The RAS proteins are the most frequently mutated oncogenes in cancer, with highest frequency found in pancreatic, lung, and colon tumors. Moreover, the activity of RAS is required for the proliferation and/or survival of these tumor cells and thus represents a high-value target for therapeutic development. Direct targeting of RAS has proven challenging for multiple reasons stemming from the biology of the protein, the complexity of downstream effector pathways and upstream regulatory networks. Thus, significant efforts have been directed at identifying downstream targets on which RAS is dependent. These efforts have proven challenging, in part due to confounding factors such as reliance on two-dimensional adherent monolayer cell cultures that inadequately recapitulate the physiologic context to which cells are exposed in vivo. To overcome these issues, we implemented a high-throughput screening (HTS) approach using a spheroid-based 3-dimensional culture format, thought to more closely reflect conditions experienced by cells in vivo. Using isogenic cell pairs, differing in the status of KRAS, we identified Proscillaridin A as a selective inhibitor of cells harboring the oncogenic KRas^G12V allele. Significantly, the identification of Proscillaridin A was facilitated by the 3D screening platform and would not have been discovered employing standard 2D culturing methods.

中文翻译：

---

一种新颖的三维高通量筛选方法，确定了突变KRAS选择性致死表型的诱导剂。

RAS蛋白是癌症中最常见的突变癌基因，在胰腺，肺和结肠肿瘤中发现的频率最高。此外，RAS的活性对于这些肿瘤细胞的增殖和/或存活是必需的，因此代表了治疗发展的高价值靶标。事实证明，直接靶向RAS具有挑战性，原因有多种，其原因在于蛋白质的生物学特性，下游效应子途径的复杂性和上游调节网络。因此，已经做出了巨大的努力来确定RAS所依赖的下游目标。这些努力已被证明具有挑战性，部分原因是由于混杂因素，例如对二维贴壁单层细胞培养的依赖性不足，无法充分概括体内暴露于细胞的生理环境。为克服这些问题，我们使用基于球体的3维培养形式实施了高通量筛选（HTS）方法，认为该方法可以更紧密地反映体内细胞所经历的状况。使用同基因细胞对，不同的KRAS状态，我们确定Proscillaridin A是具有致癌性KRas的细胞的选择性抑制剂^G12V等位基因。值得注意的是，通过3D筛选平台可以方便地鉴定Proscillaridin A，而采用标准的2D培养方法则无法发现。