Structural and dynamic determinants for highly selective RET kinase inhibition reveal cryptic druggability,Journal of Advanced Research

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Structural and dynamic determinants for highly selective RET kinase inhibition reveal cryptic druggability
Journal of Advanced Research ( IF 11.4 ) Pub Date : 2022-05-17 , DOI: 10.1016/j.jare.2022.05.004
Moustafa A Shehata ₁ , Julia Contreras ₂ , Ana Martín-Hurtado ₂ , Aurane Froux ₂ , Hossam Taha Mohamed ₃ , Ahmed A El-Sherif ₄ , Iván Plaza-Menacho ₂

Affiliation

Introduction

The structural and dynamic determinants that confer highly selective RET kinase inhibition are poorly understood.

Objectives

To explore the druggability landscape of the RET active site in order to uncover structural and dynamic vulnerabilities that can be therapeutically exploited.

Methods

We apply an integrated structural, computational and biochemical approach in order to explore the druggability landscape of the RET active site.

Results

We demonstrate that the that the druggability landscape of the RET active site is determined by the conformational setting of the ATP-binding (P-) loop and its coordination with the αC helix. Open and intermediate P-loop structures display additional druggable vulnerabilities within the active site that were not exploited by first generation RET inhibitors. We identify a cryptic pocket adjacent to the catalytic lysine formed by K758, L760, E768 and L772, that we name the post-lysine pocket, with higher druggability potential than the adenine-binding site and with important implications in the regulation of the phospho-tyrosine kinase activity. Crystal structure and simulation data show that the binding mode of highly-selective RET kinase inhibitors LOXO-292 and BLU-667 is controlled by a synchronous open P-loop and αC-in configuration that allows accessibility to the post-lysine pocket. Molecular dynamics simulations show that these inhibitors efficiently occupy the post-lysine pocket with high stability through the simulation time-scale (300 ns), with both inhibitors forming hydrophobic contacts further stabilized by pi-cation interactions with the catalytic K758. Engineered mutants targeting the post-lysine pocket impact on inhibitor binding and sensitivity, as well as RET tyrosine kinase activity.

Conclusions

The identification of the post-lysine pocket as a new druggable vulnerability in the RET kinase and its exploitation by second generation RET inhibitors have important implications for future drug design and the development of personalized therapies for patients with RET-driven cancers.

中文翻译：

高选择性 RET 激酶抑制的结构和动态决定因素揭示了神秘的成药性

介绍

赋予高度选择性 RET 激酶抑制的结构和动态决定因素知之甚少。

目标

探索 RET 活性位点的成药性景观，以发现可用于治疗的结构和动态弱点。

方法

我们应用综合结构、计算和生化方法来探索 RET 活性位点的成药性景观。

结果

我们证明 RET 活性位点的成药性景观是由 ATP 结合 (P-) 环的构象设置及其与 αC 螺旋的协调决定的。开放和中间 P-loop 结构在活性位点内显示出额外的药物漏洞，这些漏洞未被第一代 RET 抑制剂利用。我们确定了一个与由 K758、L760、E768 和 L772 形成的催化赖氨酸相邻的隐蔽口袋，我们将其命名为赖氨酸后口袋，它比腺嘌呤结合位点具有更高的成药性潜力，并且在磷酸盐的调节中具有重要意义。酪氨酸激酶活性。晶体结构和模拟数据表明，高选择性 RET 激酶抑制剂 LOXO-292 和 BLU-667 的结合模式由同步开放 P 环和 αC-in 配置控制，允许进入赖氨酸后口袋。分子动力学模拟表明，这些抑制剂在模拟时间尺度（300 ns）内有效地占据了高稳定性的赖氨酸后袋，两种抑制剂形成疏水接触，通过与催化 K758 的 pi 阳离子相互作用进一步稳定。针对赖氨酸后口袋的工程突变体会影响抑制剂结合和敏感性，以及 RET 酪氨酸激酶活性。分子动力学模拟表明，这些抑制剂在模拟时间尺度（300 ns）内有效地占据了高稳定性的赖氨酸后袋，两种抑制剂形成疏水接触，通过与催化 K758 的 pi 阳离子相互作用进一步稳定。针对赖氨酸后口袋的工程突变体会影响抑制剂结合和敏感性，以及 RET 酪氨酸激酶活性。分子动力学模拟表明，这些抑制剂在模拟时间尺度（300 ns）内有效地占据了具有高稳定性的后赖氨酸口袋，两种抑制剂形成疏水接触，通过与催化 K758 的 pi 阳离子相互作用进一步稳定。针对赖氨酸后口袋的工程突变体会影响抑制剂结合和敏感性，以及 RET 酪氨酸激酶活性。