Department of Medical Biochemistry and Molecular Biology, Faculty of Medicine, Ain Shams University, Cairo, Egypt.
Clinical Pathology-Hematology & AinShams Medical Research Institute (MASRI), Faculty of Medicine, Ain Shams University, Cairo, Egypt.
PLoS One. 2021 Jan 8;16(1):e0245349. doi: 10.1371/journal.pone.0245349. eCollection 2021.
Multiple Myeloma (MM) is a heterogeneous, hematological neoplasm that accounts 2% of all cancers. Although, autologous stem cell transplantation and chemotherapy are currently the most effective therapy, it carries a notable hazards, in addition for being non curative. Recently, the Clustered Regular Interspaced Short Palindromic Repeats (CRISPR-cas9) has been successfully tried at the experimental level, for the treatment of several hematological malignancies.
We aimed to investigate the in-vitro effect of CRISPR-cas9-mediated knock-out of V-set pre B-cell surrogate light chain 1"VPREB1" gene on the malignant proliferation of primary cultured myeloma cells.
Bioinformatics' analysis was performed to explore the gene expression profile of MM, and the VPREB1 gene was selected as a target gene for this study. We knocked-out the VPREB1 gene in primary cultured myeloma cells using CRISPR-cas9, the VPREB1 gene editing efficacy was verified by determining VPREB1 gene expression at both the mRNA and protein levels by qPCR and immunofluorescence, respectively. Furthermore, the cytotoxic effect on primary myeloma cells proliferation was evaluated using cytotoxicity assay.
There was a statistically significant reduction of both VPREB1 mRNA and protein expression levels (p<0.01). knock-out of VPREB1 gene in myeloma cell line resulted in a statistically significant reduction of myeloma cell proliferation.
CRISPR-cas9-mediated knock-out of VPREB1 gene is effective for inhibiting the proliferation of primary myeloma cells. This would provide a basis for a promising therapeutic strategy for patients with multiple myeloma.
多发性骨髓瘤（MM）是一种异质性血液病肿瘤，占所有癌症的 2%。尽管自体干细胞移植和化疗是目前最有效的治疗方法，但它存在显著的风险，并且不能治愈。最近，簇状规则间隔短回文重复（CRISPR-cas9）已在实验水平上成功用于治疗几种血液恶性肿瘤。
我们旨在研究 CRISPR-cas9 介导的 V -set 前 B 细胞替代轻链 1“VPREB1”基因敲除对原代培养骨髓瘤细胞恶性增殖的体外影响。
进行生物信息学分析以探索 MM 的基因表达谱，选择 VPREB1 基因为本研究的靶基因。我们使用 CRISPR-cas9 敲除原代培养骨髓瘤细胞中的 VPREB1 基因，通过 qPCR 和免疫荧光分别确定 VPREB1 基因在 mRNA 和蛋白水平上的表达，从而验证 VPREB1 基因编辑的效果。此外，通过细胞毒性测定评估对原代骨髓瘤细胞增殖的细胞毒性作用。
VPREB1 mRNA 和蛋白表达水平均有统计学显著降低（p<0.01）。敲除骨髓瘤细胞系中的 VPREB1 基因可导致骨髓瘤细胞增殖的统计学显著降低。
CRISPR-cas9 介导的 VPREB1 基因敲除可有效抑制原代骨髓瘤细胞的增殖。这为多发性骨髓瘤患者提供了一种有前途的治疗策略的基础。
