bims-tremyl 2022-07-03 papers

bims-tremyl

Biomed News

on Therapy resistance biology in myeloid leukemia

Issue of 2022‒07‒03
24 papers selected by
Paolo Gallipoli
Barts Cancer Institute, Queen Mary University of London

Degradation of GSPT1 causes TP53-independent cell death in leukemia whilst sparing normal hematopoietic stem cells.
Combinatorial genetics reveals the Dock1-Rac2 axis as a potential target for the treatment of NPM1;Cohesin mutated AML.
Whole-genome CRISPR screening identifies N-glycosylation as a genetic and therapeutic vulnerability in CALR-mutant MPN.
CPX-351 Yields Similar Response and Survival Outcome in Younger and Older Patients With Secondary Acute Myeloid Leukemia.
Deciphering Metabolic Adaptability of Leukemic Stem Cells.
International Consensus Classification of Myeloid Neoplasms and Acute Leukemia: Integrating Morphological, Clinical, and Genomic Data.
HERVs characterize normal and leukemia stem cells and represent a source of shared epitopes for cancer immunotherapy.
Activating STING1-dependent immune signaling in TP53 mutant and wild-type acute myeloid leukemia.
Rare germline alterations of myeloperoxidase predispose to myeloid neoplasms.
Physioxia induced Downregulation of Tet2 in Hematopoietic Stem Cells contributes to Enhanced Self-renewal.
Murine Models of Acute Myeloid Leukemia.
A dual role for the RNA helicase DHX34 in NMD and pre-mRNA splicing and its function in hematopoietic differentiation.
STAT1 is essential for HSC function and maintains MHCIIhi stem cells that resists myeloablation and neoplastic expansion.
Molecular predictors of response to venetoclax plus hypomethylating agent in treatment-naïve acute myeloid leukemia.
How I Diagnose Acute Leukemia of Ambiguous Lineage.
Reduced intensity versus non-myeloablative conditioning regimen for haploidentical transplantation and post-transplantation cyclophosphamide in complete remission acute myeloid leukemia: a study from the ALWP of the EBMT.
Poor outcome of pediatric patients with acute myeloid leukemia harboring high FLT3/ITD allelic ratios.
Outcome of human umbilical cord blood stem cell transplantation (CBT) for acute myeloid leukemia in patients achieving first complete remission after one versus two induction courses: a study from the Acute Leukemia Working Party of the European Society for Blood and Marrow Transplantation (EBMT).
Choosing between intensive and less intensive front-line treatment approaches for older patients with newly diagnosed acute myeloid leukaemia.
GATA2 deficiency and MDS/AML: Experimental strategies for disease modelling and future therapeutic prospects.
Revision of RUNX1 Variant Curation Rules.
Reengineering Ponatinib to Minimize Cardiovascular Toxicity.
Dihydroorotate dehydrogenase inhibition reveals metabolic vulnerability in chronic myeloid leukemia.
Allosteric inhibition of PPM1D serine/threonine phosphatase via an altered conformational state.

J Clin Invest. 2022 Jun 28. pii: e153514. [Epub ahead of print]

Degradation of GSPT1 causes TP53-independent cell death in leukemia whilst sparing normal hematopoietic stem cells.

Rob S Sellar, Adam S Sperling, Mikołaj Słabicki, Jessica A Gasser, Marie E McConkey, Katherine A Donovan, Nada Mageed, Dylan N Adams, Charles Zou, Peter G Miller, Ravi Kumar Dutta, Steffen Boettcher, Amy E Lin, Brittany E Sandoval, Vanessa A Quevedo Barrios, Veronica Shkolnik, Jonas Koeppel, Elizabeth K Henderson, Emma C Fink, Lu Yang, Anthony Kn Chan, Sheela Pangeni Pokharel, Erik J Bergstrom, Rajan Burt, Namrata D Udeshi, Steven A Carr, Eric S Fischer, Chun-Wei Chen, Benjamin L Ebert.

  Targeted protein degradation is a rapidly advancing and expanding therapeutic approach. Drugs that degrade GSPT1 via the CRL4CRBN ubiquitin ligase are a new class of cancer therapy in active clinical development with evidence of activity against acute myeloid leukemia in early phase trials. However, other than activation of the integrated stress response, the downstream effects of GSPT1 degradation leading to cell death are largely undefined, and no murine models are available to study these agents. We identified the domains of GSPT1 essential for cell survival and show that GSPT1 degradation leads to impaired translation termination, activation of the integrated stress response pathway, and TP53-independent cell death. CRISPR-Cas9 screens implicated decreased translation initiation as protective to GSPT1 degradation, suggesting that cells with higher levels of translation are more susceptible to GSPT1 degradation. We defined two Crbn amino acids that prevent Gspt1 degradation in mice, generated a knock-in mouse with alteration of these residues, and demonstrated the efficacy of GSPT1-degrading drugs in vivo with relative sparing of numbers and function of long-term hematopoietic stem cells. Our results provide a mechanistic basis for the use of GSPT1 degraders for the treatment of cancer, including TP53-mutant AML.

Keywords:  Hematology; Leukemias; Mouse models; Therapeutics; Ubiquitin-proteosome system

DOI:  https://doi.org/10.1172/JCI153514
Leukemia. 2022 Jul 01.

Combinatorial genetics reveals the Dock1-Rac2 axis as a potential target for the treatment of NPM1;Cohesin mutated AML.

Alison E Meyer, Cary Stelloh, Kirthi Pulakanti, Robert Burns, Joseph B Fisher, Katelyn E Heimbruch, Sergey Tarima, Quinlan Furumo, John Brennan, Yongwei Zheng, Aaron D Viny, George S Vassiliou, Sridhar Rao.

Acute myeloid leukemia (AML) is driven by mutations that occur in numerous combinations. A better understanding of how mutations interact with one another to cause disease is critical to developing targeted therapies. Approximately 50% of patients that harbor a common mutation in NPM1 (NPM1cA) also have a mutation in the cohesin complex. As cohesin and Npm1 are known to regulate gene expression, we sought to determine how cohesin mutation alters the transcriptome in the context of NPM1cA. We utilized inducible Npm1cAflox/+ and core cohesin subunit Smc3flox/+ mice to examine AML development. While Npm1cA/+;Smc3Δ/+ mice developed AML with a similar latency and penetrance as Npm1cA/+ mice, RNA-seq suggests that the Npm1cA/+; Smc3Δ/+ mutational combination uniquely alters the transcriptome. We found that the Rac1/2 nucleotide exchange factor Dock1 was specifically upregulated in Npm1cA/+;Smc3Δ/+ HSPCs. Knockdown of Dock1 resulted in decreased growth and adhesion and increased apoptosis only in Npm1cA/+;Smc3Δ/+ AML. Higher Rac activity was also observed in Npm1cA/+;Smc3Δ/+ vs. Npm1cA/+ AMLs. Importantly, the Dock1/Rac pathway is targetable in Npm1cA/+;Smc3Δ/+ AMLs. Our results suggest that Dock1/Rac represents a potential target for the treatment of patients harboring NPM1cA and cohesin mutations and supports the use of combinatorial genetics to identify novel precision oncology targets.

DOI: https://doi.org/10.1038/s41375-022-01632-y
Blood. 2022 Jun 28. pii: blood.2022015629. [Epub ahead of print]

Whole-genome CRISPR screening identifies N-glycosylation as a genetic and therapeutic vulnerability in CALR-mutant MPN.

Jonas Samuel Jutzi, Anna E Marneth, Michele Ciboddo, Angel Guerra-Moreno, María José Jiménez-Santos, Anastasia Kosmidou, James W Dressman, Hongyan Liang, Rebecca Svenja Liselotte Hamel, Patricia Rosemary Lozano, Elisa Rumi, John Doench, Jason R Gotlib, Anandi Krishnan, Shannon Elf, Fatima Al-Shahrour, Ann Mullally.

Calreticulin (CALR) mutations are frequent, disease-initiating events in myeloproliferative neoplasms (MPN). Although the biological mechanism by which CALR mutations cause MPN has been elucidated, there currently are no clonally selective therapies for CALR-mutant MPN. To identify unique genetic dependencies in CALR-mutant MPN, we performed a whole-genome CRISPR knockout depletion screen in mutant CALR-transformed hematopoietic cells. We found that genes in the N-glycosylation pathway (amongst others) were differentially depleted in mutant CALR-transformed cells as compared with control cells. Using a focused pharmacological in vitro screen targeting unique vulnerabilities uncovered in the CRISPR screen, we found that chemical inhibition of N-glycosylation impaired the growth of mutant CALR-transformed cells, through a reduction in MPL cell surface expression. We treated Calr-mutant knockin mice with the N-glycosylation inhibitor, 2-deoxy-glucose (2-DG), and found a preferential sensitivity of Calr-mutant cells to 2-DG as compared to wild-type cells, and normalization of key MPN disease features. To validate our findings in primary human cells, we performed megakaryocyte colony-forming unit (CFU-MK) assays. We found that N-glycosylation inhibition significantly reduced CFU-MK formation in patient-derived CALR-mutant bone marrow, as compared to bone marrow-derived from healthy donors. In aggregate, our findings advance the development of clonally selective treatments for CALR-mutant MPN.

DOI: https://doi.org/10.1182/blood.2022015629
Clin Lymphoma Myeloma Leuk. 2022 Jun 05. pii: S2152-2650(22)00182-3. [Epub ahead of print]

CPX-351 Yields Similar Response and Survival Outcome in Younger and Older Patients With Secondary Acute Myeloid Leukemia.

Dasom Lee, Akriti G Jain, Yehuda Deutsch, Jennifer Eatrides, Onyee Chan, Eric Padron, Andrew Kuykendall, Rami Komrokji, Jeffrey Lancet, David Sallman, Chetasi Talati, Kendra Sweet.

  BACKGROUND: CPX-351 was approved by the FDA in 2017 as frontline induction chemotherapy for patients aged ≥18 years with newly diagnosed acute myeloid leukemia (AML) which includes myelodysplasia-related changes (AML-MRC) and therapy-related acute myeloid leukemia (t-AML). The efficacy of CPX-351 among younger patients (aged <60 years) is currently unclear, as the large, randomized phase 3 study that led to approval of CPX-351 only included patients between the ages of 60 and 75 years.METHODS: We performed a retrospective study of clinical and molecular data from adult patients with newly diagnosed AML-MRC or t-AML treated with CPX-351. Patients were divided into 2 cohorts: aged <60 years (cohort A) and aged ≥60 years (cohort B). We compared overall response rate (ORR) and median overall survival (mOS) between the cohorts.
RESULTS: Of 169 evaluable patients, 21.3% were in cohort A and 78.7% were in cohort B. ORR of the entire cohort was 53.3%; ORR of cohort A was 47.2% compared with 54.9% for cohort B (P = .46). Overall, 54.4% of responding patients proceeded to allogenic stem cell transplant (allo-SCT), including 52.9% of patients in cohort A and 54.8% in cohort B (P = 1.00). At a median follow-up of 24 months, mOS of the entire cohort was 16 months and was similar between cohorts A and B (18 vs. 15 months, respectively; P = .29).
CONCLUSION: CPX-351 resulted in similar response rates and survival outcomes among both younger and older adult patients with newly diagnosed AML-MRC or t-AML.

Keywords:  AML; Acute myeloid leukemia; CPX-351; Secondary AML; Young patients

DOI:  https://doi.org/10.1016/j.clml.2022.06.001
Front Oncol. 2022 ;12 846149

Deciphering Metabolic Adaptability of Leukemic Stem Cells.

Sweta B Patel, Travis Nemkov, Angelo D'Alessandro, Robert S Welner.

  Therapeutic targeting of leukemic stem cells is widely studied to control leukemia. An emerging approach gaining popularity is altering metabolism as a potential therapeutic opportunity. Studies have been carried out on hematopoietic and leukemic stem cells to identify vulnerable pathways without impacting the non-transformed, healthy counterparts. While many metabolic studies have been conducted using stem cells, most have been carried out in vitro or on a larger population of progenitor cells due to challenges imposed by the low frequency of stem cells found in vivo. This creates artifacts in the studies carried out, making it difficult to interpret and correlate the findings to stem cells directly. This review discusses the metabolic difference seen between hematopoietic stem cells and leukemic stem cells across different leukemic models. Moreover, we also shed light on the advancements of metabolic techniques and current limitations and areas for additional research of the field to study stem cell metabolism.

Keywords:  hematopoietic stem cells; leukemia; leukemic stem cells (LSCs); metabolic techniques; metabolism

DOI:  https://doi.org/10.3389/fonc.2022.846149
Blood. 2022 Jun 29. pii: blood.2022015850. [Epub ahead of print]

International Consensus Classification of Myeloid Neoplasms and Acute Leukemia: Integrating Morphological, Clinical, and Genomic Data.

Daniel A Arber, Attilio Orazi, Robert P Hasserjian, Michael J Borowitz, Katherine R Calvo, Hans Michael Kvasnicka, Sa A Wang, Adam Bagg, Tiziano Barbui, Susan Branford, Carlos E Bueso-Ramos, Jorge Cortes, Paola Dal Cin, Courtney D DiNardo, Hervé Dombret, Eric J Duncavage, Benjamin L Ebert, Elihu Estey, Fabio Facchetti, Kathryn Foucar, Naseema Gangat, Umberto Gianelli, Lucy A Godley, Nicola Goekbuget, Jason R Gotlib, Eva Hellström-Lindberg, Gabriela Hobbs, Ronald Hoffman, Elias J Jabbour, Jean-Jacques Kiladjian, Richard A Larson, Michelle M Le Beau, Mignon L Loh, Bob Löwenberg, Elizabeth A Macintyre, Luca Malcovati, Charles G Mullighan, Charlotte M Niemeyer, Olatoyosi Odenike, Seishi Ogawa, Alberto Orfao, Elli Papaemmanuil, Francesco Passamonti, Kimmo Porkka, Ching-Hong Pui, Jerald P Radich, Andreas Reiter, María Rozman, Martina Rudelius, Michael R Savona, Charles Schiffer, Annette Schmitt-Graeff, Akiko Shimamura, Jorge Sierra, Wendy Stock, Richard M Stone, Martin S Tallman, Juergen Thiele, Hwei-Fang Tien, Alexandar Tzankov, Alessandro M Vannucchi, Paresh Vyas, Andrew H Wei, Olga K Weinberg, Agnieszka Wierzbowska, Mario Cazzola, Hartmut Döhner, Ayalew Tefferi.

The classification of myeloid neoplasms and acute leukemias was last updated in 2016 within a collaboration between the World Health Organization (WHO), the Society for Hematopathology, and the European Association for Haematopathology. This collaboration was primarily based on input from a clinical advisory committees (CAC) composed of pathologists, hematologists, oncologists, geneticists, and bioinformaticians from around the world. The recent advances in our understanding of the biology of hematologic malignancies, the experience with the use of the 2016 WHO classification in clinical practice, and the results of clinical trials have indicated the need for further revising and updating the classification. As a continuation of this CAC-based process, the authors, a group with expertise in the clinical, pathologic and genetic aspects of these disorders, developed the International Consensus Classification (ICC) of myeloid neoplasms and acute leukemias. Using a multiparameter approach, the main objective of the consensus process was the definition of real disease entities, including the introduction of new entities and refined criteria for existing diagnostic categories, based on accumulated data. The ICC is aimed at facilitating diagnosis and prognostication of these neoplasms, improving treatment of affected patients, and allowing the design of innovative clinical trials.

DOI: https://doi.org/10.1182/blood.2022015850
Am J Hematol. 2022 Jun 27.

HERVs characterize normal and leukemia stem cells and represent a source of shared epitopes for cancer immunotherapy.

Vincent Alcazer, Paola Bonaventura, Laurie Tonon, Emilie Michel, Virginie Mutez, Clémentine Fabres, Nicolas Chuvin, Rasha Boulos, Yann Estornes, Véronique Maguer-Satta, Kevin Geistlich, Alain Viari, Klaus H Metzeler, Wolfgang Hiddemann, Aarif M N Batch, Tobias Herold, Christophe Caux, Stéphane Depil.

  Human endogenous retroviruses (HERVs) represent 8% of the human genome. The expression of HERVs and their immune impact have not been extensively studied in Acute Myeloid Leukemia (AML). In this study, we used a reference of 14,968 HERV functional units to provide a thorough analysis of HERV expression in normal and AML bone marrow cells. We show that the HERV retrotranscriptome accurately characterizes normal and leukemic cell subpopulations, including leukemia stem cells, in line with different epigenetic profiles. We then show that HERV expression delineates AML subtypes with different prognoses. We finally propose a method to select and prioritize CD8+ T cell epitopes derived from AML-specific HERVs and we show that lymphocytes infiltrating patient bone marrow at diagnosis contain naturally occurring CD8+ T cells against these HERV epitopes. We also provide in vitro data supporting the functionality of HERV-specific CD8+ T-cells against AML cells. These results show that HERVs represent an important source of genetic information that can help enhancing disease stratification or biomarker identification and an important reservoir of alternative tumor-specific T cell epitopes relevant for cancer immunotherapy. This article is protected by copyright. All rights reserved.

Keywords:  Acute Myeloid Leukemia; Cancer immunotherapy; Epitopes; Human Endogenous Retroviruses; Leukemic Stem Cells

DOI:  https://doi.org/10.1002/ajh.26647
Proc Natl Acad Sci U S A. 2022 Jul 05. 119(27): e2123227119

Activating STING1-dependent immune signaling in TP53 mutant and wild-type acute myeloid leukemia.

Aksinija A Kogan, Michael J Topper, Anna J Dellomo, Lora Stojanovic, Lena J McLaughlin, T Michael Creed, Christian L Eberly, Tami J Kingsbury, Maria R Baer, Michael D Kessler, Stephen B Baylin, Feyruz V Rassool.

  DNA methyltransferase inhibitors (DNMTis) reexpress hypermethylated genes in cancers and leukemias and also activate endogenous retroviruses (ERVs), leading to interferon (IFN) signaling, in a process known as viral mimicry. In the present study we show that in the subset of acute myeloid leukemias (AMLs) with mutations in TP53, associated with poor prognosis, DNMTis, important drugs for treatment of AML, enable expression of ERVs and IFN and inflammasome signaling in a STING-dependent manner. We previously reported that in solid tumors poly ADP ribose polymerase inhibitors (PARPis) combined with DNMTis to induce an IFN/inflammasome response that is dependent on STING1 and is mechanistically linked to generation of a homologous recombination defect (HRD). We now show that STING1 activity is actually increased in TP53 mutant compared with wild-type (WT) TP53 AML. Moreover, in TP53 mutant AML, STING1-dependent IFN/inflammatory signaling is increased by DNMTi treatment, whereas in AMLs with WT TP53, DNMTis alone have no effect. While combining DNMTis with PARPis increases IFN/inflammatory gene expression in WT TP53 AML cells, signaling induced in TP53 mutant AML is still several-fold higher. Notably, induction of HRD in both TP53 mutant and WT AMLs follows the pattern of STING1-dependent IFN and inflammatory signaling that we have observed with drug treatments. These findings increase our understanding of the mechanisms that underlie DNMTi + PARPi treatment, and also DNMTi combinations with immune therapies, suggesting a personalized approach that statifies by TP53 status, for use of such therapies, including potential immune activation of STING1 in AML and other cancers.

Keywords:  AML; TP53; combination therapy; epigenetics; immune signaling

DOI:  https://doi.org/10.1073/pnas.2123227119
Leukemia. 2022 Jun 27.

Rare germline alterations of myeloperoxidase predispose to myeloid neoplasms.

Sunisa Kongkiatkamon, Laila Terkawi, Yihong Guan, Vera Adema, Metis Hasipek, Tatiana Dombrovski, Milo Co, Wencke Walter, Hassan Awada, Yvonne Parker, Stephan Hutter, Simona Pagliuca, Carmelo Gurnari, Heesun J Rogers, Manja Meggendorfer, Daniel J Lindner, Torsten Haferlach, Valeria Visconte, Thomas LaFramboise, Babal K Jha, Jaroslaw P Maciejewski.

Myeloperoxidase (MPO) gene alterations with variable clinical penetrance have been found in hereditary MPO deficiency, but their leukemia association in patients and carriers has not been established. Germline MPO alterations were found to be significantly enriched in myeloid neoplasms: 28 pathogenic/likely pathogenic variants were identified in 100 patients. The most common alterations were c.2031-2 A > C, R569W, M519fs* and Y173C accounting for about half of the cases. While functional experiments showed that the marrow stem cell pool of Mpo-/- mice was not increased, using competitive repopulation demonstrated that Mpo-/- grafts gained growth advantage over MPO wild type cells. This finding also correlated with increased clonogenic potential after serial replating in the setting of H2O2-induced oxidative stress. Furthermore, we demonstrated that H2O2-induced DNA damage and activation of error-prone DNA repair may result in secondary genetic damage potentially predisposing to leukemia leukemic evolution. In conclusion, our study for the first time demonstrates that germline MPO variants may constitute risk alleles for MN evolution.

DOI: https://doi.org/10.1038/s41375-022-01630-0
Blood. 2022 Jun 30. pii: blood.2022015499. [Epub ahead of print]

Physioxia induced Downregulation of Tet2 in Hematopoietic Stem Cells contributes to Enhanced Self-renewal.

Arafat Aljoufi, Chi Zhang, James Ropa, Wennan Chang, Lakshmi Reddy Palam, Scott Cooper, Baskar Ramdas, Maegan L Capitano, Hal E Broxmeyer, Reuben Kapur.

Hematopoietic stem cells (HSCs) manifest impaired recovery and self-renewal with a concomitant increase in differentiation when exposed to ambient air as opposed to physioxia. Mechanism(s) behind this distinction are poorly understood but have the potential to improve stem cell transplantation. Single cell RNA-seq of HSCs in physioxia revealed upregulation of HSC self-renewal genes, and downregulation of genes involved in inflammatory pathways and HSC differentiation. HSCs under physioxia also exhibited downregulation of the epigenetic modifier Tet2. Tet2 is α-ketoglutarate (α-KG), iron and oxygen-dependent dioxygenase that converts 5-mC to 5-hmC thereby promoting active transcription. We evaluated whether loss of Tet2 affects the number and function of HSCs and hematopoietic progenitor cells (HPCs) under physioxia and ambient air. In contrast to WT-HSCs, a complete non-responsiveness of Tet2-/-HSCs and HPCs to changes in oxygen tension was observed. Unlike WT-HSCs, Tet2-/-HSCs and HPCs exhibited similar numbers and function in either physioxia or ambient air. The lack of response to changes in oxygen tension in Tet2-/- HSCs was associated with similar changes in self-renewal and quiescence genes among WT-HSC-physioxia, Tet2-/-HSC-physioxia and Tet2-/-HSC-air. We define a novel molecular program involving Tet2 in regulating HSCs under physioxia.

DOI: https://doi.org/10.1182/blood.2022015499
Front Oncol. 2022 ;12 854973

Murine Models of Acute Myeloid Leukemia.

Kristen J Kurtz, Shannon E Conneely, Madeleine O'Keefe, Katharina Wohlan, Rachel E Rau.

  Acute myeloid leukemia (AML) is a phenotypically and genetically heterogeneous hematologic malignancy. Extensive sequencing efforts have mapped the genomic landscape of adult and pediatric AML revealing a number of biologically and prognostically relevant driver lesions. Beyond identifying recurrent genetic aberrations, it is of critical importance to fully delineate the complex mechanisms by which they contribute to the initiation and evolution of disease to ultimately facilitate the development of targeted therapies. Towards these aims, murine models of AML are indispensable research tools. The rapid evolution of genetic engineering techniques over the past 20 years has greatly advanced the use of murine models to mirror specific genetic subtypes of human AML, define cell-intrinsic and extrinsic disease mechanisms, study the interaction between co-occurring genetic lesions, and test novel therapeutic approaches. This review summarizes the mouse model systems that have been developed to recapitulate the most common genomic subtypes of AML. We will discuss the strengths and weaknesses of varying modeling strategies, highlight major discoveries emanating from these model systems, and outline future opportunities to leverage emerging technologies for mechanistic and preclinical investigations.

Keywords:   acute myeliod leukemia; AML; KMT2a (MLL) rearrangements; NUP98 fusion; core binding factor acute myeliod leukemia; genetically engineered mice (GEM); patient-derived xenograft (PDX); transgenic mouse

DOI:  https://doi.org/10.3389/fonc.2022.854973
RNA. 2022 Jun 29. pii: rna.079277.122. [Epub ahead of print]

A dual role for the RNA helicase DHX34 in NMD and pre-mRNA splicing and its function in hematopoietic differentiation.

Nele Hug, Stuart Aitken, Dasa Longman, Michaela Raab, Hannah Armes, Abigail R Mann, Ana Rio-Machin, Jude Fitzgibbon, Kevin Rouault-Pierre, Javier F Caceres.

  The DExD/H-box RNA helicase DHX34 is a Nonsense-mediated decay (NMD) factor that together with core NMD factors co-regulates NMD targets in nematodes and in vertebrates. Here, we show that DHX34 is also associated with the human spliceosomal catalytic C complex. Mapping of DHX34 endogenous binding sites using Cross-Linking Immunoprecipitation (CLIP) revealed that DHX34 is preferentially associated with pre-mRNAs and locates at exon-intron boundaries. Accordingly, we observed that DHX34 regulates a large number of alternative splicing (AS) events in mammalian cells in culture, establishing a dual role for DHX34 in both NMD and pre-mRNA splicing. We previously showed that germline DHX34 mutations associated to familial Myelodysplasia (MDS)/Acute Myeloid Leukemia (AML) predisposition abrogate its activity in NMD. Interestingly, we observe now that DHX34 regulates the splicing of pre-mRNAs that have been linked to AML/MDS predisposition. This is consistent with silencing experiments in hematopoietic stem/progenitor cells (HSPCs) showing that loss of DHX34 results in differentiation blockade of both erythroid and myeloid lineages, which is a hallmark of AML development. Altogether, these data unveil new cellular functions of DHX34 and suggests that alterations in the levels and/or activity of DHX34 could contribute to human disease.

Keywords:  AML; DHX34; NMD; Pre-mRNA splicing; seCLIP

DOI:  https://doi.org/10.1261/rna.079277.122
Blood. 2022 Jun 29. pii: blood.2021014009. [Epub ahead of print]

STAT1 is essential for HSC function and maintains MHCIIhi stem cells that resists myeloablation and neoplastic expansion.

Juan Li, Matthew Joseph Williams, Hyun Jung Park, Hugo P Bastos, Xiaonan Wang, Daniel Prins Prins, Nicola K Wilson, Carys Johnson, Kendig Sham, Michelle Wantoch, Sam Watcham, Sarah Jane Kinston, Dean C Pask, Tina L Hamilton, Rachel Sneade, Amie Karen Waller, Cedric Ghevaert, George S Vassiliou, Elisa Laurenti, David G Kent, Bertie Gottgens, Anthony R Green.

Adult hematopoietic stem cells (HSCs) are predominantly quiescent and can be activated in response to acute stress such as infection or cytotoxic insults. STAT1 is a pivotal downstream mediator of interferon (IFN) signaling and is required for IFN-induced HSC proliferation, but little is known about the role of STAT1 in regulating homeostatic hematopoietic stem/progenitor cells (HSPCs). Here we show that loss of STAT1 altered the steady state HSPC landscape, impaired HSC function in transplantation assays, delayed blood cell regeneration following myeloablation, and disrupted molecular programs which protect HSCs including control of quiescence. Our results also reveal STAT1-dependent functional HSC heterogeneity. A previously unrecognized subset of homeostatic HSCs with elevated MHCII expression (MHCIIhi) displayed molecular features of reduced cycling and apoptosis, and was refractory to 5-FU induced myeloablation. Conversely, MHCIIlo HSCs displayed increased megakaryocytic potential and were preferentially expanded in CALR mutant mice with thrombocytosis. Similar to mice, high MHCII expression is a feature of human HSCs residing in a deeper quiescent state. Our results therefore position STAT1 at the interface of stem cell heterogeneity and the interplay between stem cells and the adaptive immune system, areas of broad interest in the wider stem cell field.

DOI: https://doi.org/10.1182/blood.2021014009
Haematologica. 2022 Jun 30.

Molecular predictors of response to venetoclax plus hypomethylating agent in treatment-naïve acute myeloid leukemia.

Naseema Gangat, Isla Johnson, Kristen McCullough, Faiqa Farrukh, Aref Al-Kali, Hassan Alkhateeb, Kebede Begna, Abhishek Mangaonkar, Mark Litzow, William Hogan, Mithun Shah, Mrinal Patnaik, Animesh Pardanani, Ayalew Tefferi.

Not available.

DOI: https://doi.org/10.3324/haematol.2022.281214
Am J Clin Pathol. 2022 Jul 01. 158(1): 27-34

How I Diagnose Acute Leukemia of Ambiguous Lineage.

Olga K Weinberg, Daniel A Arber.

  OBJECTIVES: Classification of acute leukemia involves assigning lineage by resemblance to normal progenitor cells. This approach provides descriptive information about the blast cells that is useful for disease monitoring, provides clues to pathogenesis, and can help clinicians select effective chemotherapeutic regimens. Acute leukemias of ambiguous lineage (ALALs) are those leukemias that either fail to show evidence of myeloid, B-, or T-lymphoid lineage commitment or show evidence of commitment to more than 1 lineage. The different treatment regimens for acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) make ALAL a challenge both diagnostically and therapeutically.METHODS: Current classification criteria have reduced the reported incidence of mixed-lineage leukemias by emphasizing fewer markers and categorizing some biphenotypic leukemias with recurrent cytogenetic abnormalities as other entities. Several recent studies have explored the genomic and epigenetic landscape of mixed-phenotype acute leukemia (MPAL) and have suggested a further refinement of the World Health Organization classification to emphasize the genomic heterogeneity of MPAL.
RESULTS: Genomic and expression profile data for MPAL reveal mutations commonly seen in both AML and ALL, with T-/myeloid MPAL showing overlapping features with early T-cell precursor lymphoblastic leukemia.
CONCLUSIONS: Our review aimed to discuss the diagnostic challenges, recent genomic studies, and therapeutic strategies in this poorly understood disease.

Keywords:  Acute undifferentiated leukemia; Genetics; Mixed-phenotype acute leukemia

DOI:  https://doi.org/10.1093/ajcp/aqac070
Bone Marrow Transplant. 2022 Jun 25.

Reduced intensity versus non-myeloablative conditioning regimen for haploidentical transplantation and post-transplantation cyclophosphamide in complete remission acute myeloid leukemia: a study from the ALWP of the EBMT.

Raynier Devillier, Jacques-Emmanuel Galimard, Myriam Labopin, Didier Blaise, Anna Maria Raiola, Jiri Pavlu, Luca Castagna, Gerard Socié, Yves Chalandon, Massimo Martino, Friedrich Stölzel, Gesine Bug, Benedetto Bruno, Radovan Vrhovac, Amandine Charbonnier, Attilio Olivieri, Jacques-Olivier Bay, Herrera Arroyo, Ibrahim Yakoub-Agha, Daniele Avenoso, Andreas Neubauer, Stéphanie Nguyen, Edouard Forcade, Eolia Brissot, Bipin Savani, Arnon Nagler, Mohamad Mohty.

The optimal conditioning regimen prior haploidentical stem cell transplantation (Haplo-SCT) with post transplantation cyclophosphamide (PT-Cy) for acute myeloid leukemia (AML) remains unknown. A non-myeloablative conditioning (NMAC) regimen (cyclophosphamide + fludarabine + TBI 2 Gy [CyFluTBI]) is a safe approach, but relapse incidence remains high in this setting. Alternatively, a reduced intensity conditioning (RIC) regimen combining thiotepa and reduced-dose busulfan with fludarabine (TBF) may decrease AML relapse. However, an excess of toxicity may counterbalance this potential benefit. We retrospectively compared CyFluTBI vs. TBF in CR AML patients who underwent Haplo-SCT with PT-Cy, in two different populations based on age. We analyzed 490 patients. In patients aged <60 years (n = 203), we observed a higher RI (HR = 3.59, 95% CI = 1.75-7.37, p < 0.01), lower LFS (HR = 1.98, 95% CI = 1.22-3.22, p < 0.01) and lower OS (HR = 1.73, 95% CI = 1.04-2.88, p = 0.04) in the CyFluTBI group, without significant difference in NRM. In older patients (n = 287), we observed that conditioning regimen did not significantly influence LFS (HR = 0.90, 95% CI = 0.56-1.44, p = 0.65), OS (HR = 0.81, 95% CI = 0.49-1.32, p = 0.39) and RI (HR = 1.78, 95% CI = 0.90-3.50, p = 0.10), but showed that CyFluTBI was associated with a significantly lower risk of NRM (HR = 0.48, 95% CI = 0.25-0.92, p = 0.03). Thus, younger patients seem to benefit from conditioning intensification from CyFluTBI to TBF regimens prior PT-Cy Haplo-SCT for CR AML, while older ones do not.

DOI: https://doi.org/10.1038/s41409-022-01674-x
Nat Commun. 2022 Jun 27. 13(1): 3679

Poor outcome of pediatric patients with acute myeloid leukemia harboring high FLT3/ITD allelic ratios.

Kun-Yin Qiu, Xiong-Yu Liao, Yong Liu, Ke Huang, Yang Li, Jian-Pei Fang, Dun-Hua Zhou.

Activating FLT3 mutations are the most common mutations in acute myeloid leukemia (AML), but the optimal threshold of FLT3/ITD allelic ratio (AR) among pediatric AML patients remains controversial. Here, we present the outcome and prognostic significance of FLT3/ITD AR analysis among pediatric patients with AML from the TARGET dataset. Applying fitting curve models and threshold effect analysis using the restrictive cubic spline function following Cox proportional hazards models identifies the cut-off value of 0.5 on FLT3/ITD AR. Moreover, we observe that high FLT3/ITD AR patients have an inferior outcome when compared to low AR patients. Our study also demonstrates that stem cell transplantation may improve the outcome in pediatric AML patients with high FLT3/ITD AR and may be further improved when combined with additional therapies such as Gemtuzumab Ozogamicin. These findings underline the importance of individualized treatment of pediatric AML.

DOI: https://doi.org/10.1038/s41467-022-31489-9
Bone Marrow Transplant. 2022 Jun 30.

Outcome of human umbilical cord blood stem cell transplantation (CBT) for acute myeloid leukemia in patients achieving first complete remission after one versus two induction courses: a study from the Acute Leukemia Working Party of the European Society for Blood and Marrow Transplantation (EBMT).

Arnon Nagler, Myriam Labopin, Jan J Cornelissen, Edouard Forcade, Patrice Chevallier, Nathalie Fegueux, Jorge Sierra, Deborah Desmier, Hélène Labussière-Wallet, Jenny L Byrne, Michael Loschi, Didier Blaise, Frederic Baron, Annalisa Ruggeri, Mohamad Mohty.

We compared transplantation outcomes of adult patients with AML that underwent cord blood transplantation (CBT) in CR1 following 1 versus 2 induction courses. Study included 325 patients, 243 (75%) with 1 and 82 (25%) with 2 induction courses. Engraftment was lower for patients achieving CR1 after 1 vs. 2 induction courses: 91% vs. 99% (p = 0.02). Incidence of acute GVHD was similar, 38% and 36% (p = 0.81), as was 2-year chronic GVHD at 23.4% and 27.5%, respectively (p = 0.65). Two-year non-relapse mortality (NRM), relapse incidence (RI), leukemia-free survival (LFS), overall survival (OS) and GVHD-free, relapse-free survival (GRFS) were not statistically different between patients achieving CR1 with 1 vs. 2 induction courses with 23% vs. 24% (p = 0.87), 25% vs. 30% (p = 0.4), 52% vs. 46% (p = 0.3), 59% vs. 50% (p = 0.2), and 44% vs. 41% (p = 0.66), respectively. Results were confirmed by multivariable analysis, NRM (hazard ratio (HR) = 1.1; 95% CI, 0.6-1.8, p = 0.7), RI (HR = 1.4; 95% CI, 0.9-2.3, p = 0.1), LFS (HR = 1.3; 95% CI, 0.9-1.8, p = 0.2), OS (HR = 1.3; 95% CI, 0.9-1.9, p = 0.1), and GRFS (HR = 1.1; 95% CI, 0.8-1.5, p = 0.5). Overall, outcomes of AML patients undergoing CBT in CR1 achieved after 1 or 2 induction courses are similar.

DOI: https://doi.org/10.1038/s41409-022-01742-2
Lancet Haematol. 2022 Jul;pii: S2352-3026(22)00167-3. [Epub ahead of print]9(7): e535-e545

Choosing between intensive and less intensive front-line treatment approaches for older patients with newly diagnosed acute myeloid leukaemia.

Nicholas J Short, Hagop Kantarjian.

The outcomes of older patients with acute myeloid leukaemia are inferior to their younger counterparts, because, in part, of a more aggressive disease biology and poorer tolerance of cytotoxic chemotherapy. Although intensive chemotherapy was historically considered the only effective treatment for these patients, many older patients are not suitable for intensive chemotherapy owing to comorbidities or general frailty. Determination of patient fitness for intensive chemotherapy is imperfect, and even older patients who appear to be suitable (also known as fit) for intensive chemotherapy can have high rates of morbidity and early and late mortality with this approach. Fortunately, the outcomes of older or unfit patients with acute myeloid leukaemia have substantially improved with the use of a hypomethylating agent plus venetoclax in the front-line setting. Although the formal approval of this combination is limited to patients aged 75 years or older, or those with a clinically significant comorbidity, the high response rates and survival improvement in these patients have led many practitioners to consider this low-intensity regimen in older patients without significant comorbidities and even in younger patients with high-risk disease features for whom the expected outcomes with intensive chemotherapy are poor. Modifications to the hypomethylating agent plus venetoclax backbone might further improve the outlook for these patients, particularly in some acute myeloid leukaemia subsets with a targetable mutation. In this Viewpoint, we review the retrospective and prospective data supporting both intensive chemotherapy and low-intensity venetoclax-based approaches in older patients with acute myeloid leukaemia. We also discuss our own approach to the management of older or unfit patients with acute myeloid leukaemia, including how cytomolecular features have a role in establishing the optimal front-line therapy.

DOI: https://doi.org/10.1016/S2352-3026(22)00167-3
Br J Haematol. 2022 Jun 26.

GATA2 deficiency and MDS/AML: Experimental strategies for disease modelling and future therapeutic prospects.

Lili Kotmayer, Damia Romero-Moya, Oskar Marin-Bejar, Emilia Kozyra, Albert Català, Anna Bigas, Marcin W Wlodarski, Csaba Bödör, Alessandra Giorgetti.

  The importance of predisposition to leukaemia in clinical practice is being increasingly recognized. This is emphasized by the establishment of a novel WHO disease category in 2016 called "myeloid neoplasms with germline predisposition". A major syndrome within this group is GATA2 deficiency, a heterogeneous immunodeficiency syndrome with a very high lifetime risk to develop myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML). GATA2 deficiency has been identified as the most common hereditary cause of MDS in adolescents with monosomy 7. Allogenic haematopoietic stem cell transplantation is the only curative option; however, chances of survival decrease with progression of immunodeficiency and MDS evolution. Penetrance and expressivity within families carrying GATA2 mutations is often variable, suggesting that co-operating extrinsic events are required to trigger the disease. Predictive tools are lacking, and intrafamilial heterogeneity is poorly understood; hence there is a clear unmet medical need. On behalf of the ERAPerMed GATA2 HuMo consortium, in this review we describe the genetic, clinical, and biological aspects of familial GATA2-related MDS, highlighting the importance of developing robust disease preclinical models to improve early detection and clinical decision-making of GATA2 carriers.

Keywords:  GATA2 deficiency; acute myeloid leukaemia; blood cancer; myelodysplastic syndromes

DOI:  https://doi.org/10.1111/bjh.18330
Blood Adv. 2022 Jun 28. pii: bloodadvances.2022008017. [Epub ahead of print]

Revision of RUNX1 Variant Curation Rules.

Simone K Feurstein, Xi Luo, Mancy Shah, Taylor Walker, Nikita Mehta, David Wu, Lucy A Godley.

DOI: https://doi.org/10.1182/bloodadvances.2022008017
Cancer Res. 2022 May 31. pii: canres.CAN-21-3652-E.2021. [Epub ahead of print]

Reengineering Ponatinib to Minimize Cardiovascular Toxicity.

Anna Pavlovna Hnatiuk, Arne A N Bruyneel, Dhanir Tailor, Mallesh Pandrala, Arpit Dheeraj, Wenqi Li, Ricardo Serrano, Dries A M Feyen, Michelle M Vu, Prashila Amatya, Saloni Gupta, Yusuke Nakauchi, Isabel Morgado, Volker Wiebking, Ronglih Liao, Matthew Porteus, Ravindra Majeti, Sanjay V Malhotra, Mark Mercola.

Small molecule Tyrosine Kinase Inhibitors (TKIs) have revolutionized cancer treatment and greatly improved patient survival. However, life-threatening cardiotoxicity of many TKIs has become a major concern. Ponatinib (ICLUSIG) was developed as an inhibitor of the BCR-ABL oncogene and is among the most cardiotoxic of TKIs. Consequently, use of ponatinib is restricted to the treatment of tumors carrying T315I-mutated BCR-ABL, which occurs in chronic myeloid leukemia (CML) and confers resistance to first- and second-generation inhibitors such as imatinib and nilotinib. Through parallel screening of cardiovascular toxicity and anti-tumor efficacy assays, we engineered safer analogs of ponatinib that retained potency against T315I BCR-ABL kinase activity and suppressed T315I mutant CML tumor growth. The new compounds were substantially less toxic in human cardiac vasculogenesis and cardiomyocyte contractility assays in vitro. The compounds showed a larger therapeutic window in vivo, leading to regression of human T315I mutant CML xenografts without cardiotoxicity. Comparison of the kinase inhibition profiles of ponatinib and the new compounds suggested that ponatinib cardiotoxicity is mediated by a few kinases, some of which were previously unassociated with cardiovascular disease. Overall, the study develops an approach using complex phenotypic assays to reduce the high risk of cardiovascular toxicity that is prevalent among small molecule oncology therapeutics.

DOI: https://doi.org/10.1158/0008-5472.CAN-21-3652
Cell Death Dis. 2022 Jun 30. 13(6): 576

Dihydroorotate dehydrogenase inhibition reveals metabolic vulnerability in chronic myeloid leukemia.

Mohammad Houshmand, Nicoletta Vitale, Francesca Orso, Alessandro Cignetti, Ivan Molineris, Valentina Gaidano, Stefano Sainas, Marta Giorgis, Donatella Boschi, Carmen Fava, Alice Passoni, Marta Gai, Massimo Geuna, Federica Sora, Alessandra Iurlo, Elisabetta Abruzzese, Massimo Breccia, Olga Mulas, Giovanni Caocci, Fausto Castagnetti, Daniela Taverna, Salvatore Oliviero, Fabrizio Pane, Marco Lucio Lolli, Paola Circosta, Giuseppe Saglio.

The development of different generations of BCR-ABL1 tyrosine kinase inhibitors (TKIs) has led to the high overall survival of chronic myeloid leukemia (CML) patients. However, there are CML patients who show resistance to TKI therapy and are prone to progress to more advanced phases of the disease. So, implementing an alternative approach for targeting TKIs insensitive cells would be of the essence. Dihydroorotate dehydrogenase (DHODH) is an enzyme in the de novo pyrimidine biosynthesis pathway that is located in the inner membrane of mitochondria. Here, we found that CML cells are vulnerable to DHODH inhibition mediated by Meds433, a new and potent DHODH inhibitor recently developed by our group. Meds433 significantly activates the apoptotic pathway and leads to the reduction of amino acids and induction of huge metabolic stress in CML CD34+ cells. Altogether, our study shows that DHODH inhibition is a promising approach for targeting CML stem/progenitor cells and may help more patients discontinue the therapy.

DOI: https://doi.org/10.1038/s41419-022-05028-9
Nat Commun. 2022 Jun 30. 13(1): 3778

Allosteric inhibition of PPM1D serine/threonine phosphatase via an altered conformational state.

Peter G Miller, Murugappan Sathappa, Jamie A Moroco, Wei Jiang, Yue Qian, Sumaiya Iqbal, Qi Guo, Andrew O Giacomelli, Subrata Shaw, Camille Vernier, Besnik Bajrami, Xiaoping Yang, Cerise Raffier, Adam S Sperling, Christopher J Gibson, Josephine Kahn, Cyrus Jin, Matthew Ranaghan, Alisha Caliman, Merissa Brousseau, Eric S Fischer, Robert Lintner, Federica Piccioni, Arthur J Campbell, David E Root, Colin W Garvie, Benjamin L Ebert.

PPM1D encodes a serine/threonine phosphatase that regulates numerous pathways including the DNA damage response and p53. Activating mutations and amplification of PPM1D are found across numerous cancer types. GSK2830371 is a potent and selective allosteric inhibitor of PPM1D, but its mechanism of binding and inhibition of catalytic activity are unknown. Here we use computational, biochemical and functional genetic studies to elucidate the molecular basis of GSK2830371 activity. These data confirm that GSK2830371 binds an allosteric site of PPM1D with high affinity. By further incorporating data from hydrogen deuterium exchange mass spectrometry and sedimentation velocity analytical ultracentrifugation, we demonstrate that PPM1D exists in an equilibrium between two conformations that are defined by the movement of the flap domain, which is required for substrate recognition. A hinge region was identified that is critical for switching between the two conformations and was directly implicated in the high-affinity binding of GSK2830371 to PPM1D. We propose that the two conformations represent active and inactive forms of the protein reflected by the position of the flap, and that binding of GSK2830371 shifts the equilibrium to the inactive form. Finally, we found that C-terminal truncating mutations proximal to residue 400 result in destabilization of the protein via loss of a stabilizing N- and C-terminal interaction, consistent with the observation from human genetic data that nearly all PPM1D mutations in cancer are truncating and occur distal to residue 400. Taken together, our findings elucidate the mechanism by which binding of a small molecule to an allosteric site of PPM1D inhibits its activity and provides insights into the biology of PPM1D.

DOI: https://doi.org/10.1038/s41467-022-30463-9