Stegmaier Kimberly, Wong Jenny S, Ross Kenneth N, Chow Kwan T, Peck David, Wright Renee D, Lessnick Stephen L, Kung Andrew L, Golub Todd R
Department of Pediatric Oncology, Dana-Farber Cancer Institute and Children's Hospital Boston, Harvard Medical School, Boston, Massachusetts, United States of America.
PLoS Med. 2007 Apr;4(4):e122. doi: 10.1371/journal.pmed.0040122.
The presence of tumor-specific mutations in the cancer genome represents a potential opportunity for pharmacologic intervention to therapeutic benefit. Unfortunately, many classes of oncoproteins (e.g., transcription factors) are not amenable to conventional small-molecule screening. Despite the identification of tumor-specific somatic mutations, most cancer therapy still utilizes nonspecific, cytotoxic drugs. One illustrative example is the treatment of Ewing sarcoma. Although the EWS/FLI oncoprotein, present in the vast majority of Ewing tumors, was characterized over ten years ago, it has never been exploited as a target of therapy. Previously, this target has been intractable to modulation with traditional small-molecule library screening approaches. Here we describe a gene expression-based approach to identify compounds that induce a signature of EWS/FLI attenuation. We hypothesize that screening small-molecule libraries highly enriched for FDA-approved drugs will provide a more rapid path to clinical application.
A gene expression signature for the EWS/FLI off state was determined with microarray expression profiling of Ewing sarcoma cell lines with EWS/FLI-directed RNA interference. A small-molecule library enriched for FDA-approved drugs was screened with a high-throughput, ligation-mediated amplification assay with a fluorescent, bead-based detection. Screening identified cytosine arabinoside (ARA-C) as a modulator of EWS/FLI. ARA-C reduced EWS/FLI protein abundance and accordingly diminished cell viability and transformation and abrogated tumor growth in a xenograft model. Given the poor outcomes of many patients with Ewing sarcoma and the well-established ARA-C safety profile, clinical trials testing ARA-C are warranted.
We demonstrate that a gene expression-based approach to small-molecule library screening can identify, for rapid clinical testing, candidate drugs that modulate previously intractable targets. Furthermore, this is a generic approach that can, in principle, be applied to the identification of modulators of any tumor-associated oncoprotein in the rare pediatric malignancies, but also in the more common adult cancers.
癌症基因组中肿瘤特异性突变的存在为药物干预带来治疗益处提供了潜在机会。不幸的是，许多类别的癌蛋白（如转录因子）不适合常规小分子筛选。尽管已鉴定出肿瘤特异性体细胞突变，但大多数癌症治疗仍使用非特异性细胞毒性药物。一个典型例子是尤因肉瘤的治疗。尽管绝大多数尤因肿瘤中存在的EWS/FLI癌蛋白在十多年前就已被鉴定，但它从未被用作治疗靶点。以前，用传统小分子文库筛选方法难以对该靶点进行调控。在此，我们描述了一种基于基因表达的方法来鉴定可诱导EWS/FLI信号减弱的化合物。我们假设筛选高度富集FDA批准药物的小分子文库将为临床应用提供更快速的途径。
通过对采用EWS/FLI导向RNA干扰的尤因肉瘤细胞系进行微阵列表达谱分析，确定了EWS/FLI关闭状态的基因表达特征。使用基于荧光珠检测的高通量连接介导扩增试验，对富集FDA批准药物的小分子文库进行筛选。筛选鉴定出阿糖胞苷（ARA-C）为EWS/FLI的调节剂。ARA-C降低了EWS/FLI蛋白丰度，相应地降低了细胞活力和转化能力，并在异种移植模型中抑制了肿瘤生长。鉴于许多尤因肉瘤患者预后不佳且阿糖胞苷安全性良好，有必要开展测试阿糖胞苷的临床试验。
我们证明，基于基因表达的小分子文库筛选方法能够识别可调节先前难以处理靶点的候选药物，以便进行快速临床试验。此外，这是一种通用方法，原则上可应用于鉴定罕见儿童恶性肿瘤中任何肿瘤相关癌蛋白的调节剂，也可用于更常见的成人癌症。
