(PQ5) Investigation of intertumoral and intratumoral heterogeneity of mitochondrial apoptotic sensitivity

 描述(申请人提供)：肿瘤内和肿瘤内线粒体的异质性对癌症治疗中的化学扰动的反应在不同组织学的肿瘤之间、相同组织学的不同肿瘤之间、甚至同一肿瘤内的不同肿瘤细胞之间是不同的。癌症治疗的细胞毒性反应很大程度上是由线粒体的凋亡途径控制的。因此，我们假设，在所有这三个水平上的反应异质性是由线粒体的功能和分子异质性决定的。BH3图谱通过测量线粒体对BH3多肽的敏感性，提供了一种功能上的衡量细胞离凋亡阈值有多近的方法，我们也将这种特性称为凋亡“启动”。我们之前已经证明，在血液学和实体瘤中，患者肿瘤的预基线启动可以预测对常规化疗的临床反应。我们理解靶向治疗反应的方法是不同的。与作为传统化疗靶点的DNA和微管的无处不在的性质不同，靶向治疗攻击了某些癌细胞中选择性存在的脆弱性。为了确定这些脆弱性存在的地方，我们短暂地(不到24小时)将癌细胞暴露在药物下，并测量这些药物是否增加了细胞凋亡的启动。我们已经证明，这种方法，我们称之为动态BH3图谱(DBP)，可以在体外和体内预测血液和实体肿瘤靶向药物的肿瘤细胞死亡。与传统的细胞毒性方法相比，DBP有两个重要的优点。首先，DBP可以更有效地应用于初级患者样本。测量癌细胞细胞毒性的实验通常需要几天的培养。因为原发患者样本的长期培养是如此不可靠，所以原发肿瘤的细胞毒性测量也是不可靠的。DBP不需要超过一天的体外培养，因为我们在弗兰克细胞死亡发生之前就进行了测量，我们已经证明了它在原发液体和固体肿瘤中的预测能力。其次，有许多有用的抗癌药物，它们不会导致坦率的细胞死亡，但仍然会激发细胞凋亡信号，从而在联合疗法中促进杀戮。经典的细胞毒性测量不能识别这些，但DBP可以。我们建议研究肿瘤内和肿瘤间的异质性，基于对使线粒体对凋亡敏感的化合物的不同反应。我们将重点放在结肠癌肿瘤上，因为我们可以获得这些肿瘤的原代和PDX标本。我们的主要目标是开发一个潜在有用药物的治疗工具箱，确定如何以个性化的方式最好地结合这些药物，并了解导致这些药物反应差异的线粒体功能异质性的分子基础。

 DESCRIPTION (provided by applicant): Inter- and intra-tumoral mitochondrial heterogeneity in response to chemical perturbation in cancer Response to therapy is heterogeneous between tumors of different histology, between different tumors of the same histology, and even among different tumor cells within the same tumor. Much of the cytotoxic response to therapy in cancer is governed by the mitochondrial pathway of apoptosis. Therefore, we hypothesize that heterogeneity in response at all three of these levels is governed by functional and molecular heterogeneity of mitochondria. BH3 profiling provides a functional measure of how close a cell is to the threshold of apoptosis by measuring mitochondrial sensitivity to BH3 peptides, a property we also refer to as apoptotic "priming". We have previously shown, in both hematologic and solid tumors, that pretreatment baseline priming of patient tumors predicts clinical response to conventional chemotherapy. Our approach to understanding response to targeted therapies is distinct. In contrast to the ubiquitous nature of the DNA and microtubules that are the targets of conventional chemotherapy, targeted therapies attack vulnerabilities that are selectively present in certain cancer cells. To identify where these vulnerabilities exist, we briefly (less than 24 hours) expose cancer cells to drugs and measure whether the drugs increase apoptotic priming. We have shown that this approach, which we call dynamic BH3 profiling (DBP), can predict death of tumor cells from targeted agents in vitro and in vivo for both hematologic and solid tumors. There are two important advantages of DBP over conventional measures of cytotoxicity. First, DBP can be applied much more efficiently to primary patient samples. Experiments measuring cytotoxicity in cancer cells often require days of culture. Because the long-term culture of primary patient samples is so unreliable, cytotoxic measurements in primary tumors are unreliable. DBP requires no more than a day of ex vivo culture since we measure well before frank cell death occurs, and we have demonstrated its predictive power in primary liquid and solid tumors. Second, there are many useful anti-cancer agents that do not cause frank cell death, but which nonetheless provoke apoptotic signaling that facilitates killing in combination regimens. Classical cytotoxic measurements do not identify these, but DBP does. We propose to study intra- and inter-tumoral heterogeneity based on differential response to compounds that sensitize mitochondria for apoptosis. We will focus on colon cancer tumors, as we have access to primary and PDX specimens of these tumors. Our main goals are to develop a therapeutic toolbox of potentially useful drugs, determine how best to combine these in a personalized way, and also to understand the molecular basis of the heterogeneity of mitochondrial function that underlies differences in response to these drugs.