Reverse Engineering of Cell Senescence

细胞衰老的逆向工程

• 批准号: 10573323
• 负责人: MARC Wallace KIRSCHNER
• 金额: $ 69.39万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-15 至 2026-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10573323
• 关键词: Address; Affect; Age; Aging; Animals; Biochemical; Biological Assay; Cell Aging; Cell Culture Techniques; Cell Cycle; Cell Cycle Arrest; Cell Line; Cell Senescence Induction; Cell Size; Cell Survival; Cells; Cellular biology; Cessation of life; Clustered Regularly Interspaced Short Palindromic Repeats; Critical Pathways; Cultured Cells; DNA Damage; Dasatinib; Dependence; Development; Disease; Dryness; Event; Exposure to; FRAP1 gene; Genes; Goals; Growth; Growth Factor; Hepatocyte; Heterogeneity; Hormones; Human; Hydrogen Peroxide; Hypertrophy; Inflammatory; Insulin; Ionizing radiation; Kinetics; Laboratories; Learning; Life; Lipids; Liver; Logic; Machine Learning; Mass Spectrum Analysis; Measures; Methods; Microscopic; Microscopy; Molecular; Mus; Mutagenesis; Normal Cell; Normal tissue morphology; Oncogene Activation; Pathology; Pathway interactions; Pattern; Pharmaceutical Preparations; Pharmacologic Substance; Pharmacology; Phenotype; Phosphoproteins; Phosphorylated Peptide; Phosphorylation; Phosphorylation Site; Phosphotransferases; Physiological; Process; Proteins; Proteomics; Radiation; Reactive Oxygen Species; Research Personnel; Reverse engineering; Role; Signal Transduction; Sirolimus; Site; Small Interfering RNA; Stress; Swelling; Symptoms; Testing; Time; Tissues; cell growth; cell type; chemokine; experimental study; healthspan; insight; interest; kinase inhibitor; knock-down; mTOR inhibition; new technology; novel therapeutics; pharmacologic; phosphoproteomics; prevent; programs; proteostasis; response; senescence; stressor; tool

Abstract Cellular senescence is not just a symptom of aging, but a contributor to aging pathologies. Recent experiments in the mouse show that elimination of senescent cells (a.k.a. senolysis) can reverse several features of the aging process and extend life- and health-span. This has stimulated interest in cell senescence and in finding ways to suppress it that can be easily ported from mouse to human and made even more effective. This implies looking for such drugs as an early senolytic candidate – a kinase inhibitor dasatinib, whose mode of action is unclear. Taking senolytic therapies into human will also require a better understanding of the cell type specific progression towards senescence and diverse sub-types of senescence. In addition to aging per se cells respond to insults such as DNA damage, cell cycle arrest or oncogene activation by expressing aging phenotypes, such as hypertrophy, cell cycle arrest markers, and by secreting growth factors and chemokines, which are thought to produce degenerative responses in nearby cells. Hypertrophy may be one of the most pervasive senescent cell responses but has been hard to study, as until recently we have not had accurate enough means to measure cell mass and cell size, protein and lipid content. Yet, hypertrophy is of special interest because it is deeply connected to cell growth, which is normally under strict control in normal cells. We propose to study how cells, prompted by stressors like radiation or drugs or simply by age, become hypertrophic and how they come to express senescence markers. We have three goals: 1) to trace the progression of cell senescence in molecular terms by quantitative mass spectrometry and phospho-mass spectrometry, and in physiological terms by Raman microscopy, 2) to identify protein circuits responsible for cell senescence, and 3) to find drugs that will prevent, reverse, or eliminate senescent cells. As our tools of perturbation, we have chosen kinase inhibitors. Employing a machine learning approach, we will probe senescent cell development and senescent cell viability via a small set of well-characterized poly-specific kinase inhibitors; then regress a phenotype, such as senescence- specific signaling and senescent cell formation or death, to the activity of key kinases and their downstream circuits. Kinases implicated that way will be independently validated using siRNA knock downs and CRISPR. Additionally, extensive phospho-proteomic profiling will identify the phosphorylation sites on key proteins that most contribute to the phenotypes of senescence and which could be druggable by other means. Taken together, these methods will produce insight into the mode of action of already identified senolytic drugs and suggest new targets and new drugs for seno-therapies. This combination of pharmacology, machine learning, quantitative proteomics and new forms of microscopy can provide fresh insights into aging and suggest potential therapies for aging-related disease.

摘要 细胞衰老不仅是衰老的症状，而且是衰老病理学的一个贡献者。最近 小鼠实验显示衰老细胞（a.k.a. senolysis）可以逆转几个 它能有效地延缓衰老过程，延长寿命和健康。这激发了人们对细胞衰老的兴趣 并找到抑制它的方法，可以很容易地从老鼠移植到人类身上， 有效这意味着寻找这样的药物作为早期的衰老清除候选药物-激酶抑制剂达沙替尼， 其作用方式尚不清楚。采取senolytic疗法进入人类也将需要一个更好的 了解细胞类型特定的衰老进程和不同的衰老亚型。 除了衰老本身，细胞还对DNA损伤、细胞周期停滞或致癌基因等损伤作出反应。 通过表达衰老表型，如肥大、细胞周期阻滞标志物，以及通过分泌 生长因子和趋化因子，它们被认为在附近的细胞中产生退行性反应。 肥大可能是最普遍的衰老细胞反应之一，但一直难以研究，直到 最近，我们还没有足够精确的方法来测量细胞质量和细胞大小、蛋白质和脂质 内容然而，肥大是特别感兴趣的，因为它与细胞生长密切相关， 在正常细胞中受到严格控制。我们建议研究细胞如何受到压力刺激， 辐射或药物，或者仅仅是年龄，变得肥大，以及它们如何表达衰老标记。 我们有三个目标：1）通过定量质量在分子水平上追踪细胞衰老的进程 光谱法和磷光质谱法，以及在生理学方面通过拉曼显微镜，2）以 确定负责细胞衰老的蛋白质回路，以及3）找到能够预防，逆转，或 消灭衰老细胞作为我们的扰动工具，我们选择了激酶抑制剂。采用 通过机器学习方法，我们将通过一个小的 一组充分表征的多特异性激酶抑制剂;然后消退表型，如衰老- 特定信号传导和衰老细胞的形成或死亡，关键激酶及其下游的活性 电路.这种方式涉及的激酶将使用siRNA敲唐斯和CRISPR独立验证。 此外，广泛的磷酸化蛋白质组学分析将确定关键蛋白质上的磷酸化位点 最有助于衰老的表型，并且可以通过其他方式进行药物治疗。采取 总之，这些方法将使人们深入了解已经确定的衰老清除药物的作用方式 并为衰老疗法提供新的靶点和新药。这种结合了药理学，机器 学习，定量蛋白质组学和新形式的显微镜可以提供新的见解老化， 提示了衰老相关疾病潜在疗法。