m(6)A-dependent upregulation of DDX21 by super-enhancer-driven IGF2BP2 and IGF2BP3 facilitates progression of acute myeloid leukaemia.

Acute myeloid leukaemia (AML) is a haematological malignancy with unfavourable prognosis. Despite the effectiveness of chemotherapy and targeted therapy, relapse or drug resistance remains a major threat to AML patients. N6-methyladenosine (m6A) RNA methylation and super-enhancers (SEs) are extensively involved in the leukaemogenesis of AML. However, the potential relationship between m6A and SEs in AML has not been elaborated. Chromatin immunoprecipitation (ChIP) sequencing data from Gene Expression Omnibus (GEO) cohort were analysed to search SE-related genes. The mechanisms of m6 A-binding proteins IGF2BP2 and IGF2BP3 on DDX21 were explored via methylated RNA immunoprecipitation (MeRIP) assays, RNA immunoprecipitation (RIP) assays and luciferase reporter assays. Then we elucidated the roles of DDX21 in AML through functional assays in vitro and in vivo. Finally, co-immunoprecipitation (Co-IP) assays, RNA sequencing and ChIP assays were performed to investigate the downstream mechanisms of DDX21. We identified two SE-associated transcripts IGF2BP2 and IGF2BP3 in AML. High enrichment of H3K27ac, H3K4me1 and BRD4 was observed in IGF2BP2 and IGF2BP3, whose expression were driven by SE machinery. Then IGF2BP2 and IGF2BP3 enhanced the stability of DDX21 mRNA in an m6A-dependent manner. DDX21 was highly expressed in AML patients, which indicated a poor survival. Functionally, knockdown of DDX21 inhibited cell proliferation, promoted cell apoptosis and led to cell cycle arrest. Mechanistically, DDX21 recruited transcription factor YBX1 to cooperatively trigger ULK1 expression. Moreover, silencing of ULK1 could reverse the promoting effects of DDX21 overexpression in AML cells. Dysregulation of SE-IGF2BP2/IGF2BP3-DDX21 axis facilitated the progression of AML. Our findings provide new insights into the link between SEs and m6A modification, elucidate the regulatory mechanisms of IGF2BP2 and IGF2BP3 on DDX21, and reveal the underlying roles of DDX21 in AML.

Zhao Y ，Zhou Y ，Qian Y ，Wei W ，Lin X ，Mao S ，Sun J ，Jin J ... -  《Clinical and Translational Medicine》

KLF7 regulates super-enhancer-driven IGF2BP2 overexpression to promote the progression of head and neck squamous cell carcinoma.

Head and neck squamous carcinoma (HNSCC) is known for its high aggressiveness and susceptibility to cervical lymph node metastasis, which greatly contributes to its poor prognosis. During tumorigenesis, many types of cancer cells acquire oncogenic super-enhancers (SEs) that drive the overexpression of oncogenes, thereby maintaining malignant progression. This study aimed to identify and validate the role of oncogenic SE-associated genes in the malignant progression of HNSCC. We identified HNSCC cell-specific SE-associated genes through H3K27Ac ChIP-seq and overlapped them with HNSCC-associated genes obtained from The Cancer Genome Atlas (TCGA) dataset and Gene Expression Omnibus (GEO) datasets using weighted gene coexpression network analysis (WGCNA) to identify hub genes. The expression of IGF2BP2 and KLF7 in HNSCC was detected using clinical samples. To determine the biological role of IGF2BP2, we performed CCK-8, colony formation assay, Transwell migration assay, invasion assay, and orthotopic xenograft model experiments. Furthermore, we utilized a CRISPR/Cas9 gene-editing system, small-molecule inhibitors, ChIP-qPCR, and dual-luciferase reporter assays to investigate the molecular mechanisms of IGF2BP2 and its upstream transcription factors. Our study identified IGF2BP2 as a hub SE-associated gene that exhibited aberrant expression in HNSCC tissues. Increased expression of IGF2BP2 was observed to be linked with malignant progression and unfavorable prognosis in HNSCC patients. Both in vitro and in vivo experiments confirmed that IGF2BP2 promotes the tumorigenicity and metastasis of HNSCC by promoting cell proliferation, migration, and invasion. Mechanistically, the IGF2BP2-SE region displayed enrichment for H3K27Ac, BRD4, and MED1, which led to the inhibition of IGF2BP2 transcription and expression through deactivation of the SE-associated transcriptional program. Additionally, KLF7 was found to induce the transcription of IGF2BP2 and directly bind to its promoter and SE regions. Moreover, the abundance of KLF7 exhibited a positive correlation with the abundance of IGF2BP2 in HNSCC. Patients with high expression of both KLF7 and IGF2BP2 showed poorer prognosis. Lastly, we demonstrated that the small molecule inhibitor JQ1, targeting BRD4, attenuated the proliferation and metastatic abilities of HNSCC cells. Our study reveals the critical role of IGF2BP2 overexpression mediated by SE and KLF7 in promoting HNSCC progression. Targeting SE-associated transcriptional programs may represent a potential therapeutic strategy in managing HNSCC.

Cai H ，Liang J ，Jiang Y ，Wang Z ，Li H ，Wang W ，Wang C ，Hou J ... -  《JOURNAL OF EXPERIMENTAL & CLINICAL CANCER RESEARCH》

N6-methyladenosine-modified SENP1, identified by IGF2BP3, is a novel molecular marker in acute myeloid leukemia and aggravates progression by activating AKT signal via de-SUMOylating HDAC2.

Elevated evidence suggests that the SENPs family plays an important role in tumor progression. However, the role of SENPs in AML remains unclear. We evaluated the expression pattern of SENP1 based on RNA sequencing data obtained from OHSU, TCGA, TARGET, and MILE datasets. Clinical samples were used to verify the expression of SENP1 in the AML cells. Lentiviral vectors shRNA and sgRNA were used to intervene in SENP1 expression in AML cells, and the effects of SENP1 on AML proliferation and anti-apoptosis were detected using in vitro and in vivo models. Chip-qPCR, MERIP-qPCR, CO-IP, RNA pulldown, and dual-luciferase reporter gene assays were used to explore the regulatory mechanisms of SNEP1 in AML. SENP1 was significantly upregulated in high-risk AML patients and closely related to poor prognosis. The AKT/mTOR signaling pathway is a key downstream pathway that mediates SENP1's regulation of AML proliferation and anti-apoptosis. Mechanistically, the CO-IP assay revealed binding between SENP1 and HDAC2. SUMO and Chip-qPCR assays suggested that SENP1 can desumoylate HDAC2, which enhances EGFR transcription and activates the AKT pathway. In addition, we found that IGF2BP3 expression was upregulated in high-risk AML patients and was positively correlated with SENP1 expression. MERIP-qPCR and RIP-qPCR showed that IGF2BP3 binds SENP1 3-UTR in an m6A manner, enhances SENP1 expression, and promotes AKT pathway conduction. Our findings reveal a distinct mechanism of SENP1-mediated HDAC2-AKT activation and establish the critical role of the IGF2BP3/SENP1signaling axis in AML development.

Wen D ，Xiao H ，Gao Y ，Zeng H ，Deng J ... -  《Molecular Cancer》

LINC00987 knockdown inhibits the progression of acute myeloid leukemia by suppressing IGF2BP2-mediated PA2G4 expression.

This study aimed to investigate the role and potential mechanisms of LINC00987 in acute myeloid leukemia (AML) progression. The expression of LINC00987 in bone marrow specimens of AML patients and cell lines was measured by quantitative reverse transcription PCR (RT-qPCR). Small interfering RNA targeting LINC00987 (si-LINC00987) was transfected into AML cell lines HL-60 and KG-1, and the proliferation, invasion and apoptosis were detected with Cell Counting Kit-8 (CCK-8), Transwell and flow cytometry, respectively. Moreover, the binding between LINC00987 and insulin like growth factor 2 mRNA binding protein 2 (IGF2BP2) was validated with an RNA pull-down assay. Co-immunoprecipitation assay was used to verify the binding between IGF2BP2 and proliferation-associated 2G4 (PA2G4). Then rescue experiments were performed to explore the effects of LINC00987/IGF2BP2/PA2G4 axis on HL-60 and KG-1 cell functions. Additionally, HL-60 cells transfected with si-LINC00987 were injected into mice, followed by the evaluation of xenograft tumor growth. LINC00987 was upregulated in AML patient specimens and cell lines. LINC00987 knockdown inhibited proliferation and invasion and promoted apoptosis in AML cells. LINC00987 could bind with IGF2BP2 and promote its expression, and IGF2BP2 overexpression reversed the effects of LINC00987 knockdown on the proliferation, invasion and apoptosis in AML cells. Besides, IGF2BP2 could bind with PA2G4. IGF2BP2 knockdown inhibited proliferation and invasion, and promoted apoptosis in AML cells, whereas PA2G4 overexpression reversed these effects. Additionally, the LINC00987 knockdown inhibited the xenograft tumor growth of AML in vivo. Knockdown of LINC00987 inhibits AML cell proliferation and invasion, and promotes apoptosis in vitro and reduces tumor growth in vivo by suppressing IGF2BP2-mediated PA2G4 expression.

Liu C ，Ma Y ，Wang R ，Su G ... -  《-》

The m6A reader IGF2BP3 promotes acute myeloid leukemia progression by enhancing RCC2 stability.

N6-methyladenosine (m6A) is the most abundant posttranscriptional modification of mRNA in eukaryotes. Recent evidence suggests that dysregulated m6A-associated proteins and m6A modifications play a pivotal role in the initiation and progression of diseases such as cancer. Here, we identified that IGF2BP3 is specifically overexpressed in acute myeloid leukemia (AML), a subtype of leukemia associated with poor prognosis and high genetic risk. IGF2BP3 is required for maintaining AML cell survival in an m6A-dependent manner, and knockdown of IGF2BP3 dramatically suppresses the apoptosis, reduces the proliferation, and impairs the leukemic capacity of AML cells in vitro and in vivo. Mechanistically, IGF2BP3 interacts with RCC2 mRNA and stabilizes the expression of m6A-modified RNA. Thus, we provided compelling evidence demonstrating that the m6A reader IGF2BP3 contributes to tumorigenesis and poor prognosis in AML and can serve as a target for the development of cancer therapeutics.

Zhang N ，Shen Y ，Li H ，Chen Y ，Zhang P ，Lou S ，Deng J ... -  《-》