Faculty Bibliography

Leukemia-driving mutations are thought to arise in hematopoietic stem cells (HSC), yet the natural history of their spread is poorly understood. We genetically induced mutations within endogenous murine HSC and traced them in unmanipulated animals. In contrast to mutations associated with clonal hematopoiesis (such as Tet2 deletion), the leukemogenic KrasG12D mutation dramatically accelerated HSC contribution to all hematopoietic lineages. The acceleration was mediated by KrasG12D-expressing multipotent progenitors (MPP) that lacked self-renewal but showed increased proliferation and aberrant transcriptome. The deletion of osteopontin, a secreted negative regulator of stem/progenitor cells, delayed the early expansion of mutant progenitors. KrasG12D-carrying cells showed increased CXCR4-driven motility in the bone marrow, and the blockade of CXCR4 reduced the expansion of MPP in vivo. Finally, therapeutic blockade of KRASG12D spared mutant HSC but reduced the expansion of mutant MPP and their mature progeny. Thus, transforming mutations facilitate their own spread from stem cells by reprogramming MPP, creating a preleukemic state via a two-component stem/progenitor circuit.

Long noncoding RNAs (lncRNAs) are a significant yet largely uncharted component of the cancer transcriptome, with their isoform-specific functions remaining poorly understood. In this study, we employed RNA-targeting CRISPR-Cas13d to uncover and characterize hundreds of tumor-essential (te)-lncRNA isoforms with clinical relevance. Focusing on multiple myeloma (MM), we targeted the lncRNA transcriptome expressed in tumor cells from MM patients and revealed both MM-specific and pan-cancer dependencies across diverse cancer cell lines, which we further validated in animal models. Additionally, we mapped the subcellular localization of these te-lncRNAs, identifying over 30 cytosolic isoforms that proved essential when targeted by cytosol-localized Cas13d. Notably, a specific isoform of SNHG6, enriched in the endoplasmic reticulum, interacts with heat shock proteins to maintain cellular proteostasis. We also integrated functional and clinical data into the publicly accessible LongDEP Portal, providing a valuable resource for the research community. Our study offers a comprehensive characterization of te-lncRNAs, underscoring their oncogenic roles and therapeutic potential.

Isocitrate dehydrogenase 1/2 (IDH) mutations are early initiating events in acute myeloid leukemia (AML). The complex clonal architecture and cellular heterogeneity in IDH-mutant AML underlies the heterogeneous clinical presentation and outcomes. Integrating single-cell genotyping and transcriptomics, we demonstrate a stem-like and inflammatory phenotype of IDH-mutant AML and identify clone-specific programs associated with NPM1, NRAS, and SRSF2 co-mutations. Furthermore, these clones had distinct responses to treatment with combination IDH inhibitors and chemotherapy, including elimination, reconstitution of myeloid differentiation, or retention within progenitor populations. At relapse after IDH inhibitor monotherapy, we identify upregulated stemness, inflammation, mitochondrial metabolism, and anti-apoptotic factors, as well as downregulated major histocompatibility complex (MHC) class II antigen presentation. At the pre-leukemic stage, we observe upregulation of IDH2-associated pathways, including inflammation. We deliver a detailed phenotyping of IDH-mutant AML and a framework for dissecting contributions of recurrently mutated genes in AML at diagnosis and following therapy, with implications for precision medicine.

While the mutational landscape across early T-cell precursor acute lymphoblastic leukemia (ETP-ALL) and ETP-like leukemia is known, establishing a unified framework that activates stem cell genes characteristic of these tumors remains elusive. Using complementary mouse and human models, chromatin mapping, and enhancer profiling, we show that the coactivator ZMIZ1 promotes normal and malignant ETP population growth by inducing the transcription factor MYB in feedforward circuits to convergently activate oncogenes (MEF2C, MYCN, and BCL2) through essential enhancers. A key superenhancer, the N-Myc regulating enhancer (NMRE), drives malignant ETP population growth but is dispensable for normal lymphopoiesis. This network of stem cell superenhancers identifies treatment-resistant tumors and poor survival outcomes; unifies diverse ETP-ALLs; and contributes to cardinal features of the recently genomically identified high-risk bone marrow progenitor-like (BMP-like) ETP-ALL tumor-stem cell/myeloid gene expression, inhibited NOTCH1-induced T-cell development, aggressive clinical behavior, and venetoclax sensitivity. Since ZMIZ1 is dispensable for essential homeostasis, it might be possible to safely target this network to treat high-risk diseases.

Leukemias are a class of human cancers that originate from hematopoietic progenitors and are characterized by extensive remodeling of the immune microenvironment. Leukemic cells, on transformation, acquire the ability to evade immune recognition but, despite undergoing genetic and epigenetic changes, retain their characteristic immature immune signature. For this and other reasons, leukemias are often refractory to immune therapies. In the present Review, we cover these areas as a means of improving outcomes from a deeper understanding of immune rewiring, inflammatory signaling and the barriers to successful implementation of immune therapies.

The Hhex gene encodes a transcription factor that is important for both embryonic and post-natal development, especially of hematopoietic tissues. Hhex is one of the most common sites of retroviral integration in mouse models. We found the most common integrations in AKXD (recombinant inbred strains) T-ALLs occur 57-61kb 3' of Hhex and activate Hhex gene expression. The genomic region of murine leukemia virus (MLV) integrations has features of a developmental stage-specific cis regulatory element (CRE), as evidenced by ATAC-seq in murine progenitor cells and high H3K27 acetylation at the syntenic CRE in human hematopoietic cell lines. With ChIP-exonuclease, we describe occupancy of LIM domain binding protein 1 (LDB1), the constitutive partner of the LIM Only-2 (LMO2), GATA1, and TAL1 transcription factors at GATA sites and a composite GATA-E box within the CRE. With virtual 4C analysis, we observed looping between this +65kb CRE and the proximal intron 1 enhancer of HHEX in primary human ETP-ALLs and in normal progenitor cells. Our results show that retroviral integrations at intergenic sites can mark and take advantage of CREs. Specifically, in the case of HHEX activation, this newly described +65kb CRE is co-opted in the pathogenesis of ETP-ALL by the LMO2/LDB1 complex.

Innate-like splenic marginal zone (MZ) B (MZB) cells play unique roles in immunity due to their rapid responsiveness to blood-borne microbes. How MZB cells integrate cell-extrinsic and -intrinsic processes to achieve accelerated responsiveness is unclear. We found that Delta-like1 (Dll1) Notch ligands in splenic fibroblasts regulated MZB cell pool size, migration, and function. Dll1 could not be replaced by the alternative Notch ligand Dll4. Dll1-Notch2 signaling regulated a Myc-dependent gene expression program fostering cell growth and a Myc-independent program controlling cell-movement regulators such as sphingosine-1 phosphate receptor 1 (S1PR1). S1pr1-deficient B cells experienced Notch signaling within B cell follicles without entering the MZ and were retained in the spleen upon Notch deprivation. Key elements of the mouse B cell Notch regulome were preserved in subsets of human memory B cells and B cell lymphomas. Thus, specialized niches program the poised state and patrolling behavior of MZB cells via conserved Myc-dependent and Myc-independent Notch2-regulated mechanisms.

Cancer progression involves genetic and epigenetic changes that disrupt chromatin 3D organization, affecting enhancer-promoter interactions and promoting growth. Here, we provide an integrative approach, combining chromatin conformation, accessibility, and transcription analysis, validated by in silico and CRISPR-interference screens, to identify relevant 3D topologies in pediatric T cell leukemia (T-ALL and ETP-ALL). We characterize 3D hubs as regulatory centers for oncogenes and disease markers, linking them to biological processes like cell division, inflammation, and stress response. Single-cell mapping reveals heterogeneous gene activation in discrete epigenetic clones, aiding in patient stratification for relapse risk after chemotherapy. Finally, we identify MYB as a 3D hub regulator in leukemia cells and show that the targeting of key regulators leads to hub dissolution, thereby providing a novel and effective anti-leukemic strategy. Overall, our work demonstrates the relevance of studying oncogenic 3D hubs to better understand cancer biology and tumor heterogeneity and to propose novel therapeutic strategies.

Diverse cellular insults converge on activation of the heat shock factor 1 (HSF1), which regulates the proteotoxic stress response to maintain protein homoeostasis. HSF1 regulates numerous gene programmes beyond the proteotoxic stress response in a cell-type- and context-specific manner to promote malignancy. However, the role(s) of HSF1 in immune populations of the tumour microenvironment remain elusive. Here, we leverage an in vivo model of HSF1 activation and single-cell transcriptomic tumour profiling to show that augmented HSF1 activity in natural killer (NK) cells impairs cytotoxicity, cytokine production and subsequent anti-tumour immunity. Mechanistically, HSF1 directly binds and regulates the expression of key mediators of NK cell effector function. This work demonstrates that HSF1 regulates the immune response under the stress conditions of the tumour microenvironment. These findings have important implications for enhancing the efficacy of adoptive NK cell therapies and for designing combinatorial strategies including modulators of NK cell-mediated tumour killing.