肿瘤细胞治疗研究
英文原题：Proteomics-based approach reveals the involvement of spliceosomal components SF3B and SerpinB9 in dermatofibrosarcoma protuberans.
Proteomics-based approach reveals the involvement of spliceosomal components SF3B and SerpinB9 in dermatofibrosarcoma protuberans.
SerpinB9 过表达与 DFSP 的纤维肉瘤样特征相关，可能是一个候选预后生物标志物。
隆突性皮肤纤维肉瘤（DFSP）是一种罕见的皮肤软组织肉瘤，容易复发；其纤维肉瘤样转化型（FS-DFSP）预后更差，并具有显著转移潜能。尽管普遍认为染色体17和22区域易位导致COL1A1-PDGFB融合是DFSP的关键遗传特征，但针对伊马替尼（其靶向PDGFB通路）耐药后的挽救治疗尚未确立。本研究对DFSP肿瘤组织及其配对邻近正常皮肤进行定量蛋白质组分析，以发现新的分子机制和潜在治疗靶点；并采用免疫组化、蛋白质印迹、实时定量PCR和细胞活力检测验证结果。 结果：KEGG和基因集富集分析（GSEA）显示，剪接体通路和细胞外基质（ECM）-受体相互作用与DFSP发病机制相关。研究证实，剪接体组分SF3B以及颗粒酶B内源性抑制剂SerpinB9在肿瘤组织中上调；与经典型DFSP相比，FS-DFSP亚型中SerpinB9显著过表达。SF3B1抑制剂Pladienolide-B可显著降低肿瘤细胞活力。siRNA介导的SF3B1和SF3B2敲低以及Pladienolide-B处理均可抑制SERPINB9表达，但不影响SERPINB9前体mRNA水平，提示其受剪接依赖性调控。此外，利用MCP-counter算法定量分析肿瘤微环境细胞，提示NK细胞活化。DFSP肿瘤中CD56（NK细胞表面标志物）、肿瘤相关巨噬细胞（TAM）标志物CD14和CD163，以及与TAM浸润相关的转化生长因子β诱导蛋白（TGFBI）均呈阳性染色。SERPINB9 mRNA表达与NK细胞丰度及FCGR3A（NK细胞标志物）mRNA水平均呈正相关。 结论：SerpinB9过表达与DFSP的纤维肉瘤样特征相关，可能是候选预后生物标志物。SF3B/SERPINB9轴可能构成治疗脆弱点，尤其适用于FS-DFSP。DFSP肿瘤微环境中可见NK细胞和巨噬细胞相关特征，值得进一步开展功能研究。
BACKGROUND: Dermatofibrosarcoma protuberans (DFSP) is a rare cutaneous soft tissue sarcoma which is prone to high recurrence rate, and the fibrosarcomatous transformation of DFSP (FS-DFSP) is associated with a poorer prognosis and significant metastatic potential. Although the translocation of regions of chromosomes 17 and 22 that results in COL1A1-PDGFB fusion is generally acknowledged as a key genetic feature of DFSP, there is no established salvage treatment following resistance to imatinib, which targets the PDGFB pathway. In this study, we performed quantitative proteomics analyses of DFSP tumor tissues and paired adjacent normal skin, to uncover novel molecular mechanisms and potential therapeutic targets. Findings were validated using immunohistochemistry, Western Blot, Real-Time Quantitative PCR and cell viability assay. RESULTS: Pathway enrichment analysis including KEGG and GSEA revealed that spliceosome pathways and ECM-receptor interaction were associated with DFSP pathogenesis. The spliceosomal components SF3B, and SerpinB9, which is the endogenous inhibitor of granzyme B, were validated to be upregulated in tumor tissues. SerpinB9 was significantly overexpressed in the FS-DFSP subtype compared with the classic-type DFSP. Treatment with the SF3B1 inhibitor, Pladienolide-B, significantly reduced tumor cell viability. siRNA-mediated knockdown of SF3B1 and SF3B2, as well as Pladienolide-B treatment, inhibited SERPINB9 expression without affecting pre-SERPINB9 mRNA levels, suggesting splicing-dependent regulation. Additionally, quantitation of the tumor microenvironment cells using the MCP-counter algorithm indicated NK cells activation. The CD56 (NK cell surface marker), biomarkers of tumor-associated macrophage (TAM) such as CD14 and CD163, and transforming growth factor- induced (TGFBI) which is associated with TAM infiltration stained positive in DFSP tumors. SERPINB9 mRNA expression showed a positive correlation with both NK cell abundance and FCGR3A (NK cell marker) mRNA level. CONCLUSIONS: SerpinB9 overexpression is associated with fibrosarcomatous features in DFSP and may represent a candidate prognosis biomarker. The SF3B/SERPINB9 axis is a potential therapeutic vulnerability, particularly in FS-DFSP. NK cell- and macrophage-related signatures were evident in DFSP tumor microenvironment, warranting further functional studies.
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