Role of FH loss in development of HLRCC heriditary kidney cancer

2018年，(1)我们证明了靶向Hippo信号通路缺失治疗nf2缺陷乳头状肾癌的可行性。乳头状肾细胞癌（PRCC）是一种组织学和遗传异质性的肿瘤，占所有肾肿瘤的15-20%，可能需要多种治疗方法。编码Hippo信号通路关键调控因子的NF2肿瘤抑制基因的改变在22.5%的PRCC中被观察到。Hippo信号通路通过调节yes相关蛋白YAP1的转录活性来控制细胞增殖。NF2的缺失导致YAP1异常激活。Src家族激酶成员Yes也调节YAP1的转录活性。当我们研究YAP和Yes活性在三种nf2缺陷的PRCC细胞系中的重要性时，我们发现nf2缺陷与YAP1转录靶点的表达增加相关，而基于sirna的YAP1和Yes1的敲低下调了这一途径并显著降低了细胞活力。达沙替尼和萨拉卡替尼对YAP1有有效的抑制作用，两者均可导致YAP1转录靶点下调、细胞活力降低和G0-G1细胞周期阻滞。nf2缺陷PRCC的异种移植模型也显示，达沙替尼对肿瘤生长的反应降低。因此，抑制Yes和随后YAP1的转录活性在体外和体内都具有实质性的抗肿瘤细胞作用，可能为nf2缺陷PRCC患者提供可行的治疗方法。(2)为了更好地靶向糖酵解癌细胞的糖酵解，我们参与了一种新型的、有效的、细胞活性的吡唑基乳酸脱氢酶（LDH）抑制剂的开发。利用定量高通量筛选模式促进了命中识别，而基于结构的设计和多参数优化使得开发具有有效生化和细胞抑制LDH酶活性的化合物成为可能。在MiaPaCa2胰腺癌和A673尤因肉瘤细胞中，先导化合物对LDHA和LDHB均有低nM的抑制作用，对细胞乳酸生成有亚微摩尔的抑制作用，对糖酵解有抑制作用。此外，通过细胞热移测定（CETSA）证明了铅化合物对LDHA的强大靶向作用，并通过SPR确定了药物靶标停留时间。对这些数据的分析表明，药物靶点停留时间（off rate）可能是获得有效的基于细胞和持久的体内肿瘤代谢靶点抑制的重要因素。事实上，我们的主要LDH抑制剂在体内表现出强大的靶向活性。

In 2018, (1) we demonstrated the feasibility of targeting loss of the Hippo signaling pathway in NF2-deficient papillary kidney cancers. Papillary renal cell carcinomas (PRCC) are a histologically and genetically heterogeneous group of tumors that represent 15-20% of all kidney neoplasms and may require diverse therapeutic approaches. Alteration of the NF2 tumor suppressor gene, encoding a key regulator of the Hippo signaling pathway, is observed in 22.5% of PRCC. The Hippo signaling pathway controls cell proliferation by regulating the transcriptional activity of Yes-Associated Protein, YAP1. Loss of NF2 results in aberrant YAP1 activation. The Src family kinase member Yes also regulates YAP1 transcriptional activity. When we investigated the importance of YAP and Yes activity in three NF2-deficient PRCC cell lines, we found that NF2-deficiency correlated with increased expression of YAP1 transcriptional targets, and siRNA-based knockdown of YAP1 and Yes1 downregulated this pathway and dramatically reduced cell viability. Dasatinib and saracatinib have potent inhibitory effects on Yes and treatment with either resulted in downregulation of YAP1 transcription targets, reduced cell viability, and G0-G1 cell cycle arrest. Xenograft models for NF2-deficient PRCC also demonstrated reduced tumor growth in response to dasatinib. Thus, inhibiting Yes and the subsequent transcriptional activity of YAP1 had a substantial anti-tumor cell effect both in vitro and in vivo and may provide a viable therapeutic approach for patients with NF2-deficient PRCC. (2) In order to better target glycolysis in glycolytic cancer cells, we participated in developing a novel, potent, cell-active pyrazole-based inhibitor of lactate dehydrogenase (LDH). Utilization of a quantitative high-throughput screening paradigm facilitated hit identification, while structure-based design and multiparameter optimization enabled development of compounds with potent biochemical and cell-based inhibition of LDH enzymatic activity. Lead compounds exhibit low nM inhibition of both LDHA and LDHB, submicromolar inhibition of celular lactate production, and inhibition of glycolysis in MiaPaCa2 pancreatic cancer and A673 Ewing's sarcoma cells. Moreover, robust target engagement of LDHA by lead compounds was demonstrated using the cellular thermal shift assay (CETSA), and drug-target residence time was determined via SPR. Analysis of these data suggests that drug-target residence time (off-rate) may be an important attribute to consider for obtaining potent cell-based and durable in vivo inhibition of this cancer metabolism target. Indeed, our lead LDH inhibitor demonstrates potent in vivo on-target activity.