Proteomics-Based Approach Reveals the Involvement of SERPINB9 in Recurrent and Relapsed Multiple Myeloma

Multiple
myeloma (MM) is a common hematological malignancy with
poorly understood recurrence and relapse mechanisms. Notably, bortezomib
resistance leading to relapse makes MM treatment significantly challenging.
To clarify the drug resistance mechanism, we employed a quantitative
proteomics approach to identify differentially expressed protein candidates
implicated in bortezomib-resistant recurrent and relapsed MM (RRMM).
Bone marrow aspirates from five patients newly diagnosed with MM (NDMM)
were compared with those from five patients diagnosed with bortezomib-resistant
RRMM using tandem mass tag-mass spectrometry (TMT-MS). Subcellular
localization and functional classification of the differentially expressed
proteins were determined by gene ontology, Kyoto Encyclopedia of Genes
and Genomes pathway, and hierarchical clustering analyses. The top
candidates identified were validated with parallel reaction monitoring
(PRM) analysis using tissue samples from 11 NDMM and 8 RRMM patients,
followed by comparison with the NCBI Gene Expression Omnibus (GEO)
dataset of 10 MM patients and 10 healthy controls (accession no.:
GSE80608). Thirty-four differentially expressed proteins in RRMM,
including proteinase inhibitor 9 (SERPINB9), were identified by TMT-MS.
Subsequent functional enrichment analyses of the identified protein
candidates indicated their involvement in regulating cellular metabolism,
apoptosis, programmed cell death, lymphocyte-mediated immunity, and
defense response pathways in RRMM. The top protein candidate SERPINB9
was confirmed by PRM analysis and western blotting as well as by comparison
with an NCBI GEO dataset. We elucidated the proteome landscape of
bortezomib-resistant RRMM and identified SERPINB9 as a promising novel
therapeutic target. Our results provide a resource for future studies
on the mechanism of RRMM.

多发性骨髓瘤（MM）是一种常见的血液系统恶性肿瘤，其复发和转移的机制尚不明确。特别是，博来霉素耐药性导致的复发使得MM的治疗面临着极大的挑战。为了阐明药物耐药机制，本研究采用定量蛋白质组学方法，旨在识别参与博来霉素耐药性复发和转移性多发性骨髓瘤（RRMM）的差异性表达蛋白候选物。通过串联质谱（TMT-MS）技术，将五例新诊断的多发性骨髓瘤（NDMM）患者的骨髓穿刺液与五例博来霉素耐药性RRMM患者的骨髓穿刺液进行比较。通过基因本体、京都基因与基因组百科全书通路以及层次聚类分析，确定了差异表达蛋白的亚细胞定位和功能分类。通过平行反应监测（PRM）分析，利用11例NDMM和8例RRMM患者的组织样本进行验证，并与NCBI基因表达综合数据库（GEO）中10例MM患者和10例健康对照者的数据集（登录号：GSE80608）进行比较。串联质谱技术鉴定出34种在RRMM中差异表达的蛋白，包括蛋白酶抑制剂9（SERPINB9）。随后，对所识别的蛋白候选物进行功能富集分析，发现其在调节RRMM中的细胞代谢、细胞凋亡、程序性细胞死亡、淋巴细胞介导的免疫和防御反应通路中发挥作用。其中，最关键的蛋白候选物SERPINB9经PRM分析和蛋白质印迹法以及与NCBI GEO数据集的比较得到证实。本研究揭示了博来霉素耐药性RRMM的蛋白质组图谱，并将SERPINB9鉴定为一种有潜力的新型治疗靶点。我们的研究结果为未来关于RRMM机制的研究提供了宝贵资源。