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We hypothesized that combining adoptively transferred autologous T cells with a cancer vaccine strategy would enhance therapeutic efficacy by adding antimyeloma idiotype (Id)–keyhole limpet hemocyanin (KLH) vaccine to vaccine-specific costimulated T cells. In this randomized phase 2 trial, patients received either control (KLH only) or Id-KLH vaccine, autologous transplantation, vaccine-specific costimulated T cells expanded ex vivo, and 2 booster doses of assigned vaccine. In 36 patients (KLH, n = 20; Id-KLH, n = 16), no dose-limiting toxicity was seen. At last evaluation, 6 (30%) and 8 patients (50%) had achieved complete remission in KLH-only and Id-KLH arms, respectively (P = .22), and no difference in 3-year progression-free survival was observed (59% and 56%, respectively; P = .32). In a 594 Nanostring nCounter gene panel analyzed for immune reconstitution (IR), compared with patients receiving KLH only, there was a greater change in IR genes in T cells in those receiving Id-KLH relative to baseline. Specifically, upregulation of genes associated with activation, effector function induction, and memory CD8+ T-cell generation after Id-KLH but not after KLH control vaccination was observed. Similarly, in responding patients across both arms, upregulation of genes associated with T-cell activation was seen. At baseline, all patients had greater expression of CD8+ T-cell exhaustion markers. These changes were associated with functional Id-specific immune responses in a subset of patients receiving Id-KLH. In conclusion, in this combination immunotherapy approach, we observed significantly more robust IR in CD4+ and CD8+ T cells in the Id-KLH arm, supporting further investigation of vaccine and adoptive immunotherapy strategies. This trial was registered at www.clinicaltrials.gov as #NCT01426828.

Mutations in chromatin regulator ASXL1 are frequently identified in myeloid malignancies, in particular ∼40% of patients with chronic myelomonocytic leukemia (CMML). ASXL1 mutations are associated with poor prognosis in CMML and significantly co-occur with NRAS mutations. Here, we show that concurrent ASXL1 and NRAS mutations defined a population of CMML patients who had shorter leukemia-free survival than those with ASXL1 mutation only. Corroborating this human data, Asxl1−/− accelerated CMML progression and promoted CMML transformation to acute myeloid leukemia (AML) in NrasG12D/+ mice. NrasG12D/+;Asxl1−/− (NA) leukemia cells displayed hyperactivation of MEK/ERK signaling, increased global levels of H3K27ac, upregulation of Flt3. Moreover, we find that NA-AML cells overexpressed all the major inhibitory immune checkpoint ligands: programmed death-ligand 1 (PD-L1)/PD-L2, CD155, and CD80/CD86. Among them, overexpression of PD-L1 and CD86 correlated with upregulation of AP-1 transcription factors (TFs) in NA-AML cells. An AP-1 inhibitor or short hairpin RNAs against AP-1 TF Jun decreased PD-L1 and CD86 expression in NA-AML cells. Once NA-AML cells were transplanted into syngeneic recipients, NA-derived T cells were not detectable. Host-derived wild-type T cells overexpressed programmed cell death protein 1 (PD-1) and T-cell immunoreceptor with immunoglobulin and ITIM domains (TIGIT) receptors, leading to a predominant exhausted T-cell phenotype. Combined inhibition of MEK and BET resulted in downregulation of Flt3 and AP-1 expression, partial restoration of the immune microenvironment, enhancement of CD8 T-cell cytotoxicity, and prolonged survival in NA-AML mice. Our study suggests that combined targeted therapy and immunotherapy may be beneficial for treating secondary AML with concurrent ASXL1 and NRAS mutations.

Pro-inflammatory signaling is a hallmark feature of human cancer, including in myeloproliferative neoplasms (MPNs), most notably myelofibrosis (MF). Dysregulated inflammatory signaling contributes to fibrotic progression in MF; however, the individual cytokine mediators elicited by malignant MPN cells to promote collagen-producing fibrosis and disease evolution remain yet to be fully elucidated. Previously we identified a critical role for combined constitutive JAK/STAT and aberrant NF-kB pro-inflammatory signaling in myelofibrosis development. Using single-cell transcriptional and cytokine-secretion studies of primary MF patient cells and the hMPLW515L murine model of myelofibrosis, we extend this previous work and delineate the role of CXCL8/CXCR2 signaling in MF pathogenesis and bone marrow fibrosis progression. MF patient hematopoietic stem/progenitor cells are enriched for a CXCL8/CXCR2 gene signature and display enhanced proliferation and fitness in response to exogenous CXCL8 ligand in vitro. Genetic deletion of Cxcr2 in the hMPLW515L adoptive transfer model abrogates fibrosis and extends overall survival, and pharmacologic inhibition of the CXCR1/2 pathway improves hematologic parameters, attenuates bone marrow fibrosis, and synergizes with JAK inhibitor therapy. Our mechanistic insights provide a rationale for therapeutic targeting of the CXCL8/CXCR2 pathway in MF patients.

Leukemia cells accumulate DNA damage but altered DNA repair mechanisms protect them from apoptosis. We showed here that formaldehyde generated by serine/one-carbon cycle metabolism contributed to accumulation of toxic DNA-protein crosslinks (DPCs) in leukemia cells, especially in driver clones harboring oncogenic tyrosine kinases [OTKs: FLT3(ITD), JAK2(V617F), BCR/ABL1]. To counteract this effect, OTKs enhanced the expression of DNA polymerase theta (POLq) by ERK1/2 serine/threonine kinase-dependent inhibition of c-CBL E3 ligase-mediated ubiquitination of POLq and its proteasomal degradation. Overexpression of POLq in OTK-positive cells resulted in efficient repair of DPC-containing DNA double-strand breaks (DSBs) by POLq-mediated end-joining (TMEJ). Transforming activity of OTKs and other leukemia-inducing oncogenes, especially of those causing inhibition of BRCA1/2 -mediated homologous recombination (HR) with and without concomitant inhibition of DNA-PK -dependent non-homologous end-joining (D-NHEJ), was abrogated in Polq-/- murine bone marrow cells. Genetic and pharmacological targeting of POLq polymerase and helicase activities revealed that both activities are promising targets in leukemia cells. Moreover, OTK inhibitor or DPC-inducing drug etoposide enhanced anti-leukemia effect of POLq inhibitor (POLqi) in vitro and in vivo. In conclusion, we demonstrated that POLq plays an essential role in protecting leukemia cells from metabolically induced toxic DNA lesions triggered by formaldehyde and that it can be targeted to achieve therapeutic effect.

The successful commercialization of cell therapies requires thorough planning and consideration of product quality, cost and scale of the manufacturing process. The implementation of automation can be central to a robust and reproducible manufacturing process at industrialized scales. There have been a number of wash-and-concentrate devices developed for cell manufacturing. These technologies have arisen from transfusion medicine, hematopoietic stem cell and biologics manufacturing where operating mechanisms are distinct from manual centrifugation. This review describes the historical origin and fundamental technologies underlying each currently available wash-and-concentrate device as well as their relative advantages and disadvantages in cell therapy applications. Understanding the specific attributes and limitations of these technologies is essential to optimizing cell therapy manufacturing.

Genomic classification has improved risk assignment of pediatric, but not adult B-lineage acute lymphoblastic leukemia (B-ALL). The international UKALLXII/ECOG-ACRIN E2993 (#NCT00002514) trial accrued 1229 adolescent/adult patients with BCR-ABL1− B-ALL (aged 14 to 65 years). Although 93% of patients achieved remission, 41% relapsed at a median of 13 months (range, 28 days to 12 years). Five-year overall survival (OS) was 42% (95% confidence interval, 39, 44). Transcriptome sequencing, gene expression profiling, cytogenetics, and fusion polymerase chain reaction enabled genomic subtyping of 282 patient samples, of which 264 were eligible for trial, accounting for 64.5% of E2993 patients. Among patients with outcome data, 29.5% with favorable outcomes (5-year OS 65% to 80%) were deemed standard risk (DUX4-rearranged [9.2%], ETV6-RUNX1/-like [2.3%], TCF3-PBX1 [6.9%], PAX5 P80R [4.1%], high-hyperdiploid [6.9%]); 50.2% had high-risk genotypes with 5-year OS of 0% to 27% (Ph-like [21.2%], KMT2A-AFF1 [12%], low-hypodiploid/near-haploid [14.3%], BCL2/MYC-rearranged [2.8%]); 20.3% had intermediate-risk genotypes with 5-year OS of 33% to 45% (PAX5alt [12.4%], ZNF384/-like [5.1%], MEF2D-rearranged [2.8%]). IKZF1 alterations occurred in 86% of Ph-like, and TP53 mutations in patients who were low-hypodiploid (54%) and BCL2/MYC-rearranged (33%) but were not independently associated with outcome. Of patients considered high risk based on presenting age and white blood cell count, 40% harbored subtype-defining genetic alterations associated with standard- or intermediate-risk outcomes. We identified distinct immunophenotypic features for DUX4-rearranged, PAX5 P80R, ZNF384-R/-like, and Ph-like genotypes. These data in a large adult B-ALL cohort treated with a non–risk-adapted approach on a single trial show the prognostic importance of genomic analyses, which may translate into future therapeutic benefits.

Plasmacytoid dendritic cells (pDCs) are the principal natural type I interferon–producing dendritic cells. Neoplastic expansion of pDCs and pDC precursors leads to blastic plasmacytoid dendritic cell neoplasm (BPDCN), and clonal expansion of mature pDCs has been described in chronic myelomonocytic leukemia. The role of pDC expansion in acute myeloid leukemia (AML) is poorly studied. Here, we characterize patients with AML with pDC expansion (pDC-AML), which we observe in ∼5% of AML cases. pDC-AMLs often possess cross-lineage antigen expression and have adverse risk stratification with poor outcome. RUNX1 mutations are the most common somatic alterations in pDC-AML (>70%) and are much more common than in AML without pDC expansion and BPDCN. We demonstrate that pDCs are clonally related to, as well as originate from, leukemic blasts in pDC-AML. We further demonstrate that leukemic blasts from RUNX1-mutated AML upregulate a pDC transcriptional program, poising the cells toward pDC differentiation and expansion. Finally, tagraxofusp, a targeted therapy directed to CD123, reduces leukemic burden and eliminates pDCs in a patient-derived xenograft model. In conclusion, pDC-AML is characterized by a high frequency of RUNX1 mutations and increased expression of a pDC transcriptional program. CD123 targeting represents a potential treatment approach for pDC-AML.

MazF is an Escherichia coli-derived endoribonuclease that selectively cleaves ACA sequences of mRNA prevalent in HIV. We administered a single infusion of autologous CD4 T lymphocytes modified to express a Tat-dependent MazF transgene to 10 HIV-infected individuals (six remaining on antiretroviral therapy [ART]; four undergoing treatment interruption post-infusion) in order to provide a population of HIV-resistant immune cells. In participants who remained on ART, increases in CD4 and CD8 T cell counts of ~200 cells/mm(3) each occurred within 2 weeks of infusion and persisted for at least 6 months. Modified cells were detectable for several months in the blood and trafficked to gastrointestinal lymph tissue. HIV-1 Tat introduced ex vivo to the modified CD4(+) T cells induced MazF expression in both pre- and post-infusion samples, and MazF expression was detected in vivo post-viral-rebound during ATI. One participant experienced mild cytokine release syndrome. In sum, this study of a single infusion of MazF-modified CD4 T lymphocytes demonstrated safety of these cells, distribution to lymph tissue and maintenance of Tat-inducible MazF endoribonuclease activity, as well as sustained elevation of blood CD4 and CD8 T cell counts. Future studies to assess effects on viremia and latent proviral reservoir are warranted.

During the germinal center (GC) reaction, B cells undergo extensive redistribution of cohesin complex and three-dimensional reorganization of their genomes. Yet, the significance of cohesin and architectural programming in the humoral immune response is unknown. Herein we report that homozygous deletion of Smc3, encoding the cohesin ATPase subunit, abrogated GC formation, while, in marked contrast, Smc3 haploinsufficiency resulted in GC hyperplasia, skewing of GC polarity and impaired plasma cell (PC) differentiation. Genome-wide chromosomal conformation and transcriptional profiling revealed defects in GC B cell terminal differentiation programs controlled by the lymphoma epigenetic tumor suppressors Tet2 and Kmt2d and failure of Smc3-haploinsufficient GC B cells to switch from B cell- to PC-defining transcription factors. Smc3 haploinsufficiency preferentially impaired the connectivity of enhancer elements controlling various lymphoma tumor suppressor genes, and, accordingly, Smc3 haploinsufficiency accelerated lymphomagenesis in mice with constitutive Bcl6 expression. Collectively, our data indicate a dose-dependent function for cohesin in humoral immunity to facilitate the B cell to PC phenotypic switch while restricting malignant transformation. Melnick and colleagues show that the cohesin complex exhibits dose-dependent regulation of chromatin remodeling that accompanies the transition of germinal center B cells to plasma cells, which is necessary to prevent lymphomagenesis.

Background: ENA is an oral, selective inhibitor of IDH2 approved for the treatment (Tx) of patients (pts) with relapsed/refractory IDH2 mutated (IDH2m) AML. Here we report the results of a Phase 2 expansion and Phase 1b of the Beat AML Master Trial Phase 2/1b sub-study to assess the efficacy of Tx of newly diagnosed (ND) IDH2m AML pts ≥ 60 years of age with ENA monotherapy (ENAm) and subsequent response-driven addition of AZA Tx. (ClinicalTrials.gov NCT03013998). Methods: The study initiated with a 3-outcome, 2-stage Phase 2 design, which enrolled patients on ENAm for up to 5 cycles. Pts without CR/CRi after 5 cycles of ENAm, or progression/intolerance prior to this time, were transferred to Phase 1b to receive ENA + AZA (Figure 1). Key eligibility included ND IDH2m AML pts with age ≥ 60 years and ECOG performance status 0-2. Pts received ENAm 100 mg/day in continuous 28-day cycles and ENA + AZA (75 mg/m2 days 1-7 every 28 days) for Phase 1b. Response was assessed using 2017 ELN AML criteria. The primary endpoint was CR/CRi rate. The 2-stage design required 24 pts and tested the null hypothesis (H0) that CR/CRi rate equaled 20% vs 50% and then expanded to test a revised H0 of 30% vs 50% in 60 pts (conditional alpha=0.025, power=77%). Expansion also allowed further assessment of safety of this treatment regimen. Results: At data cut off (06/18/2020), 60 pts enrolled, received ENAm, and were evaluable for the primary endpoint. Median age was 75 years and 52% were female (Table 1). Median time on ENAm was 4.7 months (mos). At data cut off, 12 pts were still on ENAm Tx. Most common reasons for discontinuing ENAm were Tx failure (defined as no response to treatment) (23 or 38%), disease progression (loss of response to treatment) (7 or 12%) and adverse events (AEs; 6 or 10%). Five pts (8%) went to transplant. CR/CRi was achieved in 28 pts (47%; adjusted 95% CI 28-59, unadjusted exact 95% CI 34-60) (Table 2). Responses were higher (p=0.04) among the 44 pts with IDH2 R140 (55%) as compared to the 16 with IDH2 R172 mutation (25%) further supporting distinct biology between these subsets. After a median follow up of 14.6 mos, the median overall survival (mOS) was 24.4 mos (95% CI 10.6-not reached). The median duration of response was not reached with 12 mos estimation of 57% (95% CI 34-75). Overall, 20 ENA-related serious adverse events (SAEs) occurred in 15 pts, the most common was differentiation syndrome (12 or 20%) and 1 had ENA-related SAE of tumor lysis syndrome (1.7%). One pt had ENA-related Grade 5 AE (renal failure/death). Most common AEs of any grade (in ≥20%) were nausea, anemia, and low potassium (Table 3). The 7-day/30-day/60-day deaths observed with ENAm were 2%/5%/11%, respectively. Phase 1b: Seventeen pts had inadequate response to ENAm and transferred to Phase 1b to receive ENA + AZA. Median time on Tx (including ENAm) was 6.2 mos and median time on Tx after pts started ENA + AZA was 2.1 mos (Table 2). Most common reasons for discontinuing ENA + AZA included Tx failure (5 or 29%), disease progression (2 or 12%), transplant, death and AEs (each 2 or 12%). CR/CRi was 41% (exact 95% CI 18-67). After a median follow up of 12.7 mos, the mOS from start of ENA + AZA combination Tx was 8.9 mos. Four ENA-related SAEs occurred in 3 pts on ENA + AZA Tx and the most common was differentiation syndrome (2 or 12.5%). One dose-limiting toxicity (Grade 3 nausea) related to both Txs was observed. Most common AEs (≥20%) of any grade were anemia, low albumin and vomiting (Table 3). One death occurred at day 13 of ENA + AZA. Conclusions: In newly diagnosed pts ≥60 years old with IDH2m AML, ENA had a low early death rate, high CR/CRi rate (47%, adjusted 95% CI 28-59), and yielded durable remissions. The most common unique toxicity with ENA was differentiation syndrome that occurred in 20% of patients. In pts who did not achieve CR/CRi with ENAm, a subset of patients achieved CR/CRi with addition of AZA. This combined approach of serial therapy with ENA monotherapy followed by AZA addition in pts with sub-optimal response resulted in a mOS exceeding 2 years for pts enrolled on study. Further focus on improving response among patients with IDH2 R172 mutations, identifying subsets of pts not responding to ENA monotherapy, and integrating new targeted agents into this treatment regimen are warranted. Figure 1 Disclosures Stein: Syndax: Consultancy, Research Funding; Celgene Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Daiichi-Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astellas Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Seattle Genetics: Consultancy; Agios Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bayer: Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy; Biotheryx: Consultancy; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; PTC Therapeutics: Membership on an entity's Board of Directors or advisory committees; Syros: Membership on an entity's Board of Directors or advisory committees; Abbvie: Consultancy. Borate:Genentech: Membership on an entity's Board of Directors or advisory committees; Daiichi Sankyo: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Jazz Pharmaceuticals: Research Funding; AbbVie: Other: Investigator in AbbVie-funded clinical trials; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding. Baer:Takeda: Other: Institutional research funding; AbbVie: Other: Institutional research funding; Astellas: Other: Institutional research funding; Forma: Other: Institutional research funding; Kite: Other: Institutional research funding; Oscotec: Other: Institutional research funding; Incyte: Other: Institutional research funding. Kovacsovics:Agios: Honoraria; Astella: Honoraria; Pfizer: Research Funding; Novartis: Research Funding; AbbVie: Research Funding; Jazz: Honoraria. Schiller:Astellas Pharma: Honoraria, Research Funding; Celator: Research Funding; Constellation: Research Funding; Abbvie: Research Funding; Actinium: Research Funding; Ariad: Research Funding; Stemline: Speakers Bureau; Cyclacel: Research Funding; Daiichi Sankyo: Research Funding; Deciphera: Research Funding; DeltaFly: Research Funding; Bristol-Myers Squibb: Current equity holder in publicly-traded company, Research Funding; Forma: Research Funding; FujiFilm: Research Funding; Gamida: Research Funding; Genentech-Roche: Research Funding; Geron: Research Funding; Jazz Pharmaceuticals: Research Funding; Karyopharm: Research Funding; Kite Pharma: Research Funding; Mateon: Research Funding; MedImmune: Research Funding; Onconova: Research Funding; Pfizer: Current equity holder in publicly-traded company, Research Funding; Regimmune: Research Funding; Samus: Research Funding; Sangamo: Research Funding; Tolero: Research Funding; Trovagene: Research Funding; Kaiser Permanente: Consultancy; Johnson & Johnson: Current equity holder in publicly-traded company; Agios: Consultancy, Research Funding, Speakers Bureau; Amgen: Consultancy, Current equity holder in publicly-traded company, Research Funding, Speakers Bureau; AstraZeneca: Consultancy; Incyte: Consultancy, Research Funding, Speakers Bureau; Novartis: Consultancy, Research Funding; Ono Pharma: Consultancy; Celgene: Research Funding, Speakers Bureau; Sanofi: Speakers Bureau; Gilead: Speakers Bureau. Olin:Astellas: Other: Site PI; Genentech: Other: Site PI; Pfizer: Other: Site PI; Daiichi Sankyo: Other: Site PI; Genentech: Consultancy; Amgen: Consultancy. Foran:Trillium: Research Funding; Xencor: Research Funding; H3Biosciences: Research Funding; Agios: Honoraria, Research Funding; Pfizer: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees; Abbvie: Research Funding; Boehringer Ingelheim: Research Funding; Actinium: Research Funding; Aprea: Research Funding; Aptose: Research Funding; Kura Oncology: Research Funding; Takeda: Research Funding; Revolution Medicine: Consultancy; Novartis: Membership on an entity's Board of Directors or advisory committees; Servier: Membership on an entity's Board of Directors or advisory committees. Lin:Trovagene: Research Funding; Tolero Pharmaceuticals: Research Funding; Seattle Genetics: Research Funding; Prescient Therapeutics: Research Funding; Abbvie: Research Funding; Bio-Path Holdings: Research Funding; Astellas Pharma: Research Funding; Aptevo: Research Funding; Incyte: Research Funding; Pfizer: Research Funding; Mateon Therapeutics: Research Funding; Ono Pharmaceutical: Research Funding; Jazz: Research Funding; Gilead Sciences: Research Funding; Genetech-Roche: Research Funding; Celyad: Research Funding; Celgene: Research Funding. Patel:DAVA Pharmaceuticals: Honoraria; Celgene: Consultancy, Speakers Bureau; Agios: Consultancy; France Foundation: Honoraria. Foster:Daiichi Sankyo: Consultancy; Bellicum Pharmaceuticals: Research Funding; Macrogenics: Consultancy, Research Funding. Druker:Leukemia & Lymphoma Society: Research Funding; Henry Stewart Talks: Patents & Royalties; Iterion Therapeutics (formerly Beta Cat Pharmaceuticals): Membership on an entity's Board of Directors or advisory committees; Vivid Biosciences: Membership on an entity's Board of Directors or advisory committees; GRAIL: Consultancy, Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Patient True Talks: Consultancy; The RUNX1 Research Program: Membership on an entity's Board of Directors or advisory committees; Third Coast Therapeutics: Membership on an entity's Board of Directors or advisory committees; VB Therapeutics: Membership on an entity's Board of Directors or advisory committees; Gilead Sciences: Consultancy, Membership on an entity's Board of Directors or advisory committees; Millipore (formerly Upstate Biotechnology): Patents & Royalties; MolecularMD (acquired by ICON): Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Novartis Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding; McGraw Hill: Patents & Royalties; Merck & Co: Patents & Royalties; Cepheid: Consultancy, Membership on an entity's Board of Directors or advisory committees; Dana-Farber Cancer Institute: Patents & Royalties; EnLiven: Consultancy, Research Funding; Aptose Therapeutics Inc. (formerly Lorus): Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; ARIAD: Research Funding; Blueprint Medicines: Consultancy, Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; ALLCRON: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Research Funding; Aileron Therapeutics: Membership on an entity's Board of Directors or advisory committees; Pfizer: Research Funding; Oregon Health & Science University: Patents & Royalties. Byrd:Acerta Pharma: Research Funding; Syndax: Research Funding; Vincera: Research Funding; Pharmacyclics LLC, an AbbVie Company, Janssen, Novartis, Gilead, TG Therapeutics: Other; Pharmacyclics LLC, an AbbVie Company, Gilead, TG Therapeutics, Novartis, Janssen: Speakers Bureau; Novartis: Research Funding; Kartos Therapeutics: Research Funding; Trillium: Research Funding; Leukemia and Lymphoma Society: Other; Janssen: Consultancy; Pharmacyclics LLC, an AbbVie Company, Gilead, TG Therapeutics, BeiGene: Research Funding. Levine:Loxo: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Imago: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Research Funding; Qiagen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Prelude Therapeutics: Research Funding; Amgen: Honoraria; Astellas: Consultancy; Morphosys: Consultancy; Novartis: Consultancy; Isoplexis: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy; Lilly: Consultancy, Honoraria; Roche: Consultancy, Honoraria, Research Funding; Gilead: Honoraria. Mims:Abbvie: Membership on an entity's Board of Directors or advisory committees; Kura Oncology: Membership on an entity's Board of Directors or advisory committees; Leukemia and Lymphoma Society: Other: Senior Medical Director for Beat AML Study; Jazz Pharmaceuticals: Other: Data Safety Monitoring Board; Syndax Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Agios: Consultancy; Novartis: Speakers Bureau. OffLabel Disclosure: Enasidenib is not approved for the treatment of newly diagnosed AML.

Chimeric antigen receptor (CAR) T-cells directed against CD19 have drastically altered outcomes for children with relapsed and refractory acute lymphoblastic leukemia (r/r ALL). Pediatric patients with r/r ALL treated with CAR-T are at increased risk of both cytokine release syndrome (CRS) and sepsis. We sought to investigate the biologic differences between CRS and sepsis and to develop predictive models which could accurately differentiate CRS from sepsis at the time of critical illness. We identified 23 different cytokines that were significantly different between patients with sepsis and CRS. Using elastic net prediction modeling and tree classification, we identified cytokines that were able to classify subjects as having CRS or sepsis accurately. A markedly elevated interferon γ (IFNγ) or a mildly elevated IFNγ in combination with a low IL1β were associated with CRS. A normal to mildly elevated IFNγ in combination with an elevated IL1β was associated with sepsis. This combination of IFNγ and IL1β was able to categorize subjects as having CRS or sepsis with 97% accuracy. As CAR-T therapies become more common, these data provide important novel information to better manage potential associated toxicities.

STATEMENT: The International Society for Cellular and Gene Therapies (ISCT) and the International Society for Extracellular Vesicles (ISEV) recognize the potential of extracellular vesicles (EVs, including exosomes) from mesenchymal stromal cells (MSCs) and possibly other cell sources as treatments for COVID-19. Research and trials in this area are encouraged. However, ISEV and ISCT do not currently endorse the use of EVs or exosomes for any purpose in COVID-19, including but not limited to reducing cytokine storm, exerting regenerative effects or delivering drugs, pending the generation of appropriate manufacturing and quality control provisions, pre-clinical safety and efficacy data, rational clinical trial design and proper regulatory oversight. (C) 2020 International Society for Cell and Gene Therapy. This is an open access article under the CC BY-NC-ND license. (http://creativecommons.org/licenses/by-nc-nd/4.0/)

Hospital exemption (HE) is a regulated pathway that allows the use of advanced therapy medicinal products (ATMPs) within the European Union (EU) under restrictive conditions overseen by national medicine agencies. In some EU countries, HE is granted for ATMPs with no demonstrated safety and efficacy; therefore, they are equivalent to investigational drugs. In other countries, HE is granted for ATMPs with demonstrated quality, safety and efficacy and for which centralized marketing authorization has not been requested. The Committee on the Ethics of Cell and Gene Therapy of the International Society for Cell & Gene Therapy reflects here on the ethical issues concerning HE application from the perspective of the patient, including risk-benefit balance, accessibility and transparency, while providing evidence that HE must not be regarded as a conduit for unproven and unethical ATMP-based interventions. Indeed, HE represents a legal instrument under which a patient's need for access to novel ATMPs is reconciled with ethics. Moreover, for some unmet medical needs, HE is the only pathway for accessing innovative ATMPs. Nonetheless, HE harmonization across EU Member States and limitations of ATMP use under the HE rule when similar products have already been granted centralized marketing authorization to avoid a parallel regulatory pathway are controversial issues whose political and economic consequences are beyond the scope of this review. Finally, the institution of an EU registry of HE applications and outcomes represents a priority to improve transparency, reduce patient risks, increase efficiency of health systems, facilitate company awareness of business opportunities and boost progressive entry of ATMPs into the therapeutic repertoire of health systems.

The 2010 and 2017 editions of the European LeukemiaNet (ELN) recommendations for diagnosis and management of acute myeloid leukemia (AML) in adults are widely recognized among physicians and investigators. There have been major advances in our understanding of AML, including new knowledge about the molecular pathogenesis of AML, leading to an update of the disease classification, technological progress in genomic diagnostics and assessment of measurable residual disease, and the successful development of new therapeutic agents, such as FLT3, IDH1, IDH2, and BCL2 inhibitors. These advances have prompted this update that includes a revised ELN genetic risk classification, revised response criteria, and treatment recommendations.

While Ras/mitogen-activated protein kinase (MAPK) signaling is activated in most human cancers, attempts to target this pathway using kinase active site inhibitors have not typically led to durable, clinical benefit. To address this shortcoming, we sought to test the feasibility of an alternative targeting strategy, focused on the ERK2 substrate binding domains, D and DBP. We found that disabling the ERK2-DBP domain in mice caused baseline erythrocytosis. Consequently, we investigated the role of the ERK2-D and -DBP domains in disease, using a JAK2-dependent model of polycythemia vera (PV). Importantly, inactivation of the ERK2-DBP domain promoted the progression of disease from PV to myelofibrosis (MF), suggesting that the ERK2-DBP domain normally opposes progression. ERK2-DBP inactivation also prevented oncogenic JAK2 kinase (JAK2V617F) from promoting oncogene-induced senescence (OIS) in vitro. The ERK2-DBP mutation attenuated JAK2-mediated OIS by preventing the physical interaction of ERK2 with the transcription factor, Egr1. Because inactivation of the ERK2-DBP created a functional ERK2 kinase limited to binding substrates through its D-domain, these data suggested that the D-domain substrates were responsible for promoting oncogene-induced progenitor growth and tumor progression, and that pharmacologic targeting of the ERK2-D domain might attenuate cancer cell growth. Indeed, pharmacologic agents targeting the ERK2-D domain were effective in attenuating the growth of JAK2-dependent myeloproliferative neoplasm cell lines. Taken together, these data indicate that the ERK-D and -DBP domains can play distinct roles in the progression of neoplasms and that the D-domain has the potential to be potent therapeutic target in Ras/MAPK dependent cancers.

Leukemic transformation (LT) of myeloproliferative neoplasm (MPN) has a dismal prognosis and is largely fatal. Mutational inactivation of TP53 is the most common somatic event in LT; however, the mechanisms by which TP53 mutations promote LT remain unresolved. Using an allelic series of mouse models of Jak2/Trp53 mutant MPN, we identify that only biallelic inactivation of Trp53 results in LT (to a pure erythroleukemia [PEL]). This PEL arises from the megakaryocyte-erythroid progenitor population. Importantly, the bone morphogenetic protein 2/SMAD pathway is aberrantly activated during LT and results in abnormal self-renewal of megakaryocyte-erythroid progenitors. Finally, we identify that Jak2/Trp53 mutant PEL is characterized by recurrent copy number alterations and DNA damage. Using a synthetic lethality strategy, by targeting active DNA repair pathways, we show that this PEL is highly sensitive to combination WEE1 and poly(ADP-ribose) polymerase inhibition. These observations yield new mechanistic insights into the process of p53 mutant LT and offer new, clinically translatable therapeutic approaches.

Acquired somatic mutations in hematopoietic stem and progenitor cells (clonal hematopoiesis or CH) are associated with advanced age, increased risk of cardiovascular and malignant diseases, and decreased overall survival. These adverse sequelae may be mediated by altered inflammatory profiles observed in patients with CH. A pro-inflammatory immunologic profile is also associated with worse outcomes of certain infections, including SARS-CoV-2 and its associated disease Covid-19. Whether CH predisposes to severe Covid-19 or other infections is unknown. Among 525 individuals with Covid-19 from Memorial Sloan Kettering (MSK) and the Korean Clonal Hematopoiesis (KoCH) consortia, we show that CH is associated with severe Covid-19 outcomes (OR = 1.85, 95%=1.15–2.99, p = 0.01), in particular CH characterized by non-cancer driver mutations (OR = 2.01, 95% CI = 1.15–3.50, p = 0.01). We further explore the relationship between CH and risk of other infections in 14,211 solid tumor patients at MSK. CH is significantly associated with risk of Clostridium Difficile (HR = 2.01, 95% CI: 1.22–3.30, p = 6×10−3) and Streptococcus/Enterococcus infections (HR = 1.56, 95% CI = 1.15–2.13, p = 5×10−3). These findings suggest a relationship between CH and risk of severe infections that warrants further investigation., Clonal haematopoiesis (CH) has been associated with altered inflammatory profiles and increased risk of cardiovascular and malignant diseases. Here, the authors analyze patient data from two different cohorts and show that CH is associated with severe infections and severe Covid19.

Background: Measurable residual disease (MRD) is a powerful prognostic factor in AML, including in prediction of outcomes post allogeneic stem cell transplant (alloSCT). However, genomic predictors of successful MRD eradication with chemotherapy prior to alloSCT are unclear. Objectives: Here we provide an integrated analysis of 233 patients (pts) who underwent induction chemotherapy with baseline next-generation sequencing (NGS) followed by serial immunophenotypic monitoring for MRD while patients received additional therapy and alloSCT. Methods: All pts who received anthracycline + cytarabine, +/- investigational agents at Memorial Sloan Kettering Cancer Center starting in April 2014 were retrospectively studied (A). 142 out of 233 pts subsequently underwent alloSCT after induction or additional therapy (A). Immunophenotypic MRD was identified in bone marrow aspirates (BMA) by multiparameter flow cytometry. Any level of residual disease was considered MRD+. Molecular analysis was obtained from pre-induction BMA by NGS using 28 or 49 or 400 gene panels. Results: Patient and treatment characteristics for all pts are detailed in panel (B). Induction chemotherapy resulted in an MRD-CR/CRi and MRD+CR/CRi in 29% and 23% of all pts, respectively (C). Additional therapy included consolidation (n=51), intensive re-induction/salvage (n=47) and non-intensive therapy (n=9). Of 83 AML pts with persistent AML and 58 pts with MRD+CR/CRi after induction (R1), 38/141 (27%) were able to be converted to MRD-CR/CRi. While 33/38 of pts went on to alloSCT after conversion to MRD-CR/CRi, 22 and 36 pts went to alloSCT with persistent AML and MRD+CR/CRi AML, respectively. We focused on pre-induction molecular predictors for achieving an MRD-CR/CRi response prior to transplant for the 142 pts who underwent alloSCT (D). Pts with a NPM1 (79%, Odds ratio [OR] 3.7, p=0.01), IDH1 (92%, OR 3.9, p=0.01) and KRAS (100%, OR 5.0, p=0.03) mutations achieved high rates of MRD-CR/CRi prior to alloSCT. In contrast, RUNX1 (28%, OR 0.2, p=0.01), TP53 (12%, OR 0.1, p=0.02) and SF3B1 (14%, OR 0.1, p=0.04) mutations predicted decreased odds of achieving MRD-CR/CRi prior to alloSCT despite induction and post-induction therapy. AlloSCT resulted in high rates of conversion from MRD+ and persistent disease to MRD negativity. Most pts who entered transplant with CR/CRi MRD+ (28/36, 76%) or persistent AML (14/22, 64%) cleared MRD by the first post-transplant BMA at a median of 32 days (E). Post-alloSCT follow-up indicated value in converting MRD+ to MRD- prior to alloSCT. There was no significant difference in post-transplant cumulative incidence of relapse (F) and OS (G) between early MRD-CR/CRi immediately following induction versus later conversion to MRD-CR/CRi with additional therapy prior to alloSCT. Despite initial post-transplant MRD clearance, pts who entered alloSCT with persistent AML or MRD+ had higher incidence of relapse (p=0.00037, F) and poorer post-transplant OS (p=0.013, G) compared to pts who entered alloSCT with MRD-. Pts with persistent disease prior to alloSCT had shorter duration of MRD- induced by alloSCT compared to pts with MRD-CR/CRi after induction or converted MRD-CR/CRi prior to alloSCT (p=0.0042, H). Importantly, duration of MRD negativity after alloSCT for patients who achieved MRD- prior to alloSCT was not affected by whether patients received induction +/- consolidation (I: treatment type 1-3 from B) vs. induction and salvage treatment for refractory AML (I: treatment type 4-6 from B). Conclusion: We show that transplanted AML pts with specific molecular mutations (RUNX1, SF3B1, and TP53) are unlikely to achieve MRD-CR/CRi after induction, consolidation or salvage therapy, while other mutations (NPM1, IDH1, KRAS) predict high rates of MRD- prior to alloSCT. Additional post-induction therapy may be advantageous for some MRD+ pts to achieve MRD- prior to alloSCT. Post-transplant OS is improved in pts who are MRD- at time of transplant, regardless of whether they required additional therapy beyond induction to achieve this state. AlloSCT is highly effective at eradicating MRD, but post-transplant MRD- is more durable in pts who are MRD- pre-alloSCT. Our results suggest that development of MRD-eradicating therapies has the potential to improve post-transplant outcomes and argues for innovative trials for pts with adverse molecular features currently unlikely to achieve MRD- pre alloSCT. Figure Disclosures Cai: Imago Biosciences, Inc.: Consultancy, Current equity holder in private company; DAVA Oncology: Honoraria. Geyer:Amgen: Research Funding. Glass:Gerson Lehman Group: Consultancy. Stein:Syros: Membership on an entity's Board of Directors or advisory committees; PTC Therapeutics: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Biotheryx: Consultancy; Bayer: Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees; Syndax: Consultancy, Research Funding; Seattle Genetics: Consultancy; Abbvie: Consultancy; Amgen: Consultancy; Celgene Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Agios Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Astellas Pharmaceuticals: Consultancy, 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, Research Funding. Levine:Gilead: Honoraria; Isoplexis: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Qiagen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Research Funding; Janssen: Consultancy; Lilly: Consultancy, Honoraria; Imago: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy; Novartis: Consultancy; Prelude Therapeutics: Research Funding; Loxo: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria; Morphosys: Consultancy; Roche: Consultancy, Honoraria, Research Funding. Gyurkocza:Actinium: Research Funding. Perales:Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; Nektar Therapeutics: 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; MolMed: Membership on an entity's Board of Directors or advisory committees; Abbvie: Honoraria, Membership on an entity's Board of Directors or advisory committees; Medigene: Membership on an entity's Board of Directors or advisory committees, Other; Servier: Membership on an entity's Board of Directors or advisory committees, Other; Omeros: Honoraria, Membership on an entity's Board of Directors or advisory committees; Merck: Consultancy, Honoraria; NexImmune: Membership on an entity's Board of Directors or advisory committees; Cidara Therapeutics: Other; Miltenyi Biotec: Research Funding; Kite/Gilead: Honoraria, Research Funding; Incyte Corporation: Honoraria, Research Funding; Bristol Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria; Bellicum: Honoraria, Membership on an entity's Board of Directors or advisory committees. Abdel-Wahab:H3 Biomedicine Inc.: Consultancy, Research Funding; Janssen: Consultancy; Envisagenics Inc.: Current equity holder in private company; Merck: Consultancy. Papaemmanuil:Kyowa Hakko Kirin: Consultancy, Honoraria; Isabl: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; MSKCC: Patents & Royalties; Novartis: Consultancy, Honoraria; Illumina: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Prime Oncology: Consultancy, Honoraria. Giralt:KITE: Consultancy; NOVARTIS: Consultancy, Honoraria, Research Funding; OMEROS: Consultancy, Honoraria; AMGEN: Consultancy, Research Funding; TAKEDA: Research Funding; ACTINUUM: Consultancy, Research Funding; MILTENYI: Consultancy, Research Funding; CELGENE: Consultancy, Honoraria, Research Funding; JAZZ: Consultancy, Honoraria. Tallman:Glycomimetics: Research Funding; Rafael: Research Funding; Amgen: Research Funding; Bioline rx: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; KAHR: Membership on an entity's Board of Directors or advisory committees; UpToDate: Patents & Royalties; Rigel: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Membership on an entity's Board of Directors or advisory committees; Oncolyze: Membership on an entity's Board of Directors or advisory committees; Delta Fly Pharma: Membership on an entity's Board of Directors or advisory committees; BioSight: Membership on an entity's Board of Directors or advisory committees, Research Funding; ADC Therapeutics: Research Funding; Orsenix: Research Funding; Cellerant: Research Funding; Abbvie: Research Funding. Goldberg:AROG: Research Funding; Aprea: Research Funding; ADC Therapeutics: Research Funding; Genentech: Consultancy, 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, Research Funding; Celgene: Consultancy; Aptose: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Dava Oncology: Honoraria; Pfizer: Research Funding; Celularity: Research Funding.

INTRODUCTION Anti-CD19 CAR T-cell immunotherapy is promising for patients with relapsed/refractory CLL. Hypogammaglobinemia can result from normal CD19+ B-cell depletion by CAR-T cells. CLL patients are already at risk for infections due to impaired immune function, lymphodepletion prior to CAR-T cells infusion, and immunosuppressive therapies. Intravenous immunoglobulin (IVIG) is used to manage hypogammaglobulinemia, although standard criteria for IVIG administration in this setting has not been established. We studied the incidence of hypogammaglobinemia and report infectious complications, risk factors, IVIG use, and clinical outcomes for CLL patients treated with anti-CD19 CAR-T cells. METHODS Adult CLL patients who received CD19-directed CAR-T therapy in 3 clinical trials (NCT01029366, NCT01747486, NCT02640209) from July 2010 to February 2020 were included. We reviewed demographics, available IgG levels, IVIG use, and clinical outcomes with a particular focus on infectious complications. Hypogammaglobulinemia was defined as IgG RESULTS Records of 71 adult patients with CLL were reviewed; 4 were excluded from further analysis as they did not have IgG levels after CAR-T. The median age at time of CAR-T was 63 years (range 43-78 years). 31 patients (46%) were alive, 31 (46%) were deceased, and 5 (7%) were lost to follow up at time of review. Median follow up for all patients was 33 months (range 2-114 months). Of the 55 patients with an IgG level prior to CAR-T infusion, 24 (44%) had hypogammaglobulinemia at baseline After CAR-T infusion, 54 of 67 patients (81%) developed new or persistent hypogammaglobulinemia, and 40 of these patients (74%) received IVIG (Table 1). Forty-eight patients (72%) received at least one infusion of IVIG after CAR-T. Median time to initiation of IVIG after CAR-T infusion was 2.8 months (range 0.1-71.2 months). IVIG was used in 29 of 35 (83%) responders (defined as PR or CR) vs 19 of 32 (59%) in non-responders (defined as NR or PD) (p=0.056). There was no difference in survival observed based on whether or not patients had hypogammaglobulinemia (Figure). 42 patients (63%) had documented infections not related to chemotherapy-induced neutropenia. Fifteen (22%) had one documented infection, and 27 (40%) had more than one documented infection. There were 13 infections documented in patients who did not have hypogammaglobulinemia; the most common were ENT/sinus infections (5), bacteremia (2), URI (2), and other (2). There were 94 infections documented in patients who developed or had persistent hypogammaglobulinemia; the most common were lower respiratory tract infection/pneumonia (21), URI (20), and ENT/sinus infections (13). Complete and partial responders had more infections compared to non-responders or those who progressed (p = 0.01). Patients with hypogammaglobulinemia after CAR-T had an average of 1.74 (range 0-15) infections vs 1 (range 0-3) in patients without hypogammaglobulinemia. Patients who received IVIG had more infections (p=0.003); without IVIG the average number of infections was 0.58 (range 0-3), and with IVIG, the average number of infections was 2.00 (range 0-15). CONCLUSION Evaluating clinical outcomes with infections after CAR-T and potential strategies to minimize infection risk may improve morbidity and mortality in CLL patients. Use of IVIG was driven by individual practice, with heterogeneity regarding indication, frequency, and duration of treatment, though there was no difference in patient characteristics or response in patients who did or did not develop hypogammaglobulinemia. Patients who responded to CAR-T had more frequent infections, as might be expected in the setting of transient or persistent B- cell aplasia or hypoplasia. Patients who had more infections were more likely to receive IVIG. Further studies to define criteria for IVIG repletion in CLL patients treated with CD19-directed CAR-T cells may be incorporated in a standard clinical management algorithm. Table 1 Disclosures Frey: Amgen: Consultancy, Honoraria; Syntax: Consultancy, Honoraria; Kite Pharma: Consultancy, Honoraria. Davis:Tmunity Therapeutics, Inc.: Consultancy, Patents & Royalties, Research Funding; Cellares Corporation: Membership on an entity's Board of Directors or advisory committees; Novartis Institutes for Biomedical Research: Patents & Royalties. Hexner:Blueprint Medicines Corporation: Other: serves on a data safety monitoring committee, Research Funding; Novartis: Research Funding; Samus Therapeutics: Research Funding; American Board of Internal Medicine: Other: member of the hematology exam committee. Schuster:Novartis, Genentech, Inc./ F. Hoffmann-La Roche: Research Funding; AlloGene, AstraZeneca, BeiGene, Genentech, Inc./ F. Hoffmann-La Roche, Juno/Celgene, Loxo Oncology, Nordic Nanovector, Novartis, Tessa Therapeutics: Consultancy, Honoraria. Gill:Fate: Consultancy; Sensei: Consultancy; Aileron: Consultancy; Tmunity Therapeutics: Research Funding; Carisma Therapeutics: Patents & Royalties, Research Funding; Novartis: Research Funding. Grupp:Servier: Research Funding; Kite/Gilead: Research Funding; Roche: Consultancy; GlaxoSmithKline: Consultancy; Humanigen: Consultancy; CBMG: Consultancy; Jazz: Other: SSC; Adaptimmune: Other: SAB; TCR2: Other: SAB; Cellectis: Other; Juno/BMS: Other; Janssen/JnJ: Consultancy; CRISPR Therapeutics/Vertex Pharmaceuticals: Other; Allogene: Other; Novartis: Consultancy, Other: SSC, Research Funding. Melenhorst:Johnson & Johnson: Consultancy, Other: Speaker; Novartis: Other: Speaker, Research Funding; Kite Pharma: Research Funding; IASO Biotherapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Poseida Therapeutics: Consultancy; Simcere of America: Consultancy. Lacey:Novartis: Patents & Royalties: CAR T cells, Research Funding; Tmunity: Research Funding; Cabaletta: Research Funding; Carisma: Research Funding. Fraietta:Tmunity: Research Funding. Hwang:Novartis: Research Funding; Tmunity Therapeutics: Research Funding. Siegel:Novartis: Patents & Royalties; Tmunity: Patents & Royalties; Poseida: Membership on an entity's Board of Directors or advisory committees; Vetigenics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees. Levine:Terumo: Consultancy; Novartis: Consultancy, Patents & Royalties: Dr. Levine has a patent Methods for treatment of cancer (US 8906682) (US 8916381)(US 9101584) with royalties paid to University of Pennsylvania, a patent Compositions for treatment of cancer (US 8911993) (US 9102761) (US 9102760) with royalties paid to U; Lilly Asia Ventures: Consultancy; Avectas: Membership on an entity's Board of Directors or advisory committees; Patheon: Membership on an entity's Board of Directors or advisory committees; Immuneel: Membership on an entity's Board of Directors or advisory committees; Incysus: Membership on an entity's Board of Directors or advisory committees; Ori Biotech: Membership on an entity's Board of Directors or advisory committees; Vycellix: Membership on an entity's Board of Directors or advisory committees; Tmunity Therapeutics: Current equity holder in private company, Research Funding. June:Bluesphere Bio: Membership on an entity's Board of Directors or advisory committees; Cabaletta Bio: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Carisma Therapeutics: Membership on an entity's Board of Directors or advisory committees; Cellares: Membership on an entity's Board of Directors or advisory committees; Celldex: Consultancy, Membership on an entity's Board of Directors or advisory committees; DeCART Therapeutics: Membership on an entity's Board of Directors or advisory committees; Immune Design: Membership on an entity's Board of Directors or advisory committees; Kiadis Pharma: Current equity holder in private company; Novartis: Patents & Royalties, Research Funding; Tmunity Therapeutics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding; Ziopharm Oncology: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees. Porter:Tmunity: Patents & Royalties; Novartis: Honoraria, Other: Advisory board, Patents & Royalties: CAR T cells for CD19+ malignancies, Research Funding; American Board of Internal Medicine: Other: Member, exam writing committee (end date Oct 2019); National Marrow Donor Program: Membership on an entity's Board of Directors or advisory committees; Janssen: Other: Advisory board; Genentech/Roche: Current equity holder in publicly-traded company, Other: Spouse employment (ended Sept 2020); her salary includes stock/options; Glenmark: Other: Advisory board; Adicet bio: Other: Advisory board; Incyte: Other: Advisory board; Kite/Gilead: Other: Advisory board. OffLabel Disclosure: CAR T cells for CLL

Methylation profiling in myeloid malignancies such as Acute Myeloid Leukemia (AML) and Chronic Myelomonocytic Leukemia (CMML) has demonstrated the ability to define distinct biological and clinical subgroups, including predicting which patients will respond to therapy with a hypomethylating agent (HMA). The Philadelphia-chromosome negative myeloproliferative neoplasms (MPNs) carry an inherent risk of progression to an accelerated-phase disease (AP; 10-19% blasts in the peripheral blood or bone marrow), as well as to blast phase disease (BP; ≥ 20% blasts in the peripheral blood or bone marrow), which is associated with a poor prognosis. It is unknown whether the methylation profiles of MPN-AP/BP cases may further help identify distinct biological, genomic, and clinical subgroups, including identifying patients more likely to respond to HMA. We recently carried out a phase I/II study to test the safety and efficacy of combination therapy with the JAK1/2 inhibitor ruxolitinib (RUX) and the HMA Decitabine (DAC) in patients with MPN-AP/BP (MPD-RC 109 study; NCT02076191). A total of 46 patients were accrued to the phase I and II studies. 37 patients were evaluable for response. Complete response (CR) occurred in 10%, Complete Response with incomplete count recovery (CRi) in 24%, Partial Response in 24%. 42% of patients had no response to therapy. Using samples available from the MPD-RC 109 study, we sought to assess whether the baseline global methylation profile predicts for response to this regimen. Further, we sought to utilize this dataset to determine if IDH2 mutations (amongst the most common mutations in MPN-AP/BP) are associated with a distinct methylation profile, as has been demonstrated in de novo AML. We carried out a pilot study of 11 MPN-AP/BP patients from the MPD-109 phase I/II trial and performed Enhanced Reduced Representation Bisulfite Sequencing (ERRBS) for DNA methylation quantification at ~3M CpG sites across the genome. Baseline DNA methylation profiles were compared between Responder (R) and Non-responder (NR) patients. Notably, unsupervised analysis using correspondence analysis (COA) demonstrated an almost complete separation of the two groups of patients (Fig 1A), while supervised analysis using a beta binomial model identified 134 differentially methylated regions (DMRs) (FDR25%) between the two groups at diagnosis (Fig 1B). Similar to our prior observation in CMML, response-associated DMRs were depleted from promoter regions (p We next carried out a pilot study to characterize the epigenetic abnormalities of IDH2-mutant MPN-AP/BP cases. For this purpose, we compared the genome-wide DNA methylation profiles of 12 IDH2-mutant to 7 IDH1/2 wild type MPN-AP/BP cases using ERRBS. Unsupervised analysis based on the DNA methylation profiles alone showed a strong trend to naturally segregate mutant from wild-type cases, indicating strong underlying epigenetic differences (Fig.1D). A supervised analysis using the beta binomial method identified 1,477 differentially methylated regions (DMRs) between the two groups (average absolute methylation difference ≥25% and FDR Our data demonstrate that the methylation profile of MPN-AP/BP may predict for response to HMA-based therapy. Such data could be used to guide therapeutic decisions and select patient for whom HMA has the highest likelihood of procuring a response. As well, these findings indicate that IDH2-mutant MPN-AP/BP are epigenetically distinct, and given the preferred targeting of regulatory elements, these epigenetic differences may play a functional role in disease biology. Further validation of these observations is required. Updated data, including analysis of further cases, and RNA-sequencing analysis of gene-expression and pathway enrichment of genes differentially methylated between responders and non-responders, and IDH2 mutated and wildtype cases will be presented at the conference. Disclosures Rampal: Constellation: Research Funding; Pharmaessentia: Consultancy; CTI Biopharma: Consultancy; Promedior: Consultancy; Celgene: Consultancy; Incyte: Consultancy, Research Funding; Abbvie: Consultancy; Galecto: Consultancy; Jazz Pharmaceuticals: Consultancy; Blueprint: Consultancy; Stemline: Consultancy, Research Funding. Mascarenhas:Celgene, Prelude, Galecto, Promedior, Geron, Constellation, and Incyte: Consultancy; Incyte, Kartos, Roche, Promedior, Merck, Merus, Arog, CTI Biopharma, Janssen, and PharmaEssentia: Other: Research funding (institution). Levine:Morphosys: Consultancy; Prelude Therapeutics: Research Funding; Novartis: Consultancy; Amgen: Honoraria; Lilly: Consultancy, Honoraria; Janssen: Consultancy; Astellas: Consultancy; Roche: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Imago: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Isoplexis: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Loxo: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Qiagen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria. Hoffman:Novartis: Membership on an entity's Board of Directors or advisory committees; Protagonist: Consultancy; Forbius: Consultancy; Dompe: Research Funding; Abbvie: Membership on an entity's Board of Directors or advisory committees.

Thirty years after initial publications of the concept of a chimeric antigen receptor (CAR), the U.S. Food and Drug Administration (FDA) approved the first anti-CD19 CAR T-cell therapy. Unlike other immunotherapies, such as immune checkpoint inhibitors and bispecific antibodies, CAR T cells are unique as they are "living drugs," that is, gene-edited killer cells that can recognize and kill cancer. During these 30 years of development, the CAR construct, T-cell manufacturing process, and clinical patient management have gone through rounds of failures and successes that drove continuous improvement. Tisagenlecleucel was the first gene therapy to receive approval from the FDA for any indication. The initial approval was for relapsed or refractory (r/r) pediatric and young-adult B-cell acute lymphoblastic leukemia in August 2017 and in May 2018 for adult r/r diffuse large B-cell lymphoma. Here we review the preclinical and clinical development of what began as CART19 at the University of Pennsylvania and later developed into tisagenlecleucel.

Background aims Autologous cell therapy (AuCT) is an emerging therapeutic treatment that is undergoing transformation from laboratory- to industry-scale manufacturing with recent regulatory approvals. Various challenges facing the complex AuCT manufacturing and supply chain process hinder the scale out and broader application of this highly potent treatment. Methods We present a multiscale logistics simulation framework, AuCT-Sim, that integrates novel supply chain system modeling algorithms, methods, and tools. AuCT-Sim includes a single facility model and a system-wide network model. Unique challenges of the AuCT industry are analyzed and addressed in AuCT-Sim. Decision-supporting tools can be developed based on this framework to explore “what-if” manufacturing and supply chain scenarios of importance to various cell therapy stakeholder groups. Results Two case studies demonstrate the decision-supporting capability of AuCT-Sim where one investigates the optimal reagent base stocking level, and the other one simulates a reagent supply disruption event. These case studies serve as guidelines for designing computational experiments with AuCT-Sim to solve specific problems in AuCT manufacturing and supply chain. Discussion This simulation framework will be useful in understanding the impact of possible manufacturing and supply chain strategies, policies, regulations, and standards informing strategies to increase patient access to AuCT.

PURPOSE CD19-targeted chimeric antigen receptor (CAR)–modified T cells demonstrate unprecedented responses in B-cell acute lymphoblastic leukemia (B-ALL); however, relapse remains a substantial challenge. Short CAR T-cell persistence contributes to this risk; therefore, strategies to improve persistence are needed. METHODS We conducted a pilot clinical trial of a humanized CD19 CAR T-cell product (huCART19) in children and young adults with relapsed or refractory B-ALL (n = 72) or B-lymphoblastic lymphoma (n = 2), treated in two cohorts: with (retreatment, n = 33) or without (CAR-naive, n = 41) prior CAR exposure. Patients were monitored for toxicity, response, and persistence of huCART19. RESULTS Seventy-four patients 1-29 years of age received huCART19. Cytokine release syndrome developed in 62 (84%) patients and was grade 4 in five (6.8%). Neurologic toxicities were reported in 29 (39%), three (4%) grade 3 or 4, and fully resolved in all cases. The overall response rate at 1 month after infusion was 98% (100% in B-ALL) in the CAR-naive cohort and 64% in the retreatment cohort. At 6 months, the probability of losing huCART19 persistence was 27% (95% CI, 14 to 41) for CAR-naive and 48% (95% CI, 30 to 64) for retreatment patients, whereas the incidence of B-cell recovery was 15% (95% CI, 6 to 28) and 58% (95% CI, 33 to 77), respectively. Relapse-free survival at 12 and 24 months, respectively, was 84% (95% CI, 72 to 97) and 74% (95% CI, 60 to 90) in CAR-naive and 74% (95% CI, 56 to 97) and 58% (95% CI, 37 to 90) in retreatment cohorts. CONCLUSION HuCART19 achieved durable remissions with long-term persistence in children and young adults with relapsed or refractory B-ALL, including after failure of prior CAR T-cell therapy.

Leukemias bearing fusions of the AF10/MLLT10 gene are associated with poor prognosis, and therapies targeting these fusion proteins (FPs) are lacking. To understand mechanisms underlying AF10 fusion-mediated leukemogenesis, we generated inducible mouse models of acute myeloid leukemia (AML) driven by the most common AF10 FPs, PICALM/CALM-AF10 and KMT2A/MLL-AF10, and performed comprehensive characterization of the disease using transcriptomic, epigenomic, proteomic, and functional genomic approaches. Our studies provide a detailed map of gene networks and protein interactors associated with key AF10 fusions involved in leukemia. Specifically, we report that AF10 fusions activate a cascade of JAK/STAT-mediated inflammatory signaling through direct recruitment of JAK1 kinase. Inhibition of the JAK/STAT signaling by genetic Jak1 deletion or through pharmacological JAK/STAT inhibition elicited potent antioncogenic effects in mouse and human models of AF10 fusion AML. Collectively, our study identifies JAK1 as a tractable therapeutic target in AF10-rearranged leukemias.

The adoptive transfer of T lymphocytes reprogrammed to target tumor cells has demonstrated significant potential in various malignancies. However, little is known about the long-term potential and the clonal stability of the infused cells. Here, we studied the longest persisting CD19 redirected chimeric antigen receptor (CAR) T cells to date in two chronic lymphocytic leukemia (CLL) patients who achieved a complete remission in 2010. CAR T-cells were still detectable up to 10+ years post-infusion, with sustained remission in both patients. Surprisingly, a prominent, highly activated CD4+ population developed in both patients during the years post-infusion, dominating the CAR T-cell population at the late time points. This transition was reflected in the stabilization of the clonal make-up of CAR T-cells with a repertoire dominated by few clones. Single cell multi-omics profiling via Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-Seq) with TCR sequencing of CAR T-cells obtained 9.3 years post-infusion demonstrated that these long-persisting CD4+ CAR T-cells exhibited cytotoxic characteristics along with strong evidence of ongoing functional activation and proliferation. Our data provide novel insight into the CAR T-cell characteristics associated with long-term remission in leukemia.

Acute myeloid leukemia (AML) is the most common diagnosed leukemia. In older adults, AML confers an adverse outcome1,2. AML originates from a dominant mutation, then acquires collaborative transformative mutations leading to myeloid transformation and clinical/biological heterogeneity. Currently, AML treatment is initiated rapidly, precluding the ability to consider the mutational profile of a patient’s leukemia for treatment decisions. Untreated patients with AML ≥ 60 years were prospectively enrolled on the ongoing Beat AML trial (ClinicalTrials.gov NCT03013998 ), which aims to provide cytogenetic and mutational data within 7 days (d) from sample receipt and before treatment selection, followed by treatment assignment to a sub-study based on the dominant clone. A total of 487 patients with suspected AML were enrolled; 395 were eligible. Median age was 72 years (range 60–92 years; 38% ≥75 years); 374 patients (94.7%) had genetic and cytogenetic analysis completed within 7 d and were centrally assigned to a Beat AML sub-study; 224 (56.7%) were enrolled on a Beat AML sub-study. The remaining 171 patients elected standard of care (SOC) (103), investigational therapy (28) or palliative care (40); 9 died before treatment assignment. Demographic, laboratory and molecular characteristics were not significantly different between patients on the Beat AML sub-studies and those receiving SOC (induction with cytarabine + daunorubicin (7 + 3 or equivalent) or hypomethylation agent). Thirty-day mortality was less frequent and overall survival was significantly longer for patients enrolled on the Beat AML sub-studies versus those who elected SOC. A precision medicine therapy strategy in AML is feasible within 7 d, allowing patients and physicians to rapidly incorporate genomic data into treatment decisions without increasing early death or adversely impacting overall survival. Preliminary results from the Beat AML umbrella trial demonstrates the feasibility and efficacy of applying prospective genomic profiling for matching newly diagnosed patients with AML with targeted therapies.

Cancer-associated venous thromboembolism (CAT) is a well-described complication of cancer and a leading cause of death in cancer patients. The purpose of this study was to assess potential associations of molecular signatures with CAT, including tumor-specific mutations and the presence of clonal hematopoiesis. We analyzed deep-coverage targeted DNA-sequencing data of >14,000 solid tumor samples using the MSK-IMPACT™ platform to identify somatic alterations associated with VTE. Endpoint was defined as the first instance of cancer-associated pulmonary embolism and/or proximal/distal lower extremity deep vein thrombosis. Cause-specific Cox proportional hazards regression was used, adjusting for pertinent clinical covariates. Of 11,695 evaluable individuals, 72% had metastatic disease at time of IMPACT. Tumor-specific mutations in KRAS (HR=1.34 [1.09-1.64]; adjusted p=0.08), STK11 (HR=2.12 [1.55-2.89]; adjusted pKEAP1 (HR=1.84 [1.21-2.79]; adjusted p=0.07), CTNNB1 (HR=1.73 [1.15-2.60]; adjusted p=0.09), CDKN2B (HR= 1.45 [1.13-1.85], adjusted p=0.07) and MET (HR=1.83 [1.15-2.92]; adjusted p=0.09) were associated with a significantly increased risk of CAT independent of tumor type. Mutations in SETD2 were associated with a decreased risk of CAT (HR=0.35 [0.16-0.79], adjusted p=0.09). The presence of clonal hematopoiesis was not associated with an increased VTE rate. This is the first large-scale analysis to elucidate tumor-specific genomic events associated with CAT. Somatic tumor mutations of STK11, KRAS, CTNNB1, KEAP1, CDKN2B and MET were associated with an increased risk of VTE in solid tumor patients. Further analysis is needed to validate these findings and identify additional molecular signatures unique to individual tumor types.Key PointsTumor mutations in STK11, KRAS, CTNNB1, KEAP1, CDKN2B, MET and SETD2 modulate the risk of cancer-associated thrombosis.The presence of clonal hematopoiesis does not affect the risk of cancer-associated thrombosis.

Coronavirus disease 2019 (SARS-CoV2) is an active global health threat for which treatments are desperately being sought. Even though most people infected experience mild to moderate respiratory symptoms and recover with supportive care, certain vulnerable hosts develop severe clinical deterioration. While several drugs are currently being investigated in clinical trials, there are currently no approved treatments or vaccines for COVID-19 and hence there is an unmet need to explore additional therapeutic options. At least three inflammatory disorders or syndromes associated with immune dysfunction have been described in the context of cellular therapy. Specifically, Cytokine Release Syndrome (CRS), Immune Reconstitution Inflammatory Syndrome (IRIS), and Secondary Hemophagocytic Lymphohistiocytosis (sHLH) all have clinical and laboratory characteristics in common with COVID19 and associated therapies that could be worth testing in the context of clinical trials. Here we discuss these diseases, their management, and potential applications of these treatment in the context of COVID-19. We also discuss current cellular therapies that are being evaluated for the treatment of COVID-19 and/or its associated symptoms.

Genetic studies have identified recurrent somatic mutations in acute myeloid leukemia (AML) patients, including in the Wilms’ tumor 1 (WT1) gene. The molecular mechanisms by which WT1 mutations contribute to leukemogenesis have not yet been fully elucidated. We investigated the role of Wt1 gene dosage in steady-state and pathologic hematopoiesis. Wt1 heterozygous loss enhanced stem cell self-renewal in an age-dependent manner, which increased stem cell function over time and resulted in age-dependent leukemic transformation. Wt1-haploinsufficient leukemias were characterized by progressive genetic and epigenetic alterations, including those in known leukemia-associated alleles, demonstrating a requirement for additional events to promote hematopoietic transformation. Consistent with this observation, we found that Wt1 depletion cooperates with Flt3-ITD mutation to induce fully penetrant AML. Our studies provide insight into mechanisms of Wt1-loss leukemogenesis and into the evolutionary events required to induce transformation of Wt1-haploinsufficient stem/progenitor cells.

The interaction between ERK2 and its substrates is critically mediated by two domains, the common docking (CD) D-domain and DEF-binding pocket (DBP) domain. Previous studies have suggested that ERK2 is not only necessary to drive hematopoietic and myelo-erythroid development, but it is also important for the pathogenesis of hematological cancers, as revealed by recurrent ERK2 somatic mutations in many types of lymphoma and leukemia. Here we show that the activation of ERK2 in JAK2V617F-driven myeloproliferative neoplasm (MPN) enhance polycythemia vera (PV) progression from erythrocytosis to myelofibrosis when ERK2-DBP binding capacity is disabled. Conversely, targeting the ERK2 D-domain preserves ERK2 catalytic function while reducing outgrowth and proliferation of human and murine MPN cell lines. To determine whether ERK2 binding domains play a role in the pathogenesis of JAK2V617F-driven MPN, we generated an ERK2-Y261A (Erk2 Y261A) mutant knockin mouse model in which the ERK2-DBP domain was inactivated, but ERK2 kinase activity and D-domain function was preserved. We observed splenomegaly in Erk2 Y261A/Y261A, characterized by a 10-fold expansion of CD44+/Ter119+ primitive erythroblasts and Ter119+CD44+/ FSC hi immature erythroid progenitors. We then transplanted Jak2 V617F-expressing ERK2 wildtype (WT), knockout (KO) or DBP hematopoietic progenitors (HSPCs) into irradiated, immunodeficient mice (Rag2 −/−Il2rg −/−) and observed disease resembling human PV, with elevated RBC counts, hematocrit, and hemoglobin levels. ERK2-DBP mutant recipients exhibited mild but sustained erythrocytosis with a prevalence of circulating GFP+ monocytes and neutrophils. At 12-weeks post-transplantation, ERK2-DBP mutant recipients had significant splenic burden and decreased bone marrow cellularity relative to WT and KO recipients. Reticulin staining confirmed progression towards myelofibrosis with interstitial infiltration in both the BM and the spleen of ERK2 DBP mutant recipients. To understand the molecular basis of ERK-DBP domain in JAK2V617F-driven MPN, we performed in vitro colony forming unit assay to assess the clonogenic potential of JAK2V617F-expressing HSPCs in cytokine supplemented-methylcellulose supporting myeloid or erythroid progenitor development. Ectopic JAK2V617F expression markedly reduced colony formation by both ERK2 wildtype and knockout HSPCs. On the other hand, JAK2V617F expression did not reduce colony formation by ERK2-DBP HSPCs. These differences were not evident in methylcellulose cultures that supported erythroid development, suggesting that disabling ERK-DBP binding capacity primarily affects the myeloid lineage. Furthermore, we found that JAK2V617F increased senescence associated β-galactosidase (SA-βGa) activity in ERK2-WT HSPCs. SA-βGal activity was modestly affected by ERK deficiency but was strongly attenuated by the ERK2-DBP mutation, supporting the notion that ERK2-DBP domain promotes oncogene-induced senescence (OIS). Previously, we identified transcription factor, Early growth response 1 (Egr1) which interacts with ERK2-DBP domain and plays a critical role as a tumor suppressor in myeloid neoplasms. We observed that JAK2V617F expression failed to suppress colony formation and senescence by Egr1-deficient HSPCs. The ability of EGR1 to restore these effects is abrogated by inactivating the DEF motif (EGR1-Y252A). Together, this data demonstrates that the interaction between ERK2-DBP and Egr1 is required for JAK2V617F-mediated colony suppression and senescence induction. Finally, to determine whether ERK-D domain acts to promote progression by targeting substrates distinct from those that bind to the DBP domain, we developed an inhibitor (#76) of the ERK-D domain. Indeed, 76 spared ERK phosphorylation and EGR1 induction but impaired RSK phosphorylation, a critical D-domain substrate involved in cancer progression. Moreover, 76 suppressed colony formation by JAK2V61F-expressing HSPCs akin to the active site MEK/ERK inhibitor, U0126. By altering the 4-ethoxybenylidene moiety of 76, we were able to enhance anti-proliferative effects of the parent compound against human MPN cell lines, SET-2, UKE-1, and Ku812. Our findings identify ERK2-domain specific roles in the pathogenesis of JAK2V617F driven MPN and supports a novel therapeutic approach to targeting JAK2 and MAPK dependent MPN. Disclosures Levine: Celgene: Research Funding; QIAGEN: Membership on an entity's Board of Directors or advisory committees; Auron: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria; Mission Bio: Membership on an entity's Board of Directors or advisory committees; Zentalis: Membership on an entity's Board of Directors or advisory committees; Ajax: Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy; Lilly: Honoraria; Janssen: Consultancy; Astellas: Consultancy; Gilead: Honoraria; Prelude: Membership on an entity's Board of Directors or advisory committees; Isoplexis: Membership on an entity's Board of Directors or advisory committees; Morphosys: Consultancy; Roche: Honoraria, Research Funding; Imago: Membership on an entity's Board of Directors or advisory committees. Wiest: Janssen Pharmaceuticals: Research Funding.

INTRODUCTION Mixed phenotype (MP) is characteristic for de novo mixed phenotype acute leukemia (dnMPAL) but can also be seen in blast phase of myeloproliferative neoplasms (MPN-BP), myeloid/lymphoid neoplasms with eosinophilia & rearrangements, acute myeloid leukemia (AML) with recurrent cytogenetic abnormalities (AML-RCA) and secondary AML (sAML) including AML with myelodysplasia-related changes (AML-MRC) and therapy-related AML (t-AML). Although WHO classification excludes sAML with MP (sAML-MP) from dnMPAL, the significance of MP in the setting of sAML and their genetic landscape has not been studied. METHODS The MSKCC pathology data base was searched from 01/2014 to 09/2020 and a cohort of 125 patients with MP as defined per WHO 2016 classification was obtained. The clinical, morphologic, immunophenotypic, and cytogenetic/molecular results were reviewed. Patients with a diagnosis of AML-RCA, myeloid/lymphoid neoplasms with eosinophilia & rearrangements, MPN-BP, B-ALL with isolated MPO and myelodysplastic syndrome (MDS) were excluded (Fig 1). RESULTS 50 cases of sAML-MP (14 t-AML and 36 AML-MRC) were retrieved and compared to 42 dnMPAL cases and 100 sAML without MP (37 t-AML and 63 AML-MRC) (Table 1). The median age at diagnosis was 66 years which was higher than dnMPALs (44.5 yrs, p value Molecular studies (Table 2) revealed that the most commonly mutated genes in sAML-MP were RUNX1, DNMT3A, TP53 which were all present in a significantly higher frequency in comparison to dnMPAL. The frequency of RUNX1 mutation was higher in sAML-MP even in comparison to sAML without MP. Conversely, mutations in PHF6 frequently noted in dnMPALs were uncommon in sAML cohorts. Cluster analysis based on immunophenotyping of the 3 cohorts revealed 6 clusters (Fig 2) with separation between the 3 cohorts. Most of the dnMPAL with B/M phenotype formed a distinct tight cluster (cluster 3). Most of the sAML-MP were seen clustering together in cluster 4 whereas sAML without MP predominated in cluster 5 and 6. Cluster 1 revealed a mixture of dnMPAL and sAML-MP with predominantly T/M phenotype. The overall survival (OS) was inferior in the sAML-MP cohort in comparison to dnMPAL cohort (median survival: 6.77 vs 36.99 mths, p value 0.00005; Fig 3A) and was similar to the sAML without MP cohort (median survival: 6.77 vs 4.96 mths, p value 0.33; Fig 3A). In a multivariate analysis, model adjusted for the age and allotransplant did not explain difference in OS between these groups. Comparison of OS between the 6 clusters (Fig 3B) revealed better OS in clusters 2 and 3 that were enriched for dnMPAL. Cluster1 in spite of having a mixture of dnMPAL and sAML-MP did poorly. This could be driven by enrichment in DNMT3A mutations in the dnMPAL with T/M phenotype. Immunophenotypically distinct blast populations were flow cytometrically sorted in 4 cases of sAML-MP (2 each with T/M and B/M phenotype). While the majority of the genetic abnormalities were shared between populations with different lineages, a case with multiple mutations showed divergent KRAS/NRAS mutations showing clonal divergence as possible late event driving distinct lineage maturation (Table 3). CONCLUSION sAML-MP and sAML without MP are clinically similar and have biological overlaps with frequent somatic mutations in TP53, chromatin modifying genes and spliceosome-complex genes, which is different from dnMPAL. However, the frequency of RUNX1 mutation is higher in sAML-MP than sAML without MP suggesting its role in lineage infidelity. Though, cluster analysis based on immunophenotype separates the 3 cohorts into enriched clusters, there are still overlaps between sAML-MP and dnMPAL especially with a T/M phenotype as well as between sAML-MP and sAML without MP especially in TP53 mutated cases. Additional RNA seq and ATAC seq are currently being performed on flow cytometrically sorted distinct blast populations of the sAML-MP cases. Figure 1 Figure 1. Disclosures Galera: Paige.AI: Research Funding. Dogan: Physicians' Education Resource: Honoraria; Seattle Genetics: Consultancy; Takeda: Consultancy, Research Funding; EUSA Pharma: Consultancy; Roche: Consultancy, Research Funding; Peer View: Honoraria. Tallman: Syros: Membership on an entity's Board of Directors or advisory committees; NYU Grand Rounds: Honoraria; Kura: Membership on an entity's Board of Directors or advisory committees; Innate Pharma: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Biosight: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Membership on an entity's Board of Directors or advisory committees; Oncolyze: Membership on an entity's Board of Directors or advisory committees; KAHR: Membership on an entity's Board of Directors or advisory committees; Orsenix: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Rafael Pharmaceuticals: Research Funding; Glycomimetics: Research Funding; Biosight: Research Funding; Orsenix: Research Funding; Abbvie: Research Funding; Mayo Clinic: Honoraria; UC DAVIS: Honoraria; Northwell Grand Rounds: Honoraria; NYU Grand Rounds: Honoraria; Danbury Hospital Tumor Board: Honoraria; Acute Leukemia Forum: Honoraria; Miami Leukemia Symposium: Honoraria; New Orleans Cancer Symposium: Honoraria; ASH: Honoraria; NCCN: Honoraria. Levine: Celgene: Research Funding; Incyte: Consultancy; Roche: Honoraria, Research Funding; Morphosys: Consultancy; Imago: Membership on an entity's Board of Directors or advisory committees; Lilly: Honoraria; QIAGEN: Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy; Amgen: Honoraria; Gilead: Honoraria; Zentalis: Membership on an entity's Board of Directors or advisory committees; Ajax: Membership on an entity's Board of Directors or advisory committees; Prelude: Membership on an entity's Board of Directors or advisory committees; Isoplexis: Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Mission Bio: Membership on an entity's Board of Directors or advisory committees; Auron: Membership on an entity's Board of Directors or advisory committees. Roshal: Physicians' Education Resource: Other: Provision of services; Celgene: Other: Provision of services; Auron Therapeutics: Other: Ownership / Equity interests; Provision of services. Glass: GLG: Consultancy. Xiao: Stemline Therapeutics: Research Funding.

Introduction: Allogeneic stem cell transplantation (alloSCT) is the most effective post remission anti-leukemia strategy. However, the associated toxicity is a barrier for its routine adoption as standard of care in older adults. Studies of AML patients who underwent alloSCT are mainly retrospective as randomized controlled trials comparing transplantation to non-transplantation are very difficult to conduct. We herein present prospective data from a randomized controlled phase III study, E2906, designed to compare two intensive chemotherapy arms. Patients who achieved remission and had a donor were to proceed to alloSCT after induction or first consolidation, at investigator discretion. Non-transplanted patients received 2 cycles of consolidation and then underwent a second randomization between observation and decitabine maintenance. Herein are reported all patients who received alloSCT either on protocol during first remission (CR1) or at other post induction time-points. Patients and Methods: Enrolled to E2906 study were 727 AML patients age 60 years and over. AlloSCT was performed in 166 patients, of whom 71 received alloSCT as part of the study and 95 received alloSCT off protocol. 105 patients (66/71 on protocol, 39/95 off protocol) received alloSCT at CR1/CRi1/LFS1 (CR: 92, CRi: 9 LFS: 4). Patients were followed for a median of 33.6 months from diagnosis and 29.1 months from transplant. No patients received decitabine maintenance prior to alloSCT. Overall survival (OS) is defined as the time from allo transplant to death from any cause, with follow-up censored at the date of last contact. Disease free survival (DFS) is defined as the time from alloSCT transplant to relapse or death of any cause. The censored follow-up time for patients without relapse or death information is the date of last contact. Kaplan-Meier estimates were used to estimate OS and DFS. DFS and OS were compared between subgroups using log rank tests. A cumulative incidence analysis, with death without prior relapse as competing events, was performed to evaluate the subgroup effect on time to relapse after transplant. Results: Patient characteristics of those who received alloSCT at CR1/CRi1/LFS1 are similar to the general distribution of AML patients eligible for intensive chemotherapy. Median age was 66 years, 52% were male and 88% with ECOG PS of 0-1. Cytogenetic data were available for 85 patients of whom 26% presented with unfavorable cytogenetics. Minimal residual disease (MRD) status prior to alloSCT was available for 44 patients of whom 19 (43%) achieved a MRD negative state. Long-term OS and DFS rates for all 105 patients who underwent alloSCT at CR1/CRi1/LFS1 are encouraging (Figure 1). OS and DFS at two years were 56.4% and 53.6% and 49.4%, 45.6% and 42.9%, 39% at 3 and 4 years, respectively. Age above or below 65 years, gender, induction regimen (3+7 or clofarabine) and MRD status prior to transplantation had no impact on outcome. Survival curves by intermediate or adverse cytogenetic risk are not significantly different. The numbers of patients with CRi1 or LFS1 are too small to compare with patients in CR1. Nevertheless, 36/38 patients that were alive and remain in first remission at the time of data cutoff, are patients who achieved CR1 and only 2/38 are patients transplanted at CRi1 or LFS1. Notably, the non-relapse mortality (NRM) was not significantly different for patients less or over 65 years. NRM at 6 months and 2 years for patients over and under 65 years of age were 4.4%, 8.4% and 15.6%, 24.0% respectively. Relapse during the first year post alloSCT is the main barrier for longer survival, particularly in patients with adverse cytogenetics and MRD positivity prior to transplantation (Figure 2). Conclusions: Fit older patients, including those over age 65, who undergo an alloSCT in CR1, can expect a 4-year survival of 43% with an acceptable NRM rate. The NRM was, surprisingly, not higher than for a typically younger cohort of AML. Relapse within one year after transplant is the major limitation to prolonged survival and this finding reflects the biology of the leukemia in these patients. In this patient population, novel post remission strategies should either be complimentary to alloSCT or competitive with updated outcome of alloSCT. Figure 1 Figure 1. Disclosures Ofran: Medison Israel: Consultancy; Janssen: Consultancy; Pfizer: Consultancy; Astellas: Consultancy; AbbVie: Consultancy. Claxton: Astellas: Other: Clinical Trial; Novartis: Research Funding; Astex: Research Funding; Cyclacel: Research Funding; Daiichi Sankyo: Research Funding; Incyte: Research Funding. Tallman: Syros: Membership on an entity's Board of Directors or advisory committees; Kura: Membership on an entity's Board of Directors or advisory committees; NYU Grand Rounds: Honoraria; Innate Pharma: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Biosight: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Membership on an entity's Board of Directors or advisory committees; Oncolyze: Membership on an entity's Board of Directors or advisory committees; KAHR: Membership on an entity's Board of Directors or advisory committees; Orsenix: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Rafael Pharmaceuticals: Research Funding; Glycomimetics: Research Funding; Biosight: Research Funding; Orsenix: Research Funding; Abbvie: Research Funding; Mayo Clinic: Honoraria; UC DAVIS: Honoraria; Northwell Grand Rounds: Honoraria; NYU Grand Rounds: Honoraria; Danbury Hospital Tumor Board: Honoraria; Acute Leukemia Forum: Honoraria; Miami Leukemia Symposium: Honoraria; New Orleans Cancer Symposium: Honoraria; ASH: Honoraria; NCCN: Honoraria. Melnick: Constellation: Consultancy; Epizyme: Consultancy; Daiichi Sankyo: Research Funding; Sanofi: Research Funding; Janssen Pharmaceuticals: Research Funding; KDAC Pharma: Membership on an entity's Board of Directors or advisory committees. Levine: Prelude: Membership on an entity's Board of Directors or advisory committees; Ajax: Membership on an entity's Board of Directors or advisory committees; Zentalis: Membership on an entity's Board of Directors or advisory committees; Mission Bio: Membership on an entity's Board of Directors or advisory committees; QIAGEN: Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria; Lilly: Honoraria; Morphosys: Consultancy; Janssen: Consultancy; Incyte: Consultancy; Astellas: Consultancy; Imago: Membership on an entity's Board of Directors or advisory committees; Auron: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria; Celgene: Research Funding; Roche: Honoraria, Research Funding; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Isoplexis: Membership on an entity's Board of Directors or advisory committees. Lazarus: Bristol Myer Squibb: Membership on an entity's Board of Directors or advisory committees. Luger: Syros: Honoraria; Agios: Honoraria; Daiichi Sankyo: Honoraria; Jazz Pharmaceuticals: Honoraria; Brystol Myers Squibb: Honoraria; Acceleron: Honoraria; Astellas: Honoraria; Pfizer: Honoraria; Onconova: Research Funding; Celgene: Research Funding; Biosight: Research Funding; Hoffman LaRoche: Research Funding; Kura: Research Funding. Foran: gamida: Honoraria; aptose: Research Funding; syros: Honoraria; boehringer ingelheim: Research Funding; kura: Research Funding; takeda: Research Funding; abbvie: Research Funding; trillium: Research Funding; revolution medicine: Honoraria; certara: Honoraria; actinium: Research Funding; OncLive: Honoraria; pfizer: Honoraria; servier: Honoraria; bms: Honoraria; novartis: Honoraria; taiho: Honoraria; sanofi aventis: Honoraria; h3bioscience: Research Funding; aprea: Research Funding; sellas: Research Funding; stemline: Research Funding. Litzow: Jazz: Other: Advisory Board; AbbVie: Research Funding; Omeros: Other: Advisory Board; Amgen: Research Funding; Actinium: Research Funding; Astellas: Research Funding; Pluristem: Research Funding; Biosight: Other: Data monitoring committee.

Genomic studies in acute myeloid leukemia (AML) have generated a near complete catalogue of genes mutated at varying frequencies both across patients and in individual leukemias. The high variability of mutation burden within a given leukemia is suggestive of a stepwise evolutionary process composed of early, clonal, mutations and subsequent subclonal events. The receptor tyrosine kinase, FLT3, is the most commonly mutated gene in AML, with mutations frequently manifesting as internal tandem duplications (ITDs) in the juxtamembrane domain leading to constitutive kinase activation. Although FLT3 is commonly a subclonal mutational event, FLT3 ITD mutations portend a poor prognosis particularly when combined with DNMT3A and NPM1, earlier mutations that drive clonal expansion. Notwithstanding its role as a subclonal driver, previous preclinical FLT3 models have utilized retroviral overexpression or germline mutant expression at the endogenous locus precluding accurate temporal modeling of disease. These efforts have prohibited evaluation of FLT3 mutational acquisition in the context observed in AML patients. Here, we report the development of an endogenously targeted, Flp inducible, Flt3 ITD mouse allele which can be somatically activated subsequent to cooperating disease alleles. When activated with a tamoxifen inducible FlpoER, Flt3 mutant mice developed rapid leukocytosis peaking at 4-6 weeks post activation and resolving by 8-10 weeks, a finding not previously observed in constitutive models. This leukocytosis was disproportionately monocytic and accompanied by pronounced anemia and thrombocytopenia. Long term, these mice develop a myeloproliferative disease , reminiscent of previously reported constitutive alleles. In competitive transplantation studies, Flt3 mutant cells initiated disease and outcompeted wild-type cells. Despite this competitive advantage, disease was incapable of transplanting into secondary recipients. We further observed a non-cell autonomous depletion of SLAM+ LSKs suggesting the Flt3 mutant cells cannot propagate disease in self-renewing stem cells. To evaluate how this allele influenced leukemic evolution we crossed this Flt3 ITD allele to a Flp inducible Npm1 c mouse where a pulse of tamoxifen simultaneously activated both alleles. The combination of mutant Npm1 and Flt3 resulted in progressive leukocytosis which did not resolve. Within 6 weeks of mutational activation, these mice developed a lethal AML with robust anemia, thrombocytopenia, leukocytosis and expanded cKIT+ blasts in the blood. RNA-sequencing and immunophenotyping by CyTOF revealed distinct patterns of differentiation, gene-expression and downstream signaling.In an effort to model sequential mutational acquisition, we crossed the Flp Flt3 ITD allele to a Cre-inducible Dnmt3a R878H. Cre mRNA was electroporated into lineage negative bone marrow cells to activate the Dnmt3a R878H allele and transplanted into lethally irradiated recipients. Four weeks post engraftment, Flt3 ITD was activated with a pulse of tamoxifen. In contrast to the Flt3-Npm1 model, we observed an increase and subsequent decrease in WBC similar to the kinetics observed in Flt3 ITD only mice. However, by 20 weeks we observed a robust and consistent increase in WBC accompanied by an emergence of cKIT+ cells in the blood. Histopathology indicated that >50% of mice expressing both alleles in sequence developed AML marked by increased blasts in the marrow, with moderate anemia and thrombocytopenia compared to the Flt3-Npm1 models. Critically, in contrast to Flt3 ITD only mice, acquisition of the Flt3 ITD in Npm1 or Dnmt3a mutant HPSCs induced fully transplantable AML with immunophenotypic characteristics seen in human AML with these same genotypes. Collectively these results demonstrate that different co-occurring mutations are capable of transforming Flt3 ITD mutant cells, albeit with distinct latencies and mechanisms of cooperativity. In summary, our studies utilizing novel multi-recombinase models of leukemogenesis reveal new insights into the early phase of oncogene activation, and how cooperating alleles influence this response. This inducible Flt3 ITD allele represents a significant advance in modeling clonal evolution in myeloid malignancies and provides a critical isogenic platform for preclinical development of novel leukemia therapeutic regimens. Figure 1 Figure 1. Disclosures Bowman: Mission Bio: Honoraria, Speakers Bureau. Xiao: Stemline Therapeutics: Research Funding. Miles: Mission Bio: Honoraria, Speakers Bureau. Trowbridge: Fate Therapeutics: Patents & Royalties; H3 Biomedicine: Research Funding. Levine: Amgen: Honoraria; Lilly: Honoraria; Mission Bio: Membership on an entity's Board of Directors or advisory committees; Imago: Membership on an entity's Board of Directors or advisory committees; Celgene: Research Funding; Ajax: Membership on an entity's Board of Directors or advisory committees; QIAGEN: Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria; Zentalis: Membership on an entity's Board of Directors or advisory committees; Isoplexis: Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Research Funding; Janssen: Consultancy; Astellas: Consultancy; Morphosys: Consultancy; Incyte: Consultancy; Auron: Membership on an entity's Board of Directors or advisory committees; Prelude: Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees.

Introduction: The most common severe toxicity associated with chimeric antigen receptor T-cells targeting CD19 (CART19) is cytokine release syndrome (CRS; PMID: 29972754). Our group and others have published seminal observations on the biology of CRS through cytokine profiling, measuring a small number of analytes (PMID: 27076371, 33434058). Multiple biomarkers including interferon gamma (IFNG), IL-6, and IL-10 have been associated with the development of severe CRS in previous studies (PMID: 33434058). To date, the only biomarker predictive of the development of CRS prior to infusion has been disease burden. To obtain a more robust understanding of CRS biology, we performed comprehensive secretome profiling to measure more than 1400 serum analytes on serial serum samples collected from patients treated with the 41BB-containing CTL019 on two clinical trials. Methods: Serum from patients enrolled on two clinical trials of the CART19 product CTL019 (NCT01626495 & NCT02906371) were obtained serially from pre-infusion to one month post infusion. Patients were categorised as having "minimal" (no CRS, Grade 1, or Grade 2) or "severe" (Grade 3 or 4) CRS. The serum secretome was profiled using the Olink Explore 1536 Analysis platform (Olink, Upsala, Sweden). 1484 proteins were measured from serum via proximity extension assay (PEA) high-multiplex immunoassay. Differential expression analysis, correlation analyses and receiver operating characteristic (ROC) calculations were performed using R (version 4.0.4) in RStudio. Significance was based on a fold change of greater than 2 or less than -2 and a false discovery rate of less than 0.05 calculated using a Benjamini-Hochberg correction. Results: 26 patients (10 NCT01626495 & 16 NCT02906371) were included comprising 128 unique datapoints from baseline to 35 days post-infusion. Thirteen patients had minimal and 13 had severe CRS. Differentially expressed proteins between minimal and severe CRS at the peak timepoint are shown in (A; green represents IFNG responsive proteins). Not surprisingly, proteins involved in IL-6 and IFNG signalling were increased, including biomarkers of hemophagocytic lymphohistiocytosis (HLH) such as VSIG4, CXCL9, CXCL10, CD163. The IL-18 signalling axis was dysregulated at peak CRS in severe patients with markedly elevated IL18 and IL18BP, despite prior reports suggesting IL-18 up-regulation is unique to the late CRS seen with CART22 (PMID: 32925169). Soluble markers of checkpoint inhibition, including soluble PDL1 (CD274) and LAG3 were also highly elevated. Finally, biomarkers of endothelial damage, such as PLAT, TMSB10 and CALCA were significantly elevated in patients with severe CRS. Pathway analysis revealed significant dysregulation in targetable cytokine, chemokine, and signalling pathways (B). A volcano plot of differentially expressed proteins at pre-infusion (C) identified a single protein, MILR1, as a candidate biomarker that was highly differentially expressed in patients who would subsequently develop severe CRS. MILR1 expression decreased over time (D). An ROC of MILR1 as a predictor for development of severe CRS (E) demonstrated pre-infusion elevated MILR1 could accurately predict development of severe CRS (sensitivity 88%, specificity 97%, AUC=0.977). We identified correlates of MILR1 at pre-infusion and found that MILR1 correlated most highly with soluble FLT3 (R=0.86, p Conclusions: With comprehensive secretome profiling we made multiple novel insights into the biology of CRS after CART19 and identified several potentially targetable proteins and pathways that could mitigate severe CRS. Similar secretome profiling in patients who developed neurotoxicity will also be shown. We identified two novel pre-infusion biomarkers that demonstrate significant capacity to predict the development of severe CRS following CART19 infusion. The inverse relationship apparent between FLT3 and FLT3LG that persists over time is an important finding that implies a potential biological role for FLT3/FLT3 ligand in the development of severe CRS. Mechanistic studies exploring the role of MILR1 and FLT3 in the initiation of CRS are ongoing. Figure 1 Figure 1. Disclosures Lambert: Novartis, shionogi, argenx, Rigel, octapharma: Consultancy; Rigel, Novartis, Sysmex, octapharma: Research Funding. Bassiri: Kriya Therapeutics: Consultancy, Current holder of individual stocks in a privately-held company; Guidepoint Global: Consultancy. Levine: Vycellix: Membership on an entity's Board of Directors or advisory committees; Tmunity Therapeutics: Other: Co-Founder and equity holder; Ori Biotech: Membership on an entity's Board of Directors or advisory committees; Immusoft: Membership on an entity's Board of Directors or advisory committees; Immuneel: Membership on an entity's Board of Directors or advisory committees; Avectas: Membership on an entity's Board of Directors or advisory committees; Akron: Membership on an entity's Board of Directors or advisory committees; In8bio: Membership on an entity's Board of Directors or advisory committees. Maude: Wugen: Consultancy; Novartis Pharmaceuticals Corporation: Consultancy, Research Funding. June: Tmunity, DeCART, BluesphereBio, Carisma, Cellares, Celldex, Cabaletta, Poseida, Verismo, Ziopharm: Current equity holder in publicly-traded company; Novartis: Patents & Royalties; AC Immune, DeCART, BluesphereBio, Carisma, Cellares, Celldex, Cabaletta, Poseida, Verismo, Ziopharm: Consultancy. Barrett: Tmunity Therapeutics: Current Employment. Grupp: Novartis, Kite, Vertex, and Servier: Research Funding; Novartis, Roche, GSK, Humanigen, CBMG, Eureka, and Janssen/JnJ: Consultancy; Novartis, Adaptimmune, TCR2, Cellectis, Juno, Vertex, Allogene and Cabaletta: Other: Study steering committees or scientific advisory boards; Jazz Pharmaceuticals: Consultancy, Other: Steering committee, Research Funding. Teachey: Janssen: Consultancy; NeoImmune Tech: Research Funding; Sobi: Consultancy; BEAM Therapeutics: Consultancy, Research Funding.