RAS mutations prevalent in high-risk leukemia have been linked to relapse and chemotherapy resistance. Efforts to directly target RAS proteins have been largely unsuccessful. However, since RAS-mediated transformation is dependent on signaling through the RAS-related C3 botulinum toxin substrate (RAC) small GTPase, we hypothesized that targeting RAC may be an effective therapeutic approach in RAS mutated tumors. Here we describe multiple small molecules capable of inhibiting RAC activation in acute lymphoblastic leukemia cell lines. One of these, DW0254, also demonstrates promising anti-leukemic activity in RAS-mutated cells. Using chemical proteomics and biophysical methods, we identified the hydrophobic pocket of phosphodiester 6 subunit delta (PDE6D), a known RAS chaperone, as a target for this compound. Inhibition of RAS localization to the plasma membrane upon DW0254 treatment is associated with RAC inhibition through a phosphatidylinositol-3-kinase/AKT-dependent mechanism. Our findings provide new insights into the importance of PDE6D-mediated transport for RAS-dependent RAC activation and leukemic cell survival.

在高危白血病中普遍存在的 RAS 突变与复发和化疗耐药有关。直接针对 RAS 蛋白的努力基本上没有成功。然而，由于 RAS 介导的转化依赖于通过 RAS 相关 C3 肉毒杆菌毒素底物 (RAC) 小 GTPase 的信号传导，我们假设靶向 RAC 可能是 RAS 突变肿瘤的有效治疗方法。在这里，我们描述了多种能够抑制急性淋巴细胞白血病细胞系中 RAC 活化的小分子。其中之一，DW0254，在 RAS 突变细胞中也表现出有希望的抗白血病活性。使用化学蛋白质组学和生物物理方法，我们确定了磷酸二酯 6 亚基 delta (PDE6D) 的疏水口袋，一种已知的 RAS 伴侣，作为该化合物的靶标。DW0254 处理后 RAS 定位于质膜的抑制与通过磷脂酰肌醇-3-激酶/AKT 依赖性机制抑制 RAC 相关。我们的研究结果为 PDE6D 介导的转运对 RAS 依赖性 RAC 激活和白血病细胞存活的重要性提供了新的见解。