bims-tremyl Biomed News
on Therapy resistance biology in myeloid leukemia
Issue of 2021‒02‒14
twenty-five papers selected by
Paolo Gallipoli
Barts Cancer Institute, Queen Mary University of London

  1. Cancer Discov. 2021 Feb 12. pii: candisc.1353.2020. [Epub ahead of print]
    Marinaccio C, Suraneni P, Celik H, Volk A, Wen QJ, Ling T, Bulic M, Lasho T, Koche RP, Famulare CA, Farnoud N, Stein B, Schieber M, Gurbuxani S, Root DE, Younger ST, Hoffman R, Gangat N, Ntziachristos P, Chandel NS, Levine RL, Rampal RK, Challen GA, Tefferi A, Crispino JD.
      The myeloproliferative neoplasms frequently progress to blast phase disease, an aggressive form of acute myeloid leukemia. To identify genes that suppress disease progression, we performed a focused CRISPR/Cas9 screen and discovered that depletion of LKB1/Stk11 led to enhanced in vitro self-renewal of murine MPN cells. Deletion of Stk11 in a mouse MPN model caused rapid lethality with enhanced fibrosis, osteosclerosis and an accumulation immature cells in the bone marrow, as well as enhanced engraftment of primary human MPN cells in vivo. LKB1 loss was associated with increased mitochondrial ROS and stabilization of HIF1a, and downregulation of LKB1 and increased levels of HIF1a were observed in human blast phase MPN specimens. Of note, we observed strong concordance of pathways that were enriched in murine MPN cells with LKB1 loss with those enriched in blast phase MPN patient specimens, supporting the conclusion that STK11 is a tumor suppressor in the MPNs.
  2. Blood Adv. 2021 Feb 09. 5(3): 687-699
    Krutein MC, Hart MR, Anderson DJ, Jeffery J, Kotini AG, Dai J, Chien S, DelPriore M, Borst S, Maguire JA, French DL, Gadue P, Papapetrou EP, Keel SB, Becker PS, Horwitz MS.
      RUNX1 familial platelet disorder (RUNX1-FPD) is an autosomal dominant disorder caused by a monoallelic mutation of RUNX1, initially resulting in approximately half-normal RUNX1 activity. Clinical features include thrombocytopenia, platelet functional defects, and a predisposition to leukemia. RUNX1 is rapidly degraded through the ubiquitin-proteasome pathway. Moreover, it may autoregulate its expression. A predicted kinetic property of autoregulatory circuits is that transient perturbations of steady-state levels result in continued maintenance of expression at adjusted levels, even after inhibitors of degradation or inducers of transcription are withdrawn, suggesting that transient inhibition of RUNX1 degradation may have prolonged effects. We hypothesized that pharmacological inhibition of RUNX1 protein degradation could normalize RUNX1 protein levels, restore the number of platelets and their function, and potentially delay or prevent malignant transformation. In this study, we evaluated cell lines, induced pluripotent stem cells derived from patients with RUNX1-FPD, RUNX1-FPD primary bone marrow cells, and acute myeloid leukemia blood cells from patients with RUNX1 mutations. The results showed that, in some circumstances, transient expression of exogenous RUNX1 or inhibition of steps leading to RUNX1 ubiquitylation and proteasomal degradation restored RUNX1 levels, thereby advancing megakaryocytic differentiation in vitro. Thus, drugs retarding RUNX1 proteolytic degradation may represent a therapeutic avenue for treating bleeding complications and preventing leukemia in RUNX1-FPD.
  3. Cell Stem Cell. 2021 Feb 02. pii: S1934-5909(21)00011-4. [Epub ahead of print]
    Wang T, Pine AR, Kotini AG, Yuan H, Zamparo L, Starczynowski DT, Leslie C, Papapetrou EP.
      Human cancers arise through the sequential acquisition of somatic mutations that create successive clonal populations. Human cancer evolution models could help illuminate this process and inform therapeutic intervention at an early disease stage, but their creation has faced significant challenges. Here, we combined induced pluripotent stem cell (iPSC) and CRISPR-Cas9 technologies to develop a model of the clonal evolution of acute myeloid leukemia (AML). Through the stepwise introduction of three driver mutations, we generated iPSC lines that, upon hematopoietic differentiation, capture distinct premalignant stages, including clonal hematopoiesis (CH) and myelodysplastic syndrome (MDS), culminating in a transplantable leukemia, and recapitulate transcriptional and chromatin accessibility signatures of primary human MDS and AML. By mapping dynamic changes in transcriptomes and chromatin landscapes, we characterize transcriptional programs driving specific transitions between disease stages. We identify cell-autonomous dysregulation of inflammatory signaling as an early and persistent event in leukemogenesis and a promising early therapeutic target.
    Keywords:  AML; IRAK1 inhibitor; IRAK4 inhibitor; UBE2N inhibitor; clonal evolution; gene editing; hematopoietic stem/progenitor cells; inflammatory response; innate immunity; leukemogenesis
  4. Blood Adv. 2021 Feb 09. 5(3): 711-724
    Moses BS, McCullough S, Fox JM, Mott BT, Bentzen SM, Kim M, Tyner JW, Lapidus RG, Emadi A, Rudek MA, Kingsbury TJ, Civin CI.
      Artemisinins are active against human leukemia cell lines and have low clinical toxicity in worldwide use as antimalarials. Because multiagent combination regimens are necessary to cure fully evolved leukemias, we sought to leverage our previous finding that artemisinin analogs synergize with kinase inhibitors, including sorafenib (SOR), by identifying additional synergistic antileukemic drugs with low toxicity. Screening of a targeted antineoplastic drug library revealed that B-cell lymphoma 2 (BCL2) inhibitors synergize with artemisinins, and validation assays confirmed that the selective BCL2 inhibitor, venetoclax (VEN), synergized with artemisinin analogs to inhibit growth and induce apoptotic cell death of multiple acute leukemia cell lines in vitro. An oral 3-drug "SAV" regimen (SOR plus the potent artemisinin-derived trioxane diphenylphosphate 838 dimeric analog [ART838] plus VEN) killed leukemia cell lines and primary cells in vitro. Leukemia cells cultured in ART838 had decreased induced myeloid leukemia cell differentiation protein (MCL1) levels and increased levels of DNA damage-inducible transcript 3 (DDIT3; GADD153) messenger RNA and its encoded CCATT/enhancer-binding protein homologous protein (CHOP), a key component of the integrated stress response. Thus, synergy of the SAV combination may involve combined targeting of MCL1 and BCL2 via discrete, tolerable mechanisms, and cellular levels of MCL1 and DDIT3/CHOP may serve as biomarkers for action of artemisinins and SAV. Finally, SAV treatment was tolerable and resulted in deep responses with extended survival in 2 acute myeloid leukemia (AML) cell line xenograft models, both harboring a mixed lineage leukemia gene rearrangement and an FMS-like receptor tyrosine kinase-3 internal tandem duplication, and inhibited growth in 2 AML primagraft models.
  5. Blood Adv. 2021 Jan 12. 5(1): 156-166
    Lo Presti C, Fauvelle F, Jacob MC, Mondet J, Mossuz P.
      Leukemic cells display some alterations in metabolic pathways, which play a role in leukemogenesis and in patients' prognosis. To evaluate the characteristics and the impact of this metabolic reprogramming, we explore the bone marrow samples from 54 de novo acute myeloid leukemia (AML) patients, using an untargeted metabolomics approach based on proton high-resolution magic angle spinning-nuclear magnetic resonance. The spectra obtained were subjected to multivariate statistical analysis to find specific metabolome alterations and biomarkers correlated to clinical features. We found that patients display a large diversity of metabolic profiles, according to the different AML cytologic subtypes and molecular statuses. The link between metabolism and molecular status was particularly strong for the oncometabolite 2-hydroxyglutarate (2-HG), whose intracellular production is directly linked to the presence of isocitrate dehydrogenase mutations. Moreover, patients' prognosis was strongly impacted by several metabolites, such as 2-HG that appeared as a good prognostic biomarker in our cohort. Conversely, deregulations in phospholipid metabolism had a negative impact on prognosis through 2 main metabolites (phosphocholine and phosphoethanolamine), which could be potential aggressiveness biomarkers. Finally, we highlighted an overexpression of glutathione and alanine in chemoresistant patients. Overall, our results demonstrate that different metabolic pathways could be activated in leukemic cells according to their phenotype and maturation levels. This confirms that metabolic reprogramming strongly influences prognosis of patients and underscores a particular role of certain metabolites and associated pathways in AML prognosis, suggesting common mechanisms developed by leukemic cells to maintain their aggressiveness even after well-conducted induction chemotherapy.
  6. Blood Adv. 2021 Feb 09. 5(3): 760-770
    Simoes C, Paiva B, Martínez-Cuadrón D, Bergua JM, Vives S, Algarra L, Tormo M, Martinez P, Serrano J, Herrera P, Ramos F, Salamero O, Lavilla E, Gil C, Lopez JL, Vidriales MB, Labrador J, Falantes JF, Sayas MJ, Ayala R, Martinez-Lopez J, Villar S, Calasanz MJ, Prosper F, San-Miguel JF, Sanz MÁ, Montesinos P.
      The value of measurable residual disease (MRD) in elderly patients with acute myeloid leukemia (AML) is inconsistent between those treated with intensive vs hypomethylating drugs, and unknown after semi-intensive therapy. We investigated the role of MRD in refining complete remission (CR) and treatment duration in the phase 3 FLUGAZA clinical trial, which randomized 283 elderly AML patients to induction and consolidation with fludarabine plus cytarabine (FLUGA) vs 5-azacitidine. After consolidation, patients continued treatment if MRD was ≥0.01% or stopped if MRD was <0.01%, as assessed by multidimensional flow cytometry (MFC). On multivariate analysis including genetic risk and treatment arm, MRD status in patients achieving CR (N = 72) was the only independent prognostic factor for relapse-free survival (RFS) (HR, 3.45; P = .002). Achieving undetectable MRD significantly improved RFS of patients with adverse genetics (HR, 0.32; P = .013). Longer overall survival was observed in patients with undetectable MRD after induction though not after consolidation. Although leukemic cells from most patients displayed phenotypic aberrancies vs their normal counterpart (N = 259 of 265), CD34 progenitors from cases with undetectable MRD by MFC carried extensive genetic abnormalities identified by whole-exome sequencing. Interestingly, the number of genetic alterations significantly increased from diagnosis to MRD stages in patients treated with FLUGA vs 5-azacitidine (2.2-fold vs 1.1-fold; P = .001). This study supports MRD assessment to refine CR after semi-intensive therapy or hypomethylating agents, but unveils that improved sensitivity is warranted to individualize treatment and prolong survival of elderly AML patients achieving undetectable MRD.
  7. Leukemia. 2021 Feb 08.
    Patkar N, Kakirde C, Shaikh AF, Salve R, Bhanshe P, Chatterjee G, Rajpal S, Joshi S, Chaudhary S, Kodgule R, Ghoghale S, Deshpande N, Shetty D, Khizer SH, Jain H, Bagal B, Menon H, Khattry N, Sengar M, Tembhare P, Subramanian P, Gujral S.
      We accrued 201 patients of adult AML treated with conventional therapy, in morphological remission, and evaluated MRD using sensitive error-corrected next generation sequencing (NGS-MRD) and multiparameter flow cytometry (FCM-MRD) at the end of induction (PI) and consolidation (PC). Nearly 71% of patients were PI NGS-MRD+ and 40.9% PC NGS-MRD+ (median VAF 0.76%). NGS-MRD+ patients had a significantly higher cumulative incidence of relapse (p = 0.003), inferior overall survival (p = 0.001) and relapse free survival (p < 0.001) as compared to NGS-MRD- patients. NGS-MRD was predictive of inferior outcome in intermediate cytogenetic risk and demonstrated potential in favorable cytogenetic risk AML. PI NGS-MRD- patients had a significantly improved survival as compared to patients who became NGS-MRD- subsequently indicating that kinetics of NGS-MRD clearance was of paramount importance. NGS-MRD identified over 80% of cases identified by flow cytometry at PI time point whereas FCM identified 49.3% identified by NGS. Only a fraction of cases were NGS-MRD- but FCM-MRD+. NGS-MRD provided additional information of the risk of relapse when compared to FCM-MRD. We demonstrate a widely applicable, scalable NGS-MRD approach that is clinically informative and synergistic to FCM-MRD in AML treated with conventional therapies. Maximum clinical utility may be leveraged by combining FCM and NGS-MRD modalities.
  8. Mol Cancer Ther. 2021 Feb 10. pii: molcanther.MCT-20-0663-A.2020. [Epub ahead of print]
    Scarpa M, Singh P, Bailey CM, Lee JK, Kapoor S, Lapidus RG, Niyongere S, Sangodkar J, Wang Y, Perrotti D, Narla G, Baer MR.
      fms-like tyrosine like kinase 3 internal tandem duplication (FLT3-ITD) is present in acute myeloid leukemia (AML) in 30% of patients and is associated with short disease-free survival. FLT3 inhibitor efficacy is limited and transient but may be enhanced by multi-targeting of FLT3-ITD signaling pathways. FLT3-ITD drives both STAT5-dependent transcription of oncogenic Pim-1 kinase and inactivation of the tumor suppressor protein phosphatase 2A (PP2A), and FLT3-ITD, Pim-1 and PP2A all regulate the c-Myc oncogene. We studied mechanisms of action of co-treatment of FLT3-ITD-expressing cells with FLT3 inhibitors and PP2A-activating drug (PADs), which are in development. PADs, including FTY720 and DT-061, enhanced FLT3 inhibitor growth suppression and apoptosis induction in FLT3-ITD-expressing cell lines and primary AML cells in vitro and MV4-11 growth suppression in vivo. PAD and FLT3 inhibitor co-treatment independently downregulated c-Myc and Pim-1 protein through enhanced proteasomal degradation. c-Myc and Pim-1 downregulation was preceded by AKT inactivation, did not occur in cells expressing myristoylated (constitutively active) AKT1, and could be induced by AKT inhibition. AKT inactivation resulted in activation of GSK-3β, and GSK-3β inhibition blocked downregulation of both c-Myc and Pim-1 by PAD and FLT3 inhibitor co-treatment GSK-3β activation increased c-Myc proteasomal degradation through c-Myc phosphorylation on T58; infection with c-Myc with T58A substitution, preventing phosphorylation, blocked downregulation of c-Myc by PAD and FLT3 inhibitor co-treatment. GSK-3β also βphosphorylated Pim-1L/Pim-1S on S95/S4. Thus, PADs enhance efficacy of FLT3 inhibitors in FLT3-ITD-expressing cells through a novel mechanism involving AKT inhibition-dependent GSK-3β-mediated increased c-Myc and Pim-1 proteasomal degradation.
  9. Cancers (Basel). 2021 Feb 05. pii: 642. [Epub ahead of print]13(4):
    Grigsby SM, Friedman A, Chase J, Waas B, Ropa J, Serio J, Shen C, Muntean AG, Maillard I, Nikolovska-Coleska Z.
      MLL1 (KMT2a) gene rearrangements underlie the pathogenesis of aggressive MLL-driven acute leukemia. AF9, one of the most common MLL-fusion partners, recruits the histone H3K79 methyltransferase DOT1L to MLL target genes, constitutively activating transcription of pro-leukemic targets. DOT1L has emerged as a therapeutic target in patients with MLL-driven leukemia. However, global DOT1L enzymatic inhibition may lead to off-target toxicities in non-leukemic cells that could decrease the therapeutic index of DOT1L inhibitors. To bypass this problem, we developed a novel approach targeting specific protein-protein interactions (PPIs) that mediate DOT1L recruitment to MLL target genes, and compared the effects of enzymatic and PPIs inhibition on leukemic and non-leukemic hematopoiesis. MLL-AF9 cell lines were engineered to carry mutant DOT1L constructs with a defective AF9 interaction site or lacking enzymatic activity. In cell lines expressing a DOT1L mutant with defective AF9 binding, we observed complete disruption of DOT1L recruitment to critical target genes and inhibition of leukemic cell growth. To evaluate the overall impact of DOT1L loss in non-leukemic hematopoiesis, we first assessed the impact of acute Dot1l inactivation in adult mouse bone marrow. We observed a rapid reduction in myeloid progenitor cell numbers within 7 days, followed by a loss of long-term hematopoietic stem cells. Furthermore, WT and PPI-deficient DOT1L mutants but not an enzymatically inactive DOT1L mutant were able to rescue sustained hematopoiesis. These data show that the AF9-DOT1L interaction is dispensable in non-leukemic hematopoiesis. Our findings support targeting of the MLL-AF9-DOT1L interaction as a promising therapeutic strategy that is selectively toxic to MLL-driven leukemic cells.
    Keywords:  Dot1l; MLL-rearrangement leukemia; hematopoiesis; protein-protein interaction
  10. Blood Adv. 2021 Jan 12. 5(1): 39-53
    Doma E, Mayer IM, Brandstoetter T, Maurer B, Grausenburger R, Menzl I, Zojer M, Hoelbl-Kovacic A, Carlsson L, Heller G, Kollmann K, Sexl V.
      Studies of molecular mechanisms of hematopoiesis and leukemogenesis are hampered by the unavailability of progenitor cell lines that accurately mimic the situation in vivo. We now report a robust method to generate and maintain LSK (Lin-, Sca-1+, c-Kit+) cells, which closely resemble MPP1 cells. HPCLSKs reconstitute hematopoiesis in lethally irradiated recipient mice over >8 months. Upon transformation with different oncogenes including BCR/ABL, FLT3-ITD, or MLL-AF9, their leukemic counterparts maintain stem cell properties in vitro and recapitulate leukemia formation in vivo. The method to generate HPCLSKs can be applied to transgenic mice, and we illustrate it for CDK6-deficient animals. Upon BCR/ABLp210 transformation, HPCLSKsCdk6-/- induce disease with a significantly enhanced latency and reduced incidence, showing the importance of CDK6 in leukemia formation. Studies of the CDK6 transcriptome in murine HPCLSK and human BCR/ABL+ cells have verified that certain pathways depend on CDK6 and have uncovered a novel CDK6-dependent signature, suggesting a role for CDK6 in leukemic progenitor cell homing. Loss of CDK6 may thus lead to a defect in homing. The HPCLSK system represents a unique tool for combined in vitro and in vivo studies and enables the production of large quantities of genetically modifiable hematopoietic or leukemic stem/progenitor cells.
  11. Blood Adv. 2021 Jan 12. 5(1): 176-184
    Chiche E, Rahmé R, Bertoli S, Dumas PY, Micol JB, Hicheri Y, Pasquier F, Peterlin P, Chevallier P, Thomas X, Loschi M, Genthon A, Legrand O, Mohty M, Raffoux E, Auberger P, Caulier A, Joris M, Bonmati C, Roth-Guepin G, Lejeune C, Pigneux A, Vey N, Recher C, Ades L, Cluzeau T.
      CPX-351 is a liposomal formulation of cytarabine and daunorubicin approved for the treatment of adults with newly diagnosed, therapy-related acute myeloid leukemia (t-AML) or AML with myelodysplasia-related changes (MRC-AML). We retrospectively analyzed the efficacy and safety of CPX-351 in a real-world setting in 103 patients from 12 French centers, including the evaluation of molecular abnormalities at baseline and minimal residual disease (MRD) in responding patients, compared with a historical data set from Bordeaux-Toulouse DATAML registry. A favorable safety profile was observed, with a low frequency of alopecia (11%) and gastrointestinal toxicity (50%). The overall response rate after induction was 59%, and MRD <10-3 was achieved in 57% of complete response (CR)/CR with incomplete hematological recovery (CRi) patients. Only the presence of mutated TP53 (P = .02) or PTPN11 (P = .004) predicted lower response in multivariate analysis. Interestingly, high-risk molecular prognosis subgroups defined by 2017 European LeukemiaNet risk stratification, including ASXL1 and RUNX1 mutations, were not associated with a significantly lower response rate using CPX-351. With a median follow-up of 8.6 months, median overall survival (OS) was 16.1 months. Thirty-six patients underwent allogeneic stem cell transplantation with a significantly longer median OS compared with nontransplanted patients (P < .001). In multivariate analyses, only spliceosome mutations were associated with better OS (P = .04). In comparison with intensive chemotherapy, there was no difference in OS for patients <60 years. These data confirm the efficacy and safety of CPX-351 in high-risk AML (t-AML and MRC-AML) in a real-life setting. CPX-351 is a treatment of choice for patients aged ≥60 years.
  12. Cancers (Basel). 2021 Feb 05. pii: 629. [Epub ahead of print]13(4):
    Vial JP, Lechevalier N, Lacombe F, Dumas PY, Bidet A, Leguay T, Vergez F, Pigneux A, Béné MC.
      The assessment of minimal residual disease (MRD) is increasingly considered to monitor response to therapy in hematological malignancies. In acute myeloblastic leukemia (AML), molecular MRD (mMRD) is possible for about half the patients while multiparameter flow cytometry (MFC) is more broadly available. However, MFC analysis strategies are highly operator-dependent. Recently, new tools have been designed for unsupervised MFC analysis, segregating cell-clusters with the same immunophenotypic characteristics. Here, the Flow-Self-Organizing-Maps (FlowSOM) tool was applied to assess MFC-MRD in 96 bone marrow (BM) follow-up (FU) time-points from 40 AML patients with available mMRD. A reference FlowSOM display was built from 19 healthy/normal BM samples (NBM), then simultaneously compared to the patient's diagnosis and FU samples at each time-point. MRD clusters were characterized individually in terms of cell numbers and immunophenotype. This strategy disclosed subclones with varying immunophenotype within single diagnosis and FU samples including populations absent from NBM. Detectable MRD was as low as 0.09% in MFC and 0.051% for mMRD. The concordance between mMRD and MFC-MRD was 80.2%. MFC yielded 85% specificity and 69% sensitivity compared to mMRD. Unsupervised MFC is shown here to allow for an easy and robust assessment of MRD, applicable also to AML patients without molecular markers.
    Keywords:  FlowSOM; acute myeloid leukemia; minimal/measurable residual disease; molecular markers; multiparameter flow cytometry; unsupervised analysis
  13. Blood Adv. 2021 Feb 09. 5(3): 889-899
    Lawson H, Sepulveda C, van de Lagemaat LN, Durko J, Barile M, Tavosanis A, Georges E, Shmakova A, Timms P, Carter RN, Allen L, Campos J, Vukovic M, Guitart AV, Giles P, O'Shea M, Vernimmen D, Morton NM, Rodrigues NP, Göttgens B, Schofield CJ, Lengeling A, O'Carroll D, Kranc KR.
      Lifelong multilineage hematopoiesis critically depends on rare hematopoietic stem cells (HSCs) that reside in the hypoxic bone marrow microenvironment. Although the role of the canonical oxygen sensor hypoxia-inducible factor prolyl hydroxylase has been investigated extensively in hematopoiesis, the functional significance of other members of the 2-oxoglutarate (2-OG)-dependent protein hydroxylase family of enzymes remains poorly defined in HSC biology and multilineage hematopoiesis. Here, by using hematopoietic-specific conditional gene deletion, we reveal that the 2-OG-dependent protein hydroxylase JMJD6 is essential for short- and long-term maintenance of the HSC pool and multilineage hematopoiesis. Additionally, upon hematopoietic injury, Jmjd6-deficient HSCs display a striking failure to expand and regenerate the hematopoietic system. Moreover, HSCs lacking Jmjd6 lose multilineage reconstitution potential and self-renewal capacity upon serial transplantation. At the molecular level, we found that JMJD6 functions to repress multiple processes whose downregulation is essential for HSC integrity, including mitochondrial oxidative phosphorylation (OXPHOS), protein synthesis, p53 stabilization, cell cycle checkpoint progression, and mTORC1 signaling. Indeed, Jmjd6-deficient primitive hematopoietic cells display elevated basal and maximal mitochondrial respiration rates and increased reactive oxygen species (ROS), prerequisites for HSC failure. Notably, an antioxidant, N-acetyl-l-cysteine, rescued HSC and lymphoid progenitor cell depletion, indicating a causal impact of OXPHOS-mediated ROS generation upon Jmjd6 deletion. Thus, JMJD6 promotes HSC maintenance and multilineage differentiation potential by suppressing fundamental pathways whose activation is detrimental for HSC function.
  14. Blood Adv. 2021 Jan 12. 5(1): 113-121
    Hamulyák EN, Daams JG, Leebeek FWG, Biemond BJ, Te Boekhorst PAW, Middeldorp S, Lauw MN.
      Patients with myeloproliferative neoplasms (MPNs), polycythemia vera, essential thrombocythemia, and primary myelofibrosis, have an increased risk of thrombosis. Risk of recurrent thrombosis can be reduced with antithrombotic therapy and/or cytoreduction, but the optimal long-term management in patients with MPN with a history of venous thromboembolism (VTE) is unknown, and clinical practice is heterogeneous. We performed a systematic review and meta-analysis of randomized trials and observational studies evaluating anticoagulant and/or antiplatelet therapy, with or without cytoreduction, in MPN patients with a history of VTE. A total of 5675 unique citations were screened for eligibility. No randomized trials were identified. Ten observational studies involving 1295 patients with MPN were included in the analysis. Overall, 23% had an arterial or recurrent venous thrombotic event on follow-up. The recurrence risk was lowest for patients on oral anticoagulation plus cytoreduction (16%); 55 of 313 (18%) with vitamin K antagonists (VKA) and 5 of 63 (8%) with direct oral anticoagulants (DOACs). In 746 analyzed patients, the risk of recurrent VTE ranged up to 33% (median 13%) and was low in 63 DOAC plus cytoreduction-treated patients (3.2%). All types of antithrombotic treatments were associated with a lower risk of recurrent VTE when combined with cytoreduction. Most studies had a high risk of bias, whereas clinical and statistical heterogeneity led to inconsistent and imprecise findings. In summary, evidence on the optimal antithrombotic treatment of VTE in patients with MPN is based on observational studies only with low certainty for all strategies. Our data suggest that a combination of anticoagulation and cytoreduction may provide the lowest recurrence risk.
  15. Haematologica. 2021 Feb 11.
    El Hajj H, Hleihel R, El Sabban M, Bruneau J, Zaatari G, Cheminant M, Marçais A, Akkouche A, Hasegawa H, Hall W, De Thé H, Hermine O, Bazarbachi A.
      Adult T cell leukemia/lymphoma (ATL) is associated to chronic human T cell leukemia virus type 1 (HTLV-1) infection and carries a poor prognosis. Arsenic trioxide (AS) and interferon-alpha (IFNα) together selectively trigger Tax viral oncoprotein degradation and cure Tax-driven murine ATL. AS/IFNα/zidovudine treatment achieves a high response rate in patients with chronic ATL. Interleukin 10 (IL-10) is an immuno-suppressive cytokine whose expression is activated by Tax. Here we show that, in ATL, AS/IFNα-induced abrogation of leukemia initiating cell activity requires IL-10 expression shutoff. Loss of IL-10 secretion drives production of inflammatory cytokines by the microenvironment, followed by innate immunity-mediated clearance of Taxdriven leukemic cells. Accordingly, anti-IL-10 monoclonal antibodies significantly increased the efficiency of AS/IFNα therapy. These results emphasize the sequential targeting of malignant ATL cells and their immune microenvironment in leukemia initiating cell (LIC) eradication and provide a strong rational to test AS/IFNα/anti-IL10 combination in ATL.
  16. Cell Metab. 2021 Feb 04. pii: S1550-4131(21)00014-0. [Epub ahead of print]
    Kosaisawe N, Sparta B, Pargett M, Teragawa CK, Albeck JG.
      Cell-to-cell heterogeneity in metabolism plays an unknown role in physiology and pharmacology. To functionally characterize cellular variability in metabolism, we treated cells with inhibitors of oxidative phosphorylation (OXPHOS) and monitored their responses with live-cell reporters for ATP, ADP/ATP, or activity of the energy-sensing kinase AMPK. Across multiple OXPHOS inhibitors and cell types, we identified a subpopulation of cells resistant to activation of AMPK and reduction of ADP/ATP ratio. This resistant state persists transiently for at least several hours and can be inherited during cell divisions. OXPHOS inhibition suppresses the mTORC1 and ERK growth signaling pathways in sensitive cells, but not in resistant cells. Resistance is linked to a multi-factorial combination of increased glucose uptake, reduced protein biosynthesis, and G0/G1 cell-cycle status. Our results reveal dynamic fluctuations in cellular energetic balance and provide a basis for measuring and predicting the distribution of cellular responses to OXPHOS inhibition.
    Keywords:  AKT; FRET; PI3K; adenosine mono-phosphate-regulated protein kinase; electron transport chain; insulin signaling; mammalian target of rapamycin; metabolic cycle; oligomycin; oscillation; translation regulation
  17. Blood Adv. 2021 Feb 09. 5(3): 775-786
    Olofsen PA, Bosch DA, Roovers O, van Strien PMH, de Looper HWJ, Hoogenboezem RM, Barnhoorn S, Mastroberardino PG, Ghazvini M, van der Velden VHJ, Bindels EMJ, de Pater EM, Touw IP.
      Mutations in ELANE cause severe congenital neutropenia (SCN), but how they affect neutrophil production and contribute to leukemia predisposition is unknown. Neutropenia is alleviated by CSF3 (granulocyte colony-stimulating factor) therapy in most cases, but dose requirements vary between patients. Here, we show that CD34+CD45+ hematopoietic progenitor cells (HPCs) derived from induced pluripotent stem cell lines from patients with SCN that have mutations in ELANE (n = 2) or HAX1 (n = 1) display elevated levels of reactive oxygen species (ROS) relative to normal iPSC-derived HPCs. In patients with ELANE mutations causing misfolding of the neutrophil elastase (NE) protein, HPCs contained elevated numbers of promyelocyte leukemia protein nuclear bodies, a hallmark of acute oxidative stress. This was confirmed in primary bone marrow cells from 3 additional patients with ELANE-mutant SCN. Apart from responding to elevated ROS levels, PML controlled the metabolic state of these ELANE-mutant HPCs as well as the expression of ELANE, suggestive of a feed-forward mechanism of disease development. Both PML deletion and correction of the ELANE mutation restored CSF3 responses of these ELANE-mutant HPCs. These findings suggest that PML plays a crucial role in the disease course of ELANE-SCN characterized by NE misfolding, with potential implications for CSF3 therapy.
  18. Blood Adv. 2021 Feb 09. 5(3): 900-912
    Stratmann S, Yones SA, Mayrhofer M, Norgren N, Skaftason A, Sun J, Smolinska K, Komorowski J, Herlin MK, Sundström C, Eriksson A, Höglund M, Palle J, Abrahamsson J, Jahnukainen K, Munthe-Kaas MC, Zeller B, Tamm KP, Cavelier L, Holmfeldt L.
      Relapse is the leading cause of death of adult and pediatric patients with acute myeloid leukemia (AML). Numerous studies have helped to elucidate the complex mutational landscape at diagnosis of AML, leading to improved risk stratification and new therapeutic options. However, multi-whole-genome studies of adult and pediatric AML at relapse are necessary for further advances. To this end, we performed whole-genome and whole-exome sequencing analyses of longitudinal diagnosis, relapse, and/or primary resistant specimens from 48 adult and 25 pediatric patients with AML. We identified mutations recurrently gained at relapse in ARID1A and CSF1R, both of which represent potentially actionable therapeutic alternatives. Further, we report specific differences in the mutational spectrum between adult vs pediatric relapsed AML, with MGA and H3F3A p.Lys28Met mutations recurrently found at relapse in adults, whereas internal tandem duplications in UBTF were identified solely in children. Finally, our study revealed recurrent mutations in IKZF1, KANSL1, and NIPBL at relapse. All of the mentioned genes have either never been reported at diagnosis in de novo AML or have been reported at low frequency, suggesting important roles for these alterations predominantly in disease progression and/or resistance to therapy. Our findings shed further light on the complexity of relapsed AML and identified previously unappreciated alterations that may lead to improved outcomes through personalized medicine.
  19. Cancer Discov. 2021 Feb 09. pii: candisc.0564.2020. [Epub ahead of print]
    Pikman Y, Tasian SK, Sulis ML, Stevenson K, Blonquist TM, Apsel Winger B, Cooper TM, Pauly M, Maloney KW, Burke MJ, Brown PA, Gossai N, McNeer JL, Shukla NN, Cole PD, Kahn JM, Chen J, Barth MJ, Magee JA, Gennarini L, Adhav AA, Clinton CM, Ocasio-Martinez N, Gotti G, Li Y, Lin S, Imamovic A, Tognon CE, Patel T, Faust HL, Contreras CF, Cremer A, Cortopassi WA, Garrido Ruiz D, Jacobson MP, Dharia NV, Su A, Robichaud AL, Saur Conway A, Tarlock K, Stieglitz E, Place AE, Puissant A, Hunger SP, Kim AS, Lindeman NI, Gore L, Janeway KA, Silverman LB, Tyner JW, Harris MH, Loh ML, Stegmaier K.
      Despite a remarkable increase in the genomic profiling of cancer, integration of genomic discoveries into clinical care has lagged behind. We report the feasibility of rapid identification of targetable mutations in 153 pediatric patients with relapsed/refractory or high-risk leukemias enrolled on a prospective clinical trial conducted by the LEAP Consortium. Eighteen percent of patients had a high confidence, Tier 1 or 2, recommendation. We describe clinical responses in the 14% of patients with relapsed/refractory leukemia who received the matched targeted therapy. Further, in order to inform future targeted therapy for patients, we validated variants of uncertain significance (VUS), performed ex vivo drug sensitivity testing in patient leukemia samples, and identified new combinations of targeted therapies in cell lines and patient-derived xenograft models. These data and our collaborative approach should inform the design of future precision medicine trials.
  20. Haematologica. 2021 Feb 11.
    Van Leeuwen-Kerkhoff N, Westers TM, Poddighe PJ, Povoleri GAM, Timms JA, Kordasti S, De Gruijl TD, Van de Loosdrecht AA.
      In myelodysplastic syndromes (MDS) the immune system is involved in pathogenesis as well as in disease progression. Dendritic cells (DC) are key players of the immune system by serving as regulators of immune responses. Their function has been scarcely studied in MDS and most of the reported studies didn't investigate naturally occurring DC subsets. Therefore, we here examined the frequency and function of DC subsets and slan+ non-classical monocytes in various MDS risk groups. Frequencies of DC as well as of slan+ monocytes were decreased in MDS bone marrow (BM) compared to normal bone marrow (NBM) samples. Transcriptional profiling revealed down-regulation of transcripts related to pro-inflammatory pathways in MDS-derived cells as compared to NBM. Additionally, their capacity to induce T cell proliferation was impaired. Multidimensional mass cytometry showed that whereas healthy donor-derived slan+ monocytes supported Th1/Th17/Treg differentiation/expansion their MDS-derived counterparts also mediated substantial Th2 expansion. Our findings point to a role for an impaired ability of DC subsets to adequately respond to cellular stress and DNA damage in the immune escape and progression of MDS. As such, it paves the way toward potential novel immunotherapeutic interventions.