TGF-betas in breast cancer progression

在2017财年，我们继续使用我们开发的一种新的功能成像方法来研究转化生长因子-β在调节癌症干细胞(CSC)隔间中的作用，该方法允许实时和原位显示这一少数细胞群体。我们的慢病毒CSC报道使用了一种合成启动子，其中荧光蛋白的表达是由干细胞主转录因子Oct4和Sox2驱动的。使用这种方法，我们已经开发出在2D和3D培养系统中扩展单个CSC的细胞命运图谱的方法，并且我们正在与阿尔伯特·爱因斯坦医学院的John Condeelis博士的实验室合作，对原发肿瘤和肺转移部位的CSC群体进行活体成像。我们对最初的报告设计进行了各种修改，以生成改进的报告，该报告进一步优化了活体成像，并具有用于组织病理学的免疫染色标记。我们已经开始在乳腺癌模型系统中讨论转化生长因子-β对肿瘤干细胞生物学的影响，该模型系统显示了对转化生长因子-β的抑癌或促进进展反应。重要的是，我们已经证明，体内中和抗转化生长因子-β抗体的治疗将增加两种移植乳腺癌模型中的肿瘤干细胞数量，在该模型中，转化生长因子-β具有肿瘤抑制作用，而在转化生长因子-β具有促进进展作用的模型中，肿瘤干细胞数量减少。了解转化生长因子-β诱导癌症干细胞数量增加或减少的详细机制的研究正在进行中。我们正在使用集成的基因组方法，包括RNA-Seq和ATAC-Seq，以解决在疾病过程的早期和晚期，转化生长因子-β对CSC和非CSC间隔的不同影响的分子机制。了解CSCs在体内是如何调控的，对于开发更有效的癌症治疗方法至关重要，因为这些细胞在很大程度上对现有的治疗方法具有抵抗力。

In FY17, we have continued to address the role of TGF-beta in regulating the cancer stem cell (CSC) compartment using a novel functional imaging approach that we developed to allow visualization of this minority cell population in real time and in situ. Our lentiviral-based CSC reporter uses a synthetic promoter in which expression of a fluorescent protein is driven by the stem cell master transcription factors Oct4 and Sox2. Using this approach, we have developed methods that allow extended cell fate mapping of individual CSCs in 2D and 3D culture systems and we have an ongoing collaboration with the lab of Dr. John Condeelis at the Albert Einstein College of Medicine to perform intravital imaging of the CSC population in the primary tumor and at the lung metastatic site. We have made various modifications to our original reporter design to generate an improved reporter that is further optimized for intravital imaging and with an immunostainable tag for histopathology. We have begun to address the effect of TGF-beta on CSC biology in breast cancer model systems that show either tumor suppressor or pro-progression responses to TGF-beta. Importantly, we have shown that treatment with neutralizing anti-TGF-beta antibodies in vivo will increase the cancer stem cell population in two xenograft breast cancer models in which TGF-beta functions as a tumor suppressor, while decreasing it in a model in which TGF-beta has pro-progression effects. Studies to understand the detailed mechanisms by which TGF-beta induces either an expansion or a reduction in the cancer stem cell population are ongoing. We are employing integrated genomic approaches including RNA-Seq and ATAC-Seq to address the molecular mechanisms underlying differential effects of TGF-beta on CSC and non-CSC compartments at early and late stages of the disease process. Understanding how CSCs are regulated in vivo will be critical to development of more effective cancer therapies, as these cells are largely resistant to existing therapeutic approaches.