Targeting Nonsense-Mediated RNA Decay in Splicing Factor Mutant Myeloid Malignancies.

靶向剪接因子突变骨髓恶性肿瘤中无义介导的 RNA 衰变。

• 批准号: 10751386
• 负责人: Claudia Cabrera
• 金额: $ 3.36万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10751386
• 关键词: Acute Myelocytic Leukemia; Alternative Splicing; Apoptosis; Biological Assay; Biological Markers; Biological Models; Bone Marrow; Cancer cell line; Cell Death; Cell Line; Cell Survival; Cells; Chronic Myelomonocytic Leukemia; Clinical; Clonal Hematopoietic Stem Cell; Combined Modality Therapy; Congenic Mice; DNA; DNA Damage; DNA Repair; Data; Development; Disease; Dose; Drug Combinations; Dysmyelopoietic Syndromes; Dysplasia; Engraftment; Future; Gene Mutation; Genes; Genetically Engineered Mouse; Genomic Instability; Goals; Hematologic Neoplasms; Heterozygote; Hybrids; Hypersensitivity; In Vitro; Ineffective Hematopoiesis; Lead; Malignant Neoplasms; Mammals; Measures; Mediating; Messenger RNA; Modeling; Molecular; Molecular Target; Mus; Mutant Strains Mice; Mutation; Myeloproliferative disease; Pathway interactions; Patients; Pattern; Pharmaceutical Preparations; Pharmacodynamics; Pharmacological Treatment; Phosphorylation; Poly(ADP-ribose) Polymerase Inhibitor; Prevalence; Process; Production; Proliferating; Protein Isoforms; Protein Kinase; Proteins; RNA; RNA Decay; RNA Helicase; RNA Splicing; Recurrence; Reporting; Resolution; Role; SRSF2 gene; Sampling; Single-Stranded DNA; Spliceosomes; Structure; Terminator Codon; Testing; Therapeutic; Transcript; Translating; acute myeloid leukemia cell; attenuation; cancer cell; cancer therapy; candidate identification; cell type; cytopenia; efficacy study; experimental study; gene repair; genetically modified cells; in vivo; in vivo Model; inhibitor; innovation; mutant; novel; novel strategies; novel therapeutic intervention; peripheral blood; pharmacologic; pre-clinical; premature; resistance mechanism; side effect; targeted treatment; therapeutic evaluation; transcriptome sequencing

Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematopoietic stem cell disorders characterized by peripheral blood cytopenias, bone marrow dysplasia, and ineffective hematopoiesis. Approximately 50% of MDS, 60% chronic myelomonocytic leukemia (CMML), 20% of acute myeloid leukemia (AML) harbor heterozygous mutations in spliceosome factor genes such as SF3B1, U2AF1, SRSF2 and ZRSR2. Although many studies have shown that mutations in splicing factor genes lead to distinct patterns of aberrant splicing, no specific alternatively spliced isoform has been demonstrated to directly cause MDS. However, aberrations in splicing induced by splicing factor gene mutations create a vulnerability in MDS cells. Our group and others showed that cells harboring spliceosome gene mutations have increased sensitivity to pharmacological perturbation of the spliceosome by splicing modulator drugs. The sensitivity of spliceosome mutant cells to further splicing perturbations raises the possibility that they are vulnerable to accumulation of misspliced transcripts. A large portion of the misspliced RNAs caused by spliceosome mutations or splicing modulator treatment are nonsense mRNAs that harbor premature termination codons (PTCs). These nonsense mRNAs, which may cause deleterious effects if translated, are normally degraded by a RNA surveillance pathway called nonsense-mediated RNA decay (NMD). The prevalence of nonsense mRNAs in cancer cells with spliceosome mutations leads us to hypothesize that mutant cells will be more sensitive to NMD attenuation due to the role of NMD in the clearance of nonsense mRNAs that can be detrimental. Preliminary data from our group indicate that NMD disruption (using a SMG1 inhibitor [SMG1i]) preferentially kills cancer cells expressing different splicing factor gene mutations. This cell death is associated with the induction of R-loops and DNA damage. Building on preliminary studies, we propose to test the therapeutic potential of NMD inhibition in selective killing of spliceosome mutant cancer cells using in vivo models and define the underlying mechanism for the hypersensitivity of spliceosome mutant cells to NMD attenuation. In Specific Aim 1, we will test the therapeutic potential of NMD inhibition to selectively kill spliceosome mutant cancer cells using in vivo models. We will engraft primary mouse AML cells in congenic mice and test whether in vivo treatment with a highly specific inhibitor of SMG1 (SMG1i), the only known protein kinase that regulates the NMD pathway, can selectively kill cancer cells with spliceosome mutations. We will further establish the therapeutic potential of targeting NMD by combining SMG1i with ATR or PARP inhibitors, DNA damage repair proteins that are activated in splicing factor mutant cells. In Specific Aim 2, we will define the molecular mechanism for the sensitivity of spliceosome mutant cells to NMD attenuation. We will identify candidate NMD targets whose levels are modulated by SMG1i or mutant splicing factors, potentially nominating downstream functional targets that could be modulated for cancer treatment. Collectively, this project will nominate therapies to treat MDS and AML with splicing factor mutations.

骨髓增生异常综合征(MDS)是一组克隆性造血干细胞疾病 以外周血细胞减少、骨髓发育不良和无效的造血为特征。 大约50%的MDS，60%的慢性单核细胞白血病(CMML)，20%的急性髓系白血病 急性髓系白血病(AML)存在SF3B1、U2AF1、SRSF2和ZRSR2等剪接体因子基因杂合性突变。 尽管许多研究表明，剪接因子基因的突变会导致不同的异常模式 剪接，没有特定的选择性剪接异构体被证明直接导致MDS。然而， 剪接因子基因突变引起的剪接异常在MDS细胞中造成了脆弱性。我们的团队 另一些研究表明，含有剪接体基因突变的细胞对 剪接调节剂药物对剪接体的药理干扰。剪接体的敏感性 突变细胞对进一步剪接的扰动增加了它们容易受到 拼接错误的文字记录。剪接体突变或剪接引起的大部分错误剪接的RNA 调节剂治疗是含有提前终止密码子(PTC)的无意义的mRNAs。这些胡说八道 翻译后可能造成有害影响的mrna通常会被rna监控降解。 途径称为无义介导的RNA衰变(NMD)。癌症细胞中无意义的mRNAs的流行 剪接体突变使我们假设，由于NMD的衰减，突变细胞将更加敏感 NMD在清除可能有害的无稽之谈的mRNAs方面的作用。来自我们小组的初步数据 表明破坏NMD(使用SMG1抑制剂[SMG1i])优先杀死表达不同基因的癌细胞 剪接因子基因突变。这种细胞死亡与R环和DNA损伤的诱导有关。 在初步研究的基础上，我们建议测试NMD抑制在选择性杀伤中的治疗潜力 利用体内模型对剪接体突变的癌细胞进行研究，并确定其潜在机制 剪接体突变细胞对NMD衰减的超敏反应。在具体目标1中，我们将测试治疗方法 利用体内模型研究抑制NMD选择性杀死剪接体突变癌细胞的可能性。我们将嫁接 原代培养小鼠急性髓系白血病细胞，并检测体内是否有高度特异的抑制因子 SMG1(SMG1i)是已知的唯一调节NMD途径的蛋白激酶，可以选择性地杀死癌细胞 剪接体突变。我们将通过联合应用进一步确定靶向NMD的治疗潜力 带有ATR或PARP抑制剂的SMG1i，剪接因子突变体中激活的DNA损伤修复蛋白 细胞。在特定目标2中，我们将确定剪接体突变细胞敏感性的分子机制。 到NMD衰减。我们将确定其水平受SMG1i或突变体调控的候选NMD靶点 剪接因子，潜在地提名可能为癌症调节的下游功能靶点 治疗。总体而言，该项目将提名治疗带有剪接因子突变的MDS和AML的疗法。