Final analysis of survival outcomes in the randomized phase 3 FIRST trial Blood (IF 13.164) Pub Date : 2017-01-01 Thierry Facon; Meletios A. Dimopoulos; Angela Dispenzieri; John V. Catalano; Andrew Belch; Michele Cavo; Antonello Pinto; Katja Weisel; Heinz Ludwig; Nizar J. Bahlis; Anne Banos; Mourad Tiab; Michel Delforge; Jamie D. Cavenagh; Catarina Geraldes; Je-Jung Lee; Christine Chen; Albert Oriol; Javier De La Rubia; Darell White; Daniel Binder; Jin Lu; Kenneth C. Anderson; Philippe Moreau; Michel Attal; Aurore Perrot; Bertrand Arnulf; Lugui Qiu; Murielle Roussel; Eileen Boyle; Salomon Manier; Mohamad Mohty; Herve Avet-Loiseau; Xavier Leleu; Annette Ervin-Haynes; Guang Chen; Vanessa Houck; Lotfi Benboubker; Cyrille Hulin
This FIRST trial final analysis examined survival outcomes in patients with transplant-ineligible newly diagnosed multiple myeloma (NDMM) treated with lenalidomide and low-dose dexamethasone until disease progression (Rd continuous), Rd for 72 weeks (18 cycles; Rd18), or melphalan, prednisone, and thalidomide (MPT; 72 weeks). The primary endpoint was progression-free survival (PFS; primary comparison: Rd continuous vs MPT). Overall survival (OS) was a key secondary endpoint (final analysis prespecified ≥ 60 months’ follow-up). Patients were randomized to Rd continuous (n = 535), Rd18 (n = 541), or MPT (n = 547). At a median follow-up of 67 months, PFS was significantly longer with Rd continuous vs MPT (HR, 0.69; 96% CI, 0.59-0.79; P < .00001) and was similarly extended vs Rd18. Median OS was 10 months longer with Rd continuous vs MPT (59.1 vs 49.1 months; HR, 0.78; 95% CI, 0.63-0.95; P = .0144), and similar with Rd18 (62.3 months). In patients achieving complete or very good partial responses, Rd continuous had a ≈ 30-month-longer median time to next treatment vs Rd18 (69.5 vs 39.9 months). Over half of all patients who received second-line treatment were given a bortezomib-based therapy. Second-line outcomes were improved in patients receiving bortezomib after Rd continuous and Rd18 vs after MPT. No new safety concerns, including risk for secondary malignancies, were observed. Treatment with Rd continuous significantly improved survival outcomes vs MPT, supporting Rd continuous as a standard of care for patients with transplant-ineligible NDMM. Study registration is at Clinicaltrials.gov (NCT00689936) and EudraCT (2007-004823-39).
A landscape of germline mutations in a cohort of inherited bone marrow failure patients Blood (IF 13.164) Pub Date : 2017-01-01 Olivier Bluteau; Marie Sebert; Thierry Leblanc; Régis Peffault de Latour; Samuel Quentin; Elodie Lainey; Lucie Hernandez; Jean-Hugues Dalle; Flore Sicre de Fontbrune; Etienne Lengline; Raphael Itzykson; Emmanuelle Clappier; Nicolas Boissel; Naddia Vasquez; Mélanie Da Costa; Julien Masliah-Planchon; Wendy Cuccuini; Anna Raimbault; Louis De Jaegere; Lionel Adès; Pierre Fenaux; Sébastien Maury; Claudine Schmitt; Marc Muller; Carine Domenech; Nicolas Blin; Bénédicte Bruno; Isabelle Pellier; Mathilde Hunault; Stéphane Blanche; Arnaud Petit; Guy Leverger; Gérard Michel; Yves Bertrand; André Baruchel; Gérard Socié; Jean Soulier
Bone marrow failure (BMF) in children and young adults is often suspected to be inherited, but in many cases diagnosis remains uncertain. We studied a cohort of 179 patients (from 173 families) with BMF of suspected inherited origin but unresolved diagnosis after medical evaluation and Fanconi anemia exclusion. All patients had cytopenias, and 12.0% presented ≥5% bone marrow blast cells. Median age at genetic evaluation was 11 years; 20.7% of patients were aged ≤2 years and 36.9% were ≥18 years. We analyzed genomic DNA from skin fibroblasts using whole-exome sequencing, and were able to assign a causal or likely causal germline mutation in 86 patients (48.0%), involving a total of 28 genes. These included genes in familial hematopoietic disorders (GATA2, RUNX1), telomeropathies (TERC, TERT, RTEL1), ribosome disorders (SBDS, DNAJC21, RPL5), and DNA repair deficiency (LIG4). Many patients had an atypical presentation, and the mutated gene was often not clinically suspected. We also found mutations in genes seldom reported in IBMF, such as SAMD9 and SAMD9L (N=16 of the 86 patients; 18.6%), MECOM/EVI1 (N=6, 7.0%), and ERCC6L2 (N=7, 8.1%), each of which was associated with a distinct natural history; SAMD9 and SAMD9L patients often experienced transient aplasia and monosomy 7, while MECOM patients presented early-onset severe aplastic anemia, and ERCC6L2 patients, mild pancytopenia with myelodysplasia. This study broadens the molecular and clinical portrait of IBMF syndromes and sheds light on newly-recognized disease entities. Using a high-throughput sequencing screen to implement precision medicine at diagnosis can improve patient management and family counseling.
Physiological Srsf2 P95H expression causes impaired hematopoietic stem cell functions and aberrant RNA splicing in mice Blood (IF 13.164) Pub Date : 2017-01-01 Ayana Kon; Satoshi Yamazaki; Yasuhito Nannya; Keisuke Kataoka; Yasunori Ota; Masahiro Marshall Nakagawa; Kenichi Yoshida; Yusuke Shiozawa; Maiko Morita; Tetsuichi Yoshizato; Masashi Sanada; Manabu Nakayama; Haruhiko Koseki; Hiromitsu Nakauchi; Seishi Ogawa
Splicing factor (SF) mutations are characteristic of myelodysplastic syndromes (MDS) and related myeloid neoplasms and implicated in their pathogenesis, but their roles in the development of MDS have not fully been elucidated. Here, we investigated the consequence of mutant Srsf2 expression using newly generated Vav1-Cre-mediated conditional knock-in mice. Mice carrying a heterozygous Srsf2 P95H mutation showed significantly reduced numbers of hematopoietic stem and progenitor cells (HSPCs) and differentiation defects both in the steady-state condition and transplant settings. Srsf2-mutated hematopoietic stem cells (HSCs) showed impaired long-term reconstitution compared to controls in competitive repopulation assays. Although the Srsf2 mutant mice did not develop MDS under the steady-state condition, when their stem cells were transplanted into lethally irradiated mice, the recipients developed anemia, leukopenia and erythroid dysplasia, suggesting the role of replicative stress in the development of an MDS-like phenotype in Srsf2-mutated mice. RNA sequencing of the Srsf2-mutated HSPCs revealed a number of abnormal splicing events and differentially expressed genes, including several potential targets implicated in the pathogenesis of hematopoietic malignancies, such as Csf3r, Fyn, Gnas, Nsd1, Hnrnpa2b1 and Trp53bp1. Among the mutant Srsf2-associated splicing events, most commonly observed were the enhanced inclusion and/or exclusion of cassette exons, which were caused by the altered consensus motifs for the recognition of exonic splicing enhancers. Our findings suggest that the mutant Srsf2 leads to a compromised HSC function by causing abnormal RNA splicing and expression, contributing to the deregulated hematopoiesis that recapitulates the MDS phenotypes, possibly as a result of additional genetic and/or environmental insults.
Miss Piggy on the catwalk again Blood (IF 13.164) Pub Date : 2017-11-16 Anna Rita Migliaccio
In this issue of Blood , by generating a novel dual oxidase 2 ( DUOX2 ) mutation responsible for congenital hypothyroidism (CH) in pigs, [Zhang et al] identify Krüppel-like factor 9 (KLF9) as the mediator for the regulation exerted by the thyroid gland on hematopoiesis. The domestic pig
Relapsed/refractory HL: FDG-PET is the trump card Blood (IF 13.164) Pub Date : 2017-11-16 Andrea Gallamini
A number of disease- or host-related markers have been proposed to predict treatment outcome in relapsed/refractory Hodgkin lymphoma (HL). In this issue of Blood , [Moskowitz et al] demonstrate that fluorodeoxyglucose (FDG)-positron emission tomography (PET)/computed tomography (CT) performed in
Mutant RUNX1 and histone tales Blood (IF 13.164) Pub Date : 2017-11-16 Lucy A. Godley
In this issue of Blood , [Lehnertz et al] identify rare cases of histone H3 mutations at position K27 that alter methylation levels and cooperate with RUNX1 mutations, further strengthening the association between RUNX1 and polycomb repressive complex 2 (PRC2) activity in leukemic
Subphenotypes of sickle cell disease in Africa Blood (IF 13.164) Pub Date : 2017-11-16 Frédéric B. Piel; Thomas N. Williams
In this issue of Blood , [Dubert et al] present the results of a large cohort study conducted across 3 sub-Saharan African countries (Mali, Cameroon, and Ivory Coast) to quantify differences between subphenotypes of sickle cell disease (SCD) based on markers of anemia, hemolysis, and vascular
Platelet CLEC-2: a molecule with 2 faces Blood (IF 13.164) Pub Date : 2017-11-16 Lijun Xia
In this issue of Blood , [Haining et al] reveal an unexpected role for platelet C-type lectin-like receptor-2 (CLEC-2) in thrombosis, which is independent of its hemi-immunoreceptor tyrosine-based activation motif (hemITAM) signaling. ![Figure] Model showing a hemITAM-dependent
Thyroid hormone regulates hematopoiesis via the TR-KLF9 axis Blood (IF 13.164) Pub Date : 2017-11-16 Ying Zhang; Yuanyuan Xue; Chunwei Cao; Jiaojiao Huang; Qianlong Hong; Tang Hai; Qitao Jia; Xianlong Wang; Guosong Qin; Jing Yao; Xiao Wang; Qiantao Zheng; Rui Zhang; Yongshun Li; Ailing Luo; Nan Zhang; Guizhi Shi; Yanfang Wang; Hao Ying; Zhonghua Liu; Hongmei Wang; Anming Meng; Qi Zhou; Hong Wei; Feng Liu; Jianguo Zhao
Congenital hypothyroidism (CH) is one of the most prevalent endocrine diseases, for which the underlying mechanisms remain unknown; it is often accompanied by anemia and immunodeficiency in patients. Here, we created a severe CH model together with anemia and T lymphopenia to mimic the clinical features of hypothyroid patients by ethylnitrosourea (ENU) mutagenesis in Bama miniature pigs. A novel recessive c.1226A>G transition of the dual oxidase 2 (DUOX2) gene was identified as the causative mutation. This mutation hindered the production of hydrogen peroxide (H2O2) and thus contributed to thyroid hormone (TH) synthesis failure. Transcriptome sequencing analysis of the thymuses showed that Krüppel-like factor 9 (KLF9) was predominantly downregulated in hypothyroid mutants. KLF9 was verified to be directly regulated by TH in a TH receptor (TR)-dependent manner both in vivo and in vitro. Furthermore, knockdown of klf9 in zebrafish embryos impaired hematopoietic development including erythroid maturation and T lymphopoiesis. Our findings suggest that the TR-KLF9 axis is responsible for the hematopoietic dysfunction and might be exploited for the development of novel therapeutic interventions for thyroid diseases.
Procoagulant platelets: generation, function, and therapeutic targeting in thrombosis Blood (IF 13.164) Pub Date : 2017-11-16 Ejaife O. Agbani; Alastair W. Poole
Current understanding of how platelets localize coagulation to wound sites has come mainly from studies of a subpopulation of activated platelets. In this review, we summarize data from the last 4 decades that have described these platelets with a range of descriptive titles and attributes. We identify striking overlaps in the reported characteristics of these platelets, which imply a single subpopulation of versatile platelets and thus suggest that their commonality requires unification of their description. We therefore propose the term procoagulant platelet as the unifying terminology. We discuss the agonist requirements and molecular drivers for the dramatic morphological transformation platelets undergo when becoming procoagulant. Finally, we provide perspectives on the biomarker potential of procoagulant platelets for thrombotic events as well as on the possible clinical benefits of inhibitors of carbonic anhydrase enzymes and the water channel Aquaporin-1 for targeting this subpopulation of platelets as antiprocoagulant antithrombotics.
Role of anthracycline and comprehensive geriatric assessment for elderly patients with diffuse large B-cell lymphoma Blood (IF 13.164) Pub Date : 2017-11-16 Richard J. Lin; Madhusmita Behera; Catherine S. Diefenbach; Christopher R. Flowers
Survival outcome for elderly patients with newly diagnosed diffuse large B-cell lymphoma remains suboptimal in the rituximab era. In this systematic review, we summarize available evidence relevant to the inclusion of anthracycline in upfront chemoimmunotherapy for these elderly patients and highlight the need of prospective clinical trials. With limited prospective data, we find that pretreatment comprehensive geriatric assessment accurately predicts survival and treatment-related toxicities, suggesting its potential role in guiding overall treatment decision-making.
Rituximab-based first-line treatment of cGVHD after allogeneic SCT: results of a phase 2 study Blood (IF 13.164) Pub Date : 2017-11-16 Florent Malard; Myriam Labopin; Ibrahim Yakoub-Agha; Sylvain Chantepie; Thierry Guillaume; Didier Blaise; Reza Tabrizi; Leonardo Magro; Bernard Vanhove; Gilles Blancho; Philippe Moreau; Béatrice Gaugler; Patrice Chevallier; Mohamad Mohty
Chronic graft-versus-host disease (cGVHD) is the main cause of late nonrelapse mortality and morbidity after allogeneic stem cell transplantation (allo-SCT). To improve such patients’ outcomes, we conducted a phase 2, prospective, multicenter trial to test the efficacy of the addition of rituximab to corticosteroids (CSs) and cyclosporine A (CsA) as first-line therapy for newly diagnosed cGVHD after allo-SCT. Twenty-four patients (median age, 47 years) with mild (n = 2), moderate (n = 7), or severe (n = 15) cGVHD were included. All patients received rituximab 375 mg/m2 weekly for 4 weeks, followed by a second course 1 month later for patients with partial response. Twenty of 24 patients (83%) were in response at 1 year. Furthermore, among 19 evaluable patients, 14 (74%) were off CSs. The estimated 1-year overall survival was 83%, and the 1-year cumulative incidence of nonrelapse mortality was 14%. One patient died of progressive multifocal leukoencephalopathy. Although PD-L1hi naive B cells were significantly decreased at diagnosis of cGVHD, they increased after anti-CD20 B-cell depletion. In contrast, activated ICOShi PD-1hi circulating T follicular helper (Tfh) cells decreased after rituximab treatment. Overall, the addition of rituximab to corticosteroid and CsA appeared to be safe and effective for first-line treatment of cGVHD. Furthermore, our data suggest that this efficacy may be in part related to an effect on PD-L1hi B cells and Tfh cells. This study was registered at www.clinicaltrials.gov as identifier NCT01135641.
Prognostic significance of baseline metabolic tumor volume in relapsed and refractory Hodgkin lymphoma Blood (IF 13.164) Pub Date : 2017-11-16 Alison J. Moskowitz; Heiko Schöder; Somali Gavane; Katie L. Thoren; Martin Fleisher; Joachim Yahalom; Susan J. McCall; Briana R. Cadzin; Stephanie Y. Fox; John Gerecitano; Ravinder Grewal; Paul A. Hamlin; Steven M. Horwitz; Anita Kumar; Matthew Matasar; Andy Ni; Ariela Noy; M. Lia Palomba; Miguel-Angel Perales; Carol S. Portlock; Craig Sauter; David Straus; Anas Younes; Andrew D. Zelenetz; Craig H. Moskowitz
Identification of prognostic factors for patients with relapsed/refractory Hodgkin lymphoma (HL) is essential for optimizing therapy with risk-adapted approaches. In our phase 2 study of positron emission tomography (PET)–adapted salvage therapy with brentuximab vedotin (BV) and augmented ifosfamide, carboplatin, and etoposide (augICE), we assessed clinical factors, quantitative PET assessments, and cytokine and chemokine values. Transplant-eligible patients with relapsed/refractory HL received 2 (cohort 1) or 3 (cohort 2) cycles of weekly BV; PET-negative patients (Deauville score ≤2) proceeded to autologous stem cell transplantation (ASCT) whereas PET-positive patients received augICE before ASCT. Serum cytokine and chemokine levels were measured at baseline and after BV. Metabolic tumor volume (MTV) and total lesion glycolysis were measured at baseline, after BV, and after augICE. Sixty-five patients enrolled (45, cohort 1; 20, cohort 2); 49 (75%) achieved complete response and 64 proceeded to ASCT. Three-year overall survival and event-free survival (EFS) were 95% and 82%, respectively. Factors predictive for EFS by multivariable analysis were baseline MTV (bMTV) (P < .001) and refractory disease (P = .003). Low bMTV (<109.5 cm3) and relapsed disease identified a favorable group (3-year EFS, 100%). For patients who received a transplant, bMTV and pre-ASCT PET were independently prognostic; 3-year EFS for pre-ASCT PET-positive patients with low bMTV was 86%. In this phase 2 study of PET-adapted therapy with BV and augICE for relapsed/refractory HL, bMTV and refractory disease were independent prognostic factors for EFS. Furthermore, bMTV improved the predictive power of pre-ASCT PET. Future studies should optimize efficacy and tolerability of salvage therapy by stratifying patients according to risk factors such as bMTV.
H3K27M/I mutations promote context-dependent transformation in acute myeloid leukemia with RUNX1 alterations Blood (IF 13.164) Pub Date : 2017-11-16 Bernhard Lehnertz; Yu Wei Zhang; Isabel Boivin; Nadine Mayotte; Elisa Tomellini; Jalila Chagraoui; Vincent-Philippe Lavallée; Josée Hébert; Guy Sauvageau
Neomorphic missense mutations affecting crucial lysine residues in histone H3 genes significantly contribute to a variety of solid cancers. Despite the high prevalence of H3K27M mutations in pediatric glioblastoma and their well-established impact on global histone H3 lysine 27 di- and trimethylation (H3K27me2/3), the relevance of these mutations has not been studied in acute myeloid leukemia (AML). Here, we report the first identification of H3K27M and H3K27I mutations in patients with AML. We find that these lesions are major determinants of reduced H3K27me2/3 in these patients and that they are associated with common aberrations in the RUNX1 gene. We demonstrate that H3K27I/M mutations are strong disease accelerators in a RUNX1-RUNX1T1 AML mouse model, suggesting that H3K27me2/3 has an important and selective leukemia-suppressive activity in this genetic context.
Degree of anemia, indirect markers of hemolysis, and vascular complications of sickle cell disease in Africa Blood (IF 13.164) Pub Date : 2017-11-16 Marie Dubert; Jacques Elion; Aissata Tolo; Dapa Aly Diallo; Saliou Diop; Ibrahima Diagne; Ibrahima Sanogo; Suzanne Belinga; Odette Guifo; Guillaume Wamba; Françoise Ngo Sack; Kouakou Boidy; Ismael Kamara; Youssouf Traore; Cheick Oumar Diakite; Valérie Gbonon; Blaise Felix Faye; Moussa Seck; Indou Deme Ly; David Chelo; Roland N’Guetta; Ibrahima Bara Diop; Bamba Gaye; Xavier Jouven; Brigitte Ranque
The hyperhemolysis paradigm that describes overlapping “hyperhemolytic-endothelial dysfunction” and “high hemoglobin-hyperviscous” subphenotypes of sickle cell disease (SCD) patients is based on North American studies. We performed a transversal study nested in the CADRE cohort to analyze the association between steady-state hemolysis and vascular complications of SCD among sub-Saharan African patients. In Mali, Cameroon, and Ivory Coast, 2407 SCD patients (1751 SS or sickle β-zero-thalassemia [Sβ0], 495 SC, and 161 sickle β+-thalassemia [Sβ+]), aged 3 years old and over, were included at steady state. Relative hemolytic intensity was estimated from a composite index derived from principal component analysis, which included bilirubin levels or clinical icterus, and lactate dehydrogenase levels. We assessed vascular complications (elevated tricuspid regurgitant jet velocity [TRV], microalbuminuria, leg ulcers, priapism, stroke, and osteonecrosis) by clinical examination, laboratory tests, and echocardiography. After adjustment for age, sex, country, and SCD phenotype, a low hemoglobin level was significantly associated with TRV and microalbuminuria in the whole population and with leg ulcers in SS-Sβ0 adults. A high hemolysis index was associated with microalbuminuria in the whole population and with elevated TRV, microalbuminuria, and leg ulcers in SS-Sβ0 adults, but these associations were no longer significant after adjustment for hemoglobin level. In conclusion, severe anemia at steady state in SCD patients living in West and Central Africa is associated with elevated TRV, microalbuminuria, and leg ulcers, but these vascular complications are not independently associated with indirect markers of increased hemolysis. Other mechanisms leading to anemia, including malnutrition and infectious diseases, may also play a role in the development of SCD vasculopathy.
CLEC-2 contributes to hemostasis independently of classical hemITAM signaling in mice Blood (IF 13.164) Pub Date : 2017-11-16 Elizabeth J. Haining; Deya Cherpokova; Karen Wolf; Isabelle C. Becker; Sarah Beck; Johannes A. Eble; David Stegner; Steve P. Watson; Bernhard Nieswandt
C-type lectin-like receptor 2 (CLEC-2) is a platelet receptor that is critical during development in blood-lymph separation and implicated in thrombus stability in thrombosis and hemostasis. It is the only known platelet activatory receptor that participates in both of these aspects of platelet function, and it is the only one to signal through a hemi-immunoreceptor tyrosine–based activation motif (hemITAM). Current investigations into the function of CLEC-2 in vivo have focused on knockout (KO) studies in which both the receptor and its signaling are deleted, making it impossible to explore the possible signaling-independent functions of the receptor, which are indicated by its only known physiological ligand, podoplanin, being an integral membrane protein. In this report, we present a novel knockin mouse model that maintains the expression of a CLEC-2 receptor that cannot signal through its hemITAM (Y7A KI). Remarkably, this mouse phenocopies the blood-lymphatic mixing and lethality of CLEC-2 KO models, but not their hemostatic/thrombotic defect. However, treatment of Y7A KI mice with Fab′ fragments of the function-blocking anti–CLEC-2 antibody, INU1, resulted in a thrombus formation defect in vivo and ex vivo, revealing a hemITAM signaling–independent role for CLEC-2 in hemostasis and thrombosis.
A genome-wide association study of LCH identifies a variant in SMAD6 associated with susceptibility Blood (IF 13.164) Pub Date : 2017-11-16 Erin C. Peckham-Gregory; Rikhia Chakraborty; Michael E. Scheurer; John W. Belmont; Harshal Abhyankar; Amel G. Sengal; Brooks P. Scull; Olive Eckstein; Daniel J. Zinn; Louisa Mayer; Albert Shih; Miriam Merad; D. Williams Parsons; Kenneth L. McClain; Philip J. Lupo; Carl E. Allen
To the editor: Langerhans cell histiocytosis (LCH) is a hematologic disorder that presents with a wide spectrum of symptoms, ranging from focal lesions to potentially lethal multiorgan disease, affecting 4 to 8 per million children per year and 1 to 2 per million adults per year.
Leukemic presentation of diffuse large B-cell lymphoma: an unusual pattern associated with splenic involvement Blood (IF 13.164) Pub Date : 2017-11-16 Genevieve M. Crane; Archibald S. Perkins
![Figure] A 61-year-old man without significant medical history presented with a several-week history of drenching night sweats, 30-pound weight loss, diarrhea, and increasing fatigue. His evaluation in the emergency room revealed hepatosplenomegaly with elevated lactate dehydrogenase (
Nasrallah R, Fast EM, Solaimani P, et al. Identification of novel regulators of developmental hematopoiesis using Endoglin regulatory elements as molecular probes. Blood. 2016;128(15):1928-1939. Blood (IF 13.164) Pub Date : 2017-11-16 American Society of Hematology
In the Acknowledgments section on page 1938 in the 13 October 2016 issue, support from European Research Council Advanced Grant 341096 is missing. The error has been corrected in the online version, which now differs from the print version
A phase I study of romidepsin and pralatrexate reveals marked activity in relapsed and refractory T-cell lymphoma Blood (IF 13.164) Pub Date : 2017-01-01 Jennifer E. Amengual; Renee Lichtenstein; Jennifer Lue; Ahmed Sawas; Changchun Deng; Emily Lichtenstein; Karen Khan; Laine Atkins; Aishling Rada; Hye A. Kim; Codruta Chiuzan; Matko Kalac; Enrica Marchi; Lorenzo Falchi; Mark A. Francescone; Lawrence Schwartz; Serge Cremers; Owen A. O'Connor
The peripheral T-cell lymphomas (PTCL) are a group of rare malignancies characterized by chemotherapy insensitivity and poor prognosis. Romidepsin and pralatrexate were approved by the U.S. FDA for patients with relapsed/refractory PTCL, exhibiting response rates of 25% and 29% respectively. Based on synergy of the combination in preclinical models of PTCL, we initiated a phase I study of pralatrexate plus romidepsin in patients with relapsed/refractory lymphoma (ClinicalTrials.gov (NCT01947140)). This was a single institution dose-escalation phase I study of pralatrexate plus romidepsin designed to determine the dose limiting toxicities (DLT), maximum tolerated dose (MTD), pharmacokinetic profile and response rates. Patients were treated with pralatrexate 10 mg/m2 to 25 mg/m2, and romidepsin 12mg/m2 to 14 mg/m2 on one of three schedules: (1) QWx3 Q28D; (2) QWx2 Q21D; (3) QOW Q28D. Treatment continued until progression, withdrawal of consent, or medical necessity. Response was assessed using the Lugano Classification. Twenty-nine patients were enrolled and evaluable for toxicity. Co-administration of pralatrexate and romidepsin was safe and well tolerated. There were 3 DLTs consisting of 2 Grade 3 oral mucositis and 1 Grade 4 sepsis. The RP2D was defined as pralatrexate 25 mg/m2 and romidepsin 12 mg/m2 QOW. Twenty-three patients were evaluable for response. The ORR across all patients was 57% (13/23); and in PTCL was 71% (10/14). The phase I study of pralatrexate plus romidepsin resulted in a high response rate in patients with previously treated PTCL. A phase II study in PTCL will determine the efficacy of the combination on the QOW dose schedule.
Aging of hematopoietic stem cells Blood (IF 13.164) Pub Date : 2017-01-01 Gerald de Haan; Seka Lazare
Hematopoietic stem cells (HSCs) ensure a balanced production of all blood cells throughout life. As they age, HSCs gradually lose their self-renewal and regenerative potential, while the occurrence of cellular derailment strongly increases. Here we review our current understanding of the molecular mechanisms that contribute to HSC aging. We argue that most of the causes that underlie HSC aging result from cell-intrinsic pathways, and reflect on which aspects of the aging process may be reversible. As many hematological pathologies are strongly age-associated, strategies to intervene in aspects of the stem cell aging process may have significant clinical relevance.
Age related clonal hematopoiesis (ARCH) Blood (IF 13.164) Pub Date : 2017-01-01 Liran I. Shlush
Age related alterations in the human blood system occur in B cells, T cells, cells of the innate system, as well as in hematopoietic stem and progenitor cells (HSPCs). Interestingly, age related, reduced genetic diversity can be identified both at the stem cell level, but also independently in B cells and T cells. This reduced diversity is most probably related to somatic mutations, or to changes in the micro-environmental niche. Either process can select for specific clones, or cause repeated evolutionary bottlenecks. This review will discuss the age related clonal expansions in the human HSPC pool, which was termed in the past Age Related Clonal Hematopoiesis (ARCH). ARCH is defined as the gradual, clonal expansion of HSPCs carrying specific, disruptive and recurrent genetic variants, in individuals without clear diagnosis of hematological malignancies. ARCH is not just associated with chronological aging but also with several other, age-related pathological conditions, including inflammation, vascular diseases, cancer mortality and high risk for hematological malignancies. While it remains unclear whether ARCH is a marker of aging or play an active role in these various pathophysiologies it is suggested here that treating or even preventing ARCH may prove to be beneficial for human health. This review also describes a decision tree for the diagnosis and follow-up for ARCH in a research setting.
DNA damage responses and p53 in the aging process Blood (IF 13.164) Pub Date : 2017-01-01 Hui-Ling Ou; Björn Schumacher
The genome is constantly attacked by genotoxic insults. DNA damage has long been established to cause cancer development through its mutagenic consequences. Conversely, DNA damage is induced during radiation- and chemotherapy to drive cells into apoptosis or senescence as outcomes of the DNA damage response (DDR). More recently, DNA damage has been recognized as a causal factor for the aging process. The causal role of DNA damage in aging and age-related diseases is illustrated by numerous congenital progeroid syndromes that are caused by mutations in genome maintenance pathways. The past two decades have brought rapid progress in the understanding of how DDR drives cancer development and causally contributes to the aging process. The DDR factor p53 turns out to not only take the centre stage during tumour development but also to play an important role in the aging process. Studies in metazoan models ranging from C. elegans to mammalian disease models have revealed cell autonomous and systemic DDR mechanisms that orchestrate adaptive responses that augment maintenance of the aging organism amid gradually accumulating DNA damage.
Anemia at older age: etiologies, clinical implications and management Blood (IF 13.164) Pub Date : 2017-01-01 Reinhard Stauder; Peter Valent; Igor Theurl
Anemia is quite frequently diagnosed in older individuals and is a key indicator of various reactive and clonal conditions. Many underlying diseases, like the myelodysplastic syndromes (MDS), develop preferentially in elderly individuals. The prevalence of anemia at older age is increasing, and this is mainly due to more frequently applied diagnostics and demographic changes in our societies. The etiology of anemia at older age is complex and ranges from bone marrow failure syndromes to chronic kidney disease, and from nutritional deficiencies to inflammatory processes including inflammaging in immunosenescence. In a smaller number of cases, no clear-cut etiology is identified. These patients are referred to as unexplained anemia (UA) or idiopathic cytopenia of unknown significance (ICUS). In others, somatic mutations in leukocytes are found but diagnostic criteria for MDS or other hematological diseases are not fulfilled, a condition termed clonal cytopenia of uncertain significance (CCUS). Management of anemias at older age depends on i) the severity of the anemia, ii) underlying condition(s), and iii) patient-related factors, including comorbidities. Even a mild anemia may substantially affect physical and cognitive capacities and QoL. An underestimated aspect is that due to age-related changes, organ function such as erythropoietin production in the kidney may become suboptimal. Management and treatment of anemia in older patients often requires a multidisciplinary approach and detailed investigations of organ function. In this article, we review current concepts around anemias at older age, with special emphasis on etiologies, clinical implications and innovative concepts in the management of these patients.
Frailty and the management of hematologic malignancies Blood (IF 13.164) Pub Date : 2017-01-01 Gregory A. Abel; Heidi D. Klepin
The majority of blood cancers occur in the elderly. This fact conspires with an aging population in many countries to make rigorous assessment for frailty increasingly important for hematologic oncologists. In this review, we first define frailty and its relevance for patients with hematologic malignancy. Next, we review current data regarding the impact of domains of frailty on outcomes for blood cancers including myelodysplastic syndromes (MDS), acute leukemia, non-Hodgkin lymphomas such as chronic lymphocytic leukemia (CLL), and multiple myeloma. Finally, after presenting assessment and treatment options for the practicing hematologist, we propose elements of a new research agenda for geriatric hematology: the exchange of age limits for rigorous frailty screening, development of disease-specific measures, and inclusion of functional and patient-reported outcomes along with survival.
Decitabine enhances targeting of AML cells by CD34+ progenitor-derived NK cells in NOD/SCID/IL2Rgnull mice Blood (IF 13.164) Pub Date : 2017-01-01 Jeannette Cany; Mieke W.H. Roeven; Janneke S. Hoogstad-van Evert; Willemijn Hobo; Frans Maas; Rosalia Franco Fernandez; Nicole M.A. Blijlevens; Walter J. van der Velden; Gerwin Huls; Joop H. Jansen; Nicolaas P.M. Schaap; Harry Dolstra
Combining NK cell adoptive transfer with hypomethylating agents (HMA) is an attractive therapeutic approach for patients with acute myeloid leukemia (AML). However, data regarding the impact of HMA on NK cell functionality are mostly derived from in vitro studies with high non-clinical relevant drug concentrations. Here, we report a comparative study of azacitidine and decitabine in combination with allogeneic NK cells generated from CD34+ hematopoietic stem and progenitor cells (HSPC-NK cells) in in vitro and in vivo AML models. In vitro, low-dose HMA did not impair HSPC-NK cell viability. Furthermore, low-dose decitabine preserved HSPC-NK killing, proliferation and IFN-γ production capacity, while azacitidine diminished their proliferation and reactivity. Importantly, we showed that HMA and HSPC-NK cells can potently cooperate to target AML cell lines and patient AML blasts. In vivo, both agents exerted a significant delay in AML progression in NOD/SCID/IL2Rgnull mice, while the persistence of adoptively transferred HSPC-NK cells was not affected. Infused NK cells showed sustained expression of most activating receptors, up-regulated NKp44 expression and remarkable KIR acquisition. Most importantly, only decitabine potentiated HSPC-NK cell anti-leukemic activity in vivo. Besides up-regulation of NKG2D and DNAM-1 activating ligands on AML cells, decitabine enhanced mRNA expression of inflammatory cytokines, perforin, and TRAIL by HSPC-NK cells. In addition, treatment resulted in increased numbers of HSPC-NK cells in the bone marrow compartment, suggesting that decitabine could positively modulate NK cell activity, trafficking and tumor targeting. These data provide a rationale to explore combination therapy of adoptive HSPC-NK cells and decitabine in AML patients.
Immature CML cells Implement a BMP Autocrine Loop to Escape TKI Treatment Blood (IF 13.164) Pub Date : 2017-01-01 Elodie Grockowiak; Bastien Laperrousaz; Sandrine Jeanpierre; Thibault Voeltzel; Boris Guyot; Stéphanie Gobert; Franck E. Nicolini; Véronique Maguer-Satta
The BCR-ABL specific Tyrosine Kinase Inhibitors (TKI) changed the outcome of Chronic Myeloid Leukemia (CML), turning a life-threatening disease into a chronic illness. However, TKI are not yet curative, since most patients retain leukemic stem cells (LSC) and their progenitors in bone marrow and relapse following treatment cessation. At diagnosis, deregulations of the Bone Morphogenetic Proteins (BMP) pathway are involved in LSC and progenitors expansion. Here, we report that BMP pathway alterations persist in TKI-resistant patients. As compared to patients in Complete Cytogenetic Remission (CCyR), TKI-resistant LSC and progenitors display high levels of BMPR1b expression and alterations of its cellular localization. In vitro treatment of immature chronic phase (CP) CML cells with TKI alone, or in combination with IFN-α results in the preferential survival of BMPR1b+ cells. We demonstrated persistent and increasing BMP4 production by patients' mesenchymal cells with resistance. Patient follow-up revealed an increase of BMPR1b expression and also in BMP4 expression in LSC from TKI-resistant patients as compared to diagnosis, while remaining unchanged in sensitive patients. Both leukemic and non-leukemic cells exhibit higher BMP4 levels in the bone marrow of TKI-resistant patients. Exposure to BMP2/BMP4 does not alter BCR-ABL transcript expression, but is accompanied by the overexpression of TWIST-1, a transcription factor highly expressed in resistant LSC. By modulating BMP4 or BMPR1b expression, we show that these elements are involved in TKI-resistance. To summarize, we revealed that persistence of BMP alterations and existence of an autocrine loop promote CML primitive cells TKI-resistance.
Antibodies Targeting Surface Membrane Antigens in Patients with Chronic Graft-Versus-Host Disease Blood (IF 13.164) Pub Date : 2017-01-01 Kathy S. Wang; Haesook T. Kim; Sarah Nikiforow; Alexander T. Heubeck; Vincent T. Ho; John Koreth; Edwin P. Alyea; Philippe Armand; Bruce R. Blazar; Robert J. Soiffer; Joseph H. Antin; Corey S. Cutler; Jerome Ritz
Chronic graft-versus-host disease (cGVHD) after allogeneic hematopoietic stem cell transplant (HSCT) reflects a complex immune response resulting in chronic damage to multiple tissues. Previous studies indicated that donor B cells and the antibodies they produce play an important role in the development of cGVHD. To understand the pathogenic role of antibodies in cGVHD, we focused our studies on post-transplant production of IgG antibodies targeting cell surface antigens expressed in multiple cGVHD affected tissues, due to their potential functional impact on living cells in vivo. Using plate-bound cell membrane proteins as targets, we detected a significantly higher level of antibodies reactive with these membrane antigens in patients who developed cGVHD, compared to those who did not and healthy donors. Plasma reactive antibody levels increased significantly prior to the clinical diagnosis of cGVHD and were reduced following cGVHD therapies including prednisone, IL-2 or rituximab. Using cell-based immunoprecipitation with plasma from cGVHD patients and mass spectrometry, we identified 43 membrane proteins targeted by these antibodies. The presence of antibodies in cGVHD patients' plasma that specifically target 6 of these proteins was validated. Antibodies reactive with these 6 antigens were more frequently detected in patients with cGVHD compared to patients without cGVHD and healthy donors. These results indicate that antibodies that target membrane antigens of living cells frequently develop in cGVHD patients and further support a role for B cells and antibodies in the development of cGVHD.
Extracellular histones induce erythrocyte fragility and anemia Blood (IF 13.164) Pub Date : 2017-01-01 Farzaneh Kordbacheh; Connor H. O'Meara; Lucy A. Coupland; Patrick M. Lelliott; Christopher R. Parish
Extracellular histones have been shown to play an important pathogenic role in many diseases, primarily through their cytotoxicity towards nucleated cells and their ability to promote platelet activation with resultant thrombosis and thrombocytopenia. In contrast, little is known about the effect of extracellular histones on erythrocyte function. We demonstrate here that histones promote erythrocyte aggregation, sedimentation and, using a novel in vitro shear stress model, induce erythrocyte fragility and lysis in a concentration dependent manner. Furthermore, histones impair erythrocyte deformability based on reduced passage of erythrocytes through an artificial spleen. These in vitro results were mirrored in vivo with the injection of histones inducing anemia, within minutes of administration, with a concomitant increase in splenic hemoglobin content. Thrombocytopenia and leukopenia were also observed. These findings suggest that histones binding to erythrocytes may contribute to the elevated erythrocyte sedimentation rates (ESR) observed in inflammatory conditions. Furthermore, histone-induced increases in RBC lysis and splenic clearance may be a significant factor in the unexplained anemias seen in critically ill patients.
CRISPR-mediated TCR replacement generates superior anticancer transgenic T-cells Blood (IF 13.164) Pub Date : 2017-01-01 Mateusz Legut; Garry Dolton; Afsar Ali Mian; Oliver Ottmann; Andrew Sewell
Adoptive transfer of T-cells genetically modified to express a cancer-specific T-cell receptor (TCR) has shown significant therapeutic potential for both hematological and solid tumors. However, a major issue of transducing T-cells with a transgenic TCR is the pre-existing expression of TCRs in the recipient cells. These endogenous TCRs compete with the transgenic TCR for surface expression and allow mixed dimer formation. Mixed dimers, formed by mispairing between the endogenous and transgenic TCRs, may harbor autoreactive specificities. To circumvent these problems, we designed a system where the endogenous TCR-β is knocked out from the recipient cells using CRISPR/Cas9 technology, simultaneously with transduction with a cancer-reactive receptor of choice. This TCR replacement strategy resulted in markedly increased surface expression of transgenic αβ and γδ TCRs, which in turn translated to a stronger, and more polyfunctional, response of engineered T-cells to their target cancer cell lines. Additionally, the TCR+CRISPR modified T-cells were up to a thousandfold more sensitive to antigen than standard TCR-transduced T-cells or conventional model proxy systems used for studying TCR activity. Finally, transduction with a pan-cancer reactive γδ TCR used in conjunction with CRISPR/Cas9 knockout of the endogenous αβ TCR resulted in more efficient redirection of CD4+ and CD8+ T-cells against a panel of established blood cancers and primary, patient-derived B acute lymphoblastic leukemia blasts compared to standard TCR transfer. Our results suggest that TCR transfer combined with genome editing could lead to new improved generations of cancer immunotherapies.
FOXP1 expression is a prognostic biomarker in follicular lymphoma treated with rituximab-containing regimens Blood (IF 13.164) Pub Date : 2017-01-01 Anja Mottok; Vindi Jurinovic; Pedro Farinha; Andreas Rosenwald; Ellen Leich; German Ott; Heike Horn; Wolfram Klapper; Michael Boesl; Wolfgang Hiddemann; Christian Steidl; Joseph M. Connors; Laurie H. Sehn; Randy D. Gascoyne; Eva Hoster; Oliver Weigert; Robert Kridel
Follicular lymphoma (FL) is a clinically and molecularly highly heterogeneous disease, yet prognostication relies predominantly on clinical tools. We recently demonstrated that integration of mutation status of seven genes, including EZH2 and MEF2B, improves risk stratification. We mined gene expression data to uncover genes that are differentially expressed in EZH2- and MEF2B-mutated cases. We focused on FOXP1 and assessed its protein expression by immunohistochemistry (IHC) in a total of 763 tissue biopsies. For outcome correlation, a population-based training cohort of 142 FL patients treated with R-CVP, and a clinical trial validation cohort comprising 395 patients treated with CHOP +/- rituximab were used. We found FOXP1 to be significantly down-regulated in both EZH2- and MEF2B-mutated cases. By IHC, 76 specimens in the training cohort (54%) had high FOXP1 expression (>10%), which was associated with reduced five-year failure-free survival (FFS) rates (55% vs. 70%). In the validation cohort, high FOXP1 expression status was observed in 248 patients (63%) and correlated with significantly shorter FFS in patients treated with R-CHOP (HR 1.95, P=0.017), but not in patients treated with CHOP (HR 1.15, P=0.44). The impact of high FOXP1 expression on FFS in immunochemotherapy-treated patients was additional to the FLIPI. High FOXP1 expression was associated with distinct molecular features such as TP53 mutations, expression of IRF4, and gene expression signatures reminiscent of dark zone germinal centre or activated B cells. In summary, FOXP1 is a downstream phenotypic commonality of gene mutations and predicts outcome following rituximab-containing regimens.
CD38 antibodies in multiple myeloma: back to the future Blood (IF 13.164) Pub Date : 2017-01-01 Niels W.C.J. van de Donk; Paul G. Richardson; Fabio Malavasi
CD38 is highly and uniformly expressed on MM cells, and at relatively low levels on normal lymphoid and myeloid cells, and in some tissues of non-hematopoietic origin. CD38 is a transmembrane glycoprotein with ectoenzymatic activity, and also functions as receptor and adhesion molecule. Altogether, this has triggered the development of several CD38 antibodies including daratumumab (fully human), isatuximab (chimeric), and MOR202 (fully human). CD38 antibodies have pleiotropic mechanisms of action including Fc-dependent immune effector mechanisms, direct apoptotic activity, and immunomodulatory effects by the elimination of CD38-positive immunesuppressor cells. CD38-targeting antibodies are generally well tolerated and induce partial response or better in approximately 30% of heavily pretreated multiple myeloma (MM) patients as monotherapy. Based on their distinct mechanisms of action, favorable toxicity profile, and single agent activity, CD38 antibodies are attractive partners in combination regimens. Indeed, deep responses and prolonged progression-free survival can be achieved in relapsed/refractory MM patients when CD38 antibodies are combined with immunomodulatory agents or proteasome inhibitors. Infusion-related reactions, which typically occur during the first infusion, are the most frequent adverse events. Attention should also be paid to the interference of CD38 antibodies with certain laboratory assays, which may complicate response evaluation and blood compatibility testing. Several studies are currently examining the role of CD38-based therapies in newly diagnosed and high-risk smoldering MM. Furthermore, CD38 antibodies are currently also under investigation in other hematologic malignancies, including acute lymphoblastic leukemia, NK/T cell lymphoma, and acute myeloid leukemia, as well as in solid tumors.
Immunotherapy targeting 4-1BB: mechanistic rationale, clinical results, and future strategies Blood (IF 13.164) Pub Date : 2017-01-01 Cariad Chester; Miguel F. Sanmamed; Jun Wang; Ignacio Melero
4-1BB (CD137, TNFRSF9) is an inducible costimulatory receptor expressed on activated T and natural killer (NK) cells. 4-1BB ligation on T cells triggers a signaling cascade that results in upregulation of antiapoptotic molecules, cytokine secretion, and enhanced effector function. In dysfunctional T cells that have a decreased cytotoxic capacity, 4-1BB ligation demonstrates a potent ability to restore effector functions. On NK cells, 4-1BB signaling can increase antibody-dependent cell-mediated cytotoxicity. Agonistic monoclonal antibodies targeting 4-1BB have been developed to harness 4-1BB signaling for cancer immunotherapy. Preclinical results in a variety of induced and spontaneous tumor models suggest that targeting 4-1BB with agonist antibodies can lead to tumor clearance and durable anti-tumor immunity. Clinical trials of two agonist antibodies, urelumab and utomilumab, are ongoing. Despite initial signs of efficacy, clinical development of urelumab has been hampered by inflammatory liver toxicity at doses above 1 mg/kg. Utomilumab has a superior safety profile, but is a less potent 4-1BB agonist relative to urelumab. Both antibodies have demonstrated promising results in patients with lymphoma and are currently being tested in combination therapy trials with other immunomodulatory agents. In an effort to optimally leverage 4-1BB-mediated immune activation, the next generation of 4-1BB-targeting strategies attempts to decouple the observed antitumor efficacy from the on-target liver toxicity. Multiple therapeutics that attempt to restrict 4-1BB agonism to the tumor microenvironment and minimize systemic exposure have emerged. 4-1BB is a compelling target for cancer immunotherapy and future agents show great promise for achieving potent immune activation while avoiding limiting immune-related adverse events.
CTLA-4: a moving target in immunotherapy Blood (IF 13.164) Pub Date : 2017-01-01 Behzad Rowshanravan; Neil Halliday; David M. Sansom
CD28 and CTLA-4 are members of a family of Immunoglobulin-related receptors that are responsible for various aspects of T cell immune regulation. The family includes CD28, CTLA-4 and ICOS as well as other proteins including PD-1, BTLA and TIGIT. These receptors have both stimulatory (CD28, ICOS) as well as inhibitory roles (CTLA-4, PD-1, BTLA and TIGIT) in T cell function. Increasingly these pathways are targeted as part of immune modulatory strategies to treat cancers, referred to generically as immune checkpoint blockade, and conversely to treat autoimmunity and CTLA-4 deficiency. Here we focus on the biology of the CD28/CTLA-4 pathway as a framework for understanding the impacts of therapeutic manipulation of this pathway.
PD-1 expression and clinical PD-1 blockade in B-cell lymphomas Blood (IF 13.164) Pub Date : 2017-01-01 Zijun Y. Xu-Monette; Jianfeng Zhou; Ken H. Young
PD-1 blockade targeting the PD-1 immune checkpoint has demonstrated unprecedented clinical efficacy in the treatment of advanced cancers including hematologic malignancies. This article reviews the landscape of PD-1/PD-L1 expression and current PD-1 blockade immunotherapy trials in B-cell lymphomas. Most notably, in relapsed/refractory classical Hodgkin lymphoma, which frequently has increased PD-1+ tumor-infiltrating T cells, 9p24 genetic alteration and high PD-L1 expression, anti-PD-1 monotherapy has demonstrated remarkable objective response rates (ORR) of 65-87% and durable disease control in phase I/II clinical trials. The median duration of response was 16 months in a phase II trial. PD-1 blockade has also shown promise in a phase I trial of nivolumab in relapsed/refractory B-cell non-Hodgkin lymphomas, including follicular lymphoma, which often displays abundant PD-1 expression on intratumoral CD4+ T cells, and diffuse large B-cell lymphoma, which variably expresses PD-1 and PD-L1. In primary mediastinal large B-cell lymphoma, which frequently has 9p24 alterations, the ORR was 35% in a phase II trial of pembrolizumab. In contrast, the ORR with pembrolizumab was 0% in relapsed chronic lymphocytic leukemia (CLL) and 44% in CLL with Richter transformation in a phase II trial. T cells from CLL patients have elevated PD-1 expression; CLL PD-1+ T cells can exhibit a pseudo-exhaustion or a replicative senescence phenotype. PD-1 expression was also found in marginal zone lymphoma but not in mantle cell lymphoma, although no reliable PD-1 blockade data are currently available. Mechanisms of PD-1 blockade immunotherapy, toxicities, hyperprogression, predictive biomarkers and combination therapies are discussed in the context of B-cell lymphomas.
The immunobiology of CD27 and OX40 and their potential as targets for cancer immunotherapy Blood (IF 13.164) Pub Date : 2017-01-01 Sarah L. Buchan; Anne Rogel; Aymen Al-Shamkhani
In recent years monoclonal antibodies (mAbs) able to reinvigorate anti-tumor T cell immunity have heralded a paradigm shift in cancer treatment. The most high profile of these mAbs block the inhibitory checkpoint receptors PD-1 and CTLA-4 and have improved life expectancy for patients across a range of tumor types. However, it is becoming increasingly clear that failure of some patients to respond to checkpoint inhibition is due to inadequate T-cell priming. For full T-cell activation two signals must be received and ligands providing the second of these signals, termed costimulation, are often lacking in tumors. Members of the TNF receptor superfamily (TNFRSF) are key co-stimulators of T cells during infection and there has been an increasing interest in harnessing these receptors to augment tumor immunity. We here review the immunobiology of two particularly promising TNFRSF target receptors, CD27 and OX40, and their respective ligands CD70 and OX40L, focusing on their role within a tumor setting. We describe the influence of CD27 and OX40 on human T cells based on in vitro studies and on the phenotypes of several recently described individuals exhibiting natural deficiencies in CD27/CD70 and OX40. Finally we review key literature describing progress in elucidating the efficacy and mode of action of OX40- and CD27-targeting mAbs in pre-clinical models and provide an overview of current clinical trials targeting these promising receptor/ligand pairings in cancer.
Redirecting T cells to hematological malignancies with bispecific antibodies Blood (IF 13.164) Pub Date : 2017-01-01 Mireya Paulina Velasquez; Challice L. Bonifant; Stephen Gottschalk
There is a need to improve outcomes for patients with recurrent and/or refractory hematological malignancies. Immunotherapy holds the promise to meet this need since it does not rely on the cytotoxic mechanism of conventional therapies. Among different forms of immunotherapy, redirecting T cells to hematological malignancies with bispecific antibodies (BsAbs) is an attractive strategy. BsAbs are an 'off-the-shelf' product that is easily scalable in contrast to adoptive T-cell therapies. Among these, the bispecific T-cell engager (BiTE) blinatumomab has emerged as the most successful BsAb to date. It consists of two single chain variable fragments (scFvs) specific for CD19 present on B-cell malignancies, and CD3 expressed on almost all T cells. Blinatumomab has shown potent anti-tumor activity as a single agent, particularly for acute lymphoblastic leukemia (ALL) resulting in its FDA approval. However, while successful in inducing remissions, these are normally short-lived with median response durations of less than one year. Nevertheless, the success of blinatumomab has reinvigorated the BsAb field, which is bustling with preclinical and clinical studies not only for B-cell derived lymphoblastic leukemia and lymphoma, but also for acute myeloid leukemia and multiple myeloma. Here we will review successes and challenges of T-cell targeted BsAbs for the immunotherapy of hematological malignancies with special focus on conducted clinical studies and strategies to improve their efficacy.
Recombinant ADAMTS-13: goodbye, allergic reactions! Blood (IF 13.164) Pub Date : 2017-11-09 Ravi Sarode
In this issue of Blood , [Scully et al] describe very promising results of a phase 1 study of recombinant ADAMTS-13 (rADAMTS-13; a disintegrin and metalloproteinase with thrombospondin type 1 motif, member 13) in patients with congenital thrombotic thrombocytopenic purpura (cTTP) that is likely
Prognostic relevance of CCAs/Ph− in CML settled Blood (IF 13.164) Pub Date : 2017-11-09 Rüdiger Hehlmann
In this issue of Blood , [Issa et al] report that patients with chronic-phase chronic myeloid leukemia (CML) in remission after treatment with tyrosine kinase inhibitors (TKIs) with clonal chromosomal aberrations (CCAs) in Philadelphia chromosome–negative (Ph−) metaphases had a
’Tis one thing to adhere, another to migrate Blood (IF 13.164) Pub Date : 2017-11-09 Christopher J. Kuckleburg
In this issue of Blood , we review 2 studies investigating neutrophil integrin activation. [Morikis et al] describe the molecular dynamics of mechanotransduction, which underlie E-selectin–mediated neutrophil rolling and adhesion. [Bai et al] investigate CD177-induced integrin activation in
Releasing the FKBP12 brake on hepcidin Blood (IF 13.164) Pub Date : 2017-11-09 Nermi L. Parrow; Robert E. Fleming
In this issue of Blood , [Colucci et al] add to the understanding of hepcidin regulation by demonstrating a role for the immunophilin FKBP12 in dampening signaling via the bone morphogenetic protein (BMP) type I receptor ALK2. ![Figure] FKBP12 as a regulator of BMP/SMAD signaling
FcγR and SCD alloimmunization: a nonclass(ical) act Blood (IF 13.164) Pub Date : 2017-11-09 Karina Yazdanbakhsh
In this issue of Blood , [Meinderts et al] found a protective association of a genetic variant in the FCGR gene locus with decreased risk of red cell (RBC) alloimmunization in patients with sickle cell disease (SCD). If validated in a larger study, the identified polymorphism may be used as
Notching up B-cell pathology in chronic GVHD Blood (IF 13.164) Pub Date : 2017-11-09 Vedran Radojcic; Leo Luznik
In this issue of Blood , [Poe et al] identify the pathogenic and druggable B-cell receptor (BCR)–Notch axis as a feed-forward loop in patients with chronic graft-versus-host disease (cGVHD), providing a rationale and methods for its therapeutic targeting. Chronic GVHD remains one of the
Recombinant ADAMTS-13: first-in-human pharmacokinetics and safety in congenital thrombotic thrombocytopenic purpura Blood (IF 13.164) Pub Date : 2017-11-09 Marie Scully; Paul Knöbl; Karim Kentouche; Lawrence Rice; Jerzy Windyga; Reinhard Schneppenheim; Johanna A. Kremer Hovinga; Michiko Kajiwara; Yoshihiro Fujimura; Caterina Maggiore; Jennifer Doralt; Christopher Hibbard; Leah Martell; Bruce Ewenstein
Safety, tolerability, and pharmacokinetics of recombinant ADAMTS-13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13; BAX 930; SHP655) were investigated in 15 patients diagnosed with severe congenital ADAMTS-13 deficiency (plasma ADAMTS-13 activity <6%) in a prospective phase 1, first-in-human, multicenter dose escalation study. BAX 930 was well tolerated, no serious adverse events occurred, and no anti-ADAMTS-13 antibodies were observed. After single-dose administration of BAX 930 at 5, 20, or 40 U/kg body weight to adolescents and adults, there was approximate dose proportionality with respect to maximum plasma concentration (Cmax [U/mL]) and area under the concentration–time curve (AUC [h∙U/mL]). Dose-related increases of individual ADAMTS-13:Ag and activity were observed and reached a maximum within 1 hour. With escalating BAX 930 doses administered, a dose-dependent persistence of ADAMTS-13-mediated von Willebrand factor (VWF) cleavage products and reduced VWF multimeric size were observed. This study demonstrated that pharmacokinetic parameters of BAX 930 were comparable to those estimated in previous plasma infusion studies and provided evidence of pharmacodynamic activity. This study was registered at www.clinicaltrials.gov as #NCT02216084.
Philadelphia chromosome–like acute lymphoblastic leukemia Blood (IF 13.164) Pub Date : 2017-11-09 Sarah K. Tasian; Mignon L. Loh; Stephen P. Hunger
Philadelphia chromosome (Ph)-like acute lymphoblastic leukemia (ALL), also referred to as BCR-ABL1–like ALL, is a high-risk subset with a gene expression profile that shares significant overlap with that of Ph-positive (Ph+) ALL and is suggestive of activated kinase signaling. Although Ph+ ALL is defined by BCR-ABL1 fusion, Ph-like ALL cases contain a variety of genomic alterations that activate kinase and cytokine receptor signaling. These alterations can be grouped into major subclasses that include ABL-class fusions involving ABL1, ABL2, CSF1R, and PDGFRB that phenocopy BCR-ABL1 and alterations of CRLF2, JAK2, and EPOR that activate JAK/STAT signaling. Additional genomic alterations in Ph-like ALL activate other kinases, including BLNK, DGKH, FGFR1, IL2RB, LYN, NTRK3, PDGFRA, PTK2B, TYK2, and the RAS signaling pathway. Recent studies have helped to define the genomic landscape of Ph-like ALL and how it varies across the age spectrum, associated clinical features and outcomes, and genetic risk factors. Preclinical studies and anecdotal reports show that targeted inhibitors of relevant signaling pathways are active in specific Ph-like ALL subsets, and precision medicine trials have been initiated for this high-risk ALL subset.
Synergy of BCL2 and histone deacetylase inhibition against leukemic cells from cutaneous T-cell lymphoma patients Blood (IF 13.164) Pub Date : 2017-11-09 Benoit M. Cyrenne; Julia M. Lewis; Jason G. Weed; Kacie R. Carlson; Fatima N. Mirza; Francine M. Foss; Michael Girardi
The presence and degree of peripheral blood involvement in patients with cutaneous T-cell lymphoma (CTCL) portend a worse clinical outcome. Available systemic therapies for CTCL may variably decrease tumor burden and improve quality of life, but offer limited effects on survival; thus, novel approaches to the treatment of advanced stages of this non-Hodgkin lymphoma are clearly warranted. Mutational analyses of CTCL patient peripheral blood malignant cell samples suggested the antiapoptotic mediator B-cell lymphoma 2 (BCL2) as a potential therapeutic target. To test this, we developed a screening assay for evaluating the sensitivity of CTCL cells to targeted molecular agents, and compared a novel BCL2 inhibitor, venetoclax, alone and in combination with a histone deacetylase (HDAC) inhibitor, vorinostat or romidepsin. Peripheral blood CTCL malignant cells were isolated from 25 patients and exposed ex vivo to the 3 drugs alone and in combination, and comparisons were made to 4 CTCL cell lines (Hut78, Sez4, HH, MyLa). The majority of CTCL patient samples were sensitive to venetoclax, and BCL2 expression levels were negatively correlated (r = −0.52; P = .018) to 50% inhibitory concentration values. Furthermore, this anti-BCL2 effect was markedly potentiated by concurrent HDAC inhibition with 93% of samples treated with venetoclax and vorinostat and 73% of samples treated with venetoclax and romidepsin showing synergistic effects. These data strongly suggest that concurrent BCL2 and HDAC inhibition may offer synergy in the treatment of patients with advanced CTCL. By using combination therapies and correlating response to gene expression in this way, we hope to achieve more effective and personalized treatments for CTCL.
Clonal chromosomal abnormalities appearing in Philadelphia chromosome–negative metaphases during CML treatment Blood (IF 13.164) Pub Date : 2017-11-09 Ghayas C. Issa; Hagop M. Kantarjian; Graciela Nogueras Gonzalez; Gautam Borthakur; Guilin Tang; William Wierda; Koji Sasaki; Nicholas J. Short; Farhad Ravandi; Tapan Kadia; Keyur Patel; Raja Luthra; Alessandra Ferrajoli; Guillermo Garcia-Manero; Mary Beth Rios; Sara Dellasala; Elias Jabbour; Jorge E. Cortes
Clonal chromosomal abnormalities in Philadelphia chromosome-negative (CCA/Ph−) metaphases emerge as patients with chronic phase chronic myeloid leukemia (CP-CML) are treated with tyrosine kinase inhibitors (TKIs). We assessed the characteristics and prognostic impact of 598 patients with CP-CML treated on clinical trials with various TKIs. CCA/Ph− occurred in 58 patients (10%); the most common were −Y in 25 (43%) and trisomy 8 in 7 patients (12%). Response to TKI therapy was similar for patients with CCA/Ph− and those without additional chromosomal abnormalities (ACAs). We further categorized CCA/Ph− into those in which –Y was the only clonal abnormality, and all others. We found that patients with non –Y CCA/Ph− had worse failure-free survival (FFS), event-free survival (EFS), transformation-free survival (TFS), and overall survival (OS) compared with those without ACAs with the following 5-year rates: FFS (52% vs 70%, P = .02), EFS (68% vs 86%, P = .02), TFS (76% vs 94%, P < .01), and OS (79% vs 94%, P = .03). In a multivariate analysis, non –Y CCA/Ph− increased the risk of transformation or death when baseline characteristics were considered with a hazard ratio of 2.81 (95% confidence interval, 1.15-6.89; P = .02). However, this prognostic impact was not statistically significant when achieving BCR-ABL <10% at 3 months was included in the analysis. In conclusion, non –Y CCA/Ph− are associated with decreased survival when emerging in patients with chronic-phase CML across various TKIs. This trial was registered at www.clinicaltrials.gov as #NCT00048672, #NCT00038649, and #NCT00050531 (imatinib); #NCT00254423 (dasatinib); #NCT00129740 (nilotinib); and NCT01570868 (ponatinib).
CD177 modulates human neutrophil migration through activation-mediated integrin and chemoreceptor regulation Blood (IF 13.164) Pub Date : 2017-11-09 Ming Bai; Ricardo Grieshaber-Bouyer; Junxia Wang; Angela B. Schmider; Zachary S. Wilson; Liling Zeng; Olha Halyabar; Matthew D. Godin; Hung N. Nguyen; Anaïs Levescot; Pierre Cunin; Craig T. Lefort; Roy J. Soberman; Peter A. Nigrovic
CD177 is a glycosylphosphatidylinositol (GPI)-anchored protein expressed by a variable proportion of human neutrophils that mediates surface expression of the antineutrophil cytoplasmic antibody antigen proteinase 3. CD177 associates with β2 integrins and recognizes platelet endothelial cell adhesion molecule 1 (PECAM-1), suggesting a role in neutrophil migration. However, CD177pos neutrophils exhibit no clear migratory advantage in vivo, despite interruption of in vitro transendothelial migration by CD177 ligation. We sought to understand this paradox. Using a PECAM-1-independent transwell system, we found that CD177pos and CD177neg neutrophils migrated comparably. CD177 ligation selectively impaired migration of CD177pos neutrophils, an effect mediated through immobilization and cellular spreading on the transwell membrane. Correspondingly, CD177 ligation enhanced its interaction with β2 integrins, as revealed by fluorescence lifetime imaging microscopy, leading to integrin-mediated phosphorylation of Src and extracellular signal-regulated kinase (ERK). CD177-driven cell activation enhanced surface β2 integrin expression and affinity, impaired internalization of integrin attachments, and resulted in ERK-mediated attenuation of chemokine signaling. We conclude that CD177 signals in a β2 integrin-dependent manner to orchestrate a set of activation-mediated mechanisms that impair human neutrophil migration.
Selectin catch-bonds mechanotransduce integrin activation and neutrophil arrest on inflamed endothelium under shear flow Blood (IF 13.164) Pub Date : 2017-11-09 Vasilios A. Morikis; Shannon Chase; Ted Wun; Elliot L. Chaikof; John L. Magnani; Scott I. Simon
E-selectin extends from the plasma membrane of inflamed endothelium and serves to capture leukocytes from flowing blood via long-lived catch-bonds that support slow leukocyte rolling under shear stress. Its ligands are glycosylated with the tetrasaccharide sialyl Lewisx (sLex), which contributes to bond affinity and specificity. E-selectin-mediated rolling transmits signals into neutrophils that trigger activation of high-affinity β2-integrins necessary for transition to shear-resistant adhesion and transendothelial migration. Rivipansel is a glycomimetic drug that inhibits E-selectin-mediated vaso-occlusion induced by integrin-dependent sickle–red blood cell–leukocyte adhesion. How Rivipansel antagonizes ligand recognition by E-selectin and blocks outside-in signaling of integrin-mediated neutrophil arrest while maintaining rolling immune-surveillance is unknown. Here, we demonstrate that sLex expressed on human L-selectin is preferentially bound by E-selectin and, on ligation, initiates secretion of MRP8/14 that binds TLR4 to elicit the extension of β2-integrin to an intermediate affinity state. Neutrophil rolling over E-selectin at precise shear stress transmits tension and catch-bond formation with L-selectin via sLex, resulting in focal clusters that deliver a distinct signal to upshift β2-integrins to a high-affinity state. Rivipansel effectively blocked formation of selectin catch-bonds, revealing a novel mechanotransduction circuit that rapidly converts extended β2-integrins to high-affinity shear-resistant bond clusters with intracellular adhesion molecule 1 on inflamed endothelium.
The immunophilin FKBP12 inhibits hepcidin expression by binding the BMP type I receptor ALK2 in hepatocytes Blood (IF 13.164) Pub Date : 2017-11-09 Silvia Colucci; Alessia Pagani; Mariateresa Pettinato; Irene Artuso; Antonella Nai; Clara Camaschella; Laura Silvestri
The expression of the key regulator of iron homeostasis hepcidin is activated by the BMP-SMAD pathway in response to iron and inflammation and among drugs, by rapamycin, which inhibits mTOR in complex with the immunophilin FKBP12. FKBP12 interacts with BMP type I receptors to avoid uncontrolled signaling. By pharmacologic and genetic studies, we identify FKBP12 as a novel hepcidin regulator. Sequestration of FKBP12 by rapamycin or tacrolimus activates hepcidin both in vitro and in murine hepatocytes. Acute tacrolimus treatment transiently increases hepcidin in wild-type mice. FKBP12 preferentially targets the BMP receptor ALK2. ALK2 mutants defective in binding FKBP12 increase hepcidin expression in a ligand-independent manner, through BMP-SMAD signaling. ALK2 free of FKBP12 becomes responsive to the noncanonical inflammatory ligand Activin A. Our results identify a novel hepcidin regulator and a potential therapeutic target to increase defective BMP signaling in disorders of low hepcidin.
Nonclassical FCGR2C haplotype is associated with protection from red blood cell alloimmunization in sickle cell disease Blood (IF 13.164) Pub Date : 2017-11-09 Sanne M. Meinderts; Joep W. R. Sins; Karin Fijnvandraat; Sietse Q. Nagelkerke; Judy Geissler; Michael W. Tanck; Christine Bruggeman; Bart J. Biemond; Anita W. Rijneveld; Jean-Louis H. Kerkhoffs; Sadaf Pakdaman; Anoosha Habibi; Robin van Bruggen; Taco W. Kuijpers; France Pirenne; Timo K. van den Berg
Red blood cell (RBC) transfusions are of vital importance in patients with sickle cell disease (SCD). However, a major complication of transfusion therapy is alloimmunization. The low-affinity Fcγ receptors, expressed on immune cells, are important regulators of antibody responses. Genetic variation in FCGR genes has been associated with various auto- and alloimmune diseases. The aim of this study was to evaluate the association between genetic variation of FCGR and RBC alloimmunization in SCD. In this case-control study, DNA samples from 2 cohorts of transfused SCD patients were combined (France and The Netherlands). Cases had a positive history of alloimmunization, having received ≥1 RBC unit. Controls had a negative history of alloimmunization, having received ≥20 RBC units. Single nucleotide polymorphisms and copy number variation of the FCGR2/3 gene cluster were studied in a FCGR-specific multiplex ligation-dependent probe amplification assay. Frequencies were compared using logistic regression. Two hundred seventy-two patients were included (130 controls, 142 cases). The nonclassical open reading frame in the FCGR2C gene (FCGR2C.nc-ORF) was strongly associated with a decreased alloimmunization risk (odds ratio [OR] 0.26, 95% confidence [CI] 0.11-0.64). This association persisted when only including controls with exposure to ≥100 units (OR 0.30, CI 0.11-0.85) and appeared even stronger when excluding cases with Rh or K antibodies only (OR 0.19, CI 0.06-0.59). In conclusion, SCD patients with the FCGR2C.nc-ORF polymorphism have over a 3-fold lower risk for RBC alloimmunization in comparison with patients without this mutation. This protective effect was strongest for exposure to antigens other than the immunogenic Rh or K antigens.
An aberrant NOTCH2-BCR signaling axis in B cells from patients with chronic GVHD Blood (IF 13.164) Pub Date : 2017-11-09 Jonathan C. Poe; Wei Jia; Hsuan Su; Sarah Anand; Jeremy J. Rose; Prasanthi V. Tata; Amy N. Suthers; Corbin D. Jones; Pei Fen Kuan; Benjamin G. Vincent; Jonathan S. Serody; Mitchell E. Horwitz; Vincent T. Ho; Steven Z. Pavletic; Frances T. Hakim; Kouros Owzar; Dadong Zhang; Bruce R. Blazar; Christian W. Siebel; Nelson J. Chao; Ivan Maillard; Stefanie Sarantopoulos
B-cell receptor (BCR)-activated B cells contribute to pathogenesis in chronic graft-versus-host disease (cGVHD), a condition manifested by both B-cell autoreactivity and immune deficiency. We hypothesized that constitutive BCR activation precluded functional B-cell maturation in cGVHD. To address this, we examined BCR-NOTCH2 synergy because NOTCH has been shown to increase BCR responsiveness in normal mouse B cells. We conducted ex vivo activation and signaling assays of 30 primary samples from hematopoietic stem cell transplantation patients with and without cGVHD. Consistent with a molecular link between pathways, we found that BCR-NOTCH activation significantly increased the proximal BCR adapter protein BLNK. BCR-NOTCH activation also enabled persistent NOTCH2 surface expression, suggesting a positive feedback loop. Specific NOTCH2 blockade eliminated NOTCH-BCR activation and significantly altered NOTCH downstream targets and B-cell maturation/effector molecules. Examination of the molecular underpinnings of this “NOTCH2-BCR axis” in cGVHD revealed imbalanced expression of the transcription factors IRF4 and IRF8, each critical to B-cell differentiation and fate. All-trans retinoic acid (ATRA) increased IRF4 expression, restored the IRF4-to-IRF8 ratio, abrogated BCR-NOTCH hyperactivation, and reduced NOTCH2 expression in cGVHD B cells without compromising viability. ATRA-treated cGVHD B cells had elevated TLR9 and PAX5, but not BLIMP1 (a gene-expression pattern associated with mature follicular B cells) and also attained increased cytosine guanine dinucleotide responsiveness. Together, we reveal a mechanistic link between NOTCH2 activation and robust BCR responses to otherwise suboptimal amounts of surrogate antigen. Our findings suggest that peripheral B cells in cGVHD patients can be pharmacologically directed from hyperactivation toward maturity.
Protocol II vs protocol III given twice during reinduction therapy in children with medium-risk ALL Blood (IF 13.164) Pub Date : 2017-11-09 Franco Locatelli; Maria Grazia Valsecchi; Anja Möricke; Martin Zimmermann; Bernd Gruhn; Andrea Biondi; Andreas E. Kulozik; Daniela Silvestri; Nicole Bodmer; Maria Caterina Putti; Stefan Burdach; Concetta Micalizzi; Andrea Teigler-Schlegel; Jörg Ritter; Andrea Pession; Gunnar Cario; Stefan Bielack; Giuseppe Basso; Thomas Klingebiel; Luciana Vinti; Carmelo Rizzari; Andishe Attarbaschi; Nicola Santoro; Rosanna Parasole; Georg Mann; Leonid Karawajew; Oskar A. Haas; Valentino Conter; Martin Schrappe
To the editor: Reinduction therapy (also known as delayed intensification, DI) is an essential part of childhood acute lymphoblastic leukemia (ALL) treatment., In Berlin-Frankfurt-Münster (BFM) studies, it includes same/similar drugs as those employed in induction therapy. The Children
Aleukemic mast cell leukemia associated with chronic myelomonocytic leukemia and chronic lymphocytic leukemia Blood (IF 13.164) Pub Date : 2017-11-09 Zhihong Hu; Sergej Konoplev
![Figure] An 88-year-old man presented with anemia (hemoglobin, 10.3 g/dL) and thrombocytopenia (platelets, 74 × 109/L). Peripheral blood smear showed prominent leukocytosis (white blood cells, 37.3 × 109/L) with dysplastic granulocytes, monocytosis (4.1 × 109/L), eosinophilia (4.5 ×
Classical Hodgkin lymphoma arising in a patient with chronic lymphocytic leukemia (Richter syndrome) Blood (IF 13.164) Pub Date : 2017-11-09 Andrés E. Quesada; Sergej Konoplev
![Figure] A 66-year-old man was diagnosed with chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) in 2006, observed until 2011, and then treated with fludarabine/cyclophosphamide/rituximab followed by ibrutinib/venetoclax without complete remission. In 2017, an abdominal
Gain-of-function of ASXL1 truncating protein in the pathogenesis of myeloid malignancies Blood (IF 13.164) Pub Date : 2017-01-01 Hui Yang; Stefan Kurtenbach; Ying Guo; Ines Lohse; Michael A. Durante; Jianping Li; Zhaomin Li; Hassan AL-Ali; Lingxiao Li; Zizhen Chen; Matthew G. Field; Peng Zhang; Shi Chen; Shohei Yamamoto; Zhuo Li; Yuan Zhou; Stephen D. Nimer; J. William Harbour; Claes Wahlestedt; Mingjiang Xu; Feng-Chun Yang
Additional Sex Combs-Like 1 (ASXL1) is mutated at a high frequency in all forms of myeloid malignancies associated with poor prognosis. We generated a Vav1 promoter driven Flag-Asxl1Y588X transgenic mouse model, Asxl1Y588XTg, to express a truncated FLAG-ASXL1aa1-587 protein in the hematopoietic system. The Asxl1Y588XTg mice had an enlarged hematopoietic stem cell (HSC) pool, shortened survival, and predisposition to a spectrum of myeloid malignancies, thereby recapitulating the characteristics of myeloid malignancy patients with ASXL1 mutations. ATAC- and RNA-Seq analyses revealed that the ASXL1aa1-587 truncating protein expression results in more open chromatin in cKit+ cells compared to wildtype cells, accompanied by dysregulated expression of genes critical for HSC self-renewal and differentiation. Liquid chromatography-tandem mass spectrometry and co-immunoprecipitation experiments showed that ASXL1aa1-587 acquired an interaction with BRD4. An epigenetic drug screening demonstrated a hyper-sensitivity of Asxl1Y588XTg bone marrow cells to BET bromodomain inhibitors. This study demonstrates that ASXL1aa1-587 plays a gain-of-function role in promoting myeloid malignancies. Our model provides a powerful platform to test therapeutic approaches of targeting the ASXL1 truncation mutations in myeloid malignancies.
Hepatocyte nuclear factor 1A deficiency causes hemolytic anemia in mice by altering erythrocyte sphingolipid homeostasis Blood (IF 13.164) Pub Date : 2017-01-01 Karin von Wnuck Lipinski; Sarah Weske; Petra Keul; Susann Peters; Hideo A. Baba; Gerd Heusch; Markus H. Gräler; Bodo Levkau
The Hepatocyte Nuclear Factor (HNF) family regulates complex networks of metabolism and organ development. Human mutations in its prototypical member HNF1A cause maturity-onset of diabetes of the young (MODY) type 3. In this study, we identified an important role for HNF1A in the preservation of erythrocyte membrane integrity, calcium homeostasis and osmotic resistance through a yet unrecognized link of HNF1A to sphingolipid homeostasis. HNF1A-/- mice displayed microcytic hypochromic anemia with reticulocytosis that was partially compensated by avid extramedullary erythropoiesis at all erythroid stages in the spleen thereby excluding erythroid differentiation defects. Morphologically, HNF1A-/- erythrocytes resembled acanthocytes and displayed increased phosphatidylserine exposure, high intracellular calcium, and elevated osmotic fragility. Sphingolipidome analysis by mass spectrometry revealed substantial and tissue-specific sphingolipid disturbances in several tissues including erythrocytes with the accumulation of sphingosine as most prominent common feature. All HNF1A-/- erythrocyte defects could be simulated by exposure of wild type erythrocytes to sphingosine in vitro and attributed in part to sphingosine-induced suppression of the plasma-membrane Ca2+-ATPase activity. Bone marrow transplantation rescued the anemia phenotype in vivo, whereas incubation with HNF1A-/- plasma increased the osmotic fragility of wild type erythrocytes in vitro. Our data suggest a non-cell autonomous erythrocyte defect secondary to the sphingolipid changes caused by HNF1A deficiency. Transcriptional analysis revealed four important genes involved in sphingolipid metabolism to be deregulated in HNF1A deficiency: ORMLD1, sphingosine kinase-2, neutral ceramidase and ceramide synthase-5. The considerable erythrocyte defects in murine HNF1A deficiency encourage clinical studies to explore the hematological consequences of HNF1A deficiency in human MODY3 patients.
HRI coordinates translation by eIF2αP and mTORC1 to mitigate ineffective erythropoiesis in mice during iron deficiency Blood (IF 13.164) Pub Date : 2017-01-01 Shuping Zhang; Alejandra Macias-Garcia; Jason Velazquez; Elena Paltrinieri; Randal J. Kaufman; Jane-Jane Chen
Iron deficiency anemia is a prevalent disease, yet molecular mechanisms by which iron and heme regulate erythropoiesis are not completely understood. HRI, heme-regulated eIF2α kinase, is a key hemoprotein in erythroid precursors sensing intracellular heme concentrations to balance globin synthesis with the amount of heme available for hemoglobin production. HRI is activated by heme deficiency and oxidative stress, and phosphorylates eIF2α (eIF2αP), which inhibits the translation of globin mRNAs as well as enhances selectively the translation of activating transcription factor 4 (ATF4) mRNA to induce stress response genes. Here, we generated a novel mouse model (eAA) with the erythroid specific ablation of eIF2αP, and demonstrate that eIF2αP is required for induction of ATF4 protein synthesis in vivo in erythroid cells during iron deficiency (ID). We show for the first time that both eIF2αP and ATF4 are necessary to promote erythroid differentiation and to reduce oxidative stress in vivo during ID. Furthermore, HRI-eIF2αP-ATF4 pathway suppresses mTORC1 signaling specifically in the erythroid lineage. Pharmacological inhibition of mTORC1 significantly increased RBC counts and hemoglobin content in the blood, improved erythroid differentiation and reduced splenomegaly of iron deficient Hri-/- and eAA mice. However, globin inclusions and elevated oxidative stress remained, demonstrating the essential non-redundant role of HRI-eIF2αP in these processes. Dietary iron repletion completely reversed ID anemia and ineffective erythropoiesis of Hri-/-, eAA and Atf4-/- mice by inhibiting both HRI and mTORC1 signaling. Thus, HRI coordinates two key translation-regulation pathways, eIF2αP and mTORC1, to circumvent ineffective erythropoiesis, highlighting heme and translation in the regulation of erythropoiesis.
Glucocorticoid resistance is reverted by LCK inhibition in pediatric T-cell acute lymphoblastic leukemia Blood (IF 13.164) Pub Date : 2017-01-01 Valentina Serafin; Giorgia Capuzzo; Gloria Milani; Sonia Anna Minuzzo; Marica Pinazza; Roberta Bortolozzi; Silvia Bresolin; Elena Porcù; Chiara Frasson; Stefano Indraccolo; Giuseppe Basso; Benedetta Accordi
Pediatric T-acute lymphoblastic leukemia (T-ALL) patients often display resistance to glucocorticoid (GC) treatment. These patients, classified as Prednisone Poor Responders (PPR), have poorer outcome compared to the other pediatric T-ALL patients receiving a high-risk adapted therapy. Since glucocorticoids are administered to ALL patients during all the different phases of therapy, GC resistance represents an important challenge to be addressed in order to improve the outcome for these patients. Mechanisms underlying resistance are not yet fully unraveled, thus our research focused on the identification of deregulated signaling pathways to point out new targeted approaches. We first identified by Reverse Phase Protein Arrays the Lymphocyte Cell-Specific Protein-Tyrosine Kinase (LCK) as aberrantly activated in PPR patients. We showed that LCK inhibitors such as Dasatinib, Bosutinib, Nintedanib and WH-4-023 are able to induce cell death in GC resistant T-ALL cells and, remarkably, co-treatment with Dexamethasone is able to revert GC resistance even at therapeutic drug concentrations. This was confirmed by specific LCK gene silencing and ex vivo combined treatment of cells from PPR patient-derived xenografts. Moreover, we observed that LCK hyperactivation in PPR patients upregulates the CALCINEURIN/NFAT signaling triggering to IL-4 overexpression. GC sensitive cells cultured with IL-4 display an increased resistance to Dexamethasone, while the inhibition of IL-4 signaling was able to increase GC-induced apoptosis in resistant cells. Treatment with Dexamethasone and Dasatinib resulted also able to impair engraftment of leukemia cells in vivo. Our results suggest a quickly actionable approach to support conventional therapies and overcome GC resistance in pediatric T-ALL patients.
Human NOTCH4 is a key target of RUNX1 in megakaryocytic differentiation Blood (IF 13.164) Pub Date : 2017-01-01 Yueying Li; Chen Jin; Hao Bai; Yongxing Gao; Shu Sun; Lei Chen; Lei Qin; Paul P. Liu; Linzhao Cheng; Qian-Fei Wang
Megakaryocytes (MKs) in adult marrow produce platelets that play important roles in blood coagulation and hemostasis. Monoallelic mutations of the master transcription factor gene RUNX1 lead to familial platelet disorder (FPD) characterized by defective MK and platelet development. However, the molecular mechanisms of FPD remain unclear. Previously, we generated human induced pluripotent stem cells (iPSCs) from patients with FPD containing a RUNX1 nonsense mutation. Production of MKs from the FPD-iPSCs was reduced, and targeted correction of the RUNX1 mutation restored MK production. In this study, we utilized isogenic pairs of FPD-iPSCs and the MK differentiation system to identify RUNX1 target genes. Using integrative genomic analysis of hematopoietic progenitor cells generated from FPD iPSCs, and mutation-corrected isogenic controls, we identified two gene sets whose transcription is either up- or downregulated by RUNX1 in mutation-corrected iPSCs. Notably, NOTCH4 expression was negatively controlled by RUNX1 via a novel regulatory DNA element within the locus, and we examined its involvement in MK generation. Specific inactivation of NOTCH4 by an improved CRISPR-Cas9 system in human iPSCs enhanced megakaryopoiesis. Moreover, small molecules known to inhibit Notch signaling promoted MK generation from both normal human iPSCs and postnatal CD34+ hematopoietic stem and progenitor cells. Our study newly identified NOTCH4 as a RUNX1 target gene, and revealed a previously unappreciated role of NOTCH4 signaling in promoting human megakaryopoiesis. Our work suggests that human iPSCs with monogenic mutations have the potential to serve as an invaluable resource for discovery of novel druggable targets.
Some contents have been Reproduced by permission of The Royal Society of Chemistry.
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