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Development of 3-dimensional human pituitary corticotroph tumor cultures as a preclinical model for drug discovery

开发 3 维人垂体促肾上腺皮质激素肿瘤培养物作为药物发现的临床前模型

基本信息

批准号：
10448514
负责人：
ANTHONY P HEANEY
金额：
$ 34.97万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-07-09 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10448514
关键词：
3-Dimensional Adrenal Glands Architecture Automobile Driving Blood Catabolism Cell Adhesion Cell model Cells Collagen Corticotropin Development Disease Drug Interactions Drug Screening Evaluation Extracellular Matrix Genes Genetic Genomics Growth Hormone secretion Human Hydrocortisone Immune response Individual Libraries Life Maps Medical Methods Modeling Molecular Monitor Operative Surgical Procedures Pathway interactions Patients Pattern Recognition Pharmaceutical Preparations Pharmacotherapy Pituitary Gland Pituitary Gland Adenoma Pituitary Neoplasms Pituitary-dependent Cushing&apos s disease Pre-Clinical Model Primary Neoplasm Production Proteins Rare Diseases Resected Resources Screening Result Serial Passage Signal Transduction Structure System Testing Time Tissues Translational Research Tumor Tissue Validation Visualization angiogenesis biobank cheminformatics drug discovery drug sensitivity exome sequencing experience experimental study functional genomics genomic platform high throughput screening high-throughput drug screening immunocytochemistry improved individual patient inhibitor miniaturize multidisciplinary neoplastic cell novel therapeutics personalized medicine screening single-cell RNA sequencing small molecule transcriptome transcriptomics tumor

项目摘要

Abstract Cushing Disease (CD) is a life-threatening “orphan disease” caused by an adrenocorticotropic hormone (ACTH)-secreting pituitary adenoma driving excess adrenal cortisol production. There is a large unmet medical need for CD treatment. However, translational research has been greatly hampered due to unavailability of any human pituitary corticotroph tumor cell models. Using single cell RNA-sequencing (scRNAseq) and microarray transcriptome analysis of surgically resected human corticotroph tumors, we observed that loss of pituitary corticotroph tumor ACTH secretion coincided with reduced angiogenesis, survival signals and immune responses in parallel with increased collagen catabolism, cell adhesion and extracellular matrix organization. Guided by these findings, we developed a unique 3-dimensional (3D) pituitary tumor culture system and for the first time, we have been able to generate 3D human corticotroph tumor cultures that secrete ACTH >4 months. We have assembled an experienced multidisciplinary team to complete 3 focused specific aims using this first of its kind resource. Firstly, we will use whole exome sequencing to characterize the genomic landscape of our corticotroph 3D culture biobank and compare genomic and genetic fidelity between the original corticotroph tumor, normal blood and matched 3D corticotroph tumoroid cultures from the same individual patient. ScRNAseq analysis of serial passages of individual patient-derived corticotroph tumor cultures will monitor for transcriptome changes in a temporal fashion over the course of culture. The histopathological structure of our 3D corticotroph cultures at the single cell level will quantify tissue architecture so we can map corticotroph tumoroid cellular composition and distribution. A second aim will employ a miniaturized automated system to conduct a high throughput drug screen in our 3D corticotroph tumor cultures. Compounds will be subjected to rigorous evaluation to define primary “hits” and validated by re-screening in triplicate using 20 concentrations from 100µM to 20pM (2-fold dilution) to reliably calculate an EC50 for each compound. Finally, three complementary approaches, computational cheminformatic profiling, scRNAseq to delineate transcriptomic changes at the single cell level following drug treatment and functional genomics will be employed to explore the MOA of validated hit compounds. This integrated interrogation of our drug screen results and the genetic features of our patient-derived 3D tumor cultures as well as that of the original tumor tissue, will allow us to disentangle an individual drug's mode(s) of action, and directly document drug sensitivity of individualized parental corticotroph tumors. In summary, we will use our unique biobank of comprehensively molecularly characterized pituitary corticotroph tumor tissues and paired derived 3D corticotroph tumor cultures to test libraries of clincially relevant compounds. This pituitary 3D tumor culture system is transformative in the field due to the lack of any human pituitary corticotroph tumor cell models and will pave the path for much needed improved therapy for patients with this dreadfully disabling and often fatal disorder, Cushing disease.

摘要