摘要 1378：一种新型、高效的 PI3Kα 共价抑制剂可在癌细胞中解卷积 I 类 PI3K 亚型,Cancer Research

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摘要 1378：一种新型、高效的 PI3Kα 共价抑制剂可在癌细胞中解卷积 I 类 PI3K 亚型
Cancer Research ( IF 12.5 ) Pub Date : 2021-07-01 , DOI: 10.1158/1538-7445.am2021-1378
Erhan Keles ₁ , Chiara Borsari ₁ , Rohitha Sriramaratnam ₁ , Thorsten Schäfer ₁ , Matthias P. Wymann ₁

Affiliation

摘要磷酸肌醇 3-激酶 (PI3K) 产生促进细胞生长、增殖、运动和抑制细胞凋亡的中枢调节信号。PI3K 在广泛的人类肿瘤中失调。 [1] 考虑到 PI3K 信号在癌症生物学中的核心作用，该领域临床试验的成功低于预期，主要是由于代谢靶向不良反应。在这里，我们提出了一种新型的、高选择性的 PI3Kα-共价抑制剂（化合物 19），它结合了泛 PI3K 抑制剂 PQR514[2] 的特殊亲和力和一个接头部分，引导弹头通过迈克尔加成与异构体选择性反应- PI3Kα 的特异性 Cys862。结合细胞分析平台验证 19，具体而言，评估其目标选择性和占用率，并将其体外活性转化为细胞读数。总体而言，与先前报道的共价 PI3Kα 抑制剂 CNX-1351[3] 相比，19 表现出优异的细胞活性和优越的特性。NanoBRET（生物发光共振能量转移）药物靶标参与测定允许研究 HEK293 细胞的抑制功效。利用 Cys862Ser 突变 PI3Kα 蛋白的 NanoBRET 实验证实了与 Cys862 的选择性反应。各种细胞系在 PI3Kα（MCF7、T47D、SKOV3）中组成型激活或 PTEN 缺陷（PC3 和 A2058）被用于解决靶标接合问题，这证实了对蛋白激酶 B（PKB/Akt、Ser473）磷酸化的持续和长期抑制在清除了 19 种缺乏功能弹头的非共价结构类似物之后。此外，基于 FOXO1 转录因子的核质易位探针能够研究 PKB/Akt 下游的信号传播。在 PI3Kα 依赖的细胞系中，FOXO1 即使在去除 19 后仍保持主要的核定位，证实了扩展的 PI3Kα 靶向特定于该药物的共价附着。重要的是，与临床批准的非共价 PI3Kα 选择性抑制剂 BYL719 相比，19 的效力显着提高。总之，我们的结果向通过异构体选择性共价锚定增加对 PI3Kα 信号传导的局部和时间控制迈出了重要一步。该策略是朝着开发药物样、异构体选择性、共价 PI3K 抑制剂支架迈出的一步，和 19 为开发 PI3Kα 抑制剂提供了一个模板，该抑制剂在癌症治疗中总体上减少了不良副作用。此外，我们的共价抑制策略将为更好地分子理解 PI3K 亚型信号通路铺平道路。[1] Wymann MP 和 Schneiter R. Nat。莫尔牧师。细胞生物学。2008, 9, 162-176。[2] Borsari, C 等。人。ACS医学。化学。莱特。2019, 10 (10), 1473-1479。[3] 纳赫特 M 等。人。J.医学。化学。2013, 56 (3), 712-721。引文格式：Erhan Keles、Chiara Borsari、Rohitha Sriramaratnam、Thorsten Schäfer、Matthias P. Wymann。一种新型的、高效的 PI3Kα 共价抑制剂可以解卷积癌细胞中的 I 类 PI3K 亚型 [摘要]。在：美国癌症研究协会 2021 年年会论文集；2021 年 4 月 10 日至 15 日和 5 月 17 日至 21 日。费城 (PA)：AACR；癌症研究 2021；

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Abstract 1378: A novel, highly potent PI3Kα covalent inhibitor deconvolutes class I PI3K isoforms in cancer cells

Abstract Phosphoinositide 3-kinase (PI3K) imposes central regulatory signals that promote cell growth, proliferation, motility, and suppression of apoptosis. PI3K is dysregulated in a broad spectrum of human tumours.[1] Considering the central role attributed to PI3K signaling in cancer biology, the success of clinical trials in the field was below expectations, mainly due to metabolic on-target adverse effects. Here, we present a novel, highly selective PI3Kα-covalent inhibitor (compound 19), which combines the exceptional affinity of the pan-PI3K inhibitor PQR514[2] with a linker moiety, directing a warhead to selectively react via a Michael addition with isoform-specific Cys862 of PI3Kα. Cellular assay platforms were combined to validate 19, specifically, to assess its target selectivity and occupancy, and to translate its in vitro activity to cellular readouts. Overall, 19 exhibited excellent cellular activity with a superior profile compared to CNX-1351[3], a previously reported covalent PI3Kα inhibitor. A NanoBRET (Bioluminescence Resonance Energy Transfer) drug-target engagement assay allowed for an investigation of inhibitory efficacy in HEK293 cells. NanoBRET experiments exploiting the Cys862Ser mutant PI3Kα protein confirmed selective reactivity with Cys862. Various cell lines either constitutively activated in PI3Kα (MCF7, T47D, SKOV3) or deficient in PTEN (PC3 and A2058) were used to resolve target engagement, which confirmed a persistent and prolonged suppression of protein kinase B (PKB/Akt, Ser473) phosphorylation after washout of 19 over non-covalent structural analogs thereof that lack a functional warhead. Furthermore, nucleo-cytoplasmic translocation probes based on the FOXO1 transcription factor enabled the investigation of signal propagation downstream of PKB/Akt. In PI3Kα-dependent cell lines, FOXO1, maintained a predominant nuclear localization even after removal of 19, confirming extended PI3Kα targeting upon covalent attachment of specifically this drug. Importantly, 19 provided a considerable gain in potency over BYL719, a clinically approved non-covalent PI3Kα-selective inhibitor. Taken together, our results present a major step towards an increased local and temporal control of PI3Kα signaling through isoform-selective covalent anchoring. This strategy is a step towards the development of drug-like, isoform-selective, covalent PI3K inhibitor scaffolds, and 19 provides a template for the development of PI3Kα inhibitors with overall reduced adverse side effects in cancer therapy. Moreover, our covalent inhibition strategy will pave the way for a better molecular understanding of PI3K isoform signaling in general. [1] Wymann M. P. and Schneiter R. Nat. Rev. Mol. Cell Biol. 2008, 9, 162-176. [2] Borsari, C et. al. ACS Med. Chem. Lett. 2019, 10 (10), 1473-1479. [3] Nacht M et. al. J. Med. Chem. 2013, 56 (3), 712-721. Citation Format: Erhan Keles, Chiara Borsari, Rohitha Sriramaratnam, Thorsten Schäfer, Matthias P. Wymann. A novel, highly potent PI3Kα covalent inhibitor deconvolutes class I PI3K isoforms in cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1378.

更新日期：2021-07-01

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