SOD1 is a synthetic-lethal target in PPM1D-mutant leukemia cells.

The DNA damage response is critical for maintaining genome integrity and is commonly disrupted in the development of cancer. PPM1D (protein phosphatase Mg2+/Mn2+-dependent 1D) is a master negative regulator of the response; gain-of-function mutations and amplifications of PPM1D are found across several human cancers making it a relevant pharmacological target. Here, we used CRISPR/Cas9 screening to identify synthetic-lethal dependencies of PPM1D, uncovering superoxide dismutase-1 (SOD1) as a potential target for PPM1D-mutant cells. We revealed a dysregulated redox landscape characterized by elevated levels of reactive oxygen species and a compromised response to oxidative stress in PPM1D-mutant cells. Altogether, our results demonstrate a role for SOD1 in the survival of PPM1D-mutant leukemia cells and highlight a new potential therapeutic strategy against PPM1D-mutant cancers.

Increased iron uptake by splenic hematopoietic stem cells promotes TET2-dependent erythroid regeneration.

Hematopoietic stem cells (HSCs) are capable of regenerating the blood system, but the instructive cues that direct HSCs to regenerate particular lineages lost to the injury remain elusive. Here, we show that iron is increasingly taken up by HSCs during anemia and induces erythroid gene expression and regeneration in a Tet2-dependent manner. Lineage tracing of HSCs reveals that HSCs respond to hemolytic anemia by increasing erythroid output. The number of HSCs in the spleen, but not bone marrow, increases upon anemia and these HSCs exhibit enhanced proliferation, erythroid differentiation, iron uptake, and TET2 protein expression. Increased iron in HSCs promotes DNA demethylation and expression of erythroid genes. Suppressing iron uptake or TET2 expression impairs erythroid genes expression and erythroid differentiation of HSCs; iron supplementation, however, augments these processes. These results establish that the physiological level of iron taken up by HSCs has an instructive role in promoting erythroid-biased differentiation of HSCs.

SOD1 is a synthetic lethal target in PPM1D-mutant leukemia cells.

The DNA damage response is critical for maintaining genome integrity and is commonly disrupted in the development of cancer. PPM1D (protein phosphatase, Mg2+/Mn2+ dependent 1D) is a master negative regulator of the response; gain-of-function mutations and amplifications of PPM1D are found across several human cancers making it a relevant pharmacologic target. Here, we used CRISPR/Cas9 screening to identify synthetic-lethal dependencies of PPM1D, uncovering superoxide dismutase-1 (SOD1) as a potential target for PPM1D-mutant cells. We revealed a dysregulated redox landscape characterized by elevated levels of reactive oxygen species and a compromised response to oxidative stress in PPM1D-mutant cells. Altogether, our results demonstrate the protective role of SOD1 against oxidative stress in PPM1D-mutant leukemia cells and highlight a new potential therapeutic strategy against PPM1D-mutant cancers.

Applying single cell multi-omic analyses to understand treatment resistance in pediatric high grade glioma.

Despite improvements in cancer patient outcomes seen in the past decade, tumor resistance to therapy remains a major impediment to achieving durable clinical responses. Intratumoral heterogeneity related to genetic, epigenetic, transcriptomic, proteomic, and metabolic differences between individual cancer cells has emerged as a driver of therapeutic resistance. This cell to cell heterogeneity can be assessed using single cell profiling technologies that enable the identification of tumor cell clones that exhibit similar defining features like specific mutations or patterns of DNA methylation. Single cell profiling of tumors before and after treatment can generate new insights into the cancer cell characteristics that confer therapeutic resistance by identifying intrinsically resistant sub-populations that survive treatment and by describing new cellular features that emerge post-treatment due to tumor cell evolution. Integrative, single cell analytical approaches have already proven advantageous in studies characterizing treatment-resistant clones in cancers where pre- and post-treatment patient samples are readily available, such as leukemia. In contrast, little is known about other cancer subtypes like pediatric high grade glioma, a class of heterogeneous, malignant brain tumors in children that rapidly develop resistance to multiple therapeutic modalities, including chemotherapy, immunotherapy, and radiation. Leveraging single cell multi-omic technologies to analyze naïve and therapy-resistant glioma may lead to the discovery of novel strategies to overcome treatment resistance in brain tumors with dismal clinical outcomes. In this review, we explore the potential for single cell multi-omic analyses to reveal mechanisms of glioma resistance to therapy and discuss opportunities to apply these approaches to improve long-term therapeutic response in pediatric high grade glioma and other brain tumors with limited treatment options.

Using Cryo-ET to distinguish platelets during pre-acute myeloid leukemia from steady state hematopoiesis.

Early diagnosis of acute myeloid leukemia (AML) in the pre-leukemic stage remains a clinical challenge, as pre-leukemic patients show no symptoms, lacking any known morphological or numerical abnormalities in blood cells. Here, we demonstrate that platelets with structurally abnormal mitochondria emerge at the pre-leukemic phase of AML, preceding detectable changes in blood cell counts or detection of leukemic blasts in blood. We visualized frozen-hydrated platelets from mice at different time points during AML development in situ using electron cryo-tomography (cryo-ET) and identified intracellular organelles through an unbiased semi-automatic process followed by quantitative measurement. A large proportion of platelets exhibited changes in the overall shape and depletion of organelles in AML. Notably, 23% of platelets in pre-leukemic cells exhibit abnormal, round mitochondria with unfolded cristae, accompanied by a significant drop in ATP levels and altered expression of metabolism-related gene signatures. Our study demonstrates that detectable structural changes in pre-leukemic platelets may serve as a biomarker for the early diagnosis of AML.

Nuclear NAD+ homeostasis governed by NMNAT1 prevents apoptosis of acute myeloid leukemia stem cells.

Metabolic dysregulation underlies malignant phenotypes attributed to cancer stem cells, such as unlimited proliferation and differentiation blockade. Here, we demonstrate that NAD+ metabolism enables acute myeloid leukemia (AML) to evade apoptosis, another hallmark of cancer stem cells. We integrated whole-genome CRISPR screening and pan-cancer genetic dependency mapping to identify NAMPT and NMNAT1 as AML dependencies governing NAD+ biosynthesis. While both NAMPT and NMNAT1 were required for AML, the presence of NAD+ precursors bypassed the dependence of AML on NAMPT but not NMNAT1, pointing to NMNAT1 as a gatekeeper of NAD+ biosynthesis. Deletion of NMNAT1 reduced nuclear NAD+, activated p53, and increased venetoclax sensitivity. Conversely, increased NAD+ biosynthesis promoted venetoclax resistance. Unlike leukemia stem cells (LSCs) in both murine and human AML xenograft models, NMNAT1 was dispensable for hematopoietic stem cells and hematopoiesis. Our findings identify NMNAT1 as a previously unidentified therapeutic target that maintains NAD+ for AML progression and chemoresistance.

The histone H3.3 chaperone HIRA restrains erythroid-biased differentiation of adult hematopoietic stem cells.

Histone variants contribute to the complexity of the chromatin landscape and play an integral role in defining DNA domains and regulating gene expression. The histone H3 variant H3.3 is incorporated into genic elements independent of DNA replication by its chaperone HIRA. Here we demonstrate that Hira is required for the self-renewal of adult hematopoietic stem cells (HSCs) and to restrain erythroid differentiation. Deletion of Hira led to rapid depletion of HSCs while differentiated hematopoietic cells remained largely unaffected. Depletion of HSCs after Hira deletion was accompanied by increased expression of bivalent and erythroid genes, which was exacerbated upon cell division and paralleled increased erythroid differentiation. Assessing H3.3 occupancy identified a subset of polycomb-repressed chromatin in HSCs that depends on HIRA to maintain the inaccessible, H3.3-occupied state for gene repression. HIRA-dependent H3.3 incorporation thus defines distinct repressive chromatin that represses erythroid differentiation of HSCs.

Transcriptional and Microenvironmental Regulation of Lineage Ambiguity in Leukemia.

Leukemia is characterized by the uncontrolled production of leukemic cells and impaired normal hematopoiesis. Although the combination of chemotherapies and hematopoietic stem cell transplantation has significantly improved the outcome of leukemia patients, a proportion of patients still suffer from relapse after treatment. Upon relapse, a phenomenon termed "lineage switch" is observed in a subset of leukemia patients, in which conversion of lymphoblastic leukemia to myeloid leukemia or vice versa is observed. A rare entity of leukemia called mixed-phenotype acute leukemia exhibits co-expression of markers representing two or three lineages. These two phenotypes regarding the lineage ambiguity suggest that the fate of some leukemia retain or acquire a certain degree of plasticity. Studies using animal models provide insight into how lineage specifying transcription factors can enforce or convert a fate in hematopoietic cells. Modeling lineage conversion in normal hematopoietic progenitor cells may improve our current understanding of how lineage switch occurs in leukemia. In this review, we will summarize the role of transcription factors and microenvironmental signals that confer fate plasticity to normal hematopoietic progenitor cells, and their potential to regulate lineage switching in leukemias. Future efforts to uncover the mechanisms contributing to lineage conversion in both normal hematopoiesis and leukemia may pave the way to improve current therapeutic strategies.