"Starve the Tumor Cells to Death": SLC6A14 as a Drug Target for Colon Cancer

描述（由申请人提供）：结肠癌是美国第四大常见癌症。目前用于治疗结肠癌的策略多种多样，但没有一种能永久治愈这种疾病。迫切需要新的药物靶点来成功地对抗这种致命疾病。目前的提议集中在一个这样的药物靶标上。它是一种营养转运体，被称为ATB0 +，由SLC6A14基因编码，运输所有必需氨基酸和谷氨酰胺。这种转运蛋白仅在正常组织中低水平表达，但在结肠癌中被诱导数倍。我们假设肿瘤细胞上调ATB0，+以满足其对必需氨基酸和谷氨酰胺不断增加的需求。由于ATB0 +转运所有必需氨基酸和谷氨酰胺，因此肿瘤细胞只需打开一个基因即可满足其对氨基酸的需求。由于肿瘤细胞依赖ATB0 +作为必需氨基酸和谷氨酰胺的主要来源，阻断转运蛋白功能可能为癌症治疗提供一种合乎逻辑的策略。为了验证这一策略，我们提出了三个具体目标。目的1：在裸鼠异种移植瘤模型中，评估基于shrna沉默ATB0，+对结肠癌细胞系肿瘤生长和进展的影响。在这里，我们将使用分子生物学方法沉默结肠癌细胞系中的转运蛋白，然后检查转运蛋白的敲除对肿瘤生长和进展的影响，当这些细胞异种移植到裸鼠中时。目的2是通过比较一种致癌物诱导的肿瘤的形成/进展来评估ATB0 +缺失对结肠癌的影响。偶氮氧甲烷用于诱导小鼠结肠癌。这是一种删除转运蛋白编码基因的遗传方法。我们预测，转运蛋白敲除小鼠将对偶氮甲烷诱导的结肠癌产生抗性。目的3是评估1-甲基- l-色氨酸（1-MLT）阻断ATB0，+对两种不同结肠癌小鼠模型肿瘤生长的影响：人类结肠癌细胞系异种移植小鼠模型和致癌物诱导结肠癌小鼠模型。这是一种阻断转运蛋白功能的药理学方法。我们将在这些动物模型中使用1-MLT作为ATB0，+的阻滞剂来评估其预防结肠癌的能力。1-MLT也是吲哚胺双加氧酶（IDO）的有效抑制剂。已知IDO的抑制可以增强免疫系统对肿瘤的活性。因此，我们预测1-MLT将通过两种不同的机制在体内发挥抗癌作用：通过阻断肿瘤特异性营养转运体ATB0 +，从而使肿瘤细胞缺乏必需氨基酸，也通过抑制IDO，从而增强免疫系统对肿瘤的活性。这些研究将为癌症特异性氨基酸转运体ATB0，+治疗结肠癌的潜力提供重要信息。

DESCRIPTION (provided by applicant): Colon cancer is the fourth most common cancer in this country. A wide variety of strategies are currently used for treatment of colon cancer, but none of them offers a permanent cure for the disease. New drug targets are sorely needed to succeed in the fight against this deadly disease. The current proposal focuses on one such drug target. It is a nutrient transporter, known as ATB0,+ encoded by the gene SLC6A14, which transports all essential amino acids as well as glutamine. This transporter is expressed only at low levels in normal tissues, but is induced several-fold in colon cancer. We hypothesize that tumor cells up-regulate ATB0,+ to meet their increasing demand for essential amino acids and glutamine. Since ATB0,+ transports all essential amino acids as well as glutamine, it is sufficient for the tumor cells to turn on a single gene to satisfy their amino acid needs. Since tumor cells rely on ATB0,+ as the primary source of essential amino acids and glutamine, blocking the transporter function may offer a logical strategy for cancer treatment. To validate this strategy, we propose three specific aims. Aim 1 is to evaluate the effects of shRNA-based silencing of ATB0,+ on the growth and progression of tumor with colon cancer cell lines in the nude mouse xenograft model. Here we will silence the transporter in colon cancer cell lines using a molecular biological approach and then examine the effects of the knockdown of the transporter on the growth and progression of tumor when these cells are xenografted into nude mice. Aim 2 is to evaluate the effects of ATB0,+ deletion on colon cancer by comparing the formation/progression of tumors induced by a carcinogen. Azoxymethane is used to induce colon cancer in mice. This is a genetic approach to delete the gene coding for the transporter. We predict that the transporter knockout mouse will be resistant to azoxymethane-induced colon cancer. Aim 3 is to evaluate the effects of pharmacological blockade of ATB0,+ with 1-methyl-L- tryptophan (1-MLT) on tumor growth in two different mouse models of colon cancer: a xenograft mouse model with human colon cancer cell lines and a carcinogen-induced colon carcinoma mouse model. This is a pharmacological approach to block the function of the transporter. We will use 1-MLT as a blocker of ATB0,+ to evaluate its ability to prevent colon cancer in these animal models. 1-MLT is also a potent inhibitor of indoleamine dioxygenase (IDO). Inhibition of IDO is known to potentiate the activity of immune system against tumors. Therefore, we predict that 1-MLT will function as an anti-cancer agent in vivo by acting through two different mechanisms: by blocking the tumor-specific nutrient transporter ATB0,+ thereby starving the tumor cells of essential amino acids, and also by inhibiting IDO thereby enhancing the activity of the immune system against the tumor. These studies will generate important information regarding the therapeutic potential of the cancer-specific amino acid transporter ATB0,+ for the treatment of colon cancer. PUBLIC HEALTH RELEVANCE: The goal of this project is to evaluate the therapeutic potential of a cancer-specific amino acid transporter, known as SLC6A14 or ATB0,+ in the treatment of colon cancer. Studies will involve the use of a xenograft mouse model with human colon cancer cell lines and a carcinogen- induced mouse model of colon cancer. The potential of the transporter as a drug target for treatment of colon cancer will be evaluated in these models with two approaches: a genetic approach with the deletion of ATB0,+ (Slc6a14-/- mouse) and a pharmacological approach with the use of a chemical blocker of ATB0,+