S1P Lyase in colon cancer

描述(由申请人提供)： 膳食中的神经鞘脂，如神经酰胺和鞘氨醇，可促进细胞凋亡，防止肠道肿瘤的发生。然而，这些鞘磷脂可以在肠道细胞内转化为鞘氨醇-1-磷酸(S1P)，一种强大的有丝分裂素和血管生成因子。因此，肠上皮细胞中这些鞘磷脂代谢物之间存在着微妙的平衡，其变化可能会影响肠道肿瘤的发生。S1P被S1P裂解酶(SPL)不可逆地降解，S1P裂解酶促进细胞凋亡，是最大限度的凋亡反应所必需的。SPL在肠道上皮细胞中高表达，维持较低的S1P水平，促进正常的细胞更新。然而，我们发现SPL在小鼠早期息肉和人类结肠癌中表达下调。我们假设SPL下调是肠道肿瘤发生的早期事件，代表了一种关键的遗传变化，导致了有利于S1P在肠粘膜积聚的生化开关，从而激活了有助于肿瘤进展的有丝分裂和血管生成信号。为了测试这种可能性，我们提出了四个相互关联的特定目标：1.表征伴随着肠道肿瘤发生的S1P代谢的变化。我们假设，导致S1P积聚的遗传和/或表观遗传变化是肠道肿瘤发生的常见和早期事件。比较不同时期人肠息肉、结直肠癌、小鼠息肉及相应未受累组织中鞘磷脂水平及S1P代谢相关酶的活性、表达和定位；2.探讨SPL在体外对永生化和恶性结肠上皮细胞生长特性的影响。SPL在恶性和非恶性结肠上皮细胞系中的表达将通过siRNA、腺病毒和稳定表达系统来调节。将确定SPL对细胞增殖、凋亡、迁移和致瘤性的影响及其作用机制。3.确定SPL下调是否会增加肠道致瘤性并改变对膳食鞘脂的反应。将利用已有的SPL基因敲除小鼠模型进入Min小鼠背景，探讨以下假设：小鼠肠道组织中SPL表达下调和S1P水平升高将增加肠息肉的发病率、发病率、生长、血管和侵袭性，并限制或逆转膳食鞘磷脂的化学预防作用；4.确定SPL下调在肠道肿瘤发生的早期是否可逆。我们将研究SPL下调是表观遗传学改变的结果的假说，该变化涉及SPL基因组位点上的DNA甲基化。拟议的研究计划应该确定SPL下调是否是促进肠道肿瘤发生的早期和可逆事件。通过完成拟议的研究，我们将更接近我们的长期目标，即在肿瘤发生的早期采用化学预防措施来恢复微妙的鞘磷脂代谢平衡。项目叙述结肠癌是男性和女性第二常见的癌症，也是导致癌症死亡的第三大原因。据估计，美国每年用于治疗这种疾病的费用为84亿美元。识别疾病发生和发展的分子和环境决定因素是一个关键目标，这可能有助于揭示新的生物标志物，用于早期检测和治疗干预，以帮助50%或更多的晚期或难治性疾病患者。如果我们的中心假设是正确的，S1P代谢基因的状态可能成为预测结果以及对治疗和饮食方案的反应的信息生物标志物。我们的研究计划应该确立这一假说的有效性，并提供实现我们最终目标所需的信息，即识别和验证绕过或逆转鞘磷脂代谢开关并恢复癌前和/或恶性细胞中正常S1P代谢的新治疗方法。

DESCRIPTION (provided by applicant): Dietary sphingolipids such as ceramide and sphingosine promote apoptosis and protect against intestinal tumorigenesis. However, these sphingolipids can be converted within intestinal cells to sphingosine-1- phosphate (S1P), a potent mitogen and angiogenic factor. Thus, a delicate balance exists between these sphingolipid metabolites in gut epithelial cells, alterations of which may influence intestinal tumorigenesis. S1P is irreversibly degraded by the enzyme S1P lyase (SPL), which promotes apoptosis and is required for maximal apoptotic responses. SPL is highly expressed in gut epithelium, where it maintains low S1P levels and promotes normal cell turnover. However, we have found that SPL is downregulated in early Min mouse polyps and in human colon cancer. We hypothesize that SPL downregulation is an early event in intestinal tumorigenesis, representing a critical genetic change that leads to a biochemical switch favoring S1P accumulation in the intestinal mucosa, thereby activating mitogenic and angiogenic signals that contribute to tumor progression. To test this possibility, we have proposed four interrelated specific aims: 1. To characterize the changes in S1P metabolism that accompany intestinal tumorigenesis. We hypothesize that genetic and/or epigenetic changes leading to S1P accumulation are common and early events in intestinal tumorigenesis. Sphingolipid levels and the activity, expression and localization of enzymes involved in S1P metabolism will be compared in human intestinal polyps, CRC, Min mouse polyps of different stages and corresponding uninvolved tissues; 2. To establish whether SPL expression affects the growth characteristics of immortalized and malignant colonic epithelial cells in vitro. SPL expression will be modulated in malignant and nonmalignant colon epithelial cell lines using siRNA, adenoviral and stable expression systems. Effects on proliferation, apoptosis, migration and tumorigenicity and mechanism of SPL action will be determined. 3. To establish whether SPL downregulation increases intestinal tumorigenicity and alters the response to dietary sphingolipids. Available SPL knockout mouse models crossed into the Min mouse background will be used to explore the hypothesis that reduced SPL expression and increased S1P levels in mouse intestinal tissues will enhance the rate, incidence, growth, vascularity and invasive characteristics of intestinal polyposis and limit or reverse the chemopreventive effect of dietary sphingolipids; 4. To determine whether SPL downregulation is reversible early in intestinal tumorigenesis. We will investigate the hypothesis that SPL downregulation is the result of epigenetic changes involving DNA methylation at the SPL genomic locus. The proposed research plan should establish whether SPL downregulation is an early and reversible event that contributes to intestinal tumorigenesis. In accomplishing the proposed studies, we will move closer to our long term goal of employing chemopreventive measures to restore the delicate sphingolipid metabolic balance early in the process of tumorigenesis.Project Narrative Colon cancer is the second most common cancer of men and women and the third leading cause of cancer mortality. It is estimated that $8.4 billion per year is spent caring for patients with this disease in the US. Identifying the molecular and environmental determinants of disease incidence and progression is a critical goal that may help to reveal novel biomarkers for earlier detection and therapeutic intervention to help the 50% or more of patients suffering from advanced or refractory disease. If our central hypothesis is correct, the status of S1P metabolic genes may become informative biomarkers in predicting outcome and the response to therapeutic and dietary regimens. Our Research Plan should establish the validity of this hypothesis and provide information necessary for achieving our ultimate goal, which is to identify and validate novel therapeutic approaches that bypass or reverse the sphingolipid metabolic switch and restore normal S1P metabolism in premalignant and/or malignant cells.