Activating Cell Death Pathways in Breast Cancer Cells

癌细胞通过多种遗传和表观遗传机制避免细胞凋亡。肿瘤坏死因子家族死亡受体(如TNFR、Fas、DR3、TRAIL受体1和TRAIL受体2)通过募集caspase并在其各自的配体(如TNF、Fas配体和TRAIL)激活后激活，从而诱导细胞凋亡。我们正在研究TRAIL死亡受体(TNFR家族成员)及其配体(如TRAIL和激动型抗体)在乳腺癌细胞中的表达和功能，以选择性地触发癌细胞的凋亡。在我们的早期工作中，我们发现大多数乳腺癌细胞株对TRAIL诱导的细胞凋亡具有抵抗力。我们已经证明，具有三阴性/基底样特征的乳腺癌细胞亚群对TRAIL诱导的细胞凋亡非常敏感，而其他乳腺癌亚型对TRAIL诱导的细胞凋亡相对抵抗。这类乳腺癌特别具有侵袭性，最需要有针对性的治疗。此外，我们发现TRAIL受体2而不是TRAIL受体1是TRAIL诱导敏感乳腺癌细胞凋亡所必需的。后一项发现将有助于选择TRAIL激动剂用于乳腺癌患者的最终临床试验。此外，我们还发现，细胞对TRAIL诱导的细胞凋亡的抵抗可以通过与化疗药物、靶向药物如曲妥珠单抗和EGFR抑制剂共同孵育来克服。最近，我们已经证明了G2/M检查点酪氨酸激酶Wee1的抑制或丢失增强了TRAIL介导的基底样乳腺癌细胞的凋亡。总而言之，这些研究开始为TRAIL配体单独或与其他药物联合治疗乳腺癌患者的研究提供明确的临床前理由。正在进行的工作是：1)利用功能基因组学来鉴定和鉴定调控TRAIL受体激动剂诱导乳腺癌细胞凋亡的基因和蛋白。2)表征新型TRAIL受体激动剂在乳腺癌细胞中的作用；3)评价其对乳腺癌细胞免疫原性的影响。在一个副产品项目中，我们正在描述ClpP激动剂的作用机制。后一种药物被描述为通过TRAIL途径起作用。然而，我们的数据表明，它的作用与TRAIL途径无关，可以杀死乳腺癌和其他癌细胞。我们正在进行的工作包括1)表征ClpP激动剂诱导乳腺癌细胞死亡的机制；2)与NCATS合作，我们进行了一项筛选，以确定在乳腺癌细胞中与ClpP激动剂协同作用的药物，并正在验证来自该筛选的命中结果，以证明与ClpP激动剂有用的联合疗法。3)研究ClpP激动剂对乳腺癌肿瘤起始细胞的影响。

Cancer cells avoid apoptosis by a variety of genetic and epigenetic mechanisms. TNF family death receptors (e.g., TNFR, Fas, DR3, TRAIL Receptor 1, and TRAIL Receptor 2) induce apoptosis in cells by recruiting and activating caspases upon activation by their respective ligands (e.g., TNF, Fas Ligand, and TRAIL). We are investigating the expression and function of TRAIL death receptors (members of the TNFR family) and their ligands (e.g., TRAIL and agonistic antibodies) in breast cancer cells in order to selectively trigger apoptosis in the cancer cells. In our early work, we found that most breast cancer cell lines are resistant to the induction of apoptosis by TRAIL. We have demonstrated that a subset of breast cancer cells, those with triple-negative/basal-like features are very sensitive to TRAIL-induced apoptosis while other breast cancer subtypes are relatively resistant to TRAIL-induced apoptosis. This subset of breast cancers is particularly aggressive and most in need of targeted therapies. Further, we found that TRAIL Receptor 2, and not TRAIL Receptor 1, is required for TRAIL induced apoptosis in the sensitive breast cancer cells. This latter finding will help in the selection of TRAIL agonists for eventual clinical trials in breast cancer patients. In addition, we have found that resistance to TRAIL-induced apoptosis can be overcome by co-incubation of the cells with chemotherapeutic agents, targeted agents such as trastuzumab, and EGFR inhibitors. More recently, we have demonstrated that inhibition or loss of the G2/M checkpoint tyrosine kinase Wee1 enhances TRAIL-mediated apoptosis in basal-like breast cancer cells. Together these studies are beginning to provide clear preclinical rationales for studies of TRAIL ligands alone or in combination with other drugs in patients with breast cancer. Ongoing work is: 1) using functional genomics to identify and characterize the genes and proteins that regulate apoptosis induced by TRAIL receptor agonists in breast cancer cells. 2) characterizing novel TRAIL receptor agonists in breast cancer cells, 3) evaluating the effects on the immunogenecity of breast cancer cells. In a spin off projject we are characterizing the mechanism of action of ClpP agonists. The latter drug was described to work via the TRAIL pathway. However, our data suggest that it works independent of the TRAIL pathway to kill breast cancer and other cancer cells. Our ongoing work includes 1) characterizing the mchanism of death induced by ClpP agonists on breast cancer cells; 2) in collaboration with NCATS we conducted a screen to identify drugs that synergize with ClpP agonists in breast cancer cells and are validating hits from that screen to credential combination therapies that will be useful with ClpP agonists. 3) investigating the effects of ClpP agonists on breast cancer tumor initiating cells.