Elucidating cancer-intrinsic mechanisms of perineural invasion in pancreatic cancer

阐明胰腺癌神经周围浸润的癌症内在机制

基本信息

批准号：
10428889
负责人：
William L Hwang
金额：
$ 31.51万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-07-01 至 2027-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10428889
关键词：
Address Adrenergic beta-Antagonists Advisory Committees Alleles Area Biochemical CRISPR-mediated transcriptional activation Cancer Biology Cancer Etiology Candidate Disease Gene Cell Nucleus Cells Cessation of life Coculture Techniques Collaborations Committee Members Critical Pathways Custom Data Development Development Plans Epithelial Excision Fibroblasts Formalin Frequencies Future Gene Expression Gene set enrichment analysis Genes Genetic Heterogeneity Genetically Engineered Mouse Goals Guide RNA Heat-Shock Response Human Immune Immunofluorescence Immunologic Immunohistochemistry Invaded Investigation Kinetics Knock-in Knowledge Libraries Link Literature Machine Learning Malignant - descriptor Malignant Neoplasms Malignant neoplasm of pancreas Maps Mediating Mediator of activation protein Mentors Mesenchymal Metabolic Molecular Morbidity - disease rate Nature Neoplasm Metastasis Nerve Nerve Fibers Neuroglia Neurons Non-Malignant Organoids Outcome Pain Pancreas Transplantation Pancreatic Ductal Adenocarcinoma Paraffin Embedding Pathologic Pathologic Processes Patients Peripheral Nerves Phenotype Physicians Play Principal Investigator Prognosis Property Proteomics Recruitment Activity Recurrence Research Research Proposals Resolution Role SEMA3E gene Sampling Scientist Small Nuclear RNA Synaptic plasticity Techniques Tissue Microarray Tissue-Specific Gene Expression Tumor Biology United States Universities Work axon guidance base cancer cell cancer initiation career development cell type cohort density digital epidemiology study experience glial cell-line derived neurotrophic factor human data immunoregulation improved in vivo in vivo Model live cell imaging lymphatic Invasion machine learning classifier medical schools mortality mouse model multiplexed imaging nano-string neoplastic neoplastic cell neurogenesis neuronal tumor neurotransmission novel organoid transplantation perineural preclinical study preservation programs recruit spatial relationship symposium targeted treatment therapeutic development therapy development therapy resistant transcriptome transcriptome sequencing transcriptomics transplant model tumor tumor microenvironment tumorigenesis tumorigenic

项目摘要

PROJECT SUMMARY/ABSTRACT Pancreatic ductal adenocarcinoma (PDAC) is projected to become the second leading cause of cancer death in the United States by 2030. One of the differentiating hallmarks of PDAC is an exceptionally high frequency of perineural invasion (PNI), a histopathologic manifestation of tumor-nerve crosstalk whereby cancer cells recruit, migrate towards and invade peripheral nerves. These tumor-nerve interactions may be biochemical, electrical, and metabolic in nature and involve both neoplastic and non-neoplastic cells. While it has become apparent that intra-tumoral nerves play an important role in cancer initiation, progression, recurrence, treatment resistance, metastasis, and survival, the diverse molecular mechanisms underlying tumor-nerve crosstalk remain largely unknown. To address this gap in knowledge, this proposal aims to perform a comprehensive spatially-resolved whole transcriptome screen in human PDAC to identify cell-type specific genes linked to PNI and dissect their functional roles by applying CRISPR activation (CRISPRa) in a novel orthotopic organoid transplant model. It is anticipated that this work will uncover a set of previously unknown mediators of PNI and provide a high-resolution molecular interconnectivity map of the tumor-nerve interactome, which will guide therapeutic development. Prior work studying molecular mediators of PNI using bulk or single-cell/nucleus RNA-sequencing lacked spatial context. The first aim applies a transcriptomic and proteomic digital spatial profiling (DSP; Nanostring) approach optimized for formalin-fixed paraffin-embedded sections of human PDAC to preserve the spatial relationships among different cell types/states. By performing DSP on tissue microarrays created from a custom cohort of PDAC patients with tumors spatially annotated for regions with and without PNI, the ability to extract gene expression differences that associate with PNI isolated from inter-patient genetic heterogeneity is greatly enhanced. The second aim will investigate the potential causal relationships between a shortlist of ~20 candidate genes and the development of PNI by performing an arrayed in vivo CRISPRa screen involving orthotopically transplanting pancreatic cancer organoids expressing single-guide RNAs (sgRNAs) targeting one or more candidate genes (KrasLSL-G12D/+;Trp53FL/FL;Rosa26LSL-dCas9-VPR-mNeonGreen;sgRNA) into syngeneic recipients. The density and properties of intra-tumoral nerves/PNI will be assessed using multiplexed imaging and machine learning approaches. The long-term goal of this research is the successful development of therapeutics that disrupt adaptive tumor-nerve crosstalk in the tumor microenvironment. The research proposal is part of an extensive career development plan that includes formal educational opportunities and seminar/conference presentations to enable the principal investigator to become an independent physician-scientist in translational cancer biology. A diverse and experienced team of mentors, scientific advisory committee members, and collaborators at MIT, MGH, DFCI, HMS, and Columbia University School of Medicine is committed to supporting and guiding the principal investigator toward achieving this goal.

项目概要/摘要胰腺导管腺癌（PDAC）预计将成为癌症死亡的第二大原因到 2030 年，美国将实现这一目标。PDAC 的特色之一是异常高的频率神经周围侵袭（PNI），肿瘤-神经串扰的一种组织病理学表现，癌细胞通过这种方式募集，向周围神经迁移并侵入。这些肿瘤-神经相互作用可能是生化的、电的、本质上是代谢和代谢，涉及肿瘤细胞和非肿瘤细胞。虽然已经很明显的是瘤内神经在癌症的发生、进展、复发、治疗抵抗、转移和生存，肿瘤-神经串扰背后的多种分子机制在很大程度上仍然存在未知。为了解决这一知识差距，该提案旨在执行全面的空间解析在人类 PDAC 中进行全转录组筛选，以识别与 PNI 相关的细胞类型特异性基因并剖析其通过在新型原位类器官移植模型中应用 CRISPR 激活 (CRISPRa) 发挥功能作用。这是预计这项工作将揭示一组以前未知的 PNI 介体，并提供高分辨率肿瘤-神经相互作用组的分子互连图，这将指导治疗的发展。先前使用批量或单细胞/核 RNA 测序研究 PNI 分子介质的工作缺乏空间语境。第一个目标应用转录组和蛋白质组数字空间分析（DSP；Nanostring）方法针对人类 PDAC 的福尔马林固定石蜡包埋切片进行优化，以保留空间关系不同细胞类型/状态之间。通过对由定制队列创建的组织微阵列执行 DSP 患有肿瘤的 PDAC 患者对有或没有 PNI 的区域进行空间注释，提取基因的能力从患者间遗传异质性中分离出的与 PNI 相关的表达差异很大增强。第二个目标将调查约 20 个入围名单之间的潜在因果关系通过进行阵列体内 CRISPRa 筛选来开发候选基因和 PNI，其中涉及原位移植胰腺癌类器官，表达单向导RNA（sgRNA），靶向一种或更多候选基因（KrasLSL-G12D/+；Trp53FL/FL；Rosa26LSL-dCas9-VPR-mNeonGreen；sgRNA）进入同基因收件人。将使用多重成像评估肿瘤内神经/PNI 的密度和特性和机器学习方法。本研究的长期目标是成功开发破坏肿瘤微环境中适应性肿瘤神经串扰的疗法。该研究计划是广泛职业发展计划的一部分，其中包括正规教育使首席研究员成为一名转化癌症生物学领域的独立医师科学家。多元化且经验丰富的导师团队，科学顾问委员会成员以及麻省理工学院、麻省总医院、DFCI、HMS 和哥伦比亚大学的合作者医学院致力于支持和指导主要研究者实现这一目标。