Targeting Serine and Glycine Metabolic Vulnerabilities in Melanoma Brain Metastasis

项目摘要 脑转移瘤（BM）是黑色素瘤死亡率的主要原因。在44%的阶段中发现BM IV型黑色素瘤患者的中位生存期为4个月，5年生存率低于20%。 脑转移性黑色素瘤的不良预后部分是由于缺乏提供有效治疗的疗法。 持久的反应。目前还没有专门针对脑转移瘤的治疗方法。大脑 相对于血浆，微环境是独特的，因为它具有低浓度的氨基酸。 其中变化最大的氨基酸是丝氨酸和甘氨酸。丝氨酸的血浆浓度为114 µM 并在脑脊液（CSF）中降至24 µM，这是大脑中氨基酸浓度的代表。 甘氨酸水平也从血浆中的232 µM降至CSF中的5 µM。我们制造了模拟脑脊液的介质 丝氨酸和甘氨酸浓度。 我的初步数据显示，两种人类黑色素瘤细胞系，A375和SK-MEL-28， 丝氨酸和甘氨酸的合成。这表明细胞变得依赖于合成 丝氨酸和甘氨酸，并且可以靶向降低黑素瘤细胞的生存力。第一和限速 丝氨酸合成的步骤由磷酸甘油酸脱氢酶（PHGDH）催化，并且可以使用 小分子抑制剂，PH 719和PH 755。甘氨酸是由丝氨酸经丝氨酸羟甲基转移酶转化而成的 1/2（SHMT 1/2），并且可以被小分子抑制剂SHIN 1和AGF 347靶向。我最初的实验 证明黑色素瘤细胞对CSF中的这些丝氨酸和甘氨酸合成抑制剂敏感 条件这些数据表明，用PHGDH或SHMT 1/2抑制剂治疗的黑色素瘤BM小鼠 可能降低脑肿瘤负荷，这可能为BM患者提供一个新的靶点， 否则治疗选择有限。为了确定体外数据是否适用于体内，我们将 通过心内注射GFP-荧光素酶标记黑色素瘤细胞系建立黑色素瘤脑转移 NCr-Foxn 1 nu（裸）小鼠的心脏中。这些细胞在小鼠中的生长将使用 生物发光成像（IVIS）。我们将使用强力霉素诱导的PHGDH和SHMT 1/2缺失， 确定这些酶的缺失是否会减少脑损伤。 这项研究的结果将提供有关丝氨酸和甘氨酸的含义的知识， 合成途径，并提供新的治疗靶点， BM改善患者结局。

Project Summary Brain metastases (BM) are a major contributor to mortality in melanoma. BM are found in 44% of Stage IV melanoma patients resulting in a median survival of 4 months and a 5-year survival rate of less than 20%. The poor prognosis of brain metastatic melanoma is due in part to the lack of therapies that provide an effective and durable response. There are no therapies specifically designed to target brain metastases. The brain microenvironment, relative to blood plasma, is unique because it has low concentrations of amino acids. Amongst the most changed amino acids are serine and glycine. The plasma concentration of serine is 114 µM and decreases to 24 µM in the cerebrospinal fluid (CSF), a proxy for amino acid concentrations in the brain. Glycine levels also decrease from 232 µM in plasma to 5 µM in CSF. We have made media that mimics CSF serine and glycine concentrations. My preliminary data shows that two human melanoma cell lines, A375 and SK-MEL-28, have increased serine and glycine synthesis in CSF environments. This suggests that cells become dependent on synthesizing serine and glycine and can be targeted to decrease the viability of melanoma cells. The first and rate-limiting step of serine synthesis is catalyzed by phosphoglycerate dehydrogenase (PHGDH) and can be inhibited using small molecule inhibitors, PH719 and PH755. Glycine is made from serine by serine hydroxy methyltransferase 1/2 (SHMT1/2) and can be targeted by the small molecule inhibitors SHIN1 and AGF347. My initial experiments demonstrate that melanoma cells are sensitized to these serine and glycine synthesis inhibitors in CSF conditions. These data suggest that mice with melanoma BM treated with either PHGDH or SHMT1/2 inhibitors may have decreased brain tumor burden and that this could provide a novel target for patients with BM who otherwise have limited therapeutic options. To determine if our in vitro data are applicable in vivo, we will establish melanoma brain metastases by intracardiac injection of GFP-luciferase-labeled melanoma cell lines into the hearts of NCr-Foxn1nu (Nude) mice. The growth of these cells in mice will be monitored using bioluminescence imaging (IVIS). We will use doxycycline inducible deletion of PHGDH and SHMT1/2 to determine if upon deletion of these enzymes there a reduction in brain lesions. The findings from this study will provide knowledge about the implications of the serine and glycine synthesis pathway in amino acid depleted environments like the brain and provide novel therapeutic targets for BM to improve patient outcomes.