The Metabolic Regulation of Melanoma Metastasis

摘要 大多数癌症死亡是由远处转移引起的。然而，调控远处转移的机制是 人们对此知之甚少。转移是一个非常低效的过程，在这个过程中很少有播散的癌细胞存活下来，而且 扩散的更少，但原因尚不清楚。我们开发了一种患者来源的异种移植(PDX)试验 黑色素瘤非常有效地植入并自发转移。使用这种分析方法，我们发现 不同患者黑色素瘤转移潜能的内在差异。一些第三阶段 黑色素瘤是“有效的转移者”，在患者和NSG中自发形成远处转移。 而另一些则是“低效的转移者”，不会在患者或患者体内形成远处的大转移。 NSG小鼠在相同的实验条件下。使用这种分析方法，我们发现黑色素瘤细胞 在转移过程中感受到活性氧(ROS)的激增，并且远处转移是有限的 由氧化应激引起。转移成功的细胞在转移过程中经历可逆的代谢变化 这增加了他们抵抗氧化应激的能力，包括增加对叶酸途径的依赖。 然而，不同来源的黑色素瘤转移潜能不同的机制 患者的身份尚未确定。我们假设有效和低效转移的黑色素瘤 有内在的代谢差异，可以降低有效转移者的氧化应激。一个障碍是 验证这一假设的是，患者的黑色素瘤在已知的培养条件下克隆密度很低。 条件，防止某些研究癌症代谢的方法和使用CRISPR基因编辑 (因为不能筛选或扩增单个细胞来源的克隆)。我们花了数年时间发展一种文化 来自患者的单个黑色素瘤细胞形成肿瘤器官(PDO)的介质。这一能力提高了 新陈代谢和氧化应激抵抗是否在PDX和PXs中受到类似的调控 PDO。为了解决这个问题，我们将并排比较它们的生物学特性和代谢。 PDX和PDO中高效和低效转移子的调控。我们将测试是否有效地转移 黑色素瘤具有较低的ROS水平或氧化应激标志，或增加对叶酸或戊糖的使用 磷酸盐通路与低效转移剂的比较，以及这是否促进了PDX的转移 或PDO中的迁移/入侵。我们还将测试乳酸转运体MCT1是否促进肿瘤转移和 有效的转移者是否部分通过乳酸交换来减少氧化应激。最后，我们将测试是否 在高效和低效转移者之间存在线粒体功能的内在差异 减少ROS的生成。通过比较每个黑色素瘤的生物学特性和代谢调节 PDX和PDO检测，我们将评估PDX和PDO检测的优缺点以供研究 癌症患者之间的代谢和生物学差异。这些结果有可能发现新的 调节转移的机制，癌症代谢的新方面，以及阻止进展的策略。

ABSTRACT Most cancer deaths are caused by distant metastasis. Yet the mechanisms that regulate distant metastasis are poorly understood. Metastasis is a very inefficient process in which few disseminated cancer cells survive, and even fewer proliferate, but it is not known why. We developed a patient-derived xenograft (PDX) assay in which melanomas engraft very efficiently and spontaneously metastasize. Using this assay, we discovered intrinsic differences in metastatic potential among melanomas from different patients. Some stage III melanomas are “efficient metastasizers” that spontaneously form distant metastases in patients and in NSG mice while others are “inefficient metastasizers” that do not form distant macrometastases in patients or in NSG mice under the same experimental conditions. Using this assay, we discovered that melanoma cells experience a spike in reactive oxygen species (ROS) during metastasis and that distant metastasis is limited by oxidative stress. Successfully metastasizing cells undergo reversible metabolic changes during metastasis that increase their capacity to withstand oxidative stress, including increased folate pathway dependence. However, the mechanisms that confer differences in metastatic potential upon melanomas from different patients have not yet been identified. We hypothesize that efficiently and inefficiently metastasizing melanomas have intrinsic metabolic differences that reduce oxidative stress in efficient metastasizers. One impediment to testing this hypothesis is that melanomas from patients grow poorly at clonal density in known culture conditions, preventing certain approaches for studying cancer metabolism and the use of CRISPR gene editing (because single cell-derived clones could not be screened or expanded). We spent years developing a culture medium in which single melanoma cells from patients form tumor organoids (PDOs). This capability raises the general question of whether metabolism and oxidative stress resistance are regulated similarly in PDXs and in PDOs. To address this question, we will compare, side-by-side, the biological properties and metabolic regulation of efficient and inefficient metastasizers in PDXs and PDOs. We will test if efficiently metastasizing melanomas have lower ROS levels or markers of oxidative stress, or increased use of the folate or pentose phosphate pathways, as compared to inefficient metastasizers, and whether this promotes metastasis in PDXs or migration/invasion in PDOs. We will also test if MCT1, a lactate transporter, promotes metastasis and whether efficient metastasizers reduce oxidative stress partly through lactate exchange. Finally, we will test if there are intrinsic differences in mitochondrial function between efficient and inefficient metastasizers that reduce ROS generation. By comparing the biological properties and metabolic regulation of each melanoma in PDX and PDO assays, we will assess the strengths and weaknesses of PDX and PDO assays for studying cancer metabolism and biological differences among patients. These results have the potential to identify new mechanisms that regulate metastasis, new aspects of cancer metabolism, and strategies to block progression.