Gene Expression Analysis of Dog Natural Killer Cells as Immunotherapy Target

项目摘要 狗是体型较大的远缘繁殖动物，在完整的免疫系统存在的情况下会自发发生癌症。 以及在与人类共享环境暴露的背景下。人类和狗也共享一对 导致犬类和人类基因组和微生物群之间更大相似性的进化史 而不是在老鼠和人之间。总而言之，这些特点使狗成为一种有利的翻译模式 学习癌症免疫学和癌症免疫疗法。狗的临床试验使复杂免疫的研究成为可能 在治疗过程中的相互作用，同时也涉及癌症免疫疗法的长期疗效和毒性。 然而，免疫解剖需要开发可靠和有效的分析和试剂，而且 使用狗免疫疗法进行影响研究为时过早，直到对狗免疫学和 免疫生物标记物的实现，特别是对自然杀伤(NK)细胞。这项提议将建立在我们激动人心的基础上 犬NK细胞特征的初步数据，包括过继转移犬NK细胞的首个临床试验 吸入IL-15的细胞和新的细胞因子传递。在与医学院的合作下， 兽医，以及数据密集型生物实验室，进行跨物种基因组，转录， 和元基因组序列分析，我们将检查狗NK差异基因的表达 刺激和稳定条件；2)来自狗血和肿瘤生物标本；以及3)来自响应 在一项调查性狗临床试验中，用吸入的IL-15治疗无反应的狗。我们将利用多个 应用高通量测序技术(RNAseq和单细胞RNA测序)建立 来自体内和体外的静息和激活的狗NK细胞更强大和更有效的基因特征 进一步分析狗肉瘤浸润性NK细胞的条件。然后我们将比较这些基因签名 从小鼠和人类NK细胞的高通量测序数据，以弥合物种差异和 促进临床翻译。最终，关于犬NK细胞在免疫防御和癌症中的作用的知识 治疗是有限的，目前还不清楚这是由于不同物种之间的生物学差异还是有限 用于调查的试剂。这项提案的结果将扩展犬类研究提供的重要联系 在小鼠临床前研究和人类临床试验之间，一般用于癌症免疫治疗研究，但 尤其是对于NK研究，其高影响力的临床翻译仍然难以捉摸。

Project Summary Dogs are large, outbred animals that develop cancer spontaneously in the presence of an intact immune system and in the setting of shared environmental exposures with humans. Humans and dogs also share a paired evolutionary history which has led to greater similarities between canine and human genomes and microbiomes than between mouse and human. Together, these traits make dogs an advantageous translational model to study cancer immunology and cancer immunotherapy. Dog clinical trials allow for the study of complex immune interactions during therapy while also addressing long-term efficacy and toxicity of cancer immunotherapies. However, immune dissection requires the development of robust and validated assays and reagents, and high impact studies using dog immunotherapy are premature until a deeper understanding of dog immunology and immune biomarkers is realized, especially for natural killer (NK) cells. This proposal will build on our exciting preliminary data characterizing dog NK cells, including first-in-dog clinical trials of adoptive transfer of dog NK cells and novel cytokine delivery with inhaled IL-15. In collaboration with the School of Medicine, the School of Veterinary Medicine, and the Data Intensive Biology Lab which performs cross-species genomic, transcriptomic, and metagenomic sequence analyses, we will examine dog NK differential gene expression across 1) multiple stimulatory and steady-state conditions; 2) from dog blood and tumor bio-specimens; and 3) from responding and non-responding dogs treated with inhaled IL-15 on an investigational dog clinical trial. We will utilize multiple applications of high-throughput sequencing technologies (RNASeq and single cell RNA sequencing) to establish more robust and validated gene signatures of resting and activated dog NK cells from both ex vivo and in vivo conditions with further analysis of dog sarcoma-infiltrating NK cells. We will then compare these gene signatures to data from high throughput sequencing of murine and human NK cells to bridge species differences and facilitate clinical translation. Ultimately, knowledge of the role of canine NK cells in immune defense and cancer therapy is limited, and it remains unclear whether this is due to biological differences across species or limited reagents for investigation. Results from this proposal will extend the important link that canine studies provide between murine pre-clinical studies and human clinical trials, for cancer immunotherapy research in general but especially for NK studies where high impact clinical translation has remained elusive.