Transcription Factors, R-Loops and Deubiquitinating Enzymes: Emerging Targets in Myelodysplastic Syndromes and Acute Myeloid Leukemia
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cancers Review Transcription Factors, R-Loops and Deubiquitinating Enzymes: Emerging Targets in Myelodysplastic Syndromes and Acute Myeloid Leukemia Silvia M. L. Barabino , Elisabetta Citterio * and Antonella Ellena Ronchi * Department of Biotechnology and Biosciences, University of Milano-Bicocca, 20126 Milan, Italy; [email protected] * Correspondence: [email protected] (E.C.); [email protected] (A.E.R.) Simple Summary: The advent of DNA massive sequencing technologies has allowed for the first time an extensive look into the heterogeneous spectrum of genes and mutations underpinning myelodysplastic syndromes (MDSs) and acute myeloid leukemia (AML). In this review, we wish to explore the most recent advances and the rationale for the potential therapeutic interest of three main actors in myelo-leukemic transformation: transcription factors that govern myeloid differentiation; RNA splicing factors, which ensure proper mRNA maturation and whose mutations increase R-loops formation; and deubiquitinating enzymes, which contribute to genome stability in hematopoietic stem cells (HSCs). Abstract: Myeloid neoplasms encompass a very heterogeneous family of diseases characterized by the failure of the molecular mechanisms that ensure a balanced equilibrium between hematopoietic stem cells (HSCs) self-renewal and the proper production of differentiated cells. The origin of Citation: Barabino, S.M.L.; Citterio, the driver mutations leading to preleukemia can be traced back to HSC/progenitor cells. Many E.; Ronchi, A.E. Transcription Factors, properties typical to normal HSCs are exploited by leukemic stem cells (LSCs) to their advantage, R-Loops and Deubiquitinating leading to the emergence of a clonal population that can eventually progress to leukemia with Enzymes: Emerging Targets in Myelodysplastic Syndromes and variable latency and evolution. In fact, different subclones might in turn develop from the original Acute Myeloid Leukemia. Cancers malignant clone through accumulation of additional mutations, increasing their competitive fitness. 2021, 13, 3753. https://doi.org/ This process ultimately leads to a complex cancer architecture where a mosaic of cellular clones—each 10.3390/cancers13153753 carrying a unique set of mutations—coexists. The repertoire of genes whose mutations contribute to the progression toward leukemogenesis is broad. It encompasses genes involved in different cellular Academic Editor: Christopher I. Slape processes, including transcriptional regulation, epigenetics (DNA and histones modifications), DNA damage signaling and repair, chromosome segregation and replication (cohesin complex), RNA Received: 16 June 2021 splicing, and signal transduction. Among these many players, transcription factors, RNA splicing Accepted: 23 July 2021 proteins, and deubiquitinating enzymes are emerging as potential targets for therapeutic intervention. Published: 26 July 2021 Keywords: myelodysplastic syndromes (MDS); acute myeloid leukemia (AML); transcription factors; Publisher’s Note: MDPI stays neutral RNA splicing; R-loops; genome integrity; deubiquitinating enzymes (DUBs) with regard to jurisdictional claims in published maps and institutional affil- iations. 1. Introduction Hematopoietic stem cells (HSCs) ensure continuous production of all blood cell types throughout life. By means of a delicate equilibrium between self-renewal, quiescence, and Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. differentiation, HSCs maintain homeostatic conditions and dynamically respond to stress This article is an open access article stimuli. The balanced production of the different cell lineages physiologically varies during distributed under the terms and aging, when the hematopoietic potential is progressively skewed towards the myeloid conditions of the Creative Commons lineages at the expenses of immune cells [1–3]. The accumulation of heritable genetic Attribution (CC BY) license (https:// mutations in individual cells and the kinetics of their selection can lead to myeloid neo- creativecommons.org/licenses/by/ plasms, a heterogeneous group of diseases characterized by the dysfunctional production 4.0/). of myeloid cells in the bone marrow. This can manifest in cytopenia and cellular dysplasia, Cancers 2021, 13, 3753. https://doi.org/10.3390/cancers13153753 https://www.mdpi.com/journal/cancers Cancers 2021, 13, 3753 2 of 41 such as in myelodysplastic syndromes (MDSs), in the overproduction of mature clonal myeloid elements, such as in myeloproliferative neoplasms (MPN), or both, as it happens in MDS/MPN, which share different molecular and clinical traits and present both myelodys- plastic and proliferative features [4]. Myeloid neoplasms entail a high risk of developing into acute myeloid leukemia (secondary AML or s-AML). AML can also develop as a late complication in patients after leukemogenic therapies (therapy-related AML or t-AML), or without clinical history of prior MDS or known exposure to potentially leukemogenic agents (de novo AML) [5]. Driver mutations leading to a preleukemia condition originate in HSC or in hematopoietic stem-progenitor cells (HSPCs). The preleukemic clones can have a variable latency and, in some cases, can persist for years before further mutations trigger their leukemic evolution [4,6]. MDS has an estimated crude incidence of 4 to 5 cases per 100,000 persons per year. Al- though MDS occurs at all ages, the incidence is higher in elderly individuals, with a median age of 70 years old at diagnosis [4]. Careful evaluation of the individual patient prognostic risk, genetics, and age guides the clinical decision-making process [4]. Although current drugs can temporarily modulate myelodysplastic hematopoiesis, they fail in eradicating the disease [4,7]. Failure of current therapies in eradicating MDS/AML is attributed to the persis- tence of leukemic stem cells (LSCs) upon treatment and to the emergence of resistant subclones [8]. Notably, recent deep-sequencing studies revealed the possibility that relapse from chemotherapy can occur not only from LSCs that are resistant to the treatment but also from pre-leukemic mutated but not transformed HSCs, which are already present in the patient at diagnosis or during clinical remission, and which can acquire additional mutations, further highlighting the complexity and heterogeneity of the disease [9,10]. LSCs share functional properties with normal HSCs [11]. Indeed, LSCs are functionally defined as cells capable of self-renewal and of propagating the disease upon transplantation into immunodeficient mice [8]. In addition to self-renewal capacity, LSCs co-opt many survival mechanisms typical to HSCs to their advantage, including genome maintenance processes, epigenetic and stemness transcriptional programs, the pre-mRNA splicing ma- chinery, metabolic properties, interaction with the microenvironment, and inflammatory signals [11,12]. In fact, recurring mutations in regulators of gene expression, including epigenetic proteins, transcription factors (TFs), and the components of the splicing machin- ery, are found in 70% of AML patients [13,14]. Understanding the molecular mechanisms regulating HSC biology and their dysregulation in pre-leukemic HSCs and in LCSs is therefore critical for understanding the disease and for developing therapies that harness cancer HSPC-specific vulnerabilities and ultimately eradicate the disease. 2. Genetics Underlying Evolutionary Trajectories of MDS Progression MDS entails both molecular and cytogenetic complexity. Early studies and more recent analysis through whole-genome sequencing of somatic mutations in MDS patients identified more than 50 recurrently mutated genes. These genes can be broadly divided into a few main categories controlling basic cellular functions: (1) transcriptional and epigenetic control of gene expression; (2) RNA splicing; (3) genome integrity; and (4) signal transduction pathways. Notably, only six genes are most frequently mutated and found in at least 10% of patients with MDS: DNMT3A, TET2, ASXL1, SF3B1, SRSF2, and RUNX1 [13,15,16]. These studies also pointed to major mutation-driver genes acquired in each of the phases during the course of the disease, from initial clonal hematopoiesis (CH), characterized by the presence of HSC/hematopoietic precursors carrying somatic mutations, through to development of MDS and eventually progression to AML (Figure 1), suggesting that CH–MDS–AML development constitutes a continuous evolutionary process. Cancers 2021, 13, 3753 3 of 42 Cancers 2021, 13, 3753 3 of 41 No progression Normal CHIP MDS AML hematopoiesis STEM CELLS SELECTIVE ADVANTAGE INCREASED SELF-RENEWAL INHIBITION OF DIFFERENTIATION Epigenetics: Splicing: Epigenetics and metabolism: DNMT3A, TET2 SF3B1, SRSF2,U2AF1 EZH2, BCOR, IDH1, IDH2 ASXL1 Transcription factors: Genome organization: RUNX1 STAG2 Transcription factors: NMP1, GATA2, CEBPA, WT1 RECURRENT Signaling: NF1,NRAS, FLT3, KRAS, CBL SOMATIC MUTATIONS HSC pre-leukemic HSC LSC Self-renewal Self-renewal Figure 1. General model of myelodysplastic syndromes (MDS) progression.progression. MDS is a progressive disease, developing through the accumulation of mutations in hematopoietic stem cells (HSCs) or early progenitors that promotes the growth through the accumulation of mutations in hematopoietic stem cells (HSCs) or early progenitors that promotes the growth and spread of somatically mutated clones in the bone marrow (BM) and eventually leads to overt clinical disease and to and spread of somatically