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Abstract：
BACKGROUND: Gastric cancer (GC) is a prevalent malignancy with limited therapeutic options for advanced stages. This study aimed to identify novel therapeutic targets for GC by profiling HSP90 client kinases.
METHODS: We used mass spectrometry-based activity-based protein profiling (ABPP) with a desthiobiotin-ATP probe, combined with sensitivity analysis of HSP90 inhibitors, to profile kinases in a panel of GC cell lines. We identified kinases regulated by HSP90 in inhibitor-sensitive cells and investigated the impact of MASTL knockdown on GC cell behavior. Global proteomic analysis following MASTL knockdown was performed, and bioinformatics tools were used to analyze the resulting data.
RESULTS: Four kinases-MASTL, STK11, CHEK1, and MET-were identified as HSP90-regulated in HSP90 inhibitor-sensitive cells. Among these, microtubule-associated serine/threonine kinase-like (MASTL) was upregulated in GC and associated with poor prognosis. MASTL knockdown decreased migration, invasion, and proliferation of GC cells. Global proteomic profiling following MASTL knockdown revealed NEDD4-1 as a potential downstream mediator of MASTL in GC progression. NEDD4-1 was also upregulated in GC and associated with poor prognosis. Similar to MASTL inhibition, NEDD4-1 knockdown suppressed migration, invasion, and proliferation of GC cells.
CONCLUSIONS: Our multi-proteomic analyses suggest that targeting MASTL could be a promising therapy for advanced gastric cancer, potentially through the reduction of tumor-promoting proteins including NEDD4-1. This study enhances our understanding of kinase signaling pathways in GC and provides new insights for potential treatment strategies.
METHODS: We used mass spectrometry-based activity-based protein profiling (ABPP) with a desthiobiotin-ATP probe, combined with sensitivity analysis of HSP90 inhibitors, to profile kinases in a panel of GC cell lines. We identified kinases regulated by HSP90 in inhibitor-sensitive cells and investigated the impact of MASTL knockdown on GC cell behavior. Global proteomic analysis following MASTL knockdown was performed, and bioinformatics tools were used to analyze the resulting data.
RESULTS: Four kinases-MASTL, STK11, CHEK1, and MET-were identified as HSP90-regulated in HSP90 inhibitor-sensitive cells. Among these, microtubule-associated serine/threonine kinase-like (MASTL) was upregulated in GC and associated with poor prognosis. MASTL knockdown decreased migration, invasion, and proliferation of GC cells. Global proteomic profiling following MASTL knockdown revealed NEDD4-1 as a potential downstream mediator of MASTL in GC progression. NEDD4-1 was also upregulated in GC and associated with poor prognosis. Similar to MASTL inhibition, NEDD4-1 knockdown suppressed migration, invasion, and proliferation of GC cells.
CONCLUSIONS: Our multi-proteomic analyses suggest that targeting MASTL could be a promising therapy for advanced gastric cancer, potentially through the reduction of tumor-promoting proteins including NEDD4-1. This study enhances our understanding of kinase signaling pathways in GC and provides new insights for potential treatment strategies.
摘要：
背景：胃癌（GC）是一种常见的恶性肿瘤，晚期的治疗选择有限。这项研究旨在通过分析HSP90客户激酶来鉴定GC的新治疗靶标。
方法：我们使用基于质谱的基于活性的蛋白质谱分析（ABPP）与脱硫生物素-ATP探针，结合HSP90抑制剂的敏感性分析，分析一组GC细胞系中的激酶。我们在抑制剂敏感细胞中鉴定了由HSP90调节的激酶，并研究了MASTL敲低对GC细胞行为的影响。进行了MASTL敲除后的整体蛋白质组学分析，和生物信息学工具被用来分析结果数据。
结果：四种激酶-MASTL，STK11、CHEK1和MET在HSP90抑制剂敏感细胞中被鉴定为HSP90调节的。其中,微管相关丝氨酸/苏氨酸激酶样（MASTL）在GC中上调，并与不良预后相关。MASTL敲除减少迁移，入侵,和GC细胞的增殖。MASTL敲除后的整体蛋白质组学分析显示，NEDD4-1是GC进展中MASTL的潜在下游介质。NEDD4-1在GC中也上调，并与不良预后相关。类似于MASTL抑制，NEDD4-1击倒抑制了迁移，入侵,和GC细胞的增殖。
结论：我们的多蛋白质组分析表明，靶向MASTL可能是晚期胃癌的有希望的治疗方法，可能通过减少肿瘤促进蛋白，包括NEDD4-1。这项研究增强了我们对GC中激酶信号通路的理解，并为潜在的治疗策略提供了新的见解。
方法：我们使用基于质谱的基于活性的蛋白质谱分析（ABPP）与脱硫生物素-ATP探针，结合HSP90抑制剂的敏感性分析，分析一组GC细胞系中的激酶。我们在抑制剂敏感细胞中鉴定了由HSP90调节的激酶，并研究了MASTL敲低对GC细胞行为的影响。进行了MASTL敲除后的整体蛋白质组学分析，和生物信息学工具被用来分析结果数据。
结果：四种激酶-MASTL，STK11、CHEK1和MET在HSP90抑制剂敏感细胞中被鉴定为HSP90调节的。其中,微管相关丝氨酸/苏氨酸激酶样（MASTL）在GC中上调，并与不良预后相关。MASTL敲除减少迁移，入侵,和GC细胞的增殖。MASTL敲除后的整体蛋白质组学分析显示，NEDD4-1是GC进展中MASTL的潜在下游介质。NEDD4-1在GC中也上调，并与不良预后相关。类似于MASTL抑制，NEDD4-1击倒抑制了迁移，入侵,和GC细胞的增殖。
结论：我们的多蛋白质组分析表明，靶向MASTL可能是晚期胃癌的有希望的治疗方法，可能通过减少肿瘤促进蛋白，包括NEDD4-1。这项研究增强了我们对GC中激酶信号通路的理解，并为潜在的治疗策略提供了新的见解。
