Mechanisms and targeting of aberrant Gas activation in myeloid neoplasms

骨髓肿瘤中异常气体​​激活的机制和靶向

• 批准号: 10659809
• 负责人: Eirini Papapetrou
• 金额: $ 67.09万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-01 至 2028-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10659809
• 关键词: Acute Myelocytic Leukemia; Address; Apoptosis; Biochemical; Biochemical Genetics; Biological Assay; Bone Marrow; CRISPR/Cas technology; Cell physiology; Cells; Clinical; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; Cyclic Peptides; Data; Dependence; Development; Disease; Dysmyelopoietic Syndromes; FLT3 gene; Frequencies; Future; G-Protein-Coupled Receptors; GNAS gene; GTP-Binding Protein alpha Subunits, Gs; GTP-Binding Proteins; Gases; Gene Mutation; Genes; Genetic; Genetic Transcription; Goals; Hematopoietic; Hematopoietic stem cells; Human; In Vitro; Inflammatory; Knock-out; Malignant Neoplasms; Mediating; Mutate; Mutation; Myeloproliferative disease; Oncogenic; Patients; Pharmaceutical Preparations; Phase; Phenotype; Population Genetics; Process; Production; Prognosis; Protein Array; Protein Isoforms; RNA Binding; RNA Splicing; RNA-Binding Proteins; Role; SRSF2 gene; Sampling; Secondary acute myeloid leukemia; Secondary to; Signal Pathway; Signal Transduction; Signaling Molecule; Somatic Mutation; Spliced Genes; System; Testing; Therapeutic; Therapeutic Intervention; Tissue Banks; Transcript; Work; Xenograft procedure; acute myeloid leukemia cell; drug development; effective therapy; genetic approach; induced pluripotent stem cell; inhibitor; leukemic transformation; mRNA Precursor; mutant; new therapeutic target; novel; novel therapeutics; pharmacologic; programs; research clinical testing; stem cell model; therapeutic target; transcriptome sequencing; transcriptomics

PROJECT SUMMARY/ABSTRACT Myelodysplastic syndromes (MDS) are myeloid neoplasms with dismal prognosis, frequent progression to acute myeloid leukemia (AML) and no effective treatment. A decade ago, a development with transformative potential for this disease was the discovery that more than half of MDS patients have somatic mutations in genes encoding splicing factors (SFs, i.e. RNA binding proteins that regulate pre-mRNA splicing). While the high frequency and early occurrence during the disease course of SF mutations rendered them promising targets, efforts to therapeutically leverage this through splicing modulator drugs did not show promise in clinical testing and the drug development pipeline for these targets, and for MDS in general, is currently nearly empty. We developed genetically faithful human induced pluripotent stem cell (iPSC) models of SF-mutated MDS using CRISPR gene editing and performed integrative analyses of splicing (RNA-Seq) and RNA binding (eCLIP) to search for mis-spliced transcripts that are direct common targets of two of the main SF mutations (SRSF2 P95L and the U2AF1 S34F). We found that both mutant SRSF2 and mutant U2AF1 cause altered splicing of the gene GNAS, promoting the production of a longer isoform (GNAS-L), which in turn produces a longer form of the alpha subunit of the stimulatory G protein, G⍺s (G⍺s-L). G proteins are key signaling molecules involved in many important signaling pathways and cell functions, including oncogenic processes. Our preliminary data using functional, biochemical and population genetics approaches support a critical role for the long form of G⍺s (G⍺s-L) as an MDS driver and reveal a new mechanism by which SF mutations drive MDS that opens a completely novel and unexplored therapeutic avenue for MDS, AML and other cancers with SF gene mutations. The goal of this proposal is to investigate G⍺s as a therapeutic target for MDS and identify opportunities for therapeutic interventions that inhibit signaling by G⍺s-L. Specifically, we propose to: (1) Evaluate the effects of direct degradation (through a dTAG) or inhibition (by novel cyclic peptides) of G⍺s in splicing factor (SF)-mutated MDS and AML hematopoietic cells (primary and iPSC-derived) using recently developed iPSC-based models of MDS-to-sAML progression and longitudinal bone marrow samples from MDS patients who progressed to AML; (2) Characterize and target cell signaling downstream of Gs-L through candidate and unbiased approaches (Reverse Phase Protein Array, CyTOF, transcriptomics); and (3) Identify the G-protein coupled receptors (GPCRs) involved in Gs-L activation in SF-mutated MDS and AML through focused CRISPR KO screens. This study can establish a novel therapeutic target that may transform the treatment of MDS, AML and possibly other cancers.

项目总结/摘要 骨髓增生异常综合征（MDS）是一种预后不良、经常进展的骨髓肿瘤， 急性髓细胞白血病（AML），没有有效的治疗。十年前，一个具有变革性的发展 这种疾病的潜在可能性是发现超过一半的MDS患者具有体细胞突变， 编码剪接因子（SF，即调节前mRNA剪接的RNA结合蛋白）的基因。而 在疾病过程中SF突变的高频率和早期发生使其具有前景 靶点，通过剪接调节剂药物在治疗上利用这一点的努力在临床上没有显示出希望。 针对这些靶点和一般MDS的测试和药物开发管道目前几乎是空的。 我们开发了遗传上忠实的SF突变的人诱导多能干细胞（iPSC）模型， MDS使用CRISPR基因编辑并进行剪接（RNA-Seq）和RNA结合的整合分析 （eCLIP）来搜索作为两种主要SF突变的直接共同靶点的错误剪接转录物 （SRSF 2 P95 L和U2 AF 1 S34 F）。我们发现突变型SRSF 2和突变型U2 AF 1都能引起改变的 基因GNAS的剪接，促进较长同种型（GNAS-L）的产生，其反过来产生 刺激性G蛋白α亚基的较长形式，G β s（G β s-L）。G蛋白是关键信号 分子参与许多重要的信号通路和细胞功能，包括致癌过程。 我们使用功能，生物化学和群体遗传学方法的初步数据支持关键作用 对于长型G β s（G β s-L）作为MDS驱动因子，并揭示了SF突变驱动MDS的新机制， MDS为MDS、AML和其他癌症开辟了一条全新的、未经探索的治疗途径， SF基因突变。 该提案的目标是研究作为MDS治疗靶点的G β，并确定 抑制G β-L信号传导的治疗干预的机会。具体而言，我们建议：（1） 评价直接降解（通过dTAG）或抑制（通过新型环肽）G β的作用， 剪接因子（SF）突变的MDS和AML造血细胞（原代和iPSC衍生的），最近使用 开发了基于iPSC的MDS至sAML进展模型和MDS纵向骨髓样本 进展为AML的患者;（2）通过以下途径表征和靶向G β-L下游的细胞信号传导： 候选和无偏的方法（反相蛋白质阵列、CyTOF、转录组学）;和（3）鉴定 G蛋白偶联受体（GPCR）参与SF突变的MDS和AML中G蛋白-L的激活， 聚焦CRISPR KO屏幕。 这项研究可以建立一个新的治疗靶点，可能会改变MDS，AML的治疗， 可能是其他癌症。