Effect of Cyclical Intermittent Hypoxia on Lung Cancer Progression

目的：肺癌的预后仍然很差，退伍军人的发病率更高。 与平民相比，肺癌和更糟糕的预后1.阻塞性睡眠呼吸暂停(OSA)与 随着更高的癌症发病率2-5和死亡率2，6-8和再次，退伍军人有更高的OSA患病率，即 9我们的首要目标是确定周期性间歇性缺氧如何 (CIH)是OSA的主要潜在病理，促进肺癌的进展。我们的假设是 增加未成熟免疫细胞向抑制性免疫细胞的分化，然后在无意中， 保护癌症。我们已经发表了CIH加速原发肺癌进展的三倍 转基因KrasG12D+；p53fl/fl；肉豆蔻化p110fl/fl Rosa-GFP(TT-KPP)小鼠(由VA CPPF赠款资助)。这 格兰特是下一步--探索CIH如何促进癌症进展的机制。研究设计： 这项资助的技术创新之处在于，我们将率先测量轻度、中度和 严重脑出血对TT-KPP小鼠肿瘤进展的影响(目标1)。这笔赠款的概念创新在于，我们 将率先使用这一TT-KPP模型来探索系统性CIH是否会增加生产、分化 抑制型免疫细胞在原发肺癌中的招募。方法：目的1：TT-KPP小鼠 注射Ad5CC10Cre病毒将暴露在以下4种情况之一：假空气(室内空气)、轻度CIH(CIH15、 (FiO2下降0.21至FiO2 0.15[SaO2最低值为82%])中度CIH(CIH10，(FiO2下降0.21至FiO2 0.15) [SaO2最低值为61%]或重度CIH(CIH5；FiO2下降0.21至FiO2 0.05[SaO2最低值为37%])。主要 结果将是定量的肿瘤体积(每月的显微CT扫描)；次要结果将是生存。目标 2：评估CIH5(Vs Sham)对抑制性免疫的产生、分化和募集的影响 患有和未患癌症的小鼠体内的细胞。主要结果将是骨中M-MDSC//Treg的定量 早期(2周)和晚期(4个月)时间点的骨髓、脾、胸腺、左肺和右肺； 结果将是对抑制性免疫细胞的功能分析。探索性目标3：研制克隆癌细胞 从TT-KPP小鼠的肺癌中提取。此外，评估CIH5(Vs Sham)对(1)趋化因子/细胞因子的调节作用 肺癌细胞内的基因(通过早期和晚期的PCR小组)和(2)其他基因(通过rna-seq at 一个时间点)。聚合酶链式反应和rna-seq信息将一起用于未来的赠款，我们将在那里使用 CRISPR编辑克隆细胞内的重要基因并将它们注入野生型小鼠(同基因小鼠 以进一步了解CIH影响肿瘤进展的机制。调查结果：第一 呈件。临床关系：我们可能会提供证据，证明阻塞性睡眠的主要病理因素--脑出血 呼吸暂停，是一种可改变的危险因素，可以提高肺癌的存活率。影响/意义：积极的呼吸道 压力疗法是一种治疗阻塞性睡眠呼吸暂停综合征的方法，没有副作用，可能是预防肺部疾病的有效辅助疗法。 癌症进展。

Objective: Lung cancer continues to have a very poor prognosis, with Veterans having a higher incidence of lung cancer and worse outcome compared to civilians.1 Obstructive sleep apnea (OSA) has been associated with higher cancer incidence2-5 and mortality,2, 6-8 and again, Veterans have a higher prevalence of OSA that is more severe compared to civilians.9 Our overarching objective is to determine how cyclical intermittent hypoxia (CIH), a major underlying pathology of OSA, promotes lung cancer progression. Our hypothesis is that CIH increases the differentiation of immature immune cells to suppressor immune cells which then, inadvertently, protects the cancer. We have published that CIH accelerates primary lung cancer progression in Triple Transgenic KrasG12D+; p53fl/fl; myristolated p110fl/fl ROSA-gfp (TT-Kpp) mice (funded by VA CPPF grant). This grant is the next step - to explore mechanisms of HOW CIH promotes cancer progression. Research Design: The technical innovation of this grant is that we will be the first to measure the effect of mild, moderate and severe CIH on cancer progression in TT-Kpp mice (Aim 1). The conceptual innovation of this grant is that we will be the first to use this TT-Kpp model to explore whether systemic CIH increases production, differentiation and recruitment of suppressor immune cells to primary lung cancer. Methodology: Aim 1: TT-Kpp mice injected with Ad5CC10Cre virus will be exposed to one of 4 conditions: sham (room air), mild CIH (CIH15, (drops in FiO2 0.21 to FiO2 0.15 [SaO2 nadir of 82%]) moderate CIH (CIH10, (drops in FiO2 0.21 to FiO2 0.15 [SaO2 nadir of 61%]) or severe CIH (CIH5; drops in FiO2 0.21 to FiO2 0.05 [SaO2 nadir of 37%]). Primary outcome will be quantitative tumor volumes (monthly microCT scans); secondary outcome will be survival. Aim 2: Assess the effect of CIH5 (vs Sham) on production, differentiation and recruitment of suppressor immune cells in mice with and without cancer. Primary outcome will be quantitative M-MDSC/TAM/Tregs in bone marrow, spleen, thymus, left lung and right lung at early (2 weeks) and late (4 months) time points; secondary outcome will be functional assays of suppressor immune cells. Exploratory Aim 3: Develop a clone cancer cell from lung cancer of TT-Kpp mice. Also, assess the effect of CIH5 (vs Sham) regulating (1) chemokine/cytokine genes within lung cancer cell (via PCR panel at early and late time point) and (2) other genes (via RNA-seq at one time point). Together, the PCR and RNA-seq information will be utilized in a future grant where we will use CRISPR editing of significant genes within clone cells and inject them into a wild type mouse (syngeneic orthotopic model) to further understand mechanism of effect of CIH on cancer progression. Findings: First submission. Clinical Relationships: We may provide evidence that CIH, a major pathology of obstructive sleep apnea, is a modifiable risk factor that can improve lung cancer survival. Impact/Significance: Positive airway pressure, a treatment for OSA has no side effects and may be a powerful adjunct therapy to prevent lung cancer progression.