bims-tremyl Biomed News
on Therapy resistance biology in myeloid leukemia
Issue of 2021‒10‒31
thirty papers selected by
Paolo Gallipoli
Barts Cancer Institute, Queen Mary University of London

  1. Blood. 2021 Oct 26. pii: blood.2021012519. [Epub ahead of print]
      Mutations in chromatin regulator ASXL1 are frequently identified in myeloid malignancies, in particular ~40% in chronic myelomonocytic leukemia (CMML). ASXL1 mutations associate with poor prognosis in CMML and significantly co-occur with NRAS mutations. Here, we show that concurrent ASXL1 and NRAS mutations defined a population of CMML patients with shorter leukemia-free survival than those with ASXL1 mutation only. Corroborating this human data, Asxl1-/- accelerated CMML progression and promoted CMML transformation to acute myeloid leukemia (AML) in NrasG12D/+ mice. NrasG12D/+; Asxl1-/- (NA) leukemia cells displayed hyperactivation of MEK/ERK signaling, increased global level of H3K27ac, and Flt3 upregulation. Moreover, we find that NA-AML cells overexpressed all the major inhibitory immune checkpoint ligands, PD-L1/L2, CD155, and CD80/86. Among them, overexpression of PD-L1 and CD86 correlated with upregulation of AP-1 transcription factors (TFs) in NA-AML cells. An AP-1 inhibitor or shRNAs against AP-1 TF Jun decreased PD-L1 and CD86 expression in NA-AML cells. Once NA-AML cells were transplanted into syngeneic recipients, NA-derived T cells were not detectable. Host-derived wildtype T cells overexpressed PD-1 and TIGIT receptors, leading to a predominant exhausted T cell phenotype. Combined inhibition of MEK and BET led to downregulation of Flt3 and AP-1 expression, partial restoration of the immune microenvironment, enhancement of CD8+ T cell cytotoxicity, and prolonged survival in NA-AML mice. Our study suggests that combined targeted therapy and immunotherapy may be beneficial for treating secondary AML with concurrent ASXL1 and NRAS mutations.
  2. Blood. 2021 Oct 25. pii: blood.2021012778. [Epub ahead of print]
      In an effort to identify novel drugs targeting fusion-oncogene induced acute myeloid leukemia (AML), we performed high-resolution proteomic analysis. In AML1-ETO (AE) driven AML we uncovered a de-regulation of phospholipase C (PLC) signaling. We identified PLCgamma 1 (PLCG1) as a specific target of the AE fusion protein which is induced after AE binding to intergenic regulatory DNA elements. Genetic inactivation of PLCG1 in murine and human AML inhibited AML1-ETO dependent self-renewal programs, leukemic proliferation, and leukemia maintenance in vivo. In contrast, PLCG1 was dispensable for normal hematopoietic stem- and progenitor cell function. These findings are extended to and confirmed by pharmacologic perturbation of Ca++-signaling in AML1-ETO AML cells, indicating that the PLCG1 pathway poses an important therapeutic target for AML1-ETO positive leukemic stem cells.
  3. Transl Oncol. 2021 Oct 25. pii: S1936-5233(21)00236-9. [Epub ahead of print]15(1): 101244
      INTRODUCTION: FLT3-ITD mutations occur in approximately 25% of patients with acute myeloid leukemia (AML) and are associated with poor prognosis. Despite initial efficacy, short duration of response and high relapse rates limit clinical use of selective FLT3 inhibitors. Combination approaches with other targeted therapies may achieve better clinical outcomes.MATERIALS AND METHODS: Anti-leukemic activity of multikinase inhibitor olverembatinib (HQP1351), alone or in combination with BCL-2 inhibitor lisaftoclax (APG-2575), was evaluated in FLT3-ITD mutant AML cell lines in vitro and in vivo. A patient-derived FLT3-ITD mutant AML xenograft model was also used to assess the anti-leukemic activity of this combination.
    RESULTS: HQP1351 potently induced apoptosis and inhibited FLT3 signaling in FLT3-ITD mutant AML cell lines MV-4-11 and MOLM-13. HQP1351 monotherapy also significantly suppressed growth of FLT3-ITD mutant AML xenograft tumors and prolonged survival of tumor-bearing mice. HQP1351 and APG-2575 synergistically induced apoptosis in FLT3-ITD mutant AML cells and suppressed growth of MV-4-11 xenograft tumors. Combination therapy improved survival of tumor bearing-mice in a systemic MOLM-13 model and showed synergistic anti-leukemic effects in a patient-derived FLT3-ITD mutant AML xenograft model. Mechanistically, HQP1351 downregulated expression of myeloid-cell leukemia 1 (MCL-1) by suppressing FLT3-STAT5 (signal transducer and activator of transcription 5) signaling and thus enhanced APG-2575-induced apoptosis in FLT3-ITD mutant AML cells.
    CONCLUSIONS: FLT3 inhibition by HQP1351 downregulates MCL-1 and synergizes with BCL-2 inhibitor APG-2575 to potentiate cellular apoptosis in FLT3-ITD mutant AML. Our findings provide a scientific rationale for further clinical investigation of HQP1351 combined with APG-2575 in patients with FLT3-ITD mutant AML.
    Keywords:  APG-2575; Acute myeloid leukemia; BCL-2; FLT3-ITD; HQP1351
  4. Leuk Lymphoma. 2021 Oct 27. 1-8
      Cancer cells reprogram their metabolism to maintain sustained proliferation, which creates unique metabolic dependencies between malignant and healthy cells that can be exploited for therapy. In acute myeloid leukemia (AML), mitochondrial inhibitors that block tricarboxylic acid cycle enzymes or electron transport chain complexes have recently shown clinical promise. The isocitrate dehydrogenase 1 inhibitor ivosidenib, the isocitrate dehydrogenase 2 inhibitor enasidenib, and the BH3 mimetic venetoclax received FDA approval for treatment of AML in the last few years. Other mitochondrial inhibitors including CPI-613, CB-839, dihydroorotate dehydrogenase inhibitors, IACS-010759, and mubritinib, have shown encouraging preclinical efficacy and are currently being evaluated in clinical trials. In this review, we summarize recent metabolism-based therapies and their ability to target altered cancer metabolism in AML.
    Keywords:  Targeted therapy; acute myeloid leukemia; mitochondrial metabolism
  5. BMC Cancer. 2021 Oct 28. 21(1): 1153
      BACKGROUND: Disease relapse remains common following treatment of acute myeloid leukemia (AML) and is due to chemoresistance of leukemia cells with disease repopulating potential. To date, attempts to define the characteristics of in vivo resistant blasts have focused on comparisons between leukemic cells at presentation and relapse. However, further treatment responses are often seen following relapse, suggesting that most blasts remain chemosensitive. We sought to characterise in vivo chemoresistant blasts by studying the transcriptional and genetic features of blasts from before and shortly after induction chemotherapy using paired samples from six patients with primary refractory AML.METHODS: Leukemic blasts were isolated by fluorescence-activated cell sorting. Fluorescence in situ hybridization (FISH), targeted genetic sequencing and detailed immunophenotypic analysis were used to confirm that sorted cells were leukemic. Sorted blasts were subjected to RNA sequencing. Lentiviral vectors expressing short hairpin RNAs were used to assess the effect of FOXM1 knockdown on colony forming capacity, proliferative capacity and apoptosis in cell lines, primary AML cells and CD34+ cells from healthy donors.
    RESULTS: Molecular genetic analysis revealed early clonal selection occurring after induction chemotherapy. Immunophenotypic characterisation found leukemia-associated immunophenotypes in all cases that persisted following treatment. Despite the genetic heterogeneity of the leukemias studied, transcriptional analysis found concerted changes in gene expression in resistant blasts. Remarkably, the gene expression signature suggested that post-chemotherapy blasts were more proliferative than those at presentation. Resistant blasts also appeared less differentiated and expressed leukemia stem cell (LSC) maintenance genes. However, the proportion of immunophenotypically defined LSCs appeared to decrease following treatment, with implications for the targeting of these cells on the basis of cell surface antigen expression. The refractory gene signature was highly enriched with targets of the transcription factor FOXM1. shRNA knockdown experiments demonstrated that the viability of primary AML cells, but not normal CD34+ cells, depended on FOXM1 expression.
    CONCLUSIONS: We found that chemorefractory blasts from leukemias with varied genetic backgrounds expressed a common transcriptional program. In contrast to the notion that LSC quiescence confers resistance to chemotherapy we find that refractory blasts are both actively proliferating and enriched with LSC maintenance genes. Using primary patient material from a relevant clinical context we also provide further support for the role of FOXM1 in chemotherapy resistance, proliferation and stem cell function in AML.
    Keywords:  Acute myeloid leukemia; Drug resistance; FOXM1; Leukemia stem cell; Quiescence; Transcriptome
  6. Nat Commun. 2021 Oct 29. 12(1): 6233
      Acute myeloid leukemia (AML) is a hematological malignancy with an undefined heritable risk. Here we perform a meta-analysis of three genome-wide association studies, with replication in a fourth study, incorporating a total of 4018 AML cases and 10488 controls. We identify a genome-wide significant risk locus for AML at 11q13.2 (rs4930561; P = 2.15 × 10-8; KMT5B). We also identify a genome-wide significant risk locus for the cytogenetically normal AML sub-group (N = 1287) at 6p21.32 (rs3916765; P = 1.51 × 10-10; HLA). Our results inform on AML etiology and identify putative functional genes operating in histone methylation (KMT5B) and immune function (HLA).
  7. Front Oncol. 2021 ;11 754093
      ASH1L and MLL1 are two histone methyltransferases that facilitate transcriptional activation during normal development. However, the roles of ASH1L and its enzymatic activity in the development of MLL-rearranged leukemias are not fully elucidated in Ash1L gene knockout animal models. In this study, we used an Ash1L conditional knockout mouse model to show that loss of ASH1L in hematopoietic progenitor cells impaired the initiation of MLL-AF9-induced leukemic transformation in vitro. Furthermore, genetic deletion of ASH1L in the MLL-AF9-transformed cells impaired the maintenance of leukemic cells in vitro and largely blocked the leukemia progression in vivo. Importantly, the loss of ASH1L function in the Ash1L-deleted cells could be rescued by wild-type but not the catalytic-dead mutant ASH1L, suggesting the enzymatic activity of ASH1L was required for its function in promoting MLL-AF9-induced leukemic transformation. At the molecular level, ASH1L enhanced the MLL-AF9 target gene expression by directly binding to the gene promoters and modifying the local histone H3K36me2 levels. Thus, our study revealed the critical functions of ASH1L in promoting the MLL-AF9-induced leukemogenesis, which provides a molecular basis for targeting ASH1L and its enzymatic activity to treat MLL-AF9-induced leukemias.
    Keywords:  ASH1L; H3K36me2; MLL-AF9 fusion; MLL1; histone modification; leukemogenesis
  8. Blood Adv. 2021 Oct 29. pii: bloodadvances.2021004558. [Epub ahead of print]
      The transcriptional repressor, BCL11A, is involved in hematological malignancies, B-cell development, and fetal-to-adult hemoglobin switching. However, the molecular mechanism by which it promotes the development of myeloid leukemia remains largely unknown. We find that Bcl11a cooperates with the pseudokinase, Trib1, in the development of acute myeloid leukemia (AML). Bcl11a promotes the proliferation and engraftment of Trib1-expressing AML cells both in vitro and in vivo. ChIP-seq analysis showed that upon DNA-binding, Bcl11a is significantly associated with PU.1, an inducer of myeloid differentiation, and that Bcl11a represses several PU.1 target genes, such as Asb2, Clec5a, and Fcgr3. Asb2, as a Bcl11a target gene that modulates cytoskeleton and cell-cell interaction, plays a key role in Bcl11a-induced malignant progression. The repression of PU.1 target genes by Bcl11a is achieved by both sequence-specific DNA-binding activity and recruitment of corepressors by Bcl11a. Suppression of the corepressor components, HDAC and LSD1, reverses the repressive activity. Moreover, treatment of AML cells with the HDAC inhibitor, pracinostat, and LSD1 inhibitor, GSK2879552, resulted in growth inhibition both in vitro and in vivo. High BCL11A expression is associated with worse prognosis in human AML patients. Blocking of BCL11A expression upregulates the expression of PU.1 target genes, and inhibits the growth of HL-60 cells and their engraftment to the bone marrow, suggesting that BCL11A is involved in human myeloid malignancies via the suppression of PU.1 transcriptional activity.
  9. Hematol Transfus Cell Ther. 2021 Jul 30. pii: S2531-1379(21)00097-3. [Epub ahead of print]
      INTRODUCTION: Acute myeloid leukemia (AML) is a heterogeneous disease and approximately one-third of its carriers do not have evident genetic abnormalities. The mutation of specific molecular markers, such as fms-like tyrosine kinase 3 (FTL3) internal tandem duplication (ITD), FLT3 tyrosine kinase domain (TKD) and nucleophosmin (NPM1), are associated with an adverse and favorable prognosis, respectively.OBJECTIVE: The objective was to determine the prevalence of FLT3/ITD and NPM1 in Chilean patients and their association with clinical data and prognosis.
    METHOD AND RESULTS: Two hundred and thirty-two children were studied between 2011 and 2017, the median being 8.6 years (ranging from 1 to 18 months). Acute promyelocytic leukemia (APL) was diagnosed in 29%. The FLT3/ITD-mutated in non-promyelocytic AML was at 10% (14/133) and the FLT3/TKD, at 3.7% (2/54). In APL, it was at 25.4% (16/63). In non-promyelocytic AML, the FLT3/ITD-mutated was associated with a high leucocyte count, the median being 28.5 x mm3 (n = 14) versus 19.4 x mm3 (n = 119), (p = 0.25), in non-mutated cases. In APL, the median was 33.6 x mm3 (n = 15) versus 2.8 x mm3 (n = 47), (p < 0.001). The five-year overall survival (OS) in non-promyelocytic AML with non-mutated and mutated FLT3/ITD were 62.7% and 21.4%, respectively, (p < 0.001); the 5-year event-free survival (EFS) were 79.5% and 50%, respectively, (p < 0.01). The five-year OS in APL with non-mutated and mutated FLT3/ITD was 84.7% and 62.5%, respectively, (p = 0.05); the 5-year EFS was 84.7% and 68.8%, respectively, (p = 0.122). The NPM1 mutation was observed in 3.2% (5/155), all non-promyelocytic AML with the normal karyotype.
    CONCLUSION: The FLT3/ITD mutation was observed more frequently in APL and associated with a higher white cell count at diagnosis. However, the most important finding was that the FLT3/ITD mutation was associated with a shorter survival in non-promyelocytic AML.
    Keywords:  Acute myeloid leukemia; FLT3/ITD mutation; NPM1 mutation
  10. Biol Chem. 2021 Oct 26.
      Acute myeloid leukemia (AML) is a heterogeneous, highly malignant disease of the bone marrow. After decades of slow progress, recent years saw a surge of novel agents for its treatment. The most recent advancement is the registration of the Bcl-2 inhibitor ventoclax in combination with a hypomethylating agent (HMA) in the US and Europe for AML patients not eligible for intensive chemotherapy. Treatment of newly diagnosed AML patients with this combination results in remission rates that so far could only be achieved with intensive treatment. However, not all AML patients respond equally well, and some patients relapse early, while other patients experience longer periods of complete remission. A hallmark of AML is its remarkable genetic, molecular and clinical heterogeneity. Here, we review the current knowledge about molecular features of AML that help estimate the probability of response to venetoclax-containing therapies. In contrast to other newly developed AML therapies that target specific recurrent molecular alterations, it seems so far that responses are not specific for a certain subgroup. One exception is spliceosome mutations, where good response has been observed in clinical trials with venetoclax/azacitidine. These mutations are rather associated with a more unfavorable outcome with chemotherapy. In summary, venetoclax in combination with hypomethylating agents represents a significant novel option for AML patients with various molecular aberrations. Mechanisms of primary and secondary resistance seem to overlap with those towards chemotherapy.
    Keywords:  AML; azacitidine; genotype; hypomethylating agents; resistance; targeted therapy
  11. Sci Rep. 2021 Oct 27. 11(1): 21230
      CD157/BST-1 (a member of the ADP-ribosyl cyclase family) is expressed at variable levels in 97% of patients with acute myeloid leukemia (AML), and is currently under investigation as a target for antibody-based immunotherapy. We used peripheral blood and bone marrow samples from patients with AML to analyse the impact of CD157-directed antibodies in AML survival and in response to cytarabine (AraC) ex vivo. The study was extended to the U937, THP1 and OCI-AML3 AML cell lines of which we engineered CD157-low versions by shRNA knockdown. CD157-targeting antibodies enhanced survival, decreased apoptosis and reduced AraC toxicity in AML blasts and cell lines. CD157 signaling activated the PI3K/AKT/mTOR and MAPK/ERK pathways and increased expression of Mcl-1 and Bcl-XL anti-apoptotic proteins, while decreasing expression of Bax pro-apoptotic protein, thus preventing Caspase-3 activation. The primary CD157-mediated anti-apoptotic mechanism was Bak sequestration by Mcl-1. Indeed, the Mcl-1-specific inhibitor S63845 restored apoptosis by disrupting the interaction of Mcl-1 with Bim and Bak and significantly increased AraC toxicity in CD157-high but not in CD157-low AML cells. This study provides a new role for CD157 in AML cell survival, and indicates a potential role of CD157 as a predictive marker of response to therapies exploiting Mcl-1 pharmacological inhibition.
  12. Clin Cancer Res. 2021 Oct 29. pii: clincanres.2863.2021. [Epub ahead of print]
      PURPOSE: In acute myeloid leukemia (AML), recurrent DNA methyltransferase 3A (DNMT3A) mutations are associated with chemoresistance and poor prognosis, especially in advanced-age patients. Gene expression studies in DNMT3A-mutated cells identified signatures implicated in deregulated DNA damage response and replication fork integrity, suggesting sensitivity to replication stress. Here, we tested whether pharmacologically-induced replication fork stalling such as with cytarabine creates a therapeutic vulnerability in cells with DNMT3A(R882) mutations.EXPERIMENTAL DESIGN: Leukemia cell lines, genetic mouse models, and isogenic cells with and without DNMT3A(mut) were used to evaluate sensitivity to nucleoside analogs such as cytarabine in vitro and in vivo, followed by analysis of DNA damage and signaling, replication restart, and cell cycle progression on treatment and after drug removal. Transcriptome profiling identified pathways deregulated by DNMT3A(mut) expression.
    RESULTS: We found increased sensitivity to pharmacologically-induced replication stress in cells expressing DNMT3A(R882)-mutant, with persistent intra-S phase checkpoint activation, impaired PARP1 recruitment, and elevated DNA damage, which was incompletely resolved after drug removal and carried through mitosis. Pulse-chase double-labeling experiments with EdU and BrdU after cytarabine wash-out demonstrated a higher rate of fork collapse in DNMT3A(mut)-expressing cells. RNA-seq studies supported deregulated cell cycle progression and p53 activation, along with splicing, ribosome biogenesis, and metabolism.
    CONCLUSIONS: Together, our studies show that DNMT3A mutations underlie a defect in recovery from replication fork arrest with subsequent accumulation of unresolved DNA damage, which may have therapeutic tractability. These results demonstrate that, in addition to its role in epigenetic control, DNMT3A contributes to preserving genome integrity during replication stress.
  13. Am J Hematol. 2021 Oct 30.
      The progress with intensive chemotherapy and supportive care measures has improved survival in patients with newly diagnosed acute myeloid leukemia (AML). Given the recent development of effective low intensity therapies, an optimal decision on the therapy intensity may improve survival through the avoidance of early mortality. We reviewed the outcome of 3,728 patients with newly diagnosed AML who received intensive chemotherapy between August 1980 and May 2020. Intensive chemotherapy was defined as a cumulative cytarabine dose ≥700 mg/m2 during induction therapy. We divided the whole cohort into a training and validation group at a 3:1 ratio. The population was divided into a training (2,790 patients) and a validation cohort (938 patients). The median age was 55 years (range, 15-99). Among them, 442 patients (12%) had core-binding factor AML. Binary logistic regression identified older age, worse performance status, hyperbilirubinemia, elevated creatinine, hyperuricemia, cytogenetic abnormalities other than CBF and -Y, and pneumonia as adverse prognostic factors for an early 4-week mortality. This risk classification for early mortality was verified in the validation cohort of patients. In the validation cohort of more recently treated patients from 2000 to 2017, the 4-week mortality rates with intensive chemotherapy were 2%, 14%, and 50% in the low-, high-, and very high-risk group, respectively. The mortality rates with low intensity therapies were 3%, 9%, and 20%, respectively. The risk classification guides treatment intensity by the assessment of age, frailty, organ dysfunction, cytogenetic abnormality, and infection to avoid early mortality. This article is protected by copyright. All rights reserved.
  14. Cancer Discov. 2021 Oct 28. pii: candisc.0551.2021. [Epub ahead of print]
      Early T-cell acute lymphoblastic leukemia (ETP-ALL) is an aggressive hematologic malignancy associated with early relapse and poor prognosis that is genetically, immunophenotypically and transcriptionally distinct from more mature T-cell acute lymphoblastic (T-ALL) tumors. Here, we leveraged global metabolomic and transcriptomic profiling of primary ETP and T-ALL leukemia samples to identify specific metabolic circuitries differentially active in this high-risk leukemia group. ETP-ALLs showed increased biosynthesis of phospholipids and sphingolipids, and were specifically sensitive to inhibition of 3-hydroxy-3-methylglutaryl-CoA Reductase (HMGCR), the rate-limiting enzyme in the mevalonate pathway. Mechanistically, inhibition of cholesterol synthesis inhibited oncogenic AKT1 signaling and suppressed MYC expression via loss of chromatin accessibility at a leukemia stem cell-specific long range MYC enhancer. In all, these results identify the mevalonate pathway as a druggable novel vulnerability in high-risk ETP-ALL cells and uncover an unanticipated critical role for cholesterol biosynthesis in signal transduction and epigenetic circuitries driving leukemia cell growth and survival.
  15. Leuk Res. 2021 Oct 16. pii: S0145-2126(21)01728-8. [Epub ahead of print]111 106727
      Acute myeloid leukemia (AML) is the most common acute leukemia in adults that affects the myeloid lineage. The recent advances have upgraded our understanding of the cytogenetic abnormalities and molecular mutations associated with AML that further aids in prognostication and risk stratification of the disease. Based on the highly heterogeneous nature of the disease and cytogenetic profile, AML patients can be stratified into favourable, intermediate and adverse-risk groups. The recurrent genetic alterations provide novel insights into the pathogenesis, clinical characteristics and also into the overall survival of the patients. In this review we are discussing about the cytogenetics of AML and the recurrent gene alterations such us NPM1, FLT3, CEBPA, TET-2, c-KIT, DNMT3A, IDH, RUNX1, AXSL1, WT1, Ras gene mutations etc. These gene mutations serve as important prognostic markers as well as potential therapeutic targets. AML patients respond to induction chemotherapy initially and subsequently achieve complete remission (CR), eventually most of them get relapsed.
    Keywords:  AML; Prognostic markers; Relapse; Risk groups; Therapeutic targets
  16. Blood Adv. 2021 Oct 28. pii: bloodadvances.2021006139. [Epub ahead of print]
      Secondary myelodysplastic syndromes and acute myeloid leukemia (sMDS/AML) are rare in children/adolescents and have a dismal prognosis. The mainstay therapy is hematopoietic cell transplantation (HCT) but there has been no innovation in cytoreductive regimens. CPX-351, a fixed 5:1 molar ratio of liposomal cytarabine/daunorubicin, has shown favorable safety and efficacy in elderly individuals with sAML and children with relapsed de novo AML. We report the outcomes of seven young patients (six with newly diagnosed sMDS/AML and one with primary MDS/AML) uniformly treated with CPX-351. Five patients had previously received chemotherapy for osteosarcoma, Ewing sarcoma, neuroblastoma, or T-ALL; one had predisposing genomic instability disorder (Cornelia de Lange); and one MDS-related AML and multi-organ failure. The median age at diagnosis of myeloid malignancy was 17 (13-23) years. Patients received 1-3 cycles of CPX-351 (cytarabine 100mg/m2 plus daunorubicin 44mg/m2) on days 1, 3, and 5, resulting in complete morphologic remission without overt toxicity or treatment-related mortality. This approach allowed for adding FLT3 inhibitor as individualized therapy in one patient. Six patients were alive and leukemia-free at 0.5-3.3 years after HCT. One patient died from disease progression before HCT. Concluding, CPX-351 is an effective and well-tolerated regimen for cytoreduction in pediatric sMDS/AML warranting prospective studies.
  17. Lancet Haematol. 2021 Oct 20. pii: S2352-3026(21)00270-2. [Epub ahead of print]
      Research has resulted in regulatory approval of nine agents for acute myeloid leukaemia indications by the US Food and Drug Administration since 2017: the Bcl-2 inhibitor, venetoclax; two FLT3 inhibitors, midostaurin and gilteritinib; two IDH inhibitors, ivosidenib (IDH1 inhibitor) and enasidenib (IDH2 inhibitor); the anti-CD33 antibody-drug conjugate, gemtuzumab ozogamicin; the oral, poorly absorbable hypomethylating agent, azacitidine; the liposomal formulation of cytarabine and daunorubicin (5:1 ratio), CPX-351; and the hedgehog signalling pathway inhibitor, glasdegib. A 100% absorbable oral formulation of the hypomethylating agent decitabine was approved for the treatment of myelodysplastic syndrome and chronic myelomonocytic leukaemia, and might be used as an alternative to parenteral hypomethylating agents. Several of the approvals are as single-agent therapies or in specific combinations for narrow indications, thus offering poor treatment value. In this Review, we discuss ongoing research into combinations containing these commercially available targeted therapies for acute myeloid leukaemia.
  18. Eur J Haematol. 2021 Oct 29.
      OBJECTIVES AND METHODS: Intracranial hemorrhage (ICH) in acute myeloid leukemia (AML) patients is a major concern due to the increased risk of mortality. Few studies have examined ICH specifically in newly diagnosed AML patients receiving intensive induction chemotherapy (IC) and prophylactic platelet transfusions during thrombocytopenia < 10/nL. This retrospective cohort study included 423 newly diagnosed AML patients without acute promyelocytic leukemia who underwent IC between 2007 and 2019. We assessed risk factors, clinical features and outcomes of ICH.RESULTS: 17 of 423 patients (4 %) suffered ICH during hospital stay and 4 patients (24 %) died directly because of ICH despite routine prophylactic platelet transfusions. Patients with ICH had a negatively impacted overall survival (median OS, 20.1 vs. 104.8 months) and were more likely not to continue with curative treatment. Main risk factors were female gender, severe thrombocytopenia and decreased fibrinogen. Patients with subsequent ICH also had laboratory signs of liver dysfunction.
    CONCLUSIONS: ICH remains a potentially deadly complication with notable incidence despite prophylactic platelet substitution, suggesting that additional prophylactic interventions may be required to further reduce the frequency of ICH in high-risk patients. Unrecognized genetic factors may simultaneously predispose to AML and platelet dysfunction with ICH.
    Keywords:  AML; bleeding; induction chemotherapy; intracranial hemorrhage; platelet substitution
  19. Stem Cell Reports. 2021 Oct 19. pii: S2213-6711(21)00498-7. [Epub ahead of print]
      Hematopoietic stem cells (HSCs) reside at the apex of the hematopoietic differentiation hierarchy and sustain multilineage hematopoiesis. Here, we show that the transcriptional regulator CITED2 is essential for life-long HSC maintenance. While hematopoietic-specific Cited2 deletion has a minor impact on steady-state hematopoiesis, Cited2-deficient HSCs are severely depleted in young mice and fail to expand upon aging. Moreover, although they home normally to the bone marrow, they fail to reconstitute hematopoiesis upon transplantation. Mechanistically, CITED2 is required for expression of key HSC regulators, including GATA2, MCL-1, and PTEN. Hematopoietic-specific expression of anti-apoptotic MCL-1 partially rescues the Cited2-deficient HSC pool and restores their reconstitution potential. To interrogate the Cited2→Pten pathway in HSCs, we generated Cited2;Pten compound heterozygous mice, which had a decreased number of HSCs that failed to reconstitute the HSC compartment. In addition, CITED2 represses multiple pathways whose elevated activity causes HSC exhaustion. Thus, CITED2 promotes pathways necessary for HSC maintenance and suppresses those detrimental to HSC integrity.
    Keywords:  CITED2; MCL-1; PTEN; hematopoiesis; hematopoietic stem cell
  20. J Exp Med. 2021 Dec 06. pii: e20211872. [Epub ahead of print]218(12):
      Osteoporosis is caused by an imbalance of osteoclasts and osteoblasts, occurring in close proximity to hematopoietic cells in the bone marrow. Recurrent somatic mutations that lead to an expanded population of mutant blood cells is termed clonal hematopoiesis of indeterminate potential (CHIP). Analyzing exome sequencing data from the UK Biobank, we found CHIP to be associated with increased incident osteoporosis diagnoses and decreased bone mineral density. In murine models, hematopoietic-specific mutations in Dnmt3a, the most commonly mutated gene in CHIP, decreased bone mass via increased osteoclastogenesis. Dnmt3a-/- demethylation opened chromatin and altered activity of inflammatory transcription factors. Bone loss was driven by proinflammatory cytokines, including Irf3-NF-κB-mediated IL-20 expression from Dnmt3a mutant macrophages. Increased osteoclastogenesis due to the Dnmt3a mutations was ameliorated by alendronate or IL-20 neutralization. These results demonstrate a novel source of osteoporosis-inducing inflammation.
  21. Leukemia. 2021 Oct 28.
      Aberrant RHO guanine nucleotide exchange factor (RhoGEF) activation is chief mechanism driving abnormal activation of their GTPase targets in transformation and tumorigenesis. Consequently, a small-molecule inhibitor of RhoGEF can make an anti-cancer drug. We used cellular, mouse, and humanized models of RAC-dependent BCR-ABL1-driven and Ph-like acute lymphoblastic leukemia to identify VAV3, a tyrosine phosphorylation-dependent RacGEF, as the target of the small molecule IODVA1. We show that through binding to VAV3, IODVA1 inhibits RAC activation and signaling and increases pro-apoptotic activity in BCR-ABL1-transformed cells. Consistent with this mechanism of action, cellular and animal models of BCR-ABL1-induced leukemia in Vav3-null background do not respond to IODVA1. By durably decreasing in vivo RAC signaling, IODVA1 eradicates leukemic propagating activity of TKI-resistant BCR-ABL1(T315I) B-ALL cells after treatment withdrawal. Importantly, IODVA1 suppresses the leukemic burden in the treatment refractory pediatric Ph+ and TKI-resistant Ph+ B-ALL patient-derived xenograft models better than standard-of-care dasatinib or ponatinib and provides a more durable response after treatment withdrawal. Pediatric leukemia samples with diverse genetic lesions show high sensitivity to IODVA1 ex vivo and this sensitivity is VAV3 dependent. IODVA1 thus spearheads a novel class of drugs that inhibits a RacGEF and holds promise as an anti-tumor therapy.
  22. Cell Stem Cell. 2021 Oct 20. pii: S1934-5909(21)00414-8. [Epub ahead of print]
      Hematopoietic stem cells (HSCs) rely on complex regulatory networks to preserve stemness. Due to the scarcity of HSCs, technical challenges have limited our insights into the interplay between metabolites, transcription, and the epigenome. In this study, we generated low-input metabolomics, transcriptomics, chromatin accessibility, and chromatin immunoprecipitation data , revealing distinct metabolic hubs that are enriched in HSCs and their downstream multipotent progenitors. Mechanistically, we uncover a non-classical retinoic acid (RA) signaling axis that regulates HSC function. We show that HSCs rely on Cyp26b1, an enzyme conventionally considered to limit RA effects in the cell. In contrast to the traditional view, we demonstrate that Cyp26b1 is indispensable for production of the active metabolite 4-oxo-RA. Further, RA receptor beta (Rarb) is required for complete transmission of 4-oxo-RA-mediated signaling to maintain stem cells. Our findings emphasize that a single metabolite controls stem cell fate by instructing epigenetic and transcriptional attributes.
    Keywords:  4-oxo-RA; Cyp26b1; Rarb; at-RA; epigenetics; hematopoietic stem cells; metabolites; self-renewal; vitamin A
  23. Exp Hematol. 2021 Oct 20. pii: S0301-472X(21)00365-9. [Epub ahead of print]
      Adenosine triphosphate (ATP) accumulates at tissue injury and inflammation sites. The P2 × 7 receptor is an ATP-gated ion channel known for its cytotoxic activity. However, P2 × 7 receptors also play important roles in the growth of cancer and the immune regulation. Functional P2 × 7 receptor is widely expressed in murine and human hematopoietic stem cells and their lineages, including monocytes, macrophages, mast cells, and B or T lymphocytes, and participates in various physiological and pathological activities. Therefore, it is not surprising that the P2 × 7 receptor is important for the normal hematopoiesis and leukemogenesis. Here, we summarize the biological functions of P2 × 7 receptor during both normal hematopoiesis and leukemogenesis. In particular, we showed that ATP levels are dramatically increased in the leukemic bone marrow niche and the fates of leukemia-initiating cells of acute myeloid leukemia are tightly controlled by the P2 × 7 expression and ATP-P2 × 7-mediated signaling pathways. These findings strongly indicate that P2 × 7 receptor may be considered as a potential biomarker of hematological malignancies in bone marrow niches, and its antagonists may be useful for the leukemia treatment in addition to the traditional chemotherapy.
    Keywords:  Adenosine triphosphate; Hematopoiesis; Leukemia-initiating cells; Leukemogenesis; P2 × 7 receptor
  24. Nat Commun. 2021 Oct 26. 12(1): 6177
      Pulmonary hypertension (PH) is a progressive cardiopulmonary disease characterized by pulmonary arterial remodeling. Clonal somatic mutations including JAK2V617F, the most frequent driver mutation among myeloproliferative neoplasms, have recently been identified in healthy individuals without hematological disorders. Here, we reveal that clonal hematopoiesis with JAK2V617F exacerbates PH and pulmonary arterial remodeling in mice. JAK2V617F-expressing neutrophils specifically accumulate in pulmonary arterial regions, accompanied by increases in neutrophil-derived elastase activity and chemokines in chronic hypoxia-exposed JAK2V617F transgenic (JAK2V617F) mice, as well as recipient mice transplanted with JAK2V617F bone marrow cells. JAK2V617F progressively upregulates Acvrl1 (encoding ALK1) during the differentiation from bone marrow stem/progenitor cells peripherally into mature neutrophils of pulmonary arterial regions. JAK2V617F-mediated STAT3 phosphorylation upregulates ALK1-Smad1/5/8 signaling. ALK1/2 inhibition completely prevents the development of PH in JAK2V617F mice. Finally, our prospective clinical study identified JAK2V617F-positive clonal hematopoiesis is more common in PH patients than in healthy subjects. These findings indicate that clonal hematopoiesis with JAK2V617F causally leads to PH development associated with ALK1 upregulation.
  25. Cell Rep. 2021 Oct 26. pii: S2211-1247(21)01370-X. [Epub ahead of print]37(4): 109900
      Infant MLL-AF4-driven acute lymphoblastic leukemia (ALL) is a devastating disease with dismal prognosis. A lack of understanding of the unique biology of this disease, particularly its prenatal origin, has hindered improvement of survival. We perform multiple RNA sequencing experiments on fetal, neonatal, and adult hematopoietic stem and progenitor cells from human and mouse. This allows definition of a conserved fetal transcriptional signature characterized by a prominent proliferative and oncogenic nature that persists in infant ALL blasts. From this signature, we identify a number of genes in functional validation studies that are critical for survival of MLL-AF4+ ALL cells. Of particular interest are PLK1 because of the readily available inhibitor and ELOVL1, which highlights altered fatty acid metabolism as a feature of infant ALL. We identify which aspects of the disease are residues of its fetal origin and potential disease vulnerabilities.
    Keywords:  B-ALL; ELOVL1; MLL-AF4; PLK1; acute lymphoblastic leukemia; infant leukemia; onco-fetal signature
  26. iScience. 2021 Oct 22. 24(10): 103173
      Schlafen11 (SLFN11) is referred to as interferon (IFN)-inducible. Based on cancer genomic databases, we identified human acute myeloid and lymphoblastic leukemia cells with gain-of-function mutations in the Janus kinase (JAK) family as exhibiting high SLFN11 expression. In these cells, the clinical JAK inhibitors cerdulatinib, ruxolitinib, and tofacitinib reduced SLFN11 expression, but IFN did not further induce SLFN11 despite phosphorylated STAT1. We provide evidence that suppression of SLFN11 by JAK inhibitors is caused by inactivation of the non-canonical IFN pathway controlled by AKT and ERK. Accordingly, the AKT and ERK inhibitors MK-2206 and SCH77284 suppressed SLFN11 expression. Both also suppressed the E26 transformation-specific (ETS)-family genes ETS-1 and FLI-1 that act as transcription factors for SLFN11. Moreover, SLFN11 expression was inhibited by the ETS inhibitor TK216. Our study reveals that SLFN11 expression is regulated via the JAK, AKT and ERK, and ETS axis. Pharmacological suppression of SLFN11 warrants future studies.
    Keywords:  Cell biology; Immunology; Molecular biology
  27. Nat Commun. 2021 Oct 25. 12(1): 6170
      The bone marrow (BM) stroma in myeloid neoplasms is altered and it is hypothesized that this cell compartment may also harbor clonal somatically acquired mutations. By exome sequencing of in vitro expanded mesenchymal stromal cells (MSCs) from n = 98 patients with myelodysplastic syndrome (MDS) and n = 28 healthy controls we show that these cells accumulate recurrent mutations in genes such as ZFX (n = 8/98), RANK (n = 5/98), and others. MDS derived MSCs display higher mutational burdens, increased replicative stress, senescence, inflammatory gene expression, and distinct mutational signatures as compared to healthy MSCs. However, validation experiments in serial culture passages, chronological BM aspirations and backtracking of high confidence mutations by re-sequencing primary sorted MDS MSCs indicate that the discovered mutations are secondary to in vitro expansion but not present in primary BM. Thus, we here report that there is no evidence for clonal mutations in the BM stroma of MDS patients.
  28. Leukemia. 2021 Oct 25.
      The p21-Activated Kinases (PAKs) are a family of six serine/threonine kinases that were originally identified as downstream effectors of the Rho GTPases Cdc42 and Rac. Since the first PAK was discovered in 1994, studies have revealed their fundamental and biological importance in the development of physiological systems. Within the cell, PAKs also play significant roles in regulating essential cellular processes such as cytoskeletal dynamics, gene expression, cell survival, and cell cycle progression. These processes are often deregulated in numerous cancers when different PAKs are overexpressed or amplified at the chromosomal level. Furthermore, PAKs modulate multiple oncogenic signaling pathways which facilitate apoptosis escape, uncontrolled proliferation, and drug resistance. There is growing insight into the critical roles of PAKs in regulating steady-state hematopoiesis, including the properties of hematopoietic stem cells (HSC), and the initiation and progression of hematological malignancies. This review will focus on the most recent studies that provide experimental evidence showing how specific PAKs regulate the properties of leukemic stem cells (LSCs) and drug-resistant cells to initiate and maintain hematological malignancies. The current understanding of the molecular and cellular mechanisms by which the PAKs operate in specific human leukemia or lymphomas will be discussed. From a translational point of view, PAKs have been suggested to be critical therapeutic targets and potential prognosis markers; thus, this review will also discuss current therapeutic strategies against hematological malignancies using existing small-molecule PAK inhibitors, as well as promising combination treatments, to sensitize drug-resistant cells to conventional therapies. The challenges of toxicity and non-specific targeting associated with some PAK inhibitors, as well as how future approaches for PAK inhibition to overcome these limitations, will also be addressed.
  29. Nat Commun. 2021 Oct 26. 12(1): 6176
      Serine is a non-essential amino acid that is critical for tumour proliferation and depletion of circulating serine results in reduced tumour growth and increased survival in various cancer models. While many cancer cells cultured in a standard tissue culture medium depend on exogenous serine for optimal growth, here we report that these cells are less sensitive to serine/glycine depletion in medium containing physiological levels of metabolites. The lower requirement for exogenous serine under these culture conditions reflects both increased de novo serine synthesis and the use of hypoxanthine (not present in the standard medium) to support purine synthesis. Limiting serine availability leads to increased uptake of extracellular hypoxanthine, sparing available serine for other pathways such as glutathione synthesis. Taken together these results improve our understanding of serine metabolism in physiologically relevant nutrient conditions and allow us to predict interventions that may enhance the therapeutic response to dietary serine/glycine limitation.
  30. Nature. 2021 Oct 27.
      Glutathione (GSH) is a small-molecule thiol that is abundant in all eukaryotes and has key roles in oxidative metabolism1. Mitochondria, as the major site of oxidative reactions, must maintain sufficient levels of GSH to perform protective and biosynthetic functions2. GSH is synthesized exclusively in the cytosol, yet the molecular machinery involved in mitochondrial GSH import remains unknown. Here, using organellar proteomics and metabolomics approaches, we identify SLC25A39, a mitochondrial membrane carrier of unknown function, as a regulator of GSH transport into mitochondria. Loss of SLC25A39 reduces mitochondrial GSH import and abundance without affecting cellular GSH levels. Cells lacking both SLC25A39 and its paralogue SLC25A40 exhibit defects in the activity and stability of proteins containing iron-sulfur clusters. We find that mitochondrial GSH import is necessary for cell proliferation in vitro and red blood cell development in mice. Heterologous expression of an engineered bifunctional bacterial GSH biosynthetic enzyme (GshF) in mitochondria enables mitochondrial GSH production and ameliorates the metabolic and proliferative defects caused by its depletion. Finally, GSH availability negatively regulates SLC25A39 protein abundance, coupling redox homeostasis to mitochondrial GSH import in mammalian cells. Our work identifies SLC25A39 as an essential and regulated component of the mitochondrial GSH-import machinery.