Selective targeting of distinct active site nucleophiles by irreversible SRC-family kinase inhibitors.
Selective targeting of distinct active site nucleophiles by irreversible SRC-family kinase inhibitors.
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DOI：
10.1021/ja310659j
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发表时间：
2012-12-19
影响因子：
15
通讯作者：
Taunton, Jack
中科院分区：
文献类型：
--
作者：
Gushwa, Nathan N.;Kang, Sumin;Chen, Jing;Taunton, Jack
Src家族酪氨酸激酶在人类生理和疾病中起着关键作用，并且靶向该家族成员的几种药物正在临床使用中。这些药物似乎都不能区分密切相关的激酶。然而，评估它们对活细胞中内源性激酶的选择性仍然是一个重大挑战。在这里，我们报告了两个Src定向化学探针的设计，每个探针由具有5 '-亲电体的核苷支架组成。5 '-氟磺酰基苯甲酸酯（1）与保守的催化赖氨酸（Lys 295）反应，在相关激酶中几乎没有区别。相比之下，5 '-乙烯基磺酸酯（2）与在三个Src家族和六个不相关的激酶中发现的保守性差的近端半胱氨酸（Cys 277）反应。1和2都带有炔标签，并有效地标记完整细胞中各自的内源性激酶靶标。使用1作为竞争性探针，我们确定了临床Bcr-Abl抑制剂泊那替尼靶向Src家族激酶的程度。值得注意的是，虽然泊那替尼对内源性Fyn或Src几乎没有影响，但它有效地阻断了关键的T细胞激酶Lck。因此，探针1和2能够在活细胞中与不同的激酶亚群进行竞争性分析。
Src-family tyrosine kinases play pivotal roles in human physiology and disease, and several drugs that target members of this family are in clinical use. None of these drugs appear to discriminate among closely related kinases. However, assessing their selectivity toward endogenous kinases in living cells remains a significant challenge. Here, we report the design of two Src-directed chemical probes, each consisting of a nucleoside scaffold with a 5'- electrophile. A 5'-fluorosulfonylbenzoate (1) reacts with the conserved catalytic lysine (Lys295) and shows little discrimination among related kinases. By contrast, a 5'-vinylsulfonate (2) reacts with a poorly conserved, proximal cysteine (Cys277) found in three Src-family and six unrelated kinases. Both 1 and 2 bear an alkyne tag and efficiently label their respective endogenous kinase targets in intact cells. Using 1 as a competitive probe, we determined the extent to which ponatinib, a clinical Bcr-Abl inhibitor, targets Src-family kinases. Remarkably, while ponatinib had little effect on endogenous Fyn or Src, it potently blocked the critical Tcell kinase, Lck. Probes 1 and 2 thus enable competitive profiling vs. distinct kinase subsets in living cells.
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影响因子：
64.8
作者：
COURTNEIDGE, SA;SMITH, AE
通讯作者：
SMITH, AE
影响因子：
--
作者：
Patricelli MP;Nomanbhoy TK;Wu J;Brown H;Zhou D;Zhang J;Jagannathan S;Aban A;Okerberg E;Herring C;Nordin B;Weissig H;Yang Q;Lee JD;Gray NS;Kozarich JW
通讯作者：
Kozarich JW
影响因子：
--
作者：
Liu, YS;Shreder, KR;Rosenblum, JS
通讯作者：
Rosenblum, JS
影响因子：
50.3
作者：
O'Hare, Thomas;Shakespeare, William C.;Clackson, Tim
通讯作者：
Clackson, Tim
影响因子：
46.9
作者：
Bantscheff, Marcus;Eberhard, Dirk;Drewes, Gerard
通讯作者：
Drewes, Gerard