FTO degrader impairs ribosome biogenesis and protein translation in acute myeloid leukemia

Science Advances · 2025-08-15 · 9 citations

Targeting ribosome biogenesis and protein translation has emerged as a promising avenue for cancer therapy. The fat mass and obesity-associated protein (FTO), an RNA N 6 -methyladenosine (m 6 A) eraser, has been identified as an oncogenic factor in acute myeloid leukemia (AML). Here, we present the development of an FTO degrader that selectively degrades FTO in AML cells, demonstrating superior efficacy both in vitro and in vivo. We confirmed that FTO degradation increases m 6 A modifications on mRNAs associated with ribosome biogenesis, promoting their YTHDF2-mediated decay. This disruption of ribosome biogenesis and protein translation contributes to the inhibition of AML progression. Our findings highlight this FTO degrader as a valuable tool compound for elucidating the functional roles of FTO in cancer and as a potential foundation for the development of selective anticancer therapies.

Supplementary Tables S1-5 from METTL3-Mediated m<sup>6</sup>A Modification Controls Splicing Factor Abundance and Contributes to Aggressive CLL

2025-11-24

<p>Table S1 shows CLL sample information; Table S2 shows protein expression detected in CLL and normal B cells using TMT proteomics; Table S3 shows top 150 transcripts with highest m6A density in B cells; Table S4 shows sequences of shRNAs, sgRNAs, and primers used in our study; Table S5 shows cellular pathways enriched in our study.</p>

N6-methyladenosine regulation of mRNA translation is essential for early human erythropoiesis

bioRxiv (Cold Spring Harbor Laboratory) · 2025-11-12

Abstract N6-methyladenosine (m 6 A) is an abundant modification of mRNA with important regulatory roles in normal and malignant hematopoiesis. We previously reported that in human erythroid leukemia (HEL) cells, m 6 A mRNA marking selectively regulates translation of essential erythropoiesis genes required for in vitro differentiation and human erythroid colony formation. Here, we further investigated the timing and nature of requirement for m 6 A-methyltransferase (MTase) activity during human erythropoiesis, using a standardized in vitro erythroid differentiation assay for hHSPCs. We identified two critical m 6 A regulated developmental windows in BFU-E and during the transition from CFU-E to proerythroblasts. These windows of m 6 A-MTase requirement coincide with rising global m 6 A levels, which peak in proerythroblasts. After proerythroblast formation, however, m 6 A -MTase activity is dispensable for differentiation, proliferation, and survival. In BFU-E, m 6 A-MTase promotes proliferation but is dispensable for differentiation, while, in CFU-E, both m 6 A -MTase and the YTHDF family of m 6 A readers are essential for differentiation to proerythroblasts. Mechanistically, in CFU-E, m 6 A MTase activity enhances translation of ribosomal and oxidative phosphorylation (OXPHOS) genes, thereby elevating global protein synthesis rates and enabling efficient erythroblast formation. We propose that this form of translational regulation by m 6 A emerged as an evolutionary adaptation to meet the high translational demands of human erythropoiesis.

DDX6 undergoes phase separation to modulate metabolic plasticity and chemoresistance

Nature Communications · 2025-12-02 · 1 citations

Stress granules (SGs) and processing bodies (PBs), assembled via liquid-liquid phase separation (LLPS), are critical for spatial regulation of gene expression in the cytoplasm. However, their roles in tumorigenesis remain poorly understood. Here, we show DEAD-box helicase 6 (DDX6) as the most promising vulnerability in acute myeloid leukemia (AML) through in vitro and in vivo CRISPR screenings using a specialized library targeting RNA-binding proteins enriched in SGs and PBs. Knockout (KO) of DDX6 significantly delays leukemogenesis with minimal impact on normal hematopoiesis. Importantly, the functions of DDX6 in AML depend largely on its ability to trigger LLPS and PB assembly. Mechanistically, PBs serve as “reservoirs” for the mRNAs interacting directly with DDX6 and having low GC content. DDX6 KO leads to rapid PB dissolution and release of PB-enriched mRNAs, such as BCAT1, into the cytosol, where these transcripts undergo degradation. By reducing BCAT1 levels, DDX6 KO reprograms amino acid metabolism and sensitizes AML cells to cytarabine chemotherapy. DEAD-box helicase 6 (DDX6), the regulator of P-body assembly, is essential for the survival of acute myeloid leukemia (AML) cells. Here the authors report that DDX6 undergoes phase separation to preserve mRNA subsets in P-bodies, promoting branched-chain amino acid metabolism and chemoresistance in AML.

Supplementary Figures S1-8 from METTL3-Mediated m<sup>6</sup>A Modification Controls Splicing Factor Abundance and Contributes to Aggressive CLL

2025-11-24

<p>Fig S1. SF3B1 mutant CLL samples had pervasive changes in 3’ splice site. Fig S2. Omics analyses identify widespread post-transcriptional upregulation of splicing factors in CLL. Fig S3. Abundance of spliceosome complexes and RNA-binding proteins are associated with clinical outcomes in CLL. Fig S4. METTL3 is consistently upregulated along with differential m6A modification on transcripts of RNA splicing process in CLL. Fig S5. KO or pharmacological inhibition of METTL3 impacts cell growth. Fig S6. KO or pharmacological inhibition of METTL3 impacts apoptosis, cell cycle, and splicing factor abundance. Fig S7. Splicing factors are either direct or indirect targets of METTL3. Fig S8. Validation of association between m6A and splicing factor abundance using dCasRx-METTL3.</p>

MicroRNA-142 improves IL1RAP CAR-T cell activity in acute myeloid leukemia

Journal of Hematology & Oncology · 2025-11-12 · 1 citations

BACKGROUND: Interleukin-1 receptor accessory protein (IL1RAP) is selectively expressed on both bulk blasts and leukemic stem cells (LSCs) in acute myeloid leukemia (AML), while its expression is virtually absent on normal hematopoietic stem cells (HSCs), making it an appealing target for chimeric antigen receptor (CAR) T cell therapy. METHODS: We developed a novel IL1RAP-targeting CAR-T cells using a single-chain Fab (24scFab) fused to CD28 and CD3ζ costimulatory domains. CAR-T cells with a mutated IL1RAP-binding paratope were also generated as a control by introducing two point-mutations in the complementarity determining region (CDR) loops of the 24scFab domain. We tested the CAR-T cells in cell line-derived (CD) and patient-derived (PD) xenografts (X). To address persistence and activity of IL1RAP CAR-T cells, we then tested two approaches. First, we mutated two of the three immunoreceptor tyrosine-based activation motifs (ITAMs) within the CD3ζ domain (i.e., IL1RAP-1XX CAR-T). Second, we co-administered a synthetic miR-142 mimic (M-miR-142), previously shown to enhance T cell antileukemic activity, with IL1RAP CAR-T cells to AML xenografted mice. RESULTS: IL1RAP CAR-T cells demonstrated a potent antileukemic activity in both AML CDX and PDX models. Target specificity was confirmed by the complete loss of function of IL1RAP-mutated CAR-T cells. IL1RAP-1XX CAR-T cells improved T cell persistence in vitro but failed to demonstrate therapeutic benefit compared with IL1RAP CAR-T cells in vivo. We previously reported that leukemic cell growth suppresses miR-142 biogenesis, thereby hindering the metabolic switch and impairing host T cell antileukemic activity; this was rescued by administration of M-miR-142. Thus, we hypothesized a similar impact of leukemic cells on CAR-T and that M-miR-142 treatment could rescue it and enhance the IL1RAP CAR-T cell antileukemic activity. We showed that both CDXs and PDXs receiving M-miR-142 and IL1RAP CAR-T lived significantly longer than those receiving scrambled oligonucleotide and IL1RAP CAR-T or mutated CAR-T controls (median survival of PDX: 78 vs 51 vs 24 days). CONCLUSIONS: We have identified a potentially novel strategy to enhance CAR-T cell persistence and efficacy in AML by counteracting a leukemia-induced, microRNA-deficiency mediated mechanism of immune suppression.

Supplementary Tables S1-S3 from ALKBH1 Drives Tumorigenesis and Drug Resistance via tRNA-decoding Reprogramming and Codon-biased Translation

2025-11-03

<p>Supplementary Table S1: Effects of ALKBH1 KD on f5C34 in mitochondrial tRNA-Met Supplementary Table S2: Genetic background of the AML patient samples. Supplementary Table S3: List of oligonucleotide sequences</p>

Plasma ALKAL2 levels are associated with coronary atherosclerosis in patients with type 2 diabetes mellitus

Cardiovascular Diabetology · 2025-10-08

BACKGROUND: Cancer-associated signaling pathways, particularly the ALK/LTK receptor tyrosine kinases and their ligand ALKAL2, have recently been implicated in chronic inflammation and myocardial remodeling. However, the relationship between ALKAL2 and coronary artery disease (CAD) pathogenesis in type 2 diabetes mellitus (T2DM) remains undefined. METHODS: From January 2019 to December 2020, patients with type 2 diabetes mellitus (T2DM) undergoing coronary angiography were consecutively enrolled at Ruijin Hospital. Plasma ALKAL2 levels were measured using an ELISA assay. The association between ALKAL2 and CAD severity was assessed by Spearman correlation analysis. Logistic regression models were used to measure the association between ALKAL2 and CAD risk. RESULTS: 275 T2DM patients with CAD and 275 age- and sex-matched T2DM patients without CAD were included in the final analysis. Plasma ALKAL2 levels were increased in T2DM patients with CAD (0.25 [0.21, 0.37] ng/mL, median [IQR] vs. 0.18 [0.14, 0.24]ng/mL) (p < 0.001) and were positively associated with CAD severity (Spearman rho = 0.53, p < 0.001). Multivariate analysis revealed that plasma ALKAL2 levels were independently associated with the incidence of CAD after adjusting for LDL-C, hsCRP, and other traditional risk factors (OR, 2.24 [95% CI, 1.79-2.85]; p < 0.001). CONCLUSIONS: Elevated plasma ALKAL2 levels are an independent risk factor for T2DM CAD and are associated with the severity of CAD.

Mitochondrial RNA pseudouridine reprograms metabolism and induces immunosuppression in myeloid leukemia

Blood · 2025-11-03

Abstract Pseudouridine (Ψ) is the most prevalent RNA modification in human cells. Although Ψ was among the first post-transcriptional modifications discovered (1951), its biological functions remain poorly understood. More recently, Ψ was incorporated into COVID-19 mRNA vaccine, where it enhances RNA stability and reduces innate immune sensing. However, the precise roles of RNA Ψ in cancer, including leukemia, are largely unknown. Here, we identify RNA Pseudouridine Synthase D4 (RPUSD4) as a novel vulnerability in acute myeloid leukemia (AML) via directly catalyzing mitochondrial RNA (mt-RNA) Ψ formation. We further provide proof-of-concept evidence that targeting RPUSD4 suppresses leukemogenesis and stimulates anti-leukemia immunity. To investigate whether RNA Ψ dysregulation is involved in leukemia, we first compared RNA Ψ levels between primitive CD34+ cells (LSC-enriched) and CD34- bulk cells from multiple primary AML specimens. CD34+ AML cells exhibited significantly elevated RNA Ψ levels. We also examined Ψ levels across various RNA types in AML cells, identifying that mt-RNA contained much higher Ψ levels than other RNA types. These findings imply that RNA Ψ, particularly mt-RNA Ψ, may play a role in LSC biology and leukemogenesis. Further unbiased CRISPR screening revealed RPUSD4 as the most critical Ψ synthase for LSC maintenance. Large-scale transcriptomic and patient data analyses showed that RPUSD4 is highly expressed in AML specimens (P = 3.5 × 10-9), particularly within LSCs (P = 1.4 × 10-18). RPUSD4 knockout (KO) significantly suppressed AML proliferation, reduced mitochondrial OxPhos, and decreased LSC frequency in vitro and substantially delayed AML progression in vivo (MLL-r and FLT3ITD models). Of note, RPUSD4 KO-mediated anti-leukemia effects could be fully rescued by wild-type but not catalytically inactive RPUSD4 both in vitro and in vivo, demonstrating that Ψ synthase activity is essential for its function in leukemia. To profile RPUSD4-dependent Ψ sites, we applied two orthogonal technologies, direct nanopore sequencing and PRAISE sequencing (PMID: 36997645), both capable of detecting Ψ at single-base resolution. These approaches converged on a single RPUSD4-dependent Ψ at position 3,067 on mt-16S rRNA. This Ψ site is essential for maintaining rRNA stability. RPUSD4 KO induced mt-16S rRNA decay, thereby suppressing mitochondrial translation. This led to OxPhos inhibition, growth suppression, and LSC eradication in AML. Mechanistically, RPUSD4 is exclusively localized in mitochondria, where it directly interacts with the mt-RNA exonuclease PNPase. Under physiological conditions, PNPase resides within mitochondria and degrades mt-RNA to prevent accumulation of mitochondrial double-stranded RNA (mt-dsRNA). Upon RPUSD4 depletion, PNPase translocates from mitochondria to the cytosol, resulting in two major consequences: (i) mRNA decay:Cytosolic PNPase degrades mRNAs, particularly the nuclear-encoded mitochondrial mRNAs, such as DHODH(Dihydroorotate dehydrogenase). By retrogradely upregulating DHODH, RPUSD4 promotes de novo pyrimidine synthesis in AML. (ii) Aberrant mt-dsRNA accumulation: The buildup and cytoplasmic release of mt-dsRNA activates “viral mimicry” response that sensitizes AML cells to T cell cytotoxicity. Finally, to therapeutically target RPUSD4, we developed an RNA-based inhibitor, CpG-siRNARPUSD4, by conjugating a specific RPUSD4 siRNA to a Toll-like receptor (TLR9) ligand. CpG-siRNARPUSD4 is specifically internalized by TLR9+ cells, including AML cells, thereby maximizing selectivity and minimizing toxicity. Preclinical studies demonstrate that pharmacologically targeting RPUSD4 significantly prolonged survival across multiple AML models, including patient-derived xenograft (PDX) models, and rendered AML cells more susceptible to T cell–mediated cytotoxicity. Furthermore, the RPUSD4 inhibitor exhibited a promising synergistic effect with DHODH inhibitor in treating relapsed AML in vivo. For instance, in a representative relapsed PDX model, median survival times (in days) were: Ctrl – 28, RPUSD4 inhibitor – 36, DHODH inhibitor – 39, and combination – over 60. In conclusion, our findings uncover a previously unrecognized role of RPUSD4 and mt-RNA Ψ in leukemic metabolism, anti-leukemia immunity, and LSC maintenance. This study also bridges a critical knowledge gap regarding the role of RNA Ψ in leukemogenesis.

Leveraging plasma concentrations to optimize extracorporeal treatment in acute diquat poisoning: a multi-center retrospective cohort study

Clinical Toxicology · 2025-12-08

INTRODUCTION: Diquat poisoning is common in Asia and the optimal enhanced elimination strategy is unknown. This study aimed to evaluate the clinical value of plasma diquat concentrations in guiding personalized extracorporeal treatment regimens for patients with acute diquat poisoning. METHODS: This multi-center retrospective cohort study included 163 patients with acute diquat poisoning admitted between February 2022 and July 2023. Patients were divided into three groups based on plasma diquat concentrations measured upon presentation to the emergency department: low (<100 μg/L), medium (100-1,000 μg/L), and high (≥1,000 μg/L). The evaluated extracorporeal treatment regimens included hemoperfusion alone and a combination of hemoperfusion with continuous veno-venous hemodiafiltration. Kaplan-Meier survival curves were used to estimate cumulative survival probabilities, with survival probabilities compared using log-rank tests. RESULTS: All 66 patients survived in the low concentration group, regardless of the extracorporeal treatment used. In the medium and high concentration groups, five patients who refused extracorporeal treatment died, whereas 92 patients who received extracorporeal treatment had a case fatality rate of 48.9%. In the high concentration group, patients receiving hemoperfusion combined with continuous veno-venous hemodiafiltration had a case fatality rate of 76.7% and better survival probabilities, compared to hemoperfusion-only patients, which had no survivors. Additionally, among those treated with a combination of hemoperfusion and continuous veno-venous hemodiafiltration, the time interval from the end of hemoperfusion session to the initiation of continuous veno-venous hemodiafiltration was, on average, shorter for survivors than deaths (3.7 h versus 4.7 h). DISCUSSION: This retrospective observational study of diquat poisoned patients highlights the potential for personalized extracorporeal treatment regimens using an initial plasma diquat concentration. Future randomized trials are warranted to evaluate the optimal use of extracorporeal treatments. CONCLUSION: Obtaining plasma diquat concentrations may be of great value for guiding extracorporeal treatment regimens to improve prognosis in patients with acute diquat poisoning.