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Allogeneic cellular immunotherapies hold promise for broad clinical implementation but face limitations due to potential rejection of donor cells by the host immune system. Silencing of beta-2 microglobulin (B2M) expression is commonly employed to evade T cell-mediated rejection by the host, although the absence of B2M is expected to trigger missing-self responses by host natural killer (NK) cells. Here, we demonstrate that genetic deletion of the adhesion ligands CD54 and CD58 in B2M-deficient chimeric antigen receptor (CAR) T cells and multi-edited induced pluripotent stem cell (iPSC)-derived CAR NK cells reduces their susceptibility to rejection by host NK cells in vitro and in vivo. The absence of adhesion ligands limits rejection in a unidirectional manner in B2M-deficient and B2M-sufficient settings without affecting the antitumor functionality of the engineered donor cells. Thus, these data suggest that genetic ablation of adhesion ligands effectively alleviates rejection by host immune cells, facilitating the implementation of universal immunotherapy., Competing Interests: Declaration of interests K.-J.M. is a consultant and has research support from Fate Therapeutics. K.-J.M. has research support from Oncopeptides. K.-J.M. and Q.H. are consultants at Vycellix. All relationships have been approved by Oslo University Hospital, the University of Oslo, and the Karolinska Institute. R.M.M., E.V., Y.P., M.J., B.G., R.A., T.L., A.W., J.P.G., and B.V. are employees at Fate Therapeutics. Memorial Sloan Kettering has licensed IP (unrelated to this study) to Fate Therapeutics, and M.S. receives research support (for unrelated studies) from Fate Therapeutics., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Allogeneic cellular immunotherapies hold a great promise for cancer treatment owing to their potential cost-effectiveness, scalability and on-demand availability. However, immune rejection of adoptively transferred allogeneic T and natural killer (NK) cells is a substantial obstacle to achieving clinical responses that are comparable to responses obtained with current autologous chimeric antigen receptor T cell therapies. In this Perspective, we discuss strategies to confer cell-intrinsic, immune-evasive properties to allogeneic T cells and NK cells in order to prevent or delay their immune rejection, thereby widening the therapeutic window. We discuss how common viral and cancer immune escape mechanisms can serve as a blueprint for improving the persistence of off-the-shelf allogeneic cell therapies. The prospects of harnessing genome editing and synthetic biology to design cell-based precision immunotherapies extend beyond programming target specificities and require careful consideration of innate and adaptive responses in the recipient that may curtail the biodistribution, in vivo expansion and persistence of cellular therapeutics., (© 2024. Springer Nature Limited.)

The tumor microenvironment presents many obstacles to effective chimeric antigen receptor (CAR) T cell therapy, including glucose competition from tumor and myeloid cells. Using mouse models of acute lymphoblastic leukemia (ALL), renal cell carcinoma (RCC), and glioblastoma (GBM), we show that enforced expression of the glucose transporter GLUT1 enhances anti-tumor efficacy and promotes favorable CAR-T cell phenotypes for two clinically relevant CAR designs, 19-28z and IL13Rα2-BBz. In the NALM6 ALL model, 19-28z-GLUT1 promotes T stem cell-like memory formation and prolongs survival. RNA sequencing of these CAR-T cells reveals that the overexpression of GLUT1, but not GLUT3, enriches for genes involved in glycolysis, mitochondrial respiration, and memory precursor phenotypes. Extending these data, 19-28z-GLUT1 CAR-T cells improve tumor control and response to rechallenge in an RCC patient-derived xenograft model. Furthermore, IL13Rα2-BBz CAR-T cells overexpressing GLUT1 prolong the survival of mice bearing orthotopic GBMs and exhibit decreased exhaustion markers. This novel engineering approach can offer a competitive advantage to CAR-T cells in harsh tumor environments where glucose is limiting., Competing Interests: Declaration of interests The authors declare no financial competing interests. M.S. reports research funding from Takeda Pharmaceuticals, Atara Biotherapeutics, Fate Therapeutics, and Mnemo Therapeutics, all of which are unrelated to the present research. M.S. is a scientific cofounder of Mnemo Therapeutics., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

We designed a CD19-targeted CAR comprising a calibrated signaling module, termed 1XX, that differs from that of conventional CD28/CD3z and 4-1BB/CD3z CARs. Here we report the first-in-human, phase 1 clinical trial of 19(T2)28z-1XX CAR T cells in relapsed/refractory large B-cell lymphoma. We hypothesized that 1XX CAR T cells may be effective at low doses and investigated 4 doubling dose levels starting from 25×10 6 CAR T cells. The overall response rate (ORR) was 82% and complete response (CR) rate 71% in the entire cohort (n=28) and 88% ORR and 75% CR in 16 patients treated at 25×10 6 . With the median follow-up of 24 months, the 1-year EFS was 61% (95% CI: 45-82%). Overall, grade ≥3 CRS and ICANS rates were low at 4% and 7%. The calibrated potency of the 1XX CAR affords excellent efficacy at low cell doses and may benefit the treatment of other hematological malignancies, solid tumors and autoimmunity., Competing Interests: Competing Interests J.H.P. received consulting fees from Affyimmune Therapeutics, Amgen, Autolus, Be Biopharma, Beigene, Bright Pharmaceutical Services, Inc., Caribou Biosciences, Curocell, Galapagos, In8Bio, Kite, Medpace, Minerva Biotechnologies, Pfizer, Servier, Sobi, and Takeda; received honoraria from OncLive, Physician Education Resource, and MJH Life Sciences; serves on scientific advisory board of Allogene Therapeutics, Artiva Biotherapeutics and Green Cross Biopharma; and received institutional research funding from Autolus, Genentech, Fate Therapeutics, InCyte, Servier, and Takeda. M.L.P. received consulting fees from BMS, Cellectis, Kite, Mustang Bio and Synthekine. G.Sh. received research funding to the institution from Amgen, BMS, Beyond Spring, Janssen, and GPCR, and serves on DSMB for Arcellx. P.D. received consulting fees from Kite. M.Sc. served as a paid consultant for McKinsey & Company, Angiocrine Bioscience, Inc., Kite, and Omeros Corporation; received research funding from Angiocrine Bioscience, Inc., Omeros Corporation, and Amgen, Inc.; served on ad hoc advisory boards for Kite-A Gilead company; and received honoraria from i3Health, Medscape and CancerNetwork for CME-related activity. G.Sa. has received financial compensations for participating in advisory boards or consulting for Abbvie, Atbtherapeutics, Beigene, BMS, Debiopharm Genentech/Roche, Genmab, Incyte, Ipsen, Janssen, Kite/Gilead, Loxo/Lilly, Merck, Molecular Partners, Nordic Nanovector, Novartis, Nurix and Orna. He is also a shareholder of: Owkin; he received Research Support managed by his institution from Genentech, Janssen, Ipsen, Nurix. Y.K.V. received one time consulting fee from EastRx. J.M.S. is an inventor on 1XX related IP. I.R. reports grants from Takeda Pharmaceuticals and Atara, personal fees from Mnemo Therapeutics, Akron, the Centre for Commercialization of Cancer, and Oribiotech, and other support from Bristol Myers Squibb outside the submitted work. M. S. reports grants from Atara Biotherapeutics outside the submitted work, as well as patent 8389282 issued and licensed to Juno Therapeutics, patent 11242375 issued and licensed to Atara Biotherapeutics, patent 10370452 issued, licensed, and with royalties paid from Fate Therapeutics, patent 11377637 issued and licensed to Takeda Pharmaceuticals, patent 11377637 issued and licensed to Mnemo Therapeutics, and patent 11377637 issued and licensed to Minerva Biotechnologies. MSK has licensed 1XX technology to Atara Biotherapy, Fate Therapeutics, Minerva Therapeutics, and Takeda Pharmaceuticals.

Chimeric antigen receptor T cells have dramatically improved the treatment of hematologic malignancies. T cell antigen receptor (TCR)-based cell therapies are yet to achieve comparable outcomes. Importantly, chimeric antigen receptors not only target selected antigens but also reprogram T cell functions through the co-stimulatory pathways that they engage upon antigen recognition. We show here that a fusion receptor comprising the CD80 ectodomain and the 4-1BB cytoplasmic domain, termed 80BB, acts as both a ligand and a receptor to engage the CD28 and 4-1BB pathways, thereby increasing the antitumor potency of human leukocyte antigen-independent TCR (HIT) receptor- or TCR-engineered T cells and tumor-infiltrating lymphocytes. Furthermore, 80BB serves as a switch receptor that provides agonistic 4-1BB co-stimulation upon its ligation by the inhibitory CTLA4 molecule. By combining multiple co-stimulatory features in a single antigen-agnostic synthetic receptor, 80BB is a promising tool to sustain CD3-dependent T cell responses in a wide range of targeted immunotherapies., (© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Senescent cells, which accumulate in organisms over time, contribute to age-related tissue decline. Genetic ablation of senescent cells can ameliorate various age-related pathologies, including metabolic dysfunction and decreased physical fitness. While small-molecule drugs that eliminate senescent cells ('senolytics') partially replicate these phenotypes, they require continuous administration. We have developed a senolytic therapy based on chimeric antigen receptor (CAR) T cells targeting the senescence-associated protein urokinase plasminogen activator receptor (uPAR), and we previously showed these can safely eliminate senescent cells in young animals. We now show that uPAR-positive senescent cells accumulate during aging and that they can be safely targeted with senolytic CAR T cells. Treatment with anti-uPAR CAR T cells improves exercise capacity in physiological aging, and it ameliorates metabolic dysfunction (for example, improving glucose tolerance) in aged mice and in mice on a high-fat diet. Importantly, a single administration of these senolytic CAR T cells is sufficient to achieve long-term therapeutic and preventive effects., (© 2024. The Author(s).)

Suboptimal functional persistence limits the efficacy of adoptive T-cell therapies. CD28-based chimeric antigen receptors (CAR) impart potent effector function to T cells but with a limited lifespan. We show here that the genetic disruption of SUV39H1, which encodes a histone-3, lysine-9 methyl-transferase, enhances the early expansion, long-term persistence, and overall antitumor efficacy of human CAR T cells in leukemia and prostate cancer models. Persisting SUV39H1-edited CAR T cells demonstrate improved expansion and tumor rejection upon multiple rechallenges. Transcriptional and genome accessibility profiling of repeatedly challenged CAR T cells shows improved expression and accessibility of memory transcription factors in SUV39H1-edited CAR T cells. SUV39H1 editing also reduces expression of inhibitory receptors and limits exhaustion in CAR T cells that have undergone multiple rechallenges. Our findings thus demonstrate the potential of epigenetic programming of CAR T cells to balance their function and persistence for improved adoptive cell therapies., Significance: T cells engineered with CD28-based CARs possess robust effector function and antigen sensitivity but are hampered by limited persistence, which may result in tumor relapse. We report an epigenetic strategy involving disruption of the SUV39H1-mediated histone-silencing program that promotes the functional persistence of CD28-based CAR T cells. See related article by López-Cobo et al., p. 120. This article is featured in Selected Articles from This Issue, p. 5., (©2023 American Association for Cancer Research.)

Acute myeloid leukemia (AML) poses a singular challenge for chimeric antigen receptor (CAR) therapy owing to its phenotypic heterogeneity and similarity to normal hematopoietic stem/progenitor cells (HSPCs). Here we expound a CAR strategy intended to efficiently target AML while minimizing HSPC toxicity. Quantification of target expression in relapsed/refractory patient samples and normal HSPCs reveals a therapeutic window for gated co-targeting of ADGRE2 and CLEC12A: We combine an attenuated ADGRE2-CAR with a CLEC12A-chimeric costimulatory receptor (ADCLEC.syn1) to preferentially engage ADGRE2 pos CLEC12A pos leukemic stem cells over ADGRE2 low CLEC12A neg normal HSPCs. ADCLEC.syn1 prevents antigen escape in AML xenograft models, outperforms the ADGRE2-CAR alone and eradicates AML despite proximate myelopoiesis in humanized mice. Off-target HSPC toxicity is similar to that of a CD19-CAR and can be mitigated by reducing CAR T cell-derived interferon-γ. Overall, we demonstrate the ability of target density-adapted cooperative CAR targeting to selectively eliminate AML and potentially obviate the need for hematopoietic rescue., Competing Interests: Declaration of interests Memorial Sloan Kettering has submitted a patent application (WO2022232016A2) based in part on results presented in this manuscript (M.Sa., S.H., and J.M.-S. are listed among the inventors). M.Sa. and S.H. report research support and research funding from Takeda Pharmaceuticals related to the present research. M.Sa. reports research funding from Atara Biotherapeutics, Fate Therapeutics, and Mnemo Therapeutics unrelated to the present research. M.Sa. and I.R. are scientific cofounders of Mnemo Therapeutics. I.R. reports research funding from Atara Biotherapeutics, Takeda Pharmaceuticals; ownership/equity interests at Fate Therapeutics and Mnemo Therapeutics; intellectual property rights at Juno Therapeutics. K.F., M.R.N., and I.R. report employment at Takeda Pharmaceuticals. M.Su. declares the following competing interests: Novartis: Consultancy, Research Funding; Janssen: Consultancy; Seattle Genetics: Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria; Kite/Gilead: Consultancy, Honoraria, Research Funding; Roche AG: Consultancy, Research Funding. J.H.P. declares the following competing interest: research funding support from Takeda Pharmaceuticals, Fate Therapeutics, Genentech, InCyte and Servier; Consultancy from Amgen, Autolus, BMS, Curocel, Kite, Legend Biotech, Minerva, Pfizer, Servier, Sobi, and Takeda Pharmaceuticals; and serves on Scientific Advisory Board of Allogene, Artiva Biotherapeutics, and GC Cell Corporation., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Although in vivo engraftment, expansion, and persistence of chimeric antigen receptor (CAR) T cells are pivotal components of treatment efficacy, quantitative monitoring has not been implemented in routine clinical practice. We describe the development and analytical validation of a digital PCR assay for ultrasensitive detection of CAR constructs after treatment, circumventing known technical limitations of low-partitioning platforms. Primers and probes, designed for detection of axicabtagene, brexucabtagene, and Memorial Sloan Kettering CAR constructs, were employed to validate testing on the Bio-Rad digital PCR low-partitioning platform; results were compared with Raindrop, a high-partitioning system, as reference method. Bio-Rad protocols were modified to enable testing of DNA inputs as high as 500 ng. Using dual-input reactions (20 and 500 ng) and a combined analysis approach, the assay demonstrated consistent target detection around 1 × 10 -5 (0.001%) with excellent specificity and reproducibility and 100% accuracy compared with the reference method. Dedicated analysis of 53 clinical samples received during validation/implementation phases showed the assay effectively enabled monitoring across multiple time points of early expansion (day 6 to 28) and long-term persistence (up to 479 days). CAR vectors were detected at levels ranging from 0.005% to 74% (vector versus reference gene copies). The highest levels observed in our cohort correlated strongly with the temporal diagnosis of grade 2 and 3 cytokine release syndrome diagnosis (P < 0.005). Only three patients with undetectable constructs had disease progression at the time of sampling., (Copyright © 2023 Association for Molecular Pathology and American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

In preclinical models, anakinra, an IL-1 receptor antagonist (IL-1Ra), reduced immune effector cell-associated neurotoxicity syndrome (ICANS) without compromising anti-CD19 chimeric antigen receptor (CAR) T-cell efficacy. We initiated a phase 2 clinical trial of anakinra in patients with relapsed/refractory large B-cell lymphoma and mantle cell lymphoma treated with commercial anti-CD19 CAR T-cell therapy. Here we report a non-prespecified interim analysis reporting the final results from cohort 1 in which patients received subcutaneous anakinra from day 2 until at least day 10 post-CAR T-cell infusion. The primary endpoint was the rate of severe (grade ≥3) ICANS. Key secondary endpoints included the rates of all-grade cytokine release syndrome (CRS) and ICANS and overall disease response. Among 31 treated patients, 74% received axicabtagene ciloleucel, 13% received brexucabtagene ciloleucel and 4% received tisagenlecleucel. All-grade ICANS occurred in 19%, and severe ICANS occurred in 9.7% of patients. There were no grade 4 or 5 ICANS events. All-grade CRS occurred in 74%, and severe CRS occurred in 6.4% of patients. The overall disease response rate was 77% with 65% complete response rate. These initial results show that prophylactic anakinra resulted in a low incidence of ICANS in patients with lymphoma receiving anti-CD19 CAR T-cell therapy and support further study of anakinra in immune-related neurotoxicity syndromes., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Cell therapies that rely on engineered immune cells can be enhanced by achieving uniform and controlled transgene expression in order to maximize T-cell function and achieve predictable patient responses. Although they are effective, current genetic engineering strategies that use γ-retroviral, lentiviral, and transposon-based vectors to integrate transgenes, unavoidably produce variegated transgene expression in addition to posing a risk of insertional mutagenesis. In the setting of chimeric antigen receptor (CAR) therapy, inconsistent and random CAR expression may result in tonic signaling, T-cell exhaustion, and variable T-cell persistence. Here, we report and validate an algorithm for the identification of extragenic genomic safe harbors (GSH) that can be efficiently targeted for DNA integration and can support sustained and predictable CAR expression in human peripheral blood T cells. The algorithm is based on 7 criteria established to minimize genotoxicity by directing transgene integration away from functionally important genomic elements, maximize efficient CRISPR/Cas9-mediated targeting, and avert transgene silencing over time. T cells engineered to express a CD19 CAR at GSH6, which meets all 7 criteria, are curative at low cell dose in a mouse model of acute lymphoblastic leukemia, matching the potency of CAR T cells engineered at the TRAC locus and effectively resisting tumor rechallenge 100 days after their infusion. The identification of functional extragenic GSHs thus expands the human genome available for therapeutic precision engineering., (© 2023 by The American Society of Hematology.)

Pancreatic ductal adenocarcinoma (PDAC) is lethal in 88% of patients 1 , yet harbours mutation-derived T cell neoantigens that are suitable for vaccines 2,3 . Here in a phase I trial of adjuvant autogene cevumeran, an individualized neoantigen vaccine based on uridine mRNA-lipoplex nanoparticles, we synthesized mRNA neoantigen vaccines in real time from surgically resected PDAC tumours. After surgery, we sequentially administered atezolizumab (an anti-PD-L1 immunotherapy), autogene cevumeran (a maximum of 20 neoantigens per patient) and a modified version of a four-drug chemotherapy regimen (mFOLFIRINOX, comprising folinic acid, fluorouracil, irinotecan and oxaliplatin). The end points included vaccine-induced neoantigen-specific T cells by high-threshold assays, 18-month recurrence-free survival and oncologic feasibility. We treated 16 patients with atezolizumab and autogene cevumeran, then 15 patients with mFOLFIRINOX. Autogene cevumeran was administered within 3 days of benchmarked times, was tolerable and induced de novo high-magnitude neoantigen-specific T cells in 8 out of 16 patients, with half targeting more than one vaccine neoantigen. Using a new mathematical strategy to track T cell clones (CloneTrack) and functional assays, we found that vaccine-expanded T cells comprised up to 10% of all blood T cells, re-expanded with a vaccine booster and included long-lived polyfunctional neoantigen-specific effector CD8 + T cells. At 18-month median follow-up, patients with vaccine-expanded T cells (responders) had a longer median recurrence-free survival (not reached) compared with patients without vaccine-expanded T cells (non-responders; 13.4 months, P = 0.003). Differences in the immune fitness of the patients did not confound this correlation, as responders and non-responders mounted equivalent immunity to a concurrent unrelated mRNA vaccine against SARS-CoV-2. Thus, adjuvant atezolizumab, autogene cevumeran and mFOLFIRINOX induces substantial T cell activity that may correlate with delayed PDAC recurrence., (© 2023. The Author(s).)

The success of chimeric antigen receptor (CAR) T cells targeting B-cell malignancies propelled the field of synthetic immunology and raised hopes to treat solid tumors in a similar fashion. Antigen escape and the paucity of tumor-restricted CAR targets are recognized challenges to fulfilling this prospect. Recent advances in CAR T cell engineering extend the toolbox of chimeric receptors available to calibrate antigen sensitivity and combine receptors to create adapted tumor-sensing T cells. Emerging engineering strategies to lower the threshold for effective antigen recognition, when needed, and enable composite antigen recognition hold great promise for overcoming tumor heterogeneity and curbing off-tumor toxicities., Significance: Improving the clinical efficacy of CAR T cell therapies will require engineering T cells that overcome heterogeneous and low-abundance target expression while minimizing reactivity to normal tissues. Recent advances in CAR design and logic gating are poised to extend the success of CAR T cell therapies beyond B-cell malignancies., (©2023 The Authors; Published by the American Association for Cancer Research.)

Chimeric antigen receptor (CAR) T cell therapy relies on T cells that are guided by synthetic receptors to target and lyse cancer cells. CARs bind to cell surface antigens through an scFv (binder), the affinity of which is central to determining CAR T cell function and therapeutic success. CAR T cells targeting CD19 were the first to achieve marked clinical responses in patients with relapsed/refractory B cell malignancies and to be approved by the U.S. Food and Drug Administration (FDA). We report cryo-EM structures of CD19 antigen with the binder FMC63, which is used in four FDA-approved CAR T cell therapies (Kymriah, Yescarta, Tecartus, and Breyanzi), and the binder SJ25C1, which has also been used extensively in multiple clinical trials. We used these structures for molecular dynamics simulations, which guided creation of lower- or higher-affinity binders, and ultimately produced CAR T cells endowed with distinct tumor recognition sensitivities. The CAR T cells exhibited different antigen density requirements to trigger cytolysis and differed in their propensity to prompt trogocytosis upon contacting tumor cells. Our work shows how structural information can be applied to tune CAR T cell performance to specific target antigen densities.

Further advances in cell engineering are needed to increase the efficacy of chimeric antigen receptor (CAR) and other T cell-based therapies 1-5 . As T cell differentiation and functional states are associated with distinct epigenetic profiles 6,7 , we hypothesized that epigenetic programming may provide a means to improve CAR T cell performance. Targeting the gene that encodes the epigenetic regulator ten-eleven translocation 2 (TET2) 8 presents an interesting opportunity as its loss may enhance T cell memory 9,10 , albeit not cause malignancy 9,11,12 . Here we show that disruption of TET2 enhances T cell-mediated tumour rejection in leukaemia and prostate cancer models. However, loss of TET2 also enables antigen-independent CAR T cell clonal expansions that may eventually result in prominent systemic tissue infiltration. These clonal proliferations require biallelic TET2 disruption and sustained expression of the AP-1 factor BATF3 to drive a MYC-dependent proliferative program. This proliferative state is associated with reduced effector function that differs from both canonical T cell memory 13,14 and exhaustion 15,16 states, and is prone to the acquisition of secondary somatic mutations, establishing TET2 as a guardian against BATF3-induced CAR T cell proliferation and ensuing genomic instability. Our findings illustrate the potential of epigenetic programming to enhance T cell immunity but highlight the risk of unleashing unchecked proliferative responses., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

Improved identification of anti-tumor T cells is needed to advance cancer immunotherapies. CD39 expression is a promising surrogate of tumor-reactive CD8 + T cells. Here, we comprehensively profiled CD39 expression in human lung cancer. CD39 expression enriched for CD8 + T cells with features of exhaustion, tumor reactivity, and clonal expansion. Flow cytometry of 440 lung cancer biospecimens revealed weak association between CD39 + CD8 + T cells and tumoral features, such as programmed death-ligand 1 (PD-L1), tumor mutation burden, and driver mutations. Immune checkpoint blockade (ICB), but not cytotoxic chemotherapy, increased intratumoral CD39 + CD8 + T cells. Higher baseline frequency of CD39 + CD8 + T cells conferred improved clinical outcomes from ICB therapy. Furthermore, a gene signature of CD39 + CD8 + T cells predicted benefit from ICB, but not chemotherapy, in a phase III clinical trial of non-small cell lung cancer. These findings highlight CD39 as a proxy of tumor-reactive CD8 + T cells in human lung cancer., Competing Interests: Declaration of interests C.A.K. received research funding support from Kite/Gilead and Intima Bioscience; is on the Scientific and/or Clinical Advisory Boards of Achilles Therapeutics, Aleta BioTherapeutics, Bellicum Pharmaceuticals, Catamaran Bio, Obsidian Therapeutics, and T-knife; and has performed consulting services for Bristol Myers Squibb, PACT Pharma, and Roche/Genentech. C.A.K. is a co-inventor on patent applications related to TCRs targeting public neoantigens unrelated to the current work. M.D.H. received a research grant from BMS; personal fees from Achilles, Arcus, AstraZeneca, Blueprint, BMS, Genentech/Roche, Genzyme, Immunai, Instil Bio, Janssen, Merck, Mirati, Natera, Nektar, Pact Pharma, Regeneron, Shattuck Labs, and Syndax; and equity options from Arcus, Factorial, Immunai, and Shattuck Labs. A patent filed by MSKCC related to the use of tumor mutational burden to predict response to immunotherapy (PCT/US2015/062208) is pending and licensed by PGDx. J.D.W. is a consultant for Amgen, Apricity, Ascentage Pharma, Astellas, AstraZeneca, Bicara Therapeutics, Boehringer Ingelheim, Bristol Myers Squibb, CellCarta, Chugai, Daiichi Sankyo, Dragonfly, Georgiamune, Idera, Imvaq, Larkspur, Maverick Therapeutics, Merck, Psioxus, Recepta, Tizona, Trishula, Sellas, Surface Oncology, and Werewolf Therapeutics. J.D.W. receives grant/research support from Bristol Myers Squibb and Sephora. J.D.W. has equity in Apricity, Arsenal IO, Ascentage, Beigene, Imvaq, Linneaus, Georgiamune, Maverick, Tizona Pharmaceuticals, and Trieza. J.D.W. is a co-inventor on the following patent application: xenogeneic (canine) DNA vaccines, myeloid-derived suppressor cell (MDSC) assay, anti-PD1 antibody, anti-CTLA4 antibodies, anti-GITR antibodies and methods of use thereof, Newcastle disease viruses for cancer therapy, and prediction of responsiveness to treatment with immunomodulatory therapeutics and method of monitoring abscopal effects during such treatment. J.D.W. and T.M. are co-inventors on patent applications related to CD40 and in situ vaccination (PCT/US2016/045970). T.M. is a consultant for Immunos Therapeutics and Pfizer. T.M. is a cofounder of and equity holder in IMVAQ Therapeutics. T.M. receives research funding from Bristol Myers Squibb, Surface Oncology, Kyn Therapeutics, Infinity Pharmaceuticals, Peregrine Pharmaceuticals, Adaptive Biotechnologies, Leap Therapeutics, and Aprea Therapeutics. T.M. is an inventor on patent applications related to work on oncolytic viral therapy, alpha virus-based vaccine, neoantigen modeling, CD40, GITR, OX40, PD-1, and CTLA-4. C.M.R. has consulted regarding oncology drug development with AbbVie, Amgen, Ascentage, AstraZeneca, BMS, Celgene, Daiichi Sankyo, Genentech/Roche, Ipsen, Loxo, and PharmaMar and is on the scientific advisory boards of Elucida, Bridge, and Harpoon. B.Y.N. and X.G. are employees and stockholders of Genentech/Roche., (Copyright © 2022 Elsevier Inc. All rights reserved.)

The production of autologous T cells expressing a chimaeric antigen receptor (CAR) is time-consuming, costly and occasionally unsuccessful. T-cell-derived induced pluripotent stem cells (TiPS) are a promising source for the generation of 'off-the-shelf' CAR T cells, but the in vitro differentiation of TiPS often yields T cells with suboptimal features. Here we show that the premature expression of the T-cell receptor (TCR) or a constitutively expressed CAR in TiPS promotes the acquisition of an innate phenotype, which can be averted by disabling the TCR and relying on the CAR to drive differentiation. Delaying CAR expression and calibrating its signalling strength in TiPS enabled the generation of human TCR - CD8αβ + CAR T cells that perform similarly to CD8αβ + CAR T cells from peripheral blood, achieving effective tumour control on systemic administration in a mouse model of leukaemia and without causing graft-versus-host disease. Driving T-cell maturation in TiPS in the absence of a TCR by taking advantage of a CAR may facilitate the large-scale development of potent allogeneic CD8αβ + T cells for a broad range of immunotherapies., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

The globin genes are archetypal tissue-specific genes that are silent in most tissues but for late-stage erythroblasts upon terminal erythroid differentiation. The transcriptional activation of the β-globin gene is under the control of proximal and distal regulatory elements located on chromosome 11p15.4, including the β-globin locus control region (LCR). The incorporation of selected LCR elements in lentiviral vectors encoding β and β-like globin genes has enabled successful genetic treatment of the β-thalassemias and sickle cell disease. However, recent occurrences of benign clonal expansions in thalassemic patients and myelodysplastic syndrome in patients with sickle cell disease call attention to the non-erythroid functions of these powerful vectors. Here we demonstrate that lentivirally encoded LCR elements, in particular HS1 and HS2, can be activated in early hematopoietic cells including hematopoietic stem cells and myeloid progenitors. This activity is position-dependent and results in the transcriptional activation of a nearby reporter gene in these progenitor cell populations. We further show that flanking a globin vector with an insulator can effectively restrain this non-erythroid activity without impairing therapeutic globin expression. Globin lentiviral vectors harboring powerful LCR HS elements may thus expose to the risk of trans-activating cancer-related genes, which can be mitigated by a suitable insulator., Competing Interests: Declaration of interests The authors declare no financial competing interests. A patent application based in part on results presented in this manuscript has been submitted by MSKCC., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Cancer immunoediting 1 is a hallmark of cancer 2 that predicts that lymphocytes kill more immunogenic cancer cells to cause less immunogenic clones to dominate a population. Although proven in mice 1,3 , whether immunoediting occurs naturally in human cancers remains unclear. Here, to address this, we investigate how 70 human pancreatic cancers evolved over 10 years. We find that, despite having more time to accumulate mutations, rare long-term survivors of pancreatic cancer who have stronger T cell activity in primary tumours develop genetically less heterogeneous recurrent tumours with fewer immunogenic mutations (neoantigens). To quantify whether immunoediting underlies these observations, we infer that a neoantigen is immunogenic (high-quality) by two features-'non-selfness'  based on neoantigen similarity to known antigens 4,5 , and 'selfness'  based on the antigenic distance required for a neoantigen to differentially bind to the MHC or activate a T cell compared with its wild-type peptide. Using these features, we estimate cancer clone fitness as the aggregate cost of T cells recognizing high-quality neoantigens offset by gains from oncogenic mutations. With this model, we predict the clonal evolution of tumours to reveal that long-term survivors of pancreatic cancer develop recurrent tumours with fewer high-quality neoantigens. Thus, we submit evidence that that the human immune system naturally edits neoantigens. Furthermore, we present a model to predict how immune pressure induces cancer cell populations to evolve over time. More broadly, our results argue that the immune system fundamentally surveils host genetic changes to suppress cancer., (© 2022. The Author(s).)

Anti-CD19 chimeric antigen receptor (CAR) T cell therapy has led to unprecedented responses in patients with high-risk hematologic malignancies. However, up to 60% of patients still experience disease relapse and up to 80% of patients experience CAR-mediated toxicities, such as cytokine release syndrome or immune effector cell-associated neurotoxicity syndrome. We investigated the role of the intestinal microbiome on these outcomes in a multicenter study of patients with B cell lymphoma and leukemia. We found in a retrospective cohort (n = 228) that exposure to antibiotics, in particular piperacillin/tazobactam, meropenem and imipenem/cilastatin (P-I-M), in the 4 weeks before therapy was associated with worse survival and increased neurotoxicity. In stool samples from a prospective cohort of CAR T cell recipients (n = 48), the fecal microbiome was altered at baseline compared to healthy controls. Stool sample profiling by 16S ribosomal RNA and metagenomic shotgun sequencing revealed that clinical outcomes were associated with differences in specific bacterial taxa and metabolic pathways. Through both untargeted and hypothesis-driven analysis of 16S sequencing data, we identified species within the class Clostridia that were associated with day 100 complete response. We concluded that changes in the intestinal microbiome are associated with clinical outcomes after anti-CD19 CAR T cell therapy in patients with B cell malignancies., (© 2022. This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply.)

Chimeric antigen receptors (CARs) are receptors for antigen that direct potent immune responses. Tumor escape associated with low target antigen expression is emerging as one potential limitation of their efficacy. Here we edit the TRAC locus in human peripheral blood T cells to engage cell-surface targets through their T cell receptor-CD3 complex reconfigured to utilize the same immunoglobulin heavy and light chains as a matched CAR. We demonstrate that these HLA-independent T cell receptors (HIT receptors) consistently afford high antigen sensitivity and mediate tumor recognition beyond what CD28-based CARs, the most sensitive design to date, can provide. We demonstrate that the functional persistence of HIT T cells can be augmented by constitutive coexpression of CD80 and 4-1BBL. Finally, we validate the increased antigen sensitivity afforded by HIT receptors in xenograft mouse models of B cell leukemia and acute myeloid leukemia, targeting CD19 and CD70, respectively. Overall, HIT receptors are well suited for targeting cell surface antigens of low abundance., (© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.)

A paradigm shift has recently occurred in the field of cancer therapeutics. Traditional anticancer agents, such as chemotherapy, radiotherapy and small-molecule drugs targeting specific signalling pathways, have been joined by cellular immunotherapies based on T cell engineering. The rapid adoption of novel, patient-specific cellular therapies builds on scientific developments in tumour immunology, genetic engineering and cell manufacturing, best illustrated by the curative potential of chimeric antigen receptor (CAR) T cell therapy targeting CD19-expressing malignancies. However, the clinical benefit observed in many patients may come at a cost. In up to one-third of patients, significant toxicities occur that are directly associated with the induction of powerful immune effector responses. The most frequently observed immune-mediated toxicities are cytokine release syndrome and immune effector cell-associated neurotoxicity syndrome. This Review discusses our current understanding of their pathophysiology and clinical features, as well as the development of novel therapeutics for their prevention and/or management., (© 2021. Springer Nature Limited.)

Background: Waldenström macroglobulinemia (WM) is an incurable disease and, while treatable, can develop resistance to available therapies and be fatal. Chimeric antigen receptor (CAR) T cell therapy directed against the CD19 antigen has demonstrated efficacy in relapsed or refractory B lymphoid malignancies, and is now approved for B cell acute lymphoblastic leukemia and certain B cell lymphomas. However, CAR T therapy has not been evaluated for use in WM., Methods and Results: We performed preclinical studies demonstrating CAR T cell activity against WM cells in vitro, and developed an in vivo murine model of WM which demonstrated prolonged survival with use of CAR T therapy. We then report the first three patients with multiply relapsed and refractory WM treated for their disease with CD19-directed CAR T cells on clinical trials. Treatment was well tolerated, and observed toxicities were consistent with those seen in CAR T treatment for other diseases, and no grade 3 or higher cytokine release syndrome or neurotoxicity events occurred. All three patients attained at least a clinical response to treatment, including one minimal residual disease-negative complete response, though all three eventually developed recurrent disease between 3 and 26 months after initial treatment., Conclusions: This report summarizes preclinical and clinical activity of CD19-directed CAR T therapy in WM, demonstrating early tolerability and efficacy in patients with WM, and representing a possible treatment option in patients with heavily pretreated and relapsed or refractory WM. Larger studies evaluating CAR T therapy in WM are warranted, along with further evaluation into mechanisms of resistance to CAR T therapy., Competing Interests: Competing interests: MLP has received consulting fees from BeiGene, Novartis and Synthekine. An immediate family member of MLP is a Board Member with DKMS, has IP rights with Juno Therapeutics and Seres Therapeutics, and receives consulting fees from Rheos Medicines, Ceramedix, GKS, Priotera, and WindMIL Therapeutics. MR has received consulting fees from Celgene and Physicians’ Education Resource, and consulting fees plus equity interest in Auron Therapeutics. IR has received consulting fees and funding support from Juno Therapeutics, in which she had an equity interest. RJB has licensed intellectual property to and collect royalties from BMS, Caribou and Sanofi. RJB received research funding from BMS. RJB is a consultant to BMS and was a consultant for Gracell Biotechnologies but ended employment in the past 24 months. JHP received consulting fees from Amgen, Allogene, Artiva Biotherapeutics, Autolus, BMS, Curocell, GSK, Incyte, Innate Pharma, Intellia Therapeutics, Kite Pharma, Kura Oncology, Minerva Biotechnologies, Novartis, Pfizer and Servier, and serves as a DSMB member for Affyimmune and Bright Pharmaceuticals. ELS received IP or royalties from BMS, Sanofi, and is a consultant to BMS, Novartis, and Chimeric Therapeutics. DQ, SM, SS, AD, BS, XW, and MS have no conflicts of interest to disclose., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

β-Thalassemias are inherited anemias that are caused by the absent or insufficient production of the β chain of hemoglobin. Here we report 6-8-year follow-up of four adult patients with transfusion-dependent β-thalassemia who were infused with autologous CD34 + cells transduced with the TNS9.3.55 lentiviral globin vector after reduced-intensity conditioning (RIC) in a phase 1 clinical trial ( NCT01639690) . Patients were monitored for insertional mutagenesis and the generation of a replication-competent lentivirus (safety and tolerability of the infusion product after RIC-primary endpoint) and engraftment of genetically modified autologous CD34 + cells, expression of the transduced β-globin gene and post-transplant transfusion requirements (efficacy-secondary endpoint). No unexpected safety issues occurred during conditioning and cell product infusion. Hematopoietic gene marking was very stable but low, reducing transfusion requirements in two patients, albeit not achieving transfusion independence. Our findings suggest that non-myeloablative conditioning can achieve durable stem cell engraftment but underscore a minimum CD34 + cell transduction requirement for effective therapy. Moderate clonal expansions were associated with integrations near cancer-related genes, suggestive of non-erythroid activity of globin vectors in stem/progenitor cells. These correlative findings highlight the necessity of cautiously monitoring patients harboring globin vectors., (© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Despite the high remission rates achieved using T cells bearing a chimeric antigen receptor (CAR) against hematogical malignancies, there is still a considerable proportion of patients who eventually experience tumor relapse. Clinical studies have established that mechanisms of treatment failure include the down-regulation of target antigen expression and the limited persistence of effective CAR T cells. We hypothesized that dual targeting mediated by a CAR and a chimeric costimulatory receptor (CCR) could simultaneously enhance T cell cytotoxicity and improve durability. Concomitant high-affinity engagement of a CD38-binding CCR enhanced the cytotoxicity of BCMA-CAR and CD19-CAR T cells by increasing their functional binding avidity. In comparison to second-generation BCMA-CAR or CD19-CAR T cells, double-targeted CAR + CD38-CCR T cells exhibited increased sensitivity to recognize and lyse tumor variants of multiple myeloma and acute lymphoblastic leukemia with low antigen density in vitro. In addition, complimentary costimulation by 4-1BB and CD28 endodomains provided by the CAR and CCR combination conferred increased cytokine secretion and expansion and improved persistence in vivo. The cumulatively improved properties of CAR + CCR T cells enabled the in vivo eradication of antigen-low tumor clones, which were otherwise resistant to treatment with conventional CAR T cells. Therefore, multiplexing targeting and costimulation through the combination of a CAR and a CCR is a powerful strategy to improve the clinical outcomes of CAR T cells by enhancing cytotoxic efficacy and persistence, thus preventing relapses of tumor clones with low target antigen density.

Malignant pleural diseases, comprising metastatic lung and breast cancers and malignant pleural mesothelioma (MPM), are aggressive solid tumors with poor therapeutic response. We developed and conducted a first-in-human, phase I study of regionally delivered, autologous, mesothelin-targeted chimeric antigen receptor (CAR) T-cell therapy. Intrapleural administration of 0.3M to 60M CAR T cells/kg in 27 patients (25 with MPM) was safe and well tolerated. CAR T cells were detected in peripheral blood for >100 days in 39% of patients. Following our demonstration that PD-1 blockade enhances CAR T-cell function in mice, 18 patients with MPM also received pembrolizumab safely. Among those patients, median overall survival from CAR T-cell infusion was 23.9 months (1-year overall survival, 83%). Stable disease was sustained for ≥6 months in 8 patients; 2 exhibited complete metabolic response on PET scan. Combination immunotherapy with CAR T cells and PD-1 blockade agents should be further evaluated in patients with solid tumors. SIGNIFICANCE: Regional delivery of mesothelin-targeted CAR T-cell therapy followed by pembrolizumab administration is feasible, safe, and demonstrates evidence of antitumor efficacy in patients with malignant pleural diseases. Our data support the investigation of combination immunotherapy with CAR T cells and PD-1 blockade agents in solid tumors. See related commentary by Aldea et al., p. 2674 . This article is highlighted in the In This Issue feature, p. 2659 ., (©2021 American Association for Cancer Research.)

Adoptive T cell therapy (ACT) is a promising strategy for treating cancer, but it often fails because of cell intrinsic regulatory programs that limit the degree or duration of T cell function. In this study, we found that ectopic expression of microRNA-200c (miR-200c) markedly enhanced the antitumor activity of CD8 + cytotoxic T lymphocytes (CTLs) during ACT in multiple mouse models. CTLs transduced with miR-200c exhibited reduced apoptosis during engraftment and enhanced in vivo persistence, accompanied by up-regulation of the transcriptional regulator T cell factor 1 (TCF1) and the inflammatory cytokine tumor necrosis factor (TNF). miR-200c elicited these changes by suppressing the transcription factor Zeb1 and thereby inducing genes characteristic of epithelial cells. Overexpression of one of these genes, Epcam , was sufficient to augment therapeutic T cell responses against both solid and liquid tumors. These results identify the miR-200c–EpCAM axis as an avenue for improving ACT and demonstrate that select genetic perturbations can produce phenotypically distinct T cells with advantageous therapeutic properties.

Spearheaded by the therapeutic use of chimeric antigen receptors (CARs) targeting CD19, synthetic immunology has entered the clinical arena. CARs are recombinant receptors for antigen that engage cell surface molecules through the variable region of an antibody and signal through arrayed T-cell activating and costimulatory domains. CARs allow redirection of T-cell cytotoxicity against any antigen of choice, independent of MHC expression. Patient T cells engineered to express CARs specific for CD19 have yielded remarkable outcomes in subjects with relapsed/refractory B- cell malignancies, setting off unprecedented interest in T-cell engineering and cell-based cancer immunotherapy. In this review, we present the challenges to extend the use of CAR T cells to solid tumors and other pathologies. We further highlight progress in CAR design, cell manufacturing, and genome editing, which in aggregate hold the promise of generating safer and more effective genetically instructed immunity. Novel engineered cell types, including innate T-cell types, natural killer (NK) cells, macrophages, and induced pluripotent stem cell-derived immune cells, are on the horizon, as are applications of CAR T cells to treat autoimmunity, severe infections, and senescence-associated pathologies., (© 2021 Wiley-VCH GmbH.)

Background: Adequate CD34+ collection efficiency (CE) is critical to achieve target CD34+ cell doses in hematopoietic progenitor cell (HPC) collections. Autologous HPC collection in sickle cell disease (SCD) is associated with unstable collection interfaces and low CD34+ CEs. We hypothesized that variables specific to SCD, activation of blood cells and elevated viscosity, might contribute to these issues and made adjustments to the collection process and procedure to address our hypothesis., Study Design and Methods: In two patients with SCD undergoing autologous HPC collection on our clinical trial (NCT02193191), we therefore implemented adjustments to the process and procedure in the following areas: proximity of RBC exchange to HPC collection, the type of anticoagulation, and the packing factor setting., Results: There was no collection interface instability. Our CD34+ CE1s were high at 70% and 51%, and granulocyte CE, platelet CE, and product granulocyte % were remarkably low. Product hematocrits were not as high as previously reported to be required to obtain adequate CEs. Interestingly, one HPC product showed a hemoglobin S (HbS) of 91% at the same time that the peripheral blood (PB) showed a HbS of 22%., Discussion: Adjustments to the HPC collection process and procedure were associated with adequate CD34+ CEs and low granulocyte and platelet contamination in HPC products from SCD patients. Given the discrepancy in the percentage of sickle RBCs in the product versus the PB, we hypothesize that CD34+ cells and RBCs may aggregate. Our interventions and hypothesis should be further investigated in larger studies., (© 2021 AABB.)

We present data from our study of plerixafor mobilization (NCT02193191) relevant to the question of whether further dose escalation of plerixafor can address inconsistent adequacy of CD34+ mobilization for gene therapy of sickle cell disease (SCD). We found that, in the same patient, higher plerixafor dosing was associated with higher fold increases in PB CD34+ count, but not necessarily higher absolute CD34+ counts. Variation in pre-apheresis absolute CD34+ counts was related to intra-individual variation in baseline PB CD34+ counts and inter-individual variation in responsiveness to plerixafor. Overall, our results support further studies of continued dose escalation of plerixafor for autologous HPC collection in SCD., (Copyright © 2021 Elsevier Inc. All rights reserved.)

CD19-targeted chimeric antigen receptor (CAR) T-cell therapy has become a breakthrough treatment of patients with relapsed/refractory B-cell acute lymphoblastic leukemia (B-ALL). However, despite the high initial response rate, the majority of adult patients with B-ALL progress after CD19 CAR T-cell therapy. Data on the natural history, management, and outcome of adult B-ALL progressing after CD19 CAR T cells have not been described in detail. Herein, we report comprehensive data of 38 adult patients with B-ALL who progressed after CD19 CAR T therapy at our institution. The median time to progression after CAR T-cell therapy was 5.5 months. Median survival after post-CAR T progression was 7.5 months. A high disease burden at the time of CAR T-cell infusion was significantly associated with risk of post-CAR T progression. Thirty patients (79%) received salvage treatment of post-CAR T disease progression, and 13 patients (43%) achieved complete remission (CR), but remission duration was short. Notably, 7 (58.3%) of 12 patients achieved CR after blinatumomab and/or inotuzumab administered following post-CAR T failure. Multivariate analysis revealed that a longer remission duration from CAR T cells was associated with superior survival after progression following CAR T-cell therapy. In summary, overall prognosis of adult B-ALL patients progressing after CD19 CAR T cells was poor, although a subset of patients achieved sustained remissions to salvage treatments, including blinatumomab, inotuzumab, and reinfusion of CAR T cells. Novel therapeutic strategies are needed to reduce risk of progression after CAR T-cell therapy and improve outcomes of these patients., (© 2021 by The American Society of Hematology.)

T cell genome editing holds great promise to advance a range of immunotherapies but is encumbered by the dependence on difficult-to-produce and expensive viral vectors. Here, small double-stranded plasmid DNA modified to mediate high-efficiency homologous recombination is designed. The resulting chimeric antigen receptor (CAR)-T cells display a similar phenotype, transcriptional profile, and in vivo potency to CAR-T cells generated using adeno-associated viral vector. This method should simplify and accelerate the use of precision engineering to produce edited T cells for research and clinical purposes., (© 2021 Wiley-VCH GmbH.)

T cells engineered to express chimeric antigen receptors (CARs) specific for CD19 have yielded remarkable clinical outcomes in patients with refractory B-cell malignancies. The first CARs to be approved by the US Food and Drug Administration and the European Medicines Agency are CD19 CARs that comprise either CD28/CD3ζ or 4-1BB/CD3ζ dual-signalling domains. While their efficacy and safety profiles in patients with B-cell malignancies are comparable overall, the functional properties these two CAR designs impart upon engineered T cells differ significantly. Remarkably, alternative costimulatory domains have not, to date, superseded these foundational designs. Rather, recent CAR advances have focused on perfecting the original CD28- and 4-1BB-based CD19 CARs by calibrating strength of activation, pre-empting T-cell exhaustion and increasing the functional persistence of CAR T cells. This article reviews the essential biological properties of these first-in-class prototypes and their recent evolution., (© 2020 The Authors.)

Radiotherapy is potentially an important salvage strategy post-chimeric antigen receptor T cell therapy (CART), but limited data exist. We reviewed 14 patients treated with salvage radiation post-CART progression (SRT). Most received SRT for first post-CART relapse (71%) to sites previously PET-avid pre-CART (79%). Median overall survival (OS) post-SRT was 10 months. Post-SRT, six localized relapses achieved 100% response (3 = complete, 3 = partial), with improved freedom from subsequent relapse (P = 0·001) and OS (P = 0·004) compared to advanced stage relapses. Three were bridged to allogeneic transplantation; at analysis, all were alive/NED. SRT has diverse utility and can integrate with novel agents or transplantation to attempt durable remissions., (© 2020 British Society for Haematology and John Wiley & Sons Ltd.)

Cellular senescence is characterized by stable cell-cycle arrest and a secretory program that modulates the tissue microenvironment 1,2 . Physiologically, senescence serves as a tumour-suppressive mechanism that prevents the expansion of premalignant cells 3,4 and has a beneficial role in wound-healing responses 5,6 . Pathologically, the aberrant accumulation of senescent cells generates an inflammatory milieu that leads to chronic tissue damage and contributes to diseases such as liver and lung fibrosis, atherosclerosis, diabetes and osteoarthritis 1,7 . Accordingly, eliminating senescent cells from damaged tissues in mice ameliorates the symptoms of these pathologies and even promotes longevity 1,2,8-10 . Here we test the therapeutic concept that chimeric antigen receptor (CAR) T cells that target senescent cells can be effective senolytic agents. We identify the urokinase-type plasminogen activator receptor (uPAR) 11 as a cell-surface protein that is broadly induced during senescence and show that uPAR-specific CAR T cells efficiently ablate senescent cells in vitro and in vivo. CAR T cells that target uPAR extend the survival of mice with lung adenocarcinoma that are treated with a senescence-inducing combination of drugs, and restore tissue homeostasis in mice in which liver fibrosis is induced chemically or by diet. These results establish the therapeutic potential of senolytic CAR T cells for senescence-associated diseases.

Chimeric antigen receptor (CAR) T cells have demonstrated clinical benefit in patients with relapsed/refractory (R/R) B-cell acute lymphoblastic leukemia (B-ALL). We undertook a multicenter clinical trial to determine toxicity, feasibility, and response for this therapy. A total of 25 pediatric/young adult patients (age, 1-22.5 years) with R/R B-ALL were treated with 19-28z CAR T cells. Conditioning chemotherapy included high-dose (3 g/m2) cyclophosphamide (HD-Cy) for 17 patients and low-dose (≤1.5 g/m2) cyclophosphamide (LD-Cy) for 8 patients. Fifteen patients had pretreatment minimal residual disease (MRD; <5% blasts in bone marrow), and 10 patients had pretreatment morphologic evidence of disease (≥5% blasts in bone marrow). All toxicities were reversible, including severe cytokine release syndrome in 16% (4 of 25) and severe neurotoxicity in 28% (7 of 25) of patients. Treated patients were assessed for response, and, among the evaluable patients (n = 24), response and peak CAR T-cell expansion were superior in the HD-Cy/MRD cohorts, as compared with the LD-Cy/morphologic cohorts without an increase in toxicity. Our data support the safety of CD19-specific CAR T-cell therapy for R/R B-ALL. Our data also suggest that dose intensity of conditioning chemotherapy and minimal pretreatment disease burden have a positive impact on response without a negative effect on toxicity. This trial was registered at www.clinicaltrials.gov as #NCT01860937., (© 2019 by The American Society of Hematology.)

On August 30, 2017, the U.S. Food and Drug Administration (US-FDA) approved tisagenlecleucel (KYMRIAH, Novartis, Basel, Switzerland), a synthetic bioimmune product of anti-CD19 chimeric antigen receptor-T cells (CAR-T), for the treatment of children and young adults with relapsed/refractory B-cell acute lymphoblastic leukemia (B-ALL). With this new era of personalized cancer immunotherapy, multiple challenges are present ranging from implementation of a CAR-T program to safe delivery of the drug, long-term toxicity monitoring and disease assessments. To address these issues, experts representing the American Society for Blood and Marrow Transplant (ASBMT), the European Group for Blood and Marrow Transplantation (EBMT), the International Society of Cell and Gene Therapy (ISCT), and the Foundation for the Accreditation of Cellular Therapy (FACT), formed a global CAR-T task force to identify and address key questions pertinent for hematologists and transplant physicians regarding the clinical use of anti CD19 CAR-T therapy in patients with B-ALL. This article presents an initial roadmap for navigating common clinical practice scenarios that will become more prevalent now that the first commercially available CAR-T product for B-ALL has been approved.