第一部分摘要
背景：多发性骨髓瘤（MM）是一种以浆细胞恶性增殖为特征的血液肿瘤，过去20年，由于新药新技术不断研发和应用，MM患者总生存期（OS）已从3～5年延长至8～10年，但临床特征和预后仍有极大异质性，OS从几个月到十几年不等。新诊断多发性骨髓瘤（NDMM）的危险分层体系不断更新，如何定义高危患者群体仍有争议。Myc易位（Myc-R）在多个研究中被报道与NDMM的不良预后相关，但缺乏深入研究。研究目的：为探索Myc-R以及Myc其他异常对NDMM预后的影响，进一步细化危险分层体系。研究方法：本研究纳入了2009年5月至2022年9月期间在我院就诊的417名NDMM患者，使用荧光原位杂交（FISH）检测Myc异常情况，比较Myc不同异常患者之间的基线特征、一线治疗、疗效和预后差异。结果：13.7％的NDMM患者治疗前存在Myc-R，14.6％的患者检出Myc其他异常（Myc-OA）。Myc-R、Myc-OA和Myc阴性（Myc-N）患者的一线治疗中位无进展生存期（PFS）分别为15.9个月（m）、24.5m和29.8m（p=0.003）。同时，Myc-R组的中位总生存期（OS）较差（仅为25.1个月，p<0.001）。结果表明，Myc-R而不是Myc-OA是影响NDMM患者PFS和OS的独立危险因素。相较于单个传统的高危细胞遗传学异常（HRCA）患者，仅有Myc-R患者的中位PFS和OS更短（15.9m vs 28.1m，p=0.032；25.1m vs 61.2m，p=0.04）。将Myc-R或≥2个HRCAs定义为新的高危因素时，在新的危险分层模型中，高危和标危患者的中位PFS和OS分别为16.4m vs 29.8m（p=0.001）和29.3m vs 66.7m（p＜0.001）。结论：在新药治疗时代，传统的单个HRCA对NDMM患者预后的影响有所减弱。相反，Myc易位或多打击HRCAs才是不良预后的关键因素。这一发现有助于我们更准确地评估NDMM患者的治疗风险，并为更新高危预测模型提供新的视角。
第二部分摘要
背景：Myc是一种重要的原癌基因，在多个肿瘤中表达增高。我们的前期研究发现，Myc易位与NDMM不良预后密切相关，是重要的高危因素，同时Myc或通过调控多个基因如MCL1、MUC1、PYK2的表达影响骨髓瘤细胞的增殖。YM-155是一种新型小分子药物，可靶向Myc蛋白的降解，作为Myc抑制剂在临床推广，有文献报道了YM-155的抗骨髓瘤作用，但具体机制不知。研究目的：1）进一步探索YM-155是否影响MM细胞的增殖和凋亡；2)YM-155是否靶向Myc发挥抗骨髓瘤的作用及其作用机制。3）YM-155对硼替佐米耐药的MM细胞的作用。方法：YM155处理AMO-1、MM1S、RPMI8226、U266、H929骨髓瘤细胞系后CCK8检测细胞活性，流式细胞术检测凋亡细胞比例，RNAseq和生物信息技术分析调控通路，并应用qPCR和Western blot（WB）在mRNA和蛋白层面验证。结果：YM155在多个MM细胞系的IC50值为2.5-15nM，其抑制细胞增殖及促凋亡的作用呈时间和浓度依赖性，并显著下调Myc mRNA表达和蛋白水平；YM155联合硼替佐米对MM细胞的生长抑制、诱导细胞凋亡效果明显优于单药，提示二者具有协同作用。RANseq结果提示YM155处理后的细胞株Myc表达下降，P53信号通路被激活，经RT-qPCR和WB验证，其中促凋亡相关蛋白BBC3被激活，抗凋亡蛋白BCL2被抑制。JASPAR生信分析预测Myc或通过结合在BBC3的启动子区域抑制BBC3的表达，从而激活P53通路。此外，YM155在硼替佐米耐药株的IC50与非耐药株相似。结论：YM155在体外的抗骨髓瘤活性具有浓度和时间依赖性，与硼替佐米具有协同抗肿瘤及一定程度克服耐药作用。YM155通过抑制Myc上调BBC3的表达，激活P53通路发挥抗骨髓瘤作用，这为高危/难治复发MM的治疗提供了潜在依据。

Part I

Background: Multiple myeloma (MM) is a hematologic malignancy characterized by malignant proliferation of plasma cells, with prognosis varying from months to over a decade. In the era of new drugs, the prognosis of MM patients has greatly improved, and the risk stratification system for newly diagnosed multiple myeloma (NDMM) is constantly updated, with ongoing controversy regarding the definition of high-risk patients. Myc rearrangement (Myc-R) has been reported in several studies to be associated with adverse outcomes in NDMM, but there is a lack of more detailed research.Purpose: We conducted this study to investigate the impact of Myc-R on the prognosis of NDMM and to further refine the risk stratification system.Methods: A total of 417 patients were enrolled from May 2009 to September 2022. Fluorescence in situ hybridization (FISH) was used to detect abnormalities in Myc, and baseline characteristics, first-line treatments, therapeutic outcomes, and prognostic differences among different groups of Myc abnormalities were compared.Results: Patients were divided into three groups: the Myc rearrangement (Myc-R) group, the Myc-negative (Myc-N) group, and the Myc other abnormalities (Myc-OA) group. 13.7% of patients developed Myc-R and 14.6% had Myc abnormalities (Myc-OA). The median progression-free survival (PFS) for patients with Myc-R, Myc-OA and Myc-negative (Myc-N) were 15.9 months (m), 24.5m and 29.8m, respectively (p=0.003). Similarly, the median overall survival (OS) in the Myc-R group was inferior (only 25.1 months, p＜0.001). Myc-R was identified as an independent risk factor for both PFS and OS rather than Myc-OA. Patients with Myc-R alone demonstrated shorter median PFS (15.9m vs 28.1m, p=0.032) and OS (25.1m vs 61.2m, p=0.04) compared to those with traditional single HRCA. When Myc-R or ≥2 HRCAs were defined as high-risk factors in a new risk stratification model, the median PFS and OS for new high-risk and standard-risk populations were 16.4m vs 29.8m (p＜0.001) and 29.3m vs 66.7m (p＜0.001), respectively.Conclusion: Thus, it appears that traditional single HRCA may not have a substantial influence on survival. The incorporation of Myc rearrangement or traditional double/triple-hit HRCAs could potentially enhance the risk stratification model of NDMM, offering a new perspective on high-risk factors.

Part II

Background: Myc is a crucial oncogene frequently overexpressed in various tumors. Our prior studies have revealed a close association between Myc rearrangement (Myc-R) and poor prognosis in newly diagnosed multiple myeloma (NDMM), representing a significant high-risk factor. Furthermore, we have observed that Myc influences myeloma cell proliferation by regulating the expression of several genes such as MCL1, MUC1, and PYK2. YM-155 is a novel small molecule drug designed to degrade the Myc protein, considered one of the Myc inhibitors. Several studies have reported the anti-myeloma effects of YM-155, but its specific mechanism remains unknown.Objectives: 1) To further investigate the impact of YM-155 on the proliferation and apoptosis of MM cells; 2) To explore whether YM-155 targets Myc to exert antitumor effects in MM and its underlying mechanism; 3) To examine the effect of YM-155 on MM cells resistant to bortezomib.Methods: In vitro, six MM cell lines (AMO-1, MM.1S, RPMI-8226, NCI-H929, U266, and KMS-11) were used to determine the IC50 concentrations of each cell line using the CCK8 method. Cells were treated with gradient concentrations of YM-155 for 24, 48, and 72 hours to assess proliferation and apoptosis using flow cytometry. RNA- sequencing was performed on MM.1S and RPMI-8226 cells treated with the IC50 concentration of YM-155 to identify relevant regulatory pathways. Protein-protein interaction and transcription factor target prediction analyses were conducted to predict interactions between proteins and the binding sites of Myc and target genes. The identified regulatory pathways were validated at the mRNA and protein levels.Results: YM-155 demonstrated potent anti-myeloma activity in vitro, with low IC50 values (2.5-15nM) across multiple cell lines. Its ability to inhibit cell proliferation and induce apoptosis was concentration- and time-dependent. When combined with bortezomib, YM-155 exhibited significantly greater inhibition of cell growth and induction of apoptosis compared to either agent alone, indicating increased sensitivity to bortezomib. RNAseq analysis revealed activation of the P53 signaling pathway in cells treated with YM-155. Validation experiments confirmed the activation of the pro-apoptotic protein BBC3 and inhibition of the anti-apoptotic protein BCL2 in the P53 signaling pathway. JASPAR bioinformatics analysis predicted that Myc inhibits BBC3 expression by binding to the promoter region, thereby activating the P53 pathway. Additionally, YM-155 showed similar cytotoxicity in the myeloma cell line KMS11 and bortezomib-resistant strains.Conclusion: YM-155 exhibited concentration- and time-dependent cytotoxicity in vitro. It increased the sensitivity of MM cells to bortezomib and overcame bortezomib resistance. The potential mechanism of YM-155's anti-myeloma effect involves inhibiting Myc-mediated upregulation of BBC3 expression, thereby activating the P53 pathway. Our findings suggest that YM-155 may be a promising therapeutic target for high-risk/recurrent MM with Myc overexpression. These results provide a foundation for developing new treatment strategies, particularly for high-risk patients with high Myc expression and bortezomib-resistant recurrent patients.