Haematology Unit, Cancer Research Centre, Institute for Medical Research, National Institutes of Health, Ministry of Health Malaysia, 40170, Shah Alam, Selangor, Malaysia.
Faculty of Science, Institute of Biological Sciences, University of Malaya, Kuala Lumpur, Malaysia.
Genes Genomics. 2022 Aug;44(8):957-966. doi: 10.1007/s13258-022-01272-7. Epub 2022 Jun 11.
Current advances in the molecular biology of multiple myeloma (MM) are not sufficient to fully delineate the genesis and development of this disease.
This study aimed to identify molecular targets underlying MM pathogenesis.
mRNA expression profiling for 29 samples (19 MM samples, 7 MM cell lines and 3 controls) were obtained using microarray. We evaluated the in vitro effects of RAD54L gene silencing on the proliferation, apoptosis and cell cycle distribution in KMS-28BM human MM cells using siRNA approach. Cell proliferation was determined by MTS assay while apoptosis and cell cycle distribution were analysed with flow cytometry. Gene and protein expression was evaluated using RT-qPCR and ELISA, respectively.
Microarray results revealed a total of 5124 differentially expressed genes (DEGs), in which 2696 and 2428 genes were up-regulated and down-regulated in MM compared to the normal controls, respectively (fold change ≥ 2.0; P < 0.05). Up-regulated genes (RAD54L, DIAPH3, SHCBP1, SKA3 and ANLN) and down-regulated genes (HKDC1, RASGRF2, CYSLTR2) have never been reported in association with MM. Up-regulation of RAD54L was further verified by RT-qPCR (P < 0.001). In vitro functional studies revealed that RAD54L gene silencing significantly induced growth inhibition, apoptosis (small changes) and cell cycle arrest in G0/G1 phase in KMS-28BM (P < 0.05). Silencing of RAD54L also decreased its protein level (P < 0.05).
This study has identified possible molecular targets underlying the pathogenesis of MM. For the first time, we reveal RAD54L as a potential therapeutic target in MM, possibly functioning in the cell cycle and checkpoint control.
多发性骨髓瘤（MM）的分子生物学进展尚不足以充分描绘该病的发生和发展。
本研究旨在确定 MM 发病机制的分子靶标。
使用微阵列获得 29 个样本（19 个 MM 样本、7 个 MM 细胞系和 3 个对照）的 mRNA 表达谱。我们使用 siRNA 方法评估 RAD54L 基因沉默对 KMS-28BM 人 MM 细胞增殖、凋亡和细胞周期分布的体外影响。通过 MTS 测定法测定细胞增殖，通过流式细胞术分析凋亡和细胞周期分布。使用 RT-qPCR 和 ELISA 分别评估基因和蛋白表达。
微阵列结果显示，与正常对照相比，MM 中共有 5124 个差异表达基因（DEGs），其中 2696 个和 2428 个基因上调和下调（倍数变化≥2.0；P<0.05）。上调基因（RAD54L、DIAPH3、SHCBP1、SKA3 和 ANLN）和下调基因（HKDC1、RASGRF2、CYSLTR2）从未与 MM 相关报道过。RAD54L 的上调进一步通过 RT-qPCR 验证（P<0.001）。体外功能研究表明，RAD54L 基因沉默可显著诱导 KMS-28BM 生长抑制、凋亡（小变化）和 G0/G1 期细胞周期停滞（P<0.05）。RAD54L 沉默还降低了其蛋白水平（P<0.05）。
本研究确定了 MM 发病机制的潜在分子靶标。我们首次揭示 RAD54L 可能是 MM 的潜在治疗靶点，可能在细胞周期和检查点控制中发挥作用。
