Biology subjects

Wei, A. H.

Publications and source records attributed to Wei, A. H..

5 recordsLinked to original sources

bioRxiv2024

Inhibition of heme biosynthesis triggers cuproptosis in acute myeloid leukaemia.

The ubiquitous metabolite heme has diverse enzymatic and signalling functions in most mammalian cells. Cells can salvage heme from the extracellular environment or synthesise heme de novo from succinyl-CoA and glycine through a series of 8 enzymatic reactions catalysed by heme biosynthesis enzymes (HBEs) localised in the mitochondria and the cytosol1,2. Through integrated analyses of mouse models, human cell lines and primary patient samples, we identify de novo heme biosynthesis as a selective dependency in acute myeloid leukaemia (AML). The dependency is underpinned by a propensity of AML cells, and especially leukaemic stem cells (LSCs) to downregulate HBEs. The resultant low heme state upregulates self-renewal genes via the heme sensing transcription factor BACH1, but also places leukaemia cells on the threshold of heme starvation. Genetic or pharmacological inhibition of HBEs induces cuproptosis, a form of programmed cell death caused by copper accumulation and oligomerisation of lipoylated proteins3. Moreover, we identify pathways that are synthetic lethal with heme biosynthesis, including glycolysis, which can be leveraged for combination strategies. Altogether, our work uncovers a heme rheostat that controls gene expression and drug sensitivity in AML and implicates HBE inhibition as a novel cuproptosis trigger.

cancer biology↗

bioRxiv2024

Adaptive resistance to FLT3 inhibitors is potentiated by ROS-driven DNA repair signalling

Alterations in the FMS-like tyrosine kinase 3 (FLT3) gene are the most frequent driver mutations in acute myeloid leukaemia (AML), linked to a high risk of relapse in patients with internal tandem duplications (FLT3-ITD). Tyrosine kinase inhibitors (TKIs) targeting the FLT3 protein are approved for clinical use, yet resistance often emerges. This resistance is mainly seen following the acquisition of additional point mutations in the tyrosine kinase domain (TKD), resulting in a double mutant FLT3-ITD/TKD, which sustains cell signalling and survival despite the presence of FLT3 inhibitors. Here, we developed a FLT3-mutant AML model with adaptive resistance to type II TKIs, sorafenib, and quizartinib by in vitro drug selection. Through global multiomic profiling, we identified upregulation of proteins involved in reactive oxygen species (ROS) production, particularly NADPH-oxidases, driving cellular ROS-addiction, with resistant cells relying on ROS for survival, and genome fidelity preserved by ATM-driven DNA repair. Transcriptomic analysis of adult and paediatric AML (pAML) patients identified high ATM expression as a biomarker for shorter median overall survival in both the de novo and relapsed settings. Inhibition of ATM with clinically relevant therapy WSD-0628 effectively killed TKI- and chemotherapy-resistant AML cells in vitro and significantly extended the survival of mice with sorafenib- and quizartinib-resistant FLT3-ITD AML in vivo. We propose a new treatment strategy to improve survival of patients who develop resistance to sorafenib and quizartinib, as well as relapsed and refractory pAML, exploiting resistance mechanisms to precision therapies and cell-intrinsic features of high-risk cases, highlighting a clinically relevant salvage strategy.

cancer biology↗

bioRxiv2024

Single-cell Rapid Capture Hybridization sequencing (scRaCH-seq) to reliably detect isoform usage and coding mutations in targeted genes at a single-cell level

Single-cell long-read sequencing has transformed our understanding of isoform usage and the mutation heterogeneity between cells. Despite unbiased in-depth analysis, the low sequencing throughput often results in insufficient read coverage thereby limiting our ability to perform mutation calling for specific genes. Here, we developed a single-cell Rapid Capture Hybridization sequencing (scRaCH-seq) method that demonstrated high specificity and efficiency in capturing targeted transcripts using long-read sequencing, allowing an in-depth analysis of mutation status and transcript usage for genes of interest. The method includes creating a probe panel for transcript capture, using barcoded primers for pooling and efficient sequencing via Oxford Nanopore Technologies platforms. scRaCH-seq is applicable to stored and indexed single-cell cDNA which allows analysis to be combined with existing short-read RNA-seq datasets. In our investigation of BTK and SF3B1 genes in samples from patients with chronic lymphocytic leukaemia (CLL), we detected SF3B1 isoforms and mutations with high sensitivity. Integration with short-read scRNA-seq data revealed significant gene expression differences in SF3B1-mutated CLL cells, though it did not impact the sensitivity of the anti-cancer drug venetoclax. scRaCH-seqs capability to study long-read transcripts of multiple genes makes it a powerful tool for single-cell genomics.

molecular biology↗

bioRxiv2022

Blockade of redox second messengers inhibits JAK/STAT and MEK/ERK signaling sensitizing FLT3-mutant acute myeloid leukemia to targeted therapies

FLT3-mutations are diagnosed in 25-30% of patients with acute myeloid leukemia (AML) and are associated with a poor prognosis. AML is associated with the overproduction of reactive oxygen species (ROS), which drives genomic instability through the oxidation of DNA bases, promoting clonal evolution, treatment resistance and poor outcomes. ROS are also important second messengers, triggering cysteine oxidation in redox sensitive signaling proteins, however, the specific pathways influenced by ROS in AML remain enigmatic. Here we have surveyed the posttranslational architecture of primary AML patient samples and assessed oncogenic second messenger signaling. Signaling proteins responsible for growth and proliferation were differentially oxidized and phosphorylated between patient subtypes either harboring recuring mutation in FLT3 compared to patients expressing the wildtype-FLT3 receptor, particularly those mapping to the Src family kinases (SFKs). Patients harboring FLT3-mutations also showed increased oxidative posttranslational modifications in the GTPase Rac activated-NADPH oxidase-2 (NOX2) complex to drive autocratic ROS production. Pharmacological and molecular inhibition of NOX2 was cytotoxic specifically to FLT3-mutant AMLs, and reduced phosphorylation of the critical hematopoietic transcription factor STAT5 and MAPK/ERK to synergistically increase sensitivity to FLT3-inhibitors. NOX2 inhibition also reduced phosphorylation and cysteine oxidation of FLT3 in patient derived xenograft mouse models in vivo, highlighting an important link between oxidative stress and oncogenic signaling. Together, these data raise the promising possibility of targeting NOX2 in combination with FLT3-inhibitors to improve treatment of FLT3-mutant AML. One Sentence SummaryFLT3-precision therapies have entered the clinic for AML however, their durability is limited. Here we identify the Rac-NOX2 complex as the major driver of redox second messenger signaling in FLT3-mutant AML. Molecular and pharmacological inhibition of NOX2 decreased FLT3, STAT5 and MEK/ERK signaling to delay leukemia progression, and synergistically combined with FLT3 inhibitors.

cancer biology↗

bioRxiv2020

Reprogramming of serine metabolism is an actionable vulnerability in FLT3-ITD driven acute myeloid leukaemia

Activating FMS-like tyrosine kinase 3 (FLT3) mutations occur in approximately 30% of all acute myeloid leukaemias (AMLs) and are associated with poor prognosis. The limited clinical efficacy of FLT3 inhibitor monotherapy has highlighted the need for alternative therapeutic targets and treatments for FLT3-mutant AML. Using human and murine models of MLL-rearranged AML harbouring FLT3 internal tandem duplication (FLT3-ITD) and primary patient samples, we have demonstrated that FLT3-ITD promotes serine uptake and serine synthesis via transcriptional regulation of neutral amino acid transporters (SLC1A4 and SLC1A5) and genes in the de novo serine synthesis pathway (PHGDH and PSAT1). Mechanistically, dysregulation of serine metabolism in FLT3-mutant AML is dependent on the mTORC1-ATF4 axis, that drives RNA-Pol II occupancy at PHGDH, PSAT1, SLC1A4 and SLC1A5. Genetic or pharmacological inhibition of the de novo serine synthesis pathway selectively inhibited the proliferation of FLT3-ITD AML cells, and this was potentiated by withdrawal of exogenous serine. Purine supplementation effectively rescued the antiproliferative effect of inhibiting de novo serine synthesis, consistent with the idea that serine fuels purine nucleotide synthesis in FLT3-mutant AML. Pharmacological inhibition of the de novo serine synthesis pathway, using the PHGDH inhibitor WQ-2101, sensitises FLT3-mutant AML cells to the standard of care chemotherapy agent cytarabine via exacerbation of DNA damage. Collectively, these data reveal new insights as to how FLT3 mutations reprogram metabolism in AML, and reveal a combination therapy strategy to improve the treatment of FLT3-mutant AML. Statement of SignificanceFLT3 mutations are common in AML and are associated with poor prognosis. We show that FLT3-ITD stimulates serine metabolism, thereby rendering FLT3-ITD leukemias dependent on serine for proliferation and survival. This metabolic dependency can be exploited pharmacologically to sensitize FLT3-mutant AML to chemotherapy.

cancer biology↗