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Substrate Specificity Determinants in Nutrient Solute Carrier Transporters

营养溶质载体转运蛋白的底物特异性决定因素

基本信息

批准号：
10735432
负责人：
Avner Schlessinger
金额：
$ 43.78万
依托单位：
ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-07-10 至 2028-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10735432
关键词：
Alanine Allosteric Site Amino Acid Transporter Amino Acids Antiviral Agents Autoimmune Diseases Basic Amino Acid Transport Systems Binding Binding Sites Biochemical Biochemical Pathway Biological Biological Assay Biology Biomass Biophysics Cancer cell line Cell Death Cell Differentiation process Cell Line Cell Proliferation Cells Cellular Metabolic Process Chemicals Collaborations Computing Methodologies Coupled Cryoelectron Microscopy Cysteine Data Development Diabetes Mellitus Disease Docking Drug Delivery Systems Drug Design Drug Targeting Electrophysiology (science)Electrostatics Family Free Energy Future Glutamine Goals Grant Health Human Hydrophobicity Hyperactivity Ion Transport Ions Kinetics Knowledge Laboratories Leucine Libraries Ligands Malignant Neoplasms Malignant neoplasm of prostate Mediating Membrane Membrane Transport Proteins Metabolic Diseases Metabolism Methods Modeling Molecular Molecular Conformation Myocardial Ischemia Neurotransmitters Nutrient Organic Synthesis Pathology Pharmaceutical Preparations Pharmacology Physiological Play Prodrugs Proliferating Proteins Role Serine Shapes Signaling Molecule Site-Directed Mutagenesis Specificity Starvation Structural Models Structure Structure-Activity Relationship Substrate Specificity Sugar Acids Synthesis Chemistry T-Lymphocyte Testing Toxin Work amino acid metabolism analog biophysical techniques cancer cell cancer type computational chemistry design drug discovery inhibitor innovation melanoma neoplastic cell nervous system disorder novel novel strategies rapid growth rational design small molecule solute structural biology therapeutic target tool triple-negative invasive breast carcinoma uptake virtual

项目摘要

SUMMARY Alterations in cell metabolism support rapid growth and proliferation of cells in pathologies such as cancer, au- toimmune disease, and heart ischemia, resulting in increased reliance on the metabolism of amino acids such as glutamine and leucine. Nutrient Solute Carrier (SLC) transporters play a major role in reprogrammed meta- bolic networks by supplying cells with nutrients that are used to build biomass, serve as signaling molecules that enhance cell proliferation and differentiation, or regulate cell death. Our broad goal is to describe the sub- strate and inhibitor specificity determinants in disease-related nutrient SLC transporters and develop unique strategies to modulate their functions. We take an integrative approach that includes computational chemistry methods, coupled with biochemical and biophysical approaches and disease-related cell lines, to char- acterize two amino acid transporters that play a key role in metabolism of rapidly-growing cells: the Alanine- Serine-Cysteine Transporter (SLC1A5, ASCT2), a Na+-dependent amino acid exchanger that modulates intra- cellular glutamine levels, and the Amino Acid Transporter B0+ (ATB0+, SLC6A14), a neutral and cationic amino acid transporter, driven by Na+ and Cl- co-transport. In Aim 1 of this project, we will continue characterizing ASCT2, a well-validated drug target for various patholo- gies (eg triple negative breast cancer and prostate cancer). Despite recent advancements in our understanding of ASCT2 structure and function made by us and others, many aspects of its biology are highly unexplored. We will rationally design chemical tools that modulate the activity of ASCT2 using unique mechanisms, including: (A) allosteric inhibitors interacting with a recently identified allosteric site; (B) covalent inhibitors targeting a unique cysteine residue in the substrate binding site of ASCT2; and (C) conformation-specific small molecule modulators targeting specific subpockets in the substrate binding site. In Aim 2, we will characterize SLC6A14, an understudied transporter involved in cancer and metabolic diseases. We will develop structural models of SLC6A14 in different conformations. We will describe biophysical features of the models’ substrate binding site, including electrostatic potential, size, shape, and hydrophobicity, to develop hypotheses for the substrate and inhibitor specificity determinants in SLC6A14. We will use this knowledge to guide the development of inhibitors and substrates, including photoactivatable compounds, to directly test inhibitor-binding site interaction(s). Successful completion of this project will provide a greater understanding of mechanisms of transport and inhi- bition of nutrient transporters, as well as novel chemical tools to further characterize their role in disease. Notably, we will test an emerging and innovative approach to transporter drug discovery that targets allosteric modulation and covalent inhibition via small molecules, to deprive hyper-proliferating cells of nutrients, potentially expanding future applications to treat other diseases that involve SLCs.

概括细胞代谢的改变支持癌症、癌症等疾病中细胞的快速生长和增殖。免疫性疾病和心脏缺血，导致对氨基酸代谢的依赖增加，例如如谷氨酰胺和亮氨酸。营养溶质载体（SLC）转运蛋白在重编程元- 代谢网络通过为细胞提供用于构建生物质的营养物质，作为信号分子增强细胞增殖和分化，或调节细胞死亡。我们的总体目标是描述子疾病相关营养 SLC 转运蛋白的策略和抑制剂特异性决定因素并开发调节其功能的独特策略。我们采取综合方法，包括计算化学方法，加上生物化学和生物物理方法以及与疾病相关的细胞系，以表征表征两种氨基酸转运蛋白，它们在快速生长细胞的新陈代谢中发挥关键作用：丙氨酸- 丝氨酸-半胱氨酸转运蛋白（SLC1A5、ASCT2），一种 Na+ 依赖性氨基酸交换剂，可调节内部细胞谷氨酰胺水平，以及氨基酸转运蛋白 B0+（ATB0+、SLC6A14）（一种中性阳离子氨基）酸转运蛋白，由 Na+ 和 Cl- 共同转运驱动。在该项目的目标 1 中，我们将继续表征 ASCT2，这是一种经过充分验证的药物靶点，适用于各种病理学（例如三阴性乳腺癌和前列腺癌）。尽管我们的理解最近取得了进展尽管我们和其他人对 ASCT2 的结构和功能进行了研究，但其生物学的许多方面尚未得到探索。我们将合理设计化学工具，利用独特的机制调节 ASCT2 的活性，包括： (A) 与最近确定的变构位点相互作用的变构抑制剂； (B) 共价抑制剂 ASCT2底物结合位点中独特的半胱氨酸残基； (C) 构象特异性小分子调节剂靶向底物结合位点中的特定子口袋。在目标 2 中，我们将表征 SLC6A14，一种与癌症和代谢疾病有关的尚未被研究的转运蛋白。我们将开发结构模型 SLC6A14 具有不同的构象。我们将描述模型底物结合位点的生物物理特征，包括静电势、尺寸、形状和疏水性，以提出基材和 SLC6A14 中的抑制剂特异性决定因素。我们将利用这些知识来指导抑制剂的开发和底物，包括光活化化合物，以直接测试抑制剂结合位点相互作用。该项目的成功完成将加深对运输和抑制机制的了解。营养转运蛋白的咬合，以及新的化学工具，以进一步表征其在疾病中的作用。尤其，我们将测试一种针对变构调节的新兴创新转运蛋白药物发现方法和通过小分子的共价抑制，剥夺过度增殖细胞的营养，从而可能扩大未来用于治疗涉及 SLC 的其他疾病的应用。