheral blood (PB) mononuclear cells are collected via leukapheresis on 2 consecutive days after mobilization with granulocyte-colony stimulating factor (G-CSF) and plerixafor. CD34+ HSPCs are enriched, transduced with a lentiviral vector encoding for the FANCA gene (PGK-FANCA-WPRE) and infused without cryopreservation or conditioning. Patients are followed for 3 years post-infusion for safety assessments (replication competent lentivirus [RCL], insertion site analysis [ISA]) and to ascertain evidence of efficacy (increasing PB and BM vector copy number [VCN] and mitomycin-C [MMC] resistance in BM colony forming units [CFUs]), along with stabilization/correction of cytopenias.Results: As of March 2021, 9 pts (aged 2 to 6 years) have received RP-L102. Evidence of engraftment has been identified in 6 pts with ≥6 months of follow up as indicated by PB VCN. 2 of 3 pts with follow up of ≥12 months have shown increased MMC resistance in BM CFUs. 1 pt developed BMF requiring alloHSCT after influenza B infection. 1 pt had a serious Grade 2 transient RP-L102 infusion-related reaction. Updated safety and efficacy data for pts with ≥12 months of follow-up will be presented.Conclusions: RP-L102's safety profile remains favorable. Increasing evidence of engraftment has been confirmed in 6 subjects as demonstrated by PB VCN; without conditioning, 12+ months of follow up is likely required to observe the proliferative advantage of transduced HSPCs. DisclosuresCzechowicz: Editas Medicine: Current equity holder in publicly-traded company; Rocket Pharmaceuticals, Inc.: Research Funding; Global Blood Therapeutics: Current equity holder in publicly-traded company; Stemodontics: Consultancy, Current equity holder in publicly-traded company; GV: Consultancy, Current equity holder in publicly-traded company; Spotlight Therapeutics: Consultancy, Current equity holder in publicly-traded company; Beam Therapeutics: Consultancy, Current equity holder in publicly-traded company; Magenta Therapeutics: Current equity holder in publicly-traded company, Other: Intellectual property rights; Decibel Therapeutics: Current equity holder in publicly-traded company; Forty Seven Inc.: Divested equity in a private or publicly-traded company in the past 24 months; Gilead Sciences: Other: intellectual property rights; Jasper Therapeutics: Other: Intellectual property rights. Sevilla: Rocket Pharmaceuticals, Inc.: Consultancy, Other: J.Sevilla is an inventor on patents on lentiviral vectors filed by CIEMAT, CIBERER and Fundación Jiménez Díaz, and may be entitled to receive financial benefits from the licensing of such patents., Patents & Royalties: J. Sevilla is a consultant and has licensed medical products from Rocket Pharmaceuticals, Inc. ; Amgen: Consultancy; Novartis: Consultancy; Miltenyi: Consultancy; SOBI: Consultancy. Booth: GSK: Honoraria; Orchard Therapeutics: Consultancy, Honoraria; SOBI: Consultancy, Honoraria; Takeda: Honoraria; Rocket Pharmaceuticals, Inc.: Consultancy. Río: Rocket Pharmaceuticals: Current equity holder in publicly-traded company, Other: Paula Rio has licensed medicinal products and receives research funding and equity from Rocket Pharmaceuticals, Inc., Research Funding. Navarro: Rocket Pharmaceuticals, Inc.: Current equity holder in publicly-traded company, Other: Dr. Navarro has licensed medicinal products and receives research funding and equity from Rocket Pharmaceuticals, Inc., Patents & Royalties, Research Funding. Beard: Rocket Pharmaceuticals, Inc.: Current Employment, Current equity holder in publicly-traded company. Law: Rocket Pharmaceuticals, Inc.: Current Employment, Current equity holder in publicly-traded company. Choi: Rocket Pharmaceuticals, Inc.: Current Employment, Current equity holder in publicly-traded company. Zeini: Rocket Pharmaceuticals, Inc.: Current Employment, Current equity holder in publicly-traded company. Nicoletti: Rocket Pharmaceuticals, Inc.: Current Employment, Current equity holder in publicly-traded company. Wagner: Magenta Therapeutics: Consultancy; Rocket Pharmaceuticals, Inc.: Consultancy, Current equity holder in publicly-traded company; Vertex Pharmaceuticals: Consultancy; ASC Therapeutics: Consultancy; Bluebird Bio: Consultancy. Rao: Rocket Pharmaceuticals, Inc.: Current Employment, Current equity holder in publicly-traded company. Thrasher: Orchard Therapeutics: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; 4Bio Capital: Consultancy, Membership on an entity's Board of Directors or advisory committees; Generation bio: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Rocket Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees. Schwartz: Rocket Pharmaceuticals, Inc.: Current Employment, Current equity holder in publicly-traded company. Bueren: Rocket Pharmaceuticals, Inc.: Consultancy, Other: J.Bueren is an inventor on patents on lentiviral vectors filed by CIEMAT, CIBERER and Fundación Jiménez Díaz, may be entitled to receive financial benefits from the licensing of such patents and receives funding for research., Patents & Royalties, Research Funding.

Awareness, Knowledge, and Preferences of United States (US) Patients with Chronic Lymphocytic Leukemia (CLL) and Their Caregivers Related to Finite Duration (FD) Therapy and Minimal (Measurable) Residual Disease (MRD)

Introduction: The management of CLL patients historically utilized FD chemoimmunotherapy (CIT), including fludarabine, cyclophosphamide and rituximab (FCR), or bendamustine and rituximab (BR). That changed with the introduction of Bruton Tyrosine Kinase inhibitors (BTKi) as an option for oral targeted monotherapy delivered continuously until disease progression or intolerance. Recently, novel combinations that include a B-cell lymphoma 2 inhibitor (BCL2i), such as venetoclax, have been developed as FD therapy. In addition, though not FDA approved, MRD assessment is emerging as a potential tool to guide FD therapy. Understanding the patient and their caregiver perspectives on FD and MRD is critical to providing the best care.Methods: CLL Society developed a web-based survey instrument of multiple-choice questions to assess patient and caregiver awareness, understanding, and preferences with MRD and FD therapies, where FD referred to both time and MRD guided limited duration therapies. Patients and caregivers were invited to participate via message boards, CLL Society's website, email, and online communities.Results: A total of 607 (96%) self-identified US patients and 36 (5%) caregivers, who answered on behalf of patients in their care, completed the survey between 31/AUG/2020 and 31/JAN/2021. The mean patient age was 64 (range, 30-90) years, 54% were female, 169 patients (27%) treatment-naïve, 239 (38%) with one line of prior therapy, and 214 (34%) with two or more. Targeted agents (e.g., venetoclax, ibrutinib, acalabrutinib, idelalisib, and duvelisib) had been used by 401 (64%). FD use with some novel therapies was known of by 565 (90%) patients, while 588 (93%) knew some were used as continuous daily treatment until disease progression or intolerance. If effectiveness and side effects were assumed similar, 485 (77%) preferred FD, and 398 (63%) wanted that finite treatment to be guided by MRD status. Only 42 (7%) preferred continuous therapy until disease progression or intolerance. The chance for continued remission while off treatment was ranked as a very important potential benefit after stopping a FD therapy by 581 (92%) patients. When considering therapy, 589 (93%), 575 (91%), 447 (71%), 415 (66%), 302 (48%), and 288 (46%) rated having overall survival, having future therapeutic options, avoiding chemotherapy, the ability to reach undetectable (u)MRD, minimal lab and office visits, and FD therapy as very important, respectively (Fig. 1). Being tested for MRD at least once was reported by 215 (34%), while 327 (52%) had not been tested, and 88 (14%) were unsure. There were 562 (89%) who were somewhat or very familiar with the term MRD, while 450 (71%) were confident of understanding what MRD testing measures, and 446 (70%) understood its role in CLL. Only 225 (36%) stated they understood different testing methods. While self-rated understanding of the indications for MRD testing was relatively high, some wanted testing done when not clinically indicated, such as when there was persistent CLL by routine laboratory findings, or at the time of diagnosis (Fig. 2). Respondents were mixed in their stated understanding as to which therapies could result in uMRD, with 241 (38%), 197 (32%), and 195 (32%) not knowing if CIT, BCL2i, and BTKi, could lead to uMRD, respectively. Two hundred and sixty-five (42%), 84 (13%), and 57 (9%) believed it was possible for most on a BCL2i, BTKi, or CIT to achieve uMRD, respectively (Fig. 3). If patients had evidence of MRD at the end of treatment, 365 (58%) would request frequent monitoring but would make no immediate treatment decisions based on MRD status, 279 (44%) were unsure, and only 88 (14%) would want further treatment immediately.Conclusions: The results from this patient-based study reveal a high awareness of FD therapies and MRD testing, as well as a clear preference for FD among internet-active patients and caregivers. However, there are gaps in the knowledge base, thus significant educational opportunities. Limitations included the opt-in sample where the survey results may not be projectable to the total CLL population. Moreover, the survey was not designed to assess preferences if either FD or continuous therapy offered improved outcomes. Nonetheless, as the potential role of MRD and FD therapy grows in CLL it will be incumbent on clinicians to ensure patients are well informed and engaged to help them better share therapeutic decisions. [Display omitted] DisclosuresKoffman: AbbVie: Current equity holder in publicly-traded company; AstraZeneca: Current equity holder in publicly-traded company, Honoraria; Beigene: Current equity holder in publicly-traded company; Bristol Myer Squibb: Current equity holder in publicly-traded company, Honoraria; Gilead: Current equity holder in publicly-traded company; Johnson & Johnson: Current equity holder in publicly-traded company, Honoraria; Sunesis Pharmaceuticals: Current equity holder in publicly-traded company; TG Therapeutics: Current equity holder in publicly-traded company; Regeneron: Current equity holder in publicly-traded company; Miragen Therapeutics: Current equity holder in publicly-traded company; Novartis: Membership on an entity's Board of Directors or advisory committees; Portola Pharma: Current equity holder in publicly-traded company; MEI Pharma: Current equity holder in publicly-traded company; Merck: Current equity holder in publicly-traded company. Stewart: Johnson & Johnson: Current equity holder in publicly-traded company; Novartis: Current equity holder in publicly-traded company; Dr. Reddy's Laboratories Limited: Current equity holder in publicly-traded company; Adaptive Biotechnologies: Current equity holder in publicly-traded company; Pacific Biosciences: Current equity holder in publicly-traded company. Byrd: Novartis, Trillium, Astellas, AstraZeneca, Pharmacyclics, Syndax: Consultancy, Honoraria; Vincerx Pharmaceuticals: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Newave: Membership on an entity's Board of Directors or advisory committees. Danilov: Bayer Oncology: Consultancy, Honoraria, Research Funding; Genentech: Consultancy, Honoraria, Research Funding; Takeda Oncology: Research Funding; Rigel Pharm: Honoraria; TG Therapeutics: Consultancy, Research Funding; Abbvie: Consultancy, Honoraria; Beigene: Consultancy, Honoraria; Pharmacyclics: Consultancy, Honoraria; Gilead Sciences: Research Funding; Bristol-Meyers-Squibb: Honoraria, Research Funding; SecuraBio: Research Funding; Astra Zeneca: Consultancy, Honoraria, Research Funding. Davids: BeiGene: Consultancy; Janssen: Consultancy; Ascentage Pharma: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Astra-Zeneca: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding; BMS: Consultancy, Research Funding; MEI Pharma: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; Adaptive Biotechnologies: Consultancy; AbbVie: Consultancy; Surface Oncology: Research Funding; TG Therapeutics: Consultancy, Research Funding; Celgene: Consultancy; Verastem: Consultancy, Research Funding; Eli Lilly and Company: Consultancy; MEI Pharma: Consultancy; Merck: Consultancy; Research to Practice: Consultancy; Takeda: Consultancy. Furman: Beigene: Consultancy; Acerta/AstraZeneca: Consultancy; Janssen: Consultancy, Honoraria; Sunesis: Consultancy; Genentech: Consultancy; Incyte: Consultancy; Pharmacyclics: Consultancy; TG Therapeutics: Consultancy; Loxo Oncology: Consultancy; Sanofi: Consultancy; Morphosys: Consultancy; Verastem: Consultancy; X4 Pharmaceuticals: Consultancy; Oncotracker: Consultancy; Abbvie: Consultancy, Honoraria, Other: Expert testimony; AstraZeneca: Honoraria. Jain: Bristol Myers Squibb: Honoraria, Research Funding; Genentech: Honoraria, Research Funding; AstraZeneca: Honoraria, Research Funding; Fate Therapeutics: Research Funding; Beigene: Honoraria; AbbVie: Honoraria, Research Funding; Pfizer: Research Funding; TG Therapeutics: Honoraria; Cellectis: Honoraria, Research Funding; Precision Biosciences: Honoraria, Research Funding; ADC Therapeutics: Honoraria, Research Funding; Aprea Therapeutics: Research Funding; Adaptive Biotechnologies: Honoraria, Research Funding; Servier: Honoraria, Research Funding; Janssen: Honoraria; Incyte: Research Funding; Pharmacyclics: Research Funding. Kay: Oncotracker: Membership on an entity's Board of Directors or advisory committees; Genentech: Research Funding; Targeted Oncology: Membership on an entity's Board of Directors or advisory committees; Morpho-sys: Membership on an entity's Board of Directors or advisory committees; Acerta Pharma: Research Funding; MEI Pharma: Research Funding; Behring: Membership on an entity's Board of Directors or advisory committees; Pharmacyclics: Membership on an entity's Board of Directors or advisory committees, Research Funding; AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Rigel: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees; Dava Oncology: Membership on an entity's Board of Directors or advisory committees; Tolero Pharmaceuticals: Research Funding; CytomX Therapeutics: Membership on an entity's Board of Directors or advisory committees; Bristol Meyer Squib: Membership on an entity's Board of Directors or advisory committees, Research Funding; Agios Pharm: Membership on an entity's Board of Directors or advisory committees; TG Therapeutics: Research Funding; Sunesis: Research Funding; Juno Therapeutics: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding. Lamanna: Juno Therapeutics, Inc.: Research Funding; BeiGene: Consultancy; AstraZeneca: Consultancy, Research Funding; AbbVie: Consultancy, Research Funding; MingSight Pharmaceuticals, Inc.: Research Funding; Genentech, Inc.: Consultancy, Research Funding; Gilead Sciences, Inc.: Consultancy; TG Therapeutics, Inc: Research Funding; Verastem Oncology: Research Funding; Celgene Corporation: Consultancy; Oncternal Therapeutics: Research Funding; Pharmacyclics: Consultancy; Janssen Pharmaceuticals, Inc.: Consultancy. Mato: MSKCC: Current Employment; Pharmacyclics LLC, an AbbVie Company: Consultancy, Research Funding; TG Therapeutics: Consultancy, Other: DSMB, Research Funding; Nurix: Research Funding; BeiGene: Consultancy, Research Funding; Janssen: Consultancy, Research Funding; LOXO: Consultancy, Research Funding; Genmab: Research Funding; AstraZeneca: Consultancy; Sunesis: Consultancy, Research Funding; DTRM BioPharma: Consultancy, Research Funding; Acerta/AstraZeneca: Consultancy, Research Funding; Johnson and Johnson: Consultancy, Research Funding; AbbVie: Consultancy, Research Funding; Adaptive Biotechnologies: Consultancy, Research Funding; Genentech: Consultancy, Research Funding. Ujjani: Adaptive Biotechnologies: Research Funding; Atara Bio: Consultancy; Janssen: Consultancy; Epizyme: Consultancy, Membership on an entity's Board of Directors or advisory committees; TG Therapeutics: Honoraria; Gilead: Honoraria; ACDT: Honoraria; Kite, a Gilead Company: Honoraria; Loxo: Research Funding; AstraZeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding. Pagel: Kite, a Gilead Company: Consultancy; Incyte/MorphoSys: Consultancy; AstraZeneca: Consultancy; Actinium Pharmaceuticals: Consultancy; Gilead: Consultancy; Pharmacyclics/AbbVie: Consultancy; MEI Pharma: Consultancy; Epizyme: Consultancy; BeiGene: Consultancy.

Engineered Hypoimmune Allogeneic CAR T Cells Exhibit Innate and Adaptive Immune Evasion Even after Sensitization in Humanized Mice and Retain Potent Anti-Tumor Activity

Off-the-shelf CAR T cells may offer advantages over autologous strategies, including ease of manufacturing, improved quality control with avoidance of malignant contamination and T cell dysfunction as well as the ability to generate a final product from healthy T cells. While TCR editing can effectively prevent graft-versus-host reactions, the significant host-versus-graft immune response against histoincompatible T cells prevents the expansion and persistence of allogeneic CAR T cells and mitigates the efficacy of this approach. The goal is to achieve improved rates of durable complete remissions by improving allogeneic CD19CAR persistence since it has been shown that autologous CAR T cells have greater durability over years than allogeneic CAR T cells (N Engl J Med. 2021;384(7):673-674).We describe here the engineering of human immune evasive CAR T cells based on our previously described hypoimmune technology (Nat Biotechnol 2019;37(3):252-258 and Proc Natl Acad Sci U S A 2021;118(28):e2022091118).A major challenge is that, while HLA deletion can result in adaptive immune evasion, innate reactivity is enhanced by this strategy. Since CD47 overexpression can block both NK cell and macrophage killing (J Exp Med 2021;218(3):e20200839), we hypothesized that T cells would lose their immunogenicity when human leukocyte antigen (HLA) class I and II genes are inactivated and CD47 is over-expressed.Human T cells from healthy donors were obtained by leukapheresis. To generate hypoimmune CD19CAR T cells, gene editing was used to delete b2m, CIITA, and TCR expression and lentiviral transduction was used to overexpress CD47 and CD19CAR containing a 4-1BB costimulatory domain to generate hypoimmune CAR T cells. Control T cells were unmanipulated except for lentiviral transduction used to overexpress the same CD19CAR and the deletion of the TCR. When transplanted into allogeneic humanized mice, hypoimmune CD19CAR T cells evade immune recognition by T cells even in previously sensitized animals as evidenced by a lack of T cell activation measured using ELISPOT analysis. In contrast, transplantation of non-hypoimmune-edited CD19CAR T cells generated from the same human donor resulted in a significant T cell activation (see figure: mean 59 and 558 spot frequencies for hypoimmune CD19CAR T cells and non-edited CD19CAR T cells, respectively; p<0.0001 unpaired T-test).In addition to evading T cells, immune cell assays show that CD47 overexpression protects hypoimmune CD19CAR T cells from NK cell and macrophage killing in vitro and in vivo. Relative CD47 expression levels were analyzed to understand the relevance of CD47 for protection from macrophage and NK cell killing. A blocking antibody against CD47 made the hypoimmune CAR T cells susceptible to macrophage and NK cell killing in vitro and in vivo, confirming the importance of CD47 overexpression to evade innate immune clearance.The hypoimmune CD19 CAR T cells retained their antitumor activity in both the Daudi and Nalm-6 B cell leukemia models, in vitro and in vivo. This indicated that the hypoimmune technology-i.e. isolated CD47 overexpression, deletion of b2m, CIITA, and TCR- did not show any effect on the cytotoxic potential of CD19 CAR T cells (see figure). These studies demonstrate that in vivo clearance of leukemic cells in NSG mice occurs across a range of tumor cell toCD19 CAR T cell ratios in a manner comparable to control, unedited CD19 CAR T cells (see figure). This result was validated using T cells from 3 different donorsThese findings show that, in these models, hypoimmune CD19 CAR T cells are functionally immune evasive in allogeneic humanized mouse recipients and have cytotoxic anti-tumor capacity. They suggest that hypoimmune CAR T cells could provide universal CAR T cells that are able to persist without immunosuppression. Furthermore, these data suggest that hypoimmune CD19 CAR T cells can be used in sensitized patients and for re-dosing strategies. [Display omitted] DisclosuresHu: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Dao: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. White: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Gattis: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Clarke: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Landry: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Basco: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Tham: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Tucker: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Luo: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Bandoro: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Chu: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Young: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Foster: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Dowdle: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Rebar: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Fry: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company. Schrepfer: Sana Biotechnology: Current Employment, Current equity holder in publicly-traded company.

Exploring Alternative Dosing Regimens of Single-Agent Belantamab Mafodotin on Safety and Efficacy in Patients with Relapsed or Refractory Multiple Myeloma: DREAMM-14

Introduction: Belantamab mafodotin is a first-in-class, monomethyl auristatin F (MMAF)-containing, B-cell maturation antigen (BCMA)-directed antibody-drug conjugate (ADC) that is approved in the United States and European Union for adult patients with relapsed/refractory multiple myeloma (RRMM). In the pivotal Phase II DREAMM-2 study, single-agent belantamab mafodotin (2.5 mg/kg administered intravenously every 3 weeks [Q3W]) demonstrated an objective response rate of 32% with a manageable safety profile in triple-class refractory adult patients with RRMM (Lonial et al. Lancet Oncol. 2020). At 13 months of follow-up, responses were durable, with a median duration of response of 11 months and an overall survival of 13.7 months (Lonial et al. ASH 2020, Poster 1417).Corneal events are common and expected with belantamab mafodotin and other MMAF-containing ADCs. In DREAMM-2, keratopathy (a pathological eye exam finding) presented as superficial punctate keratopathy and/or microcyst-like epithelial changes. Ocular symptoms, such as decline in best-corrected visual acuity or patient-reported adverse events (eg, blurred vision or dry eye), were also common during treatment. Corneal events with or without symptoms were managed with dose modifications (delays and reductions), and clinically meaningful responses were observed even with prolonged treatment-free intervals; this suggests that alternative dosing regimens of belantamab mafodotin may lower rates of corneal events without compromising efficacy.The goal of the DREAMM-14 study is to investigate whether an improved overall benefit/risk profile of single-agent belantamab mafodotin can be achieved by modifying the belantamab mafodotin dose, schedule, or both relative to the approved dosing regimen (2.5 mg/kg Q3W).Methods: This Phase II, 5-arm, randomized, parallel, open-label multicenter study will include patients with RRMM who have received at least 3 prior lines of therapy including an anti-CD38 monoclonal antibody (mAb), an immunomodulatory agent, and a proteasome inhibitor. Patients aged ≥18 years with Eastern Cooperative Oncology Group Performance Status ≤2 and who provide informed consent will be eligible. Patients with corneal epithelial disease (except mild punctate keratopathy) or prior exposure to BCMA-targeted therapies or ADCs will be excluded.Patients will be randomized into Arms A to D (n=40 each) and arm E (n=20) in parallel and stratified by the International Staging System (ISS) for MM (I vs II vs III) and prior lines of therapy (3 vs ≥4). Single-agent belantamab mafodotin will be administered at doses and schedules as follows-Arm A: 2.5 mg/kg Q3W (control); Arm B: 1.9 mg/kg Q3W; Arm C: 2.5 mg/kg Q6W; Arm D: 1.9 mg/kg Q6W; Arm E: 1.9 mg/kg Q6W with dose modifications based on oncology staff assessment of ocular symptoms (patient-reported symptoms using the Ocular Surface Disease Index) and visual acuity. Participants in all arms will have Q3W response assessments and Q3W ophthalmic examinations and monitoring by qualified eye care specialists. Ocular event-related dose modifications for all arms except Arm E will be guided by the Keratopathy and Visual Acuity (KVA) scale. Participants in all arms will be treated until progressive disease, unacceptable toxicity, or death.The primary endpoint will be incidence of Grade ≥2 ocular adverse events according to the KVA scale. Key secondary endpoints include ocular safety and tolerability, pharmacokinetics, and efficacy outcomes of belantamab mafodotin in all arms. Follow-up for progression-free survival will be Q3W until progressive disease, start of new anticancer therapy, withdrawal of consent, end of study, or death. Follow-up for overall survival will be Q12W from treatment discontinuation. The duration of this study will be approximately 22 months. The study is planned to start in the first quarter of 2022.Funding: GSK (Study 209628); drug linker technology licensed from Seagen; mAb produced using POTELLIGENT Technology licensed from BioWa. DisclosuresHultcrantz: GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees, Research Funding; Curio Science LLC: Consultancy; Intellisphere LLC: Consultancy; Amgen: Research Funding; Daiichi Sankyo: Research Funding. Kleinman: Calm Water Therapeutics LLC: Current Employment; GlaxoSmithKline: Consultancy; Eyeon Therapeutics Inc.: Current holder of individual stocks in a privately-held company; ONL Therapeutics: Consultancy, Current holder of individual stocks in a privately-held company; Triphase Accelerator: Consultancy; Helixmith Co., Ltd: Consultancy; Aprea Therapeutics: Consultancy; Editas Medicine, Inc.: Consultancy; Olema Pharmaceuticals, Inc.: Consultancy. Ghataorhe: GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company. McKeown: GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company; AstraZeneca: Current equity holder in publicly-traded company; Novartis: Current equity holder in publicly-traded company. He: GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company. Ling: GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company. Jewell: Alcon: Current equity holder in publicly-traded company; Novartis: Current equity holder in publicly-traded company; GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company. Brunner: GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company. Byrne: GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company; Adaptimmune: Current equity holder in publicly-traded company; Novartis: Current equity holder in publicly-traded company. Eliason: GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company. Scott: GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company. Opalinska: GlaxoSmithKline: Current Employment, Current equity holder in publicly-traded company.

Nx-5948, a Selective Degrader of BTK with Activity in Preclinical Models of Hematologic and Brain Malignancies

Bruton's tyrosine kinase (BTK) plays a key role in cell survival in B cell malignancies, and covalent inhibitors of BTK, such as ibrutinib and acalabrutinib, have proven efficacious in chronic lymphocytic leukemia (CLL), mantle cell lymphoma (MCL), marginal zone lymphoma (MZL), and Waldenstrom's macroglobulinemia (WM). BTK inhibitors have also demonstrated clinical activity in small trials of patients with relapsed/refractory primary central nervous system lymphoma. The long-term efficacy of BTK inhibitors is limited by the emergence of resistance mutations, most commonly at C481 of BTK. These mutations preclude formation of a covalent bond with BTK and lead to diminished efficacy and disease progression. Several noncovalent BTK inhibitors, which do not require covalent binding to C481, are currently being investigated in clinical trials as potential therapies for patients with relapsed and refractory disease. However, other mutations have been shown to decrease the in vitro activity of these non-covalent BTK inhibitors, suggesting that mutations may ultimately limit the effectiveness of these compounds as well.Small molecule-induced protein degradation offers a unique approach to target BTK for the treatment of B-cell malignancies. Chimeric Targeting Molecules (CTMs) catalyze ubiquitylation and proteasomal degradation of target proteins and are comprised of a target binding element (“hook”), a linker, and ubiquitin ligase binding element (“harness”). NX-5948 is a CTM that contains a BTK hook linked to a cereblon (CRBN) harness. While some CRBN-binding drugs, such as lenalidomide and pomalidomide, promote the degradation of neo-substrate proteins, such as the transcription factors Aiolos and Ikaros, NX-5948 has been engineered to avoid Aiolos and Ikaros degradation and therefore does not possess IMiD activity.Preclinical studies have shown that NX-5948 catalyzes the degradation of 50% of cellular BTK (DC 50) at < 1 nM concentrations in lymphoma cell lines and peripheral blood mononuclear cells (PBMCs). NX-5948 impairs viability in the BTK-dependent ABC-DLBCL cell line, TMD8 (EC 50: < 10 nM after 72 hours). Importantly, NX-5948 induces degradation of the mutated BTK-C481S in cells and inhibits proliferation of BTK-C481S mutant TMD8 cells more effectively than ibrutinib (NX-5948 EC 50 values of < 10 nM versus > 1 μM for ibrutinib). NX-5948 also catalyzes selective degradation of BTK. In an unbiased proteomics experiment, NX-5948 treatment of TMD8 cells demonstrates robust BTK degradation without significant downregulation of other off-target proteins.Oral administration of NX-5948 in mice leads to dose-proportional exposure in plasma and BTK degradation to <10% of baseline levels in circulating and splenic B cells. In a TMD8 xenograft model in mice containing the BTK-C481S mutation, daily oral administration of NX-5948 resulted in superior tumor growth inhibition (TGI) as compared to ibrutinib. Following a single oral dose of NX-5948 in cynomolgus monkeys, BTK degradation to <10% of baseline levels is observed in the circulating B cells.In addition, NX-5948 penetrates the central nervous system (CNS) and demonstrates activity in a model of brain malignancies. Following oral dosing of NX-5948 to mice, NX-5948 was detectable in the cerebrospinal fluid at a comparable level to the unbound concentration of NX-5948 in mouse plasma. In an intracranial TMD8 xenograft model, treatment with NX-5948 resulted in degradation of BTK in intracranial TMD8 cells, decreased intracranial tumor burden and improved survival relative to treatment with vehicle.The potent BTK degradation activity of NX-5948 and the ability of this molecule to penetrate the CNS supports its development for the treatment of B-cell malignancies, including CNS lymphoma. DisclosuresRobbins: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Noviski: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Rountree: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Tan: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Brathaban: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Ingallinera: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Karr: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Kelly: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Konst: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Ma: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Tenn-McClellan: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. McKinnell: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Perez: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Guiducci: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Hansen: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company. Sands: Nurix Therapeutics: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees.

Memory Phenotype in Allogeneic Anti-BCMA CAR-T Cell Therapy (P-BCMA-ALLO1) Correlates with In Vivo Tumor Control

The emergence of CAR-T cell therapy has transformed the treatment of the previously refractory/relapsed multiple myeloma (MM). Yet, autologous CAR-T cells suffer from inconsistent manufacturing, long manufacturing timelines, and high cost, which can limit patient accessibility. To address these issues, we engineered a fully allogeneic anti-BCMA CAR-T cell candidate for MM from healthy donors (P-BCMA-ALLO1). Herein, we demonstrate that P-BCMA-ALLO1 maintains a T stem cell memory phenotype (T SCM) through genetic editing, which correlates with antitumor efficacy.Using Poseida's proprietary non-viral piggyBac® (PB) DNA Delivery System, in combination with the high-fidelity Cas-CLOVER™ (CC) Site-Specific Gene Editing System and a proprietary “booster molecule”, we generated P-BCMA-ALLO1 from healthy donor T cells. We used CC to eliminate surface expression of TCR and MHC class I to make fully allogeneic CAR-T cells. In addition to the CAR molecule, PB enables the delivery of a selectable marker allowing the generation of a final cell product that is >95% CAR-positive. The inclusion of the “booster molecule” in the manufacturing process improves the expansion of gene-edited cells without compromising memory phenotype or function. This process can produce up to hundreds of patient doses from a single manufacturing run using one healthy donor, thereby significantly reducing manufacturing cost per dose. We characterized the phenotype and functionality of P-BCMA-ALLO1 using flow cytometry and Nanostring to assess their memory phenotype at both the protein and mRNA levels. Also analyzed was antitumor toxicity and proliferative capacity through multiple rounds of activation using in vitro co-culture assays and serial restimulation, respectively. The relationship of all characterizations with in vivo efficacy was then determined, as defined by control of tumor growth in an immunodeficient RPMI-8226 subcutaneous murine tumor model.We found that P-BCMA-ALLO1 is comprised of a high frequency of T SCM after editing (Fig. 1), and the maintenance of that memory phenotype correlates with antitumor efficacy. In vivo, these CAR-T cells are potent in controlling tumor growth, comparable to or better than autologous anti-BCMA CAR-T cells. Our analysis revealed that the expression of memory markers at the surface protein level (CD27, CD62L, CD127, CCR7) and mRNA level significantly correlate with in vivo tumor control. Conversely, suboptimal research products with worse in vivo outcomes express a more exhausted gene expression profile. We reveal from our analysis that the most effective P-BCMA-ALLO1 in vivo share similar characteristics: (1) these products were a result of efficient manufacturing, with >90% CAR+ and >99% TCR-; (2) they carry a memory phenotype, with 50-70% T scm and high proliferative capacity after multiple rounds of restimulation; (3) they are >90% viable; and (4) they show strong antitumor efficacy both in vitro and in vivo.We demonstrate that Tscm percentage in the final product correlates with antitumor activity. P-BCMA-ALLO1 is advancing rapidly towards the clinic (NCT04960579) to positively impact the outcomes of CAR-T therapy for MM patients.Figure 1: Memory composition of P-BCMA-ALLO1 research products. P-BCMA-ALLO1 consists mostly of stem cell memory (T scm) and central memory (T cm) T cells that are CD62L + as opposed to effector memory (T em) and effector (T eff) T cells. [Display omitted] DisclosuresTseng: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Zhang: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Cranert: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Richter: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Marquez: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Qiu: Poseida Therapeutics: Current equity holder in publicly-traded company, Ended employment in the past 24 months. Cho: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Tan: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Tong: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Domingo: Poseida Therapeutics: Current equity holder in publicly-traded company, Ended employment in the past 24 months. Weiss: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Argus: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Sparks: Poseida Therapeutics: Current equity holder in publicly-traded company, Ended employment in the past 24 months. Ostertag: Poseida: Current Employment, Current equity holder in publicly-traded company. Coronella: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company. Shedlock: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company.

Knock out of CD123 or CLL-1 By CRISPR-Cas9 Editing from Human Hematopoietic Stem Cell Transplants Provide New Possibilities for Increasing Therapeutic Index and Safety for AML Treatment

▪Acute myeloid leukemia (AML) is a clonal disorder of hematopoiesis and the most common form of acute leukemia in adults. Most patients with AML relapse despite intensive chemotherapy. Allogeneic hematopoietic cell transplant (HCT) has become the standard of care for patients with intermediate or adverse genetics, with >3,500 transplants performed annually in the US. However, leukemia relapse post-HCT occurs in ~40% of these patients with a 2-year survival rate at <20%, necessitating new approaches to reduce relapse and improve overall outcomes.Antigen-specific immunotherapies require cell surface antigens to be uniquely expressed on cancer cells to minimize “on-target, off-tumor” toxicity. CD123 and CLL-1 are highly expressed on normal myeloid cells, thus impeding the use of therapies targeting these antigens due to myelotoxicity. To circumvent such toxicity, we aim to create CD123 or CLL-1 negative human hematopoietic stem and progenitor cells (hHSPCs) for HCT such that this can be combined with subsequent use of targeted therapy against these antigens to prevent post-HCT relapse. Here, we present the pre-clinical evaluation of engineered hHSPCs, derived from mobilized peripheral blood of healthy donors, where CD123 or CLL-1 proteins were ablated by CRISPR/Cas9 gene editing. We have identified highly efficient guide RNAs that result in >80% on-target editing and can achieve greater than 90% biallelic gene knockout (KO). Deep sequencing followed by hybrid capture of up to 1000 potential off-target sites predicted to contain a maximum of 5 mismatches revealed minimal to no detectable off-target editing events. CD123 or CLL-1 KO hHSPCs showed >85% cell viability post-editing. Loss of CD123 or CLL-1 protein did not impact the differentiation of hHSPCs into granulocytic, monocytic, or erythroid lineages in vitro. Additionally, myeloid cells derived from CD123 or CLL-1 KO hHSPCs also retained their function, demonstrating similar phagocytotic capacity and the production of inflammatory cytokines in response to TLR agonists compared to unedited control cells. Importantly, CD123 or CLL-1 KO hHSPCs xenotransplanted into NSG or NBSGW mice showed no defect in long-term engraftment (human chimerism in NBSGW mice 16 weeks post transplant: 92.3±5.1% for CD123 KO, 91.2±4.1% for CLL-1 KO, and 93.4±3.1% unedited hHSPC engrafted mice, n=15 each). This in vivo model confirmed no observable impact on multilineage differentiation after CD123 or CLL-1 deletion either, supported by equal distribution of 10 hematopoietic lineages between groups. Bone marrow analyses revealed persistence of high gene editing frequency after 16 weeks (71±10% from CD123 KO and 86±6% from CLL-1 KO engrafted mice, n=15 each, compared to original 83% input CD123 KO and 86% input CLL-1 hHSPCs). Sorted subpopulations, including granulocytes, monocytes, dendritic cells, and mast cells, from the bone marrow similarly retained high levels of editing. Together these data suggest that there was no counter-selection against CD123 or CLL-1 KO cells, no outgrowth of any cells with particular edits or hematopoietic lineages to warrant tumorigenic concerns. Most notably, proof-of-concept experiments showed that cells depleted of CD123 or CLL-1 protein are indeed protected from the cytotoxicity of CD123 or CLL-1 targeted CAR-T cells.In conclusion, we demonstrate that CD123 or CLL-1 negative human HSPCs can successfully carry out functional hematopoiesis that is resistant to CD123 or CLL-1 targeted therapies. Our findings provide a next-generation HCT strategy that supports the safe and effective use of antigen-directed immunotherapy treatments for patients with AML. DisclosuresLuo: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Angelini: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Krishnamurthy: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Lisle: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Isik: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Ghdossi: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Cummins: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Pettiglio: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Hazelbaker: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Ge: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Tavares: Vor Biopharma: Ended employment in the past 24 months. Nikam: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Paik: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Lydeard: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Lin: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Chakraborty: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company.

A First-in-Class STAT3 Degrader KT-333 in Development for Treatment of Hematologic Cancers

Signal Transducer and Activator of Transcription 3 (STAT3) plays important roles in the transduction of signals from growth factors and cytokines in both normal and malignant cells. Upon activation, STAT3 controls expression of genes that regulate cell growth, survival, differentiation, stemness and cell-cell interactions. Aberrant activation of STAT3 has been observed in many cancers including lymphoma and leukemias through activating mutations, hyper-signaling through upstream regulators or loss of negative feedback regulation. Additionally, STAT3-mediated cross-talk in the tumor microenvironment results in suppression of immune surveillance compromising anti-tumor immunity.STAT3 has been historically considered “undruggable”. Heterobifunctional degraders that recruit endogenous E3 ligases to ubiquitinate substrate proteins leading to their degradation by the proteosome have emerged as a novel therapeutic modality with great potentials to drug undrugged protein targets. Here we introduce a first-in-class, potent and selective STAT3 degrader KT-333 that is being developed for the treatment of hematologic malignancies and solid tumors.KT-333 is a small molecule targeted protein degrader of STAT3 and was discovered by an iterative medicinal chemistry SAR campaign improving parameters such as potency, selectivity and ADME profile. It has high solubility in PBS buffer (103 mg/mL, pH7.4). low to moderate clearance in rat, dog and monkey, and moderate plasma half lives across pre-clinical species.KT-333 exhibits potent and selective degradation of STAT3 with DC 50 of 2.5 -11.8 nM in four anaplastic T cell lymphoma (ALCL) lines. Selective degradation of STAT3 over nearly 9000 proteins, including other STAT family members, was demonstrated by mass spectrometry in human PBMCs . Tumor cells expressing NPM-ALK fusion oncoproteins have activated STAT3 and dependency on STAT3 signaling for survival KT-333 treatment led to rapid apoptosis of these ALK+ALCL cells. To better understand the cellular responses to STAT3 degradation, time-course transcriptomic and proteomic analyses were performed using SU-DHL-1 cells treated with KT-333. Time-dependent changes in both mRNA and protein levels of STAT3-regulated genes and pathways were observed: at 8 h only STAT3 showed significant depletion, at 24 h canonical STAT3 downstream genes including SOCS3, IL-2RA and GRZMB showed modulation and at 48 h, pathways including interferon response and cell cycle were significantly enriched. A comparison of the transcriptomic and proteomic data sets showed high correlation. Furthermore, changes in mRNA and protein levels of bona fide STAT3 regulated targets showed a high degree of concordance between SU-DHL-1 cells and SU-DHL-1 xenograft tumors.Preclinical in vitro experiments have demonstrated that approximately 90% degradation of STAT3 for 48 h was sufficient to drive ALK+ ALCL lines into irreversible cell growth inhibition and death. In vivo xenograft studies of SU-DHL-1 and SUP-M2 ALCL models demonstrated dose-dependent tumor growth suppression. An indirect response model that linked exposure to target degradation was established to describe the relationship between exposure and target degradation in the tumor. Consistent with the in vitro data, PK-PD-efficacy analysis in the SU-DHL-1 xenograft model showed that tumor regression could be achieved with intermittent dosing schedules that achieved ~90% degradation of STAT3 for about 48 h over a weekly or bi-weekly dosing interval. . Based on preclinical ADME and PK/PD/efficacy relationship, KT-333 is predicted to drive tumor regression using weekly doses.In conclusion, we have identified a potent STAT3 degrader as development candidate that can be administered parenterally in the clinic on an intermittent dosing regimen that is predicted to be both efficacious and tolerated in STAT3-dependent hematologic malignancies. DisclosuresLiu: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Dixit: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Mayo: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Dey: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Yuan: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Karnik: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Walther: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Shi: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Sharma: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Rong: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Yang: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Gollerkeri: BEAM Therapeutics: Current Employment, Current equity holder in publicly-traded company; Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. Gollob: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company. DeSavi: Kymera Therapeutics: Current Employment, Current equity holder in publicly-traded company.

Multiplex Engineering of Human CD34+ HSPCs Enables Dual Gene Knock-out While Maintaining High Engraftment Potential and Safety

Genome engineering of allogeneic hematopoietic stem cell transplants (HSCT), in which leukemia-specific cell surface antigens are removed by editing from the donor graft, may provide high therapeutic index and safety in combination with post-transplant targeted therapeutics. This strategy allows the therapeutics to specifically target leukemic cells by sparing the gene-edited graft, and thereby enable the next generation of HSCTs for treatment of acute myeloid leukemia (AML). So far, no single target antigen is known to be expressed in 100% of AML blasts, leukemic stem cells, or even in affected patients. Therefore, use of combinatorial therapies targeting multiple antigens may represent a potential paradigm shift in the standard of care for AML. Multiplex editing of HSCT in which two or more target antigens are removed from the same cell would enable combinatorial targeting while simultaneously protecting the engineered graft from on-target, off-tumor toxicity. Here we report proof-of-concept data that multiplex editing with two independent CRISPR/Cas9 ribonucleoproteins (RNPs) targeting genetic loci on chromosomes 11 and 19 robustly maintains long-term in vivo engraftment potential with persistent gene edits at both target sites, with minimally detectable translocations in bone marrow samples 16 weeks after engraftment in NSG mice. The overall viability of dual-edited input CD34+ HSPCs post gene editing was >75% and the proportion of CD34+CD38-CD45RA-CD90+CD49f+ putative long-term hematopoietic stem cells (LT-HSCs) in input cells were similar (1-2%) between unedited and multiplex-edited cells. In vivo engraftment potential of multiplex-edited cells was maintained long-term, with average chimerism of hCD45+ cells in mice transplanted with dual knock-out (KO) HSPCs similar to those transplanted with unedited HSPCs (53.36±11.80% versus 47.45±18.03%, respectively). Dual KO cells also maintained normal differentiation potential as judged by multilineage reconstitution in vivo and in vitro colony forming unit (CFU) assay, indicating the dual RNP delivery process does not lead to any observable impact on the multipotency of the engineered graft. Quantification of on-target editing by multiplexed NGS amplicon sequencing revealed no reduction in total editing between dual KO input and bone marrow cells, indicating the gene modifications in dual engineered cells persist post-engraftment.‡equal contribution, Nipul Patel and Michael A. Pettiglio DisclosuresPettiglio: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Cummins: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Pimentel: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Kapuria: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Ghodssi: Vor Biopharma: Current Employment, Current holder of stock options in a privately-held company. Isik: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Falla: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Amunugama: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Lisle: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Wang: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Lin: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Lydeard: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Ge: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Hazelbaker: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Paik: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Chakraborty: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company.

Predictive Biomarkers of Response to the Polo-like Kinase 1 (PLK1) Inhibitor, Onvansertib, in Combination with Decitabine in Relapsed or Refractory Acute Myeloid Leukemia (R/R AML)

Background: PLK1 is a serine/threonine kinase master regulator of the cell cycle, overexpressed in AML blasts. Onvansertib (ONV) is an oral, highly selective next-generation PLK1 inhibitor that demonstrated anti-tumor activity in AML preclinical models. The safety and efficacy of ONV in combination with decitabine (DAC) in patients with R/R AML is under study in an ongoing phase 1b/2 clinical trial (NCT03303339). Here we report results of correlative studies to identify molecular predictors of response to the ONV-DAC combination.Methods: R/R AML patients were treated for 5 days with ONV (orally) and DAC (20 mg/m 2 IV qd x 5d) in 21- or 28-day cycles. Patients who completed at least 1 cycle of treatment were considered evaluable for efficacy. At baseline, bone marrow (BM) samples were collected for targeted next-generation sequencing (Myeloid VariantPlex, Archer) and blood samples for RNA-sequencing.Results: As of July 7, 2021, 23 patients were treated with ONV (12 - 90 mg/m 2) + DAC in Phase 1b and 32 patients with ONV 60 mg/m 2 + DAC in Phase 2. Nine (20%) of the 46 patients that were evaluable for efficacy achieved a complete remission with or without complete hematologic recovery (CR/CRi) as their best response (7 CR and 2 CRi). Another 6 patients had ≥50% BM blast decrease from baseline at one point on trial therapy. Those 6 patients plus the 9 patients who achieved CR/CRi are referred to as BM responders (n=15). RNA-seq was performed on blood collected from 32 patients, including 12 BM responders. Differential gene expression, gene-set, and machine learning analyses of BM responders versus non-responders identified a gene expression signature predictive of response to the ONV-DAC combination. No association between the ONV-DAC signature and DAC response was observed after applying the ONV-DAC signature to gene expression profiles from patients treated with DAC as a single agent in a multicenter Phase 2 study (NCT00866073), indicating that the gene signature is specific to the combination. The ONV-DAC signature was also applied to the BeatAML cohort (n=399) and identified 241 predicted responders. Mutation analysis of the BeatAML cohort revealed a positive association between predicted ONV-DAC response and mutations in splicing factors (SF) such as SRSF2, suggesting that SF mutations may serve as a response biomarker for the ONV-DAC combination. Indeed, genomic analysis of baseline BM samples from patients evaluable for efficacy in our Phase 1b/2 (n=46), showed a significant increase in clinical responses in patients with SRSF2 and SF3B1 mutations versus patients with other alterations (Figure 1 and Table 1). In addition, biomarkers reported to be positively correlated with DAC responses (TP53 and TET2 mutations, high fetal hemoglobin expression) were not associated with ONV-DAC clinical responses (Figure 1).Conclusion: We identified a gene expression signature associated with clinical response to the ONV-DAC combination, that did not predict response to DAC single agent. The ONV-DAC predictive signature was found to be correlated with SF mutations in the BeatAML cohort. In our cohort, we confirmed the association between clinical response to ONV-DAC and SF mutations. These analyses support further investigation of the ONV-DAC combination in R/R AML patients with SF mutations and the potential use of ONV-DAC for other hematological diseases with high prevalence of SF mutations such as myelodysplastic syndrome and chronic myelomonocytic leukemia. [Display omitted] DisclosuresZeidan: Janssen: Consultancy; Astellas: Consultancy; Astex: Research Funding; Genentech: Consultancy; Daiichi Sankyo: Consultancy; Aprea: Consultancy, Research Funding; Boehringer Ingelheim: Consultancy, Research Funding; Gilead: Consultancy, Other: Clinical Trial Committees; Epizyme: Consultancy; BeyondSpring: Consultancy; Novartis: Consultancy, Other: Clinical Trial Committees, Travel support, Research Funding; Pfizer: Other: Travel support, Research Funding; BioCryst: Other: Clinical Trial Committees; BMS: Consultancy, Other: Clinical Trial Committees, Research Funding; Cardiff Oncology: Consultancy, Other: Travel support, Research Funding; Geron: Other: Clinical Trial Committees; Kura: Consultancy, Other: Clinical Trial Committees; Ionis: Consultancy; Incyte: Consultancy, Research Funding; AstraZeneca: Consultancy; Jasper: Consultancy; Loxo Oncology: Consultancy, Other: Clinical Trial Committees; Jazz: Consultancy; Amgen: Consultancy, Research Funding; Agios: Consultancy; ADC Therapeutics: Research Funding; Acceleron: Consultancy, Research Funding; AbbVie: Consultancy, Other: Clinical Trial Committees, Research Funding. Ridinger: Cardiff Oncology: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Croucher: Cardiff Oncology: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Samuëlsz: Cardiff Oncology: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Erlander: Cardiff Oncology: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Ruffner: Cardiff Oncology: Current Employment, Current equity holder in publicly-traded company; ALX Oncology: Current equity holder in publicly-traded company; Pfizer: Current equity holder in publicly-traded company; Gilead: Current equity holder in publicly-traded company; BMS: Current equity holder in publicly-traded company; Anthem INC: Current equity holder in publicly-traded company; Cardinal Health: Current equity holder in publicly-traded company; Change Healthcare: Current equity holder in publicly-traded company; CVS Health Corp: Current equity holder in publicly-traded company; Merck: Current equity holder in publicly-traded company; Organon: Current equity holder in publicly-traded company. Wang: GlaxoSmithKline: Consultancy, Honoraria, Other: Advisory Board; Jazz Pharmaceuticals: Consultancy, Honoraria, Other: Advisory Board; Mana Therapeutics: Consultancy, Honoraria; Kite Pharmaceuticals: Consultancy, Honoraria, Other: Advisory Board; Pfizer: Consultancy, Honoraria, Other: Advisory Board, Speakers Bureau; Stemline Therapeutics: Consultancy, Honoraria, Other: Advisory board, Speakers Bureau; Takeda: Consultancy, Honoraria, Other: Advisory board; DAVA Oncology: Consultancy, Speakers Bureau; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees; BMS/Celgene: Membership on an entity's Board of Directors or advisory committees; Genentech: Membership on an entity's Board of Directors or advisory committees; Kura Oncology: Consultancy, Honoraria, Other: Advisory board, steering committee, Speakers Bureau; Novartis: Consultancy, Honoraria, Other: Advisory Board; Rafael Pharmaceuticals: Other: Data safety monitoring committee; Gilead: Consultancy, Honoraria, Other: Advisory board; Daiichi Sankyo: Consultancy, Honoraria, Other: Advisory board; PTC Therapeutics: Consultancy, Honoraria, Other: Advisory board; Genentech: Consultancy; MacroGenics: Consultancy.

G-CSF/Plerixafor Dual-Mobilized Donor Derived CD33CAR T-Cells As Potent and Effective AML Therapy in Pre-Clinical Models

There are currently no known acute myeloid leukemia (AML) specific antigens. Genetic ablation of CD33 using CRISPR/Cas9 engineering of the hematopoietic stem cell (HSC) transplant (VOR33) represents a synthetic biology approach to generating a leukemia-specific antigen in the transplant recipient. VOR33 enables anti-CD33 CAR-T cell killing of AML cells while sparing normal myeloid lineage development and function, thereby potentially avoiding myelosuppression and increasing the therapeutic index of anti-CD33 CAR-T therapy. Mobilized leukapheresis product represents an attractive starting material for the generation of both a CD33 null HSC transplant and a complementary CD33CAR T-cell product. In this study, we sought to determine the impact of dual mobilization with Granulocyte-Colony Stimulating Factor (G-CSF) and plerixafor (mozobil) on immune cell composition, T cell phenotype, and the functionality of these T cells to control AML tumor growth upon chimeric antigen receptor (CAR) transduction.Mobilized (mob) peripheral blood mononuclear cells (PBMCs) were collected from healthy donors injected with G-CSF (10µg/kg/day, 5 consecutive days) and plerixafor (240µg/kg, on day 4 and 5). Non-mobilized (non-mob) PBMCs, collected from the same donors, were used as controls. Cells were analyzed by flow cytometry for immunophenotyping and T cell characterization including differentiation and bone marrow homing markers, as well as responses to T cell activation with anti-CD3 (OKT3) and IL-2. Non-/mob PBMC populations were also analyzed by single-cell next generation sequencing (CITEseq) using 127 immune cell phenotypic markers in combination with extensive transcriptome and T cell receptor repertoire analysis. In addition, lentiviral transduction of anti-CD33 CAR constructs enabled functional comparisons of mob- and non-mob-CAR T-cells in AML cell co-cultures as well as AML mouse models.Ex vivo immunophenotyping of PBMC from a total of over 30 healthy donor samples showed that mobilization decreases the overall percentage of CD3 + T cells but increases that of naïve T cells (CD45RA +/CCR7 +), at the expense of T effector-memory (CD45RA -/CCR7 -) and central-memory (CD45RA -/CCR7 +) populations. Bone marrow homing factors (e.g.: CXCR4) were increased in mob compared to non-mob T cell samples. As expected, higher percentages of monocytes (CD14 +) were detected in mob compared to non-mob donor samples, but this difference disappeared after culture under T cell activation conditions. T cell activation also led to similar increases in CD25, CD69 and CD137 expression, and a decrease in CD62L expression. Single cell sequencing analyses confirmed mobilization-induced increases in naïve T cells as well as changes in monocytes/macrophages, CD4 + T cells and NK cells percentages. Notably, functional in vitro cytotoxic assays demonstrated that mob-CD33-CAR T-cells are as effective as non-mob-CD33-CAR T-cells in killing CD33 + AML cells, with reduced ‘bystander’ activation of non-transduced T cells. Furthermore, results from in vivo AML mouse models indicate that mob-CD33-CAR T-cells are equally effective in clearing CD33 + tumors as non-mob-CD33-CAR T-cells.Our analysis showed phenotypical ex vivo differences between mob and non-mob PBMCs, which disappeared upon activation, indicating similar responses to T cell-specific stimulation. These findings are corroborated by similar in vitro cytotoxicity profiles of non-/mob-CAR T-cells. Non-transduced T cells in the mob-CAR T-cell population showed limited ‘bystander’ activation, indicating a potentially favorable clinical toxicity profile. Additional in vivo assessment of mob-CAR T-cell function shows effective tumor clearance, which supports further efforts towards their clinical use in combination with engineered HSCs for the treatment of AML patients. DisclosuresCanesin: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Hoyt: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Williams: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Silva: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Chng: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Cummins: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Ung: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Qiu: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Shin: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Hu: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Ge: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Scherer: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Chakraborty: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company. Kassim: Vor Biopharma: Current Employment, Current equity holder in publicly-traded company.