Biology subjects

Gan, O. I.

Publications and source records attributed to Gan, O. I..

7 recordsLinked to original sources

bioRxiv2024

FLT3 is genetically essential for ITD-mutated leukemic stem cells but dispensable for human hematopoietic stem cells

Leukemic stem cells (LSCs) fuel acute myeloid leukemia (AML) growth and relapse, but therapies tailored towards eradicating LSCs without harming healthy hematopoietic stem cells (HSCs) are lacking. FLT3 is considered an important therapeutic target due to frequent mutation in AML and association with relapse. However, there has been limited clinical success with FLT3 targeting, suggesting either that FLT3 is not a vulnerability in LSC, or that more potent inhibition is required, a scenario where HSC toxicity could become limiting. We tested these possibilities by ablating FLT3 using CRISPR/Cas9-mediated FLT3 knock-out (FLT3-KO) in human LSCs and HSCs followed by functional xenograft assays. FLT3-KO in LSCs from FLT3-ITD mutated, but not FLT3-WT AMLs, resulted in short-term leukemic grafts of FLT-3-KO edited cells that disappeared by 12 weeks. By contrast, FLT3-KO in HSCs from fetal liver, cord blood and adult bone marrow did not impair multilineage hematopoiesis in primary and secondary xenografts. Our study establishes FLT3 as an ideal therapeutic target where ITD+ LSC are eradicated upon FLT3 deletion, while HSCs are spared. These findings support the development of more potent FLT3-targeting drugs or gene-editing approaches for LSC eradication to improve clinical outcomes. KEY POINTSThe FLT3 gene is essential for ITD-mutated leukemic stem cells (LSCs) to establish and propagate leukemia. Normal human hematopoietic stem cells (HSCs) do not require FLT3 to engraft and sustain hematopoiesis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=179 SRC="FIGDIR/small/601363v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@b77f56org.highwire.dtl.DTLVardef@109011dorg.highwire.dtl.DTLVardef@282e04org.highwire.dtl.DTLVardef@1e51e3b_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2022

KDM6A loss sensitizes human acute myeloid leukemia to PARP and BCL2 inhibition

Acute myeloid leukemia (AML) is a heterogeneous, aggressive malignancy with dismal prognosis and with limited availability of targeted therapies. AML exhibits epigenetic deregulation and transcriptional plasticity that contributes to pathogenesis. KDM6 proteins are histone-3 lysine-27 demethylases that play major context dependent roles in AML evolution and therapy resistance. Here, we demonstrate that KDM6 demethylase function critically regulates DNA damage repair (DDR) gene expression programs in AML. Mechanistically, KDM6 family protein expression is regulated by genotoxic stress, with deficiency of KDM6A (UTX) and KDM6B (JMJD3) impairing DDR transcriptional activation and compromising repair potential. Acquired KDM6A loss-of-function mutations have been implicated in chemoresistance, although a significant percentage of relapsed AML have upregulated KDM6A. Based on these mechanistic findings, olaparib treatment significantly reduced engraftment of patient-derived xenografts. Thus KDM6A-mutant human primary AML samples have increased susceptibility to Poly-(ADP-ribose)-polymerase (PARP) inhibition in vivo. Crucially, a higher KDM6A expression is correlated with venetoclax tolerance. Loss of KDM6A increased mitochondrial activity, BCL2 expression, and sensitized AML cells to venetoclax. Additionally, KDM6A loss was accompanied with a downregulated BCL2A1, which is commonly associated with venetoclax resistance. Corroborating these results, dual targeting of PARP and BCL2 was superior to PARP or BCL2 inhibitor monotherapy in inducing AML apoptosis, and primary AML cells carrying acquired KDM6A-domain mutations were even more sensitive to the combination. Together, our study illustrates a mechanistic rationale in support for a novel combination targeted therapy for human AML based on subtype heterogeneity, and establishes KDM6A as an important molecular regulator for determining therapeutic efficacy.

cancer biology↗

bioRxiv2021

Identification of the Global miR-130a Targetome Reveals a Novel Role for TBL1XR1 in Hematopoietic Stem Cell Self-Renewal and t(8;21) AML

Gene expression profiling and proteome analysis of normal and malignant hematopoietic stem cells (HSC) point to shared core stemness properties. However, discordance between mRNA and protein signatures underscores an important role for post-transcriptional regulation by miRNAs in governing this critical nexus. Here, we identified miR-130a as a regulator of HSC self-renewal and differentiation. Enforced expression of miR-130a impaired B lymphoid differentiation and expanded long-term HSC. Integration of protein mass spectrometry and chimeric AGO2 eCLIP-seq identified TBL1XR1 as a primary miR-130a target, whose loss of function phenocopied miR-130a overexpression. Moreover, we found that miR-130a is highly expressed in t(8;21) AML where it is critical for maintaining the oncogenic molecular program mediated by AML1-ETO. Our study establishes that identification of the comprehensive miRNA targetome within primary cells enables discovery of novel genes and molecular networks underpinning stemness properties of normal and leukemic cells. HIGHLIGHTSO_LImiR-130a is a regulator of HSC self-renewal and lineage commitment C_LIO_LITBL1XR1 is a principal target of miR-130a C_LIO_LITBL1XR1 loss of function in HSPC phenocopies enforced expression of miR-130a C_LIO_LIElevated miR-130a levels maintain the AML1-ETO repressive program in t(8;21) AML C_LI

cell biology↗

bioRxiv2021

Dichotomous regulation of lysosomes by MYC and TFEB controls hematopoietic stem cell fate

It is critical to understand how quiescent long-term hematopoietic stem cells (LT-HSC) sense demand from daily and stress-mediated cues and transition into bioenergetically active progeny to differentiate and meet these cellular needs. Here, we show that lysosomes, which are sophisticated nutrient sensing and signaling centers, are dichotomously regulated by the Transcription Factor EB (TFEB) and MYC to balance catabolic and anabolic processes required for activating LT-HSC and guiding their lineage fate. TFEB-mediated induction of the endolysosomal pathway causes membrane receptor degradation, limiting LT-HSC metabolic and mitogenic activation, which promotes quiescence, self-renewal and governs erythroid-myeloid commitment. By contrast, MYC engages biosynthetic processes while repressing lysosomal catabolism to drive LT-HSC activation. Collectively, our study identifies lysosomes as a central regulatory hub for proper and coordinated stem cell fate determination.

cell biology↗

bioRxiv2020

Mapping the Cellular Origin and Early Evolution of Leukemia in Down Syndrome

Children with Down syndrome have a 150-fold increased risk of developing myeloid leukemia, but the mechanism of predisposition is unclear. As Down syndrome leukemogenesis initiates during fetal development, we characterized the cellular context of preleukemic initiation and leukemic progression using gene editing in human disomic and trisomic fetal liver hematopoietic cells and xenotransplantation. GATA1 mutations caused transient preleukemia only when introduced into trisomy 21 long-term hematopoietic stem cells, where a subset of chromosome 21 miRNAs triggers predisposition to preleukemia. By contrast, progression to leukemia was independent of trisomy 21 and originated in various stem and progenitor cells through additional mutations in cohesin genes. CD117+/KIT cells mediated the propagation of preleukemia and leukemia, and functional KIT inhibition targeted preleukemic stem cells, blocking progression to leukemia.

cancer biology↗

bioRxiv2019

Genetic predisposition to myeloproliferative neoplasms implicates hematopoietic stem cell biology

Myeloproliferative neoplasms (MPNs) are blood cancers characterized by excessive production of mature myeloid cells that result from the acquisition of somatic driver mutations in hematopoietic stem cells (HSCs)1. While substantial progress has been made to define the causal somatic mutation profile for MPNs2, epidemiologic studies indicate a significant heritable component for the disease that is among the highest known for all cancers3. However, only a limited set of genetic risk loci have been identified, and the underlying biological mechanisms leading to MPN acquisition remain unexplained. Here, to define the inherited risk profile, we conducted the largest genome-wide association study of MPNs to date (978,913 individuals with 3,224 cases) and identified 14 genome-wide significant loci, as well as a polygenic signature that increases the odds for disease acquisition by nearly 3-fold between the top and median deciles. Interestingly, we find a shared genetic architecture between MPN risk and several hematopoietic traits spanning distinct lineages, as well as an association between increased MPN risk and longer leukocyte telomere length, collectively implicating HSC function and self-renewal. Strikingly, we find a significant enrichment for risk variants mapping to accessible chromatin in HSCs compared with other hematopoietic populations. Finally, gene mapping identifies modulators of HSC biology and targeted variant-to-function analyses suggest likely roles for CHEK2 and GFI1B in altering HSC function to confer disease risk. Overall, we demonstrate the power of human genetic studies to illuminate a previously unappreciated mechanism for MPN risk through modulation of HSC function.

genetics↗

bioRxiv2019

Functional Profiling of Single CRISPR/Cas9-Edited Human Long-Term Hematopoietic Stem Cells

In the human hematopoietic system, rare self-renewing multi-potent long-term hematopoietic stem cells (LT-HSCs) are responsible for the lifelong production of mature blood cells and are the rational target for clinical regenerative therapies. However, the heterogeneity in the hematopoietic stem cell compartment and variable outcomes of CRISPR/Cas9 editing make functional interrogation of rare LT-HSCs challenging. Here, we report high efficiency LT-HSC editing at single cell resolution using electroporation of modified synthetic gRNAs and Cas9 protein. Targeted short isoform expression of the GATA1 transcription factor elicited distinct differentiation and proliferation effects in single LT-HSC when analyzed with functional in vitro differentiation and long-term repopulation xenotransplantation assays. Our method represents a blueprint for systematic genetic analysis of complex tissue hierarchies at single cell level.

genetics↗