Targeting the Brk-p27-cdk4 axis in cancer

该项目将利用最近发现的细胞周期控制细胞周期的致癌机制来控制细胞周期蛋白D。 CDK4(DK4)由p27Kip1及其激活物--乳腺肿瘤相关激酶(BRK)--靶向 癌细胞。虽然历史上被认为是DK4组装因子和CDK2抑制因子，但p27也起CDK4的作用 开/关“开关。”P27(PY88)门的酪氨酸(Y)磷酸化维持ATP结合和CAK CDK4‘S T环的磷酸化，对DK4的激活是必不可少的。我们证明了BRK是生理上的 磷酸化p27的激酶，通过这样做，增加CDK4的活性，细胞周期进展，以及 化学抑制剂Palbociclib对CDK4特异性抑制的抵抗力，目前正在进行乳房临床试验 癌症。细胞周期蛋白D和CDK4在肿瘤中过度表达，但它们的水平不是可靠的生物标志物 由于p27对它们的复合体具有激活作用，因此具有致癌的潜能。我们的数据预测， BRK-p27-DK4轴在DK4依赖性肿瘤的转化中起重要作用，并可解释肿瘤耐药性 对CDK4的特异性抑制。我们确定p27通过SH3：PxxP接触与BRK相互作用，特定于 BRK的SH3域。这种相互作用的重要性通过对自然选择的描述来验证 BRK剪接变异体BRK(ALT)，仅包含BRK的SH3结构域，在 活着。因此，阻断BRK：p27的相互作用是抑制CDK4活性的可行策略，而CDK4应该是 以治疗的方式进行探索。我们的具体目标：1)研究Py作为Palbociclib反应的标志物 组织培养和患者材料。我们将对培养的细胞株进行特征分析，以从统计上确定 Py作为CDK4的一项指标，与Palbociclib敏感性相关，可用于预测疗效。在……里面 与我们的部门合作。在病理学方面，我们将分析乳腺癌档案样本和原发患者 资料，包括ER/PR+，HER-乳腺肿瘤，直接关联Palbociclib反应性和Py。2)至 证明ALT和Py阻断疗法可以在组织培养模型中防止增殖。 使用乳腺癌组织培养模型，我们将展示ALT和帕博西利介导的停滞在 在结果方面以及在分子和细胞生物学水平上。我们的数据将显示使用ALT阻止Py 是阻止癌细胞增殖的有效方法，因为它同时抑制CDK4和CDK2，诱导 促进细胞衰老，防止抗药性。3)丙氨酸氨基转移酶和丙氨酸转氨酶阻断疗法 可以在动物模型中阻止肿瘤的进展。我们将证明在小鼠异种移植中阻断p27 Py 乳腺癌模型可以防止癌症进展和耐药性。

This project will exploit a recently discovered oncogenic mechanism of cell cycle control exerted on cyclin D- cdk4 (DK4) by p27Kip1 and its activator, the Breast tumor Related Kinase (Brk), to target vulnerabilities in cancer cells. Although historically known as a DK4 assembly factor and cdk2 inhibitor, p27 also acts as a cdk4 ON/OFF “switch.” Tyrosine (Y) phosphorylation of p27 (pY88) gatekeeps both ATP binding and CAK phosphorylation of cdk4's T loop, essential for DK4 activation. We demonstrated that Brk is the physiological kinase that phosphorylates p27, and by doing so, increases cdk4 activity, cell cycle progression, and resistance to cdk4-specific inhibition by the chemical inhibitor Palbociclib, currently in clinical trials for breast cancer. Cyclin D and cdk4 are overexpressed in tumors, but their levels are not reliable biomarkers of oncogenic tumor potential because of the activating role p27 has on their complex. Our data predict that the Brk-p27-DK4 axis is important for transformation in DK4-dependent tumors and can explain tumor resistance to cdk4-specific inhibition. We determined that p27 interacts with Brk through a SH3:PxxP contact, specific for Brk's SH3 domain. The importance of this interaction is validated by characterization of a natural ALTernatively spliced Brk variant Brk (ALT), which contains only Brk's SH3 domain, and functions as a competitive inhibitor in vivo. Thus, blocking the Brk:p27 interaction is a viable strategy to inhibit cdk4 activity, which should be explored therapeutically. Our specific aims: 1) To examine pY as a marker of Palbociclib response in tissue culture and patient material. We will characterize cultured cell lines to statistically establish whether pY, as a measure of cdk4, correlates with Palbociclib sensitivity and could be used to predict response. In conjunction with our Dept. of Pathology, we will analyze archival breast cancer samples and primary patient material, including ER/PR+, Her- breast tumors, to correlate Palbociclib responsiveness and pY directly. 2) To demonstrate that ALT and pY blockage therapy can prevent proliferation in tissue culture models. Using breast cancer tissue culture models, we will show that ALT and Palbociclib-mediated arrest is different in terms of outcome and at the molecular and cell biological level. Our data will show that blocking pY using ALT is a powerful approach to block cancer cell proliferation, because it inhibits both cdk4 and cdk2, induces cellular senescence and prevents drug resistance. 3) To demonstrate that ALT and pY blockage therapy can prevent tumor progression in animal models. We will show that blocking p27 pY in mouse xenografts breast cancer models can prevent cancer progression and drug resistance.