背景:嗜铬细胞瘤和副神经节瘤(PPGL)是罕见的神经内分泌肿瘤。新的药物靶点和蛋白质有助于敏感的PPGL成像,可以改善PPGL患者的治疗和生活质量。即患有复发性或转移性疾病的患者。使用组合的蛋白质组学策略,我们在肿瘤细胞表面上调的整合膜蛋白(IMP)和PPGL中的非膜药物酶中寻找此类临床相关靶点.
方法:我们对22个充分表征的人PPGL样品和来自肾上腺髓质的正常嗜铬细胞组织进行了详细的蛋白质组学分析。肿瘤裂解物的标准定量蛋白质组学分析,主要提供非膜蛋白的信息,伴随着特定的膜蛋白质组目标方法,即使用凝集素亲和力富集糖肽,通过酰肼化学捕获糖肽,和富集膜嵌入的疏水跨膜片段。
结果:该研究发现,与对照嗜铬细胞组织相比,PPGL中67种细胞表面整合膜蛋白强烈上调。我们根据已经记录的蛋白质在癌细胞生长或进展中的直接作用来优先考虑这些蛋白质。使用特异性抗体证实七个最有希望的药物靶标(CD146、CD171、ANO1、CD39、ATP8A1、ACE和SLC7A1)的表达增加。我们的实验策略还提供了可溶性蛋白质的表达数据。在PPGL中上调的可药用非膜酶中,我们确定了三个潜在的药物靶点(SHMT2,ARG2和自分泌运动因子),并验证了它们的上调表达.
结论:应用最近被称为“干草叉”的组合蛋白质组学策略可以对两者进行定量分析,膜和非膜蛋白质组,并在人类PPGL中鉴定出10个潜在的药物靶标。鉴定了位于细胞表面的7种膜蛋白和3种非膜可药物酶蛋白,并验证了它们在PPGL中的显着上调。所有的蛋白质以前都被证明在几种人类癌症中上调,并在癌症进展中起直接作用。这些蛋白质的明显上调以及它们的定位和在肿瘤进展中确立的直接作用使这些分子有希望候选作为敏感PPGL成像的药物靶标或蛋白质。
BACKGROUND: Pheochromocytomas and paragangliomas (PPGLs) are rare neuroendocrine tumors. New drug targets and proteins that would assist sensitive PPGL imagining could improve therapy and quality of life of patients with PPGL, namely those with recurrent or metastatic disease. Using a combined proteomic strategy, we looked for such clinically relevant targets among integral membrane proteins (IMPs) upregulated on the surface of tumor cells and non-membrane druggable enzymes in PPGL.
METHODS: We conducted a detailed proteomic analysis of 22 well-characterized human PPGL samples and normal chromaffin tissue from adrenal medulla. A standard quantitative proteomic analysis of tumor lysate, which provides information largely on non-membrane proteins, was accompanied by specific membrane proteome-aimed methods, namely glycopeptide enrichment using lectin-affinity, glycopeptide capture by hydrazide chemistry, and enrichment of membrane-embedded hydrophobic transmembrane segments.
RESULTS: The study identified 67 cell surface integral membrane proteins strongly upregulated in PPGL compared to control chromaffin tissue. We prioritized the proteins based on their already documented direct role in cancer cell growth or progression. Increased expression of the seven most promising drug targets (CD146, CD171, ANO1, CD39, ATP8A1, ACE and SLC7A1) were confirmed using specific antibodies. Our experimental strategy also provided expression data for soluble proteins. Among the druggable non-membrane enzymes upregulated in PPGL, we identified three potential drug targets (SHMT2, ARG2 and autotaxin) and verified their upregulated expression.
CONCLUSIONS: Application of a combined proteomic strategy recently presented as \"Pitchfork\" enabled quantitative analysis of both, membrane and non-membrane proteome, and resulted in identification of 10 potential drug targets in human PPGL. Seven membrane proteins localized on the cell surface and three non-membrane druggable enzymes proteins were identified and verified as significantly upregulated in PPGL. All the proteins have been previously shown to be upregulated in several human cancers, and play direct role in cancer progression. Marked upregulation of these proteins along with their localization and established direct roles in tumor progression make these molecules promising candidates as drug targets or proteins for sensitive PPGL imaging.