Dendritic Cell Proteoglycans and Reprogramming Cancer Immunity

在癌症生长的早期，转移和树突状细胞(DC)流向引流淋巴(DLN)。在肺里 癌症和其他癌症，免疫抑制主导的肿瘤微环境的特征是 未成熟的DC，弱的T效应反应，T调节细胞(Tregs)的增殖，以及过度产生 免疫抑制细胞因子。这严重抑制了抗肿瘤免疫。具有高度影响力的新领域 科学发现是免疫疗法。我们在一些晚期癌症中发现了显著的临床反应 患者通过阻断抑制效应性T细胞。然而，这类患者中只有一小部分(25%)有反应 有持久的减刑。初步研究表明，一种独特的被称为硫酸乙酰肝素(HS)的葡聚糖 推动颠覆性的DC流量和DC不成熟。虽然我们已经通过以淋巴为目标来抑制这种DC流量 内皮HS，新的工作表明DC特异性HS的改变可能调节病理性趋化因子- 依赖DC流量以及DC成熟度和功能。新的研究还表明，这些特性可能 有利影响抗肿瘤T细胞功能，抑制肿瘤生长和进展。在这里，我们的目标是 在研究免疫功能和分子机制的同时，在肿瘤和细胞研究中使用这种葡聚糖。 这一建议解决了肺癌患者树突状细胞表面HS糖链靶向的假设 通过基因手段和新型抑制剂抑制耐受树突状细胞对DLN的侵袭，提高抗DLN的能力 肿瘤免疫。更成熟的DC降低了免疫耐受性，提高了肿瘤抗原反应， 随着T细胞诱导的改善，将导致一种新的内源性抗肿瘤状态。为了测试这一点，我们建议： (1)在模型抗原和自发性肺中表征肿瘤生长和抗肿瘤免疫 携带DC-多糖改变的小鼠的肿瘤模型。我们将评估DC靶向突变在密钥中是如何 硫酸化HS生物合成酶(NDST1)对荷瘤小鼠胸椎DLN T细胞免疫功能的影响 原位表达卵白蛋白的Lewis肺癌(LLC-OVA)，包括OVA特异性免疫和 对肿瘤生长的影响。还将研究KRAS转基因突变模型中的免疫和肿瘤生长。 (2)研究荷肺癌小鼠体外制剂抗肿瘤DC和T细胞的功能。 HS生物合成中DC特异性的改变。DC的成熟、抗原提呈和DC的能力 LLC-OVA肿瘤生长在DC靶向HS突变体中以激活OVA致敏的T细胞，也将被检查 荷瘤突变小鼠DLN分离的CD8+T细胞对肿瘤细胞的杀伤作用研究 还将包括突变对浆细胞样树突状细胞对DLN定植的影响及其功能能力。 (3)在DC靶向改变的背景下评估趋化因子-受体的相互作用和信号机制 HS生物合成；以及新型HS抑制剂对肿瘤生长和免疫的影响。我们会研究房协如何 突变影响淋巴趋化因子CCL21、CXCL12或CX3CL1与同源细胞的DC表面结合 感受器。HS突变对趋化因子诱导的DC迁移和成熟信号的影响 评估，包括通过新的TLR依赖机制激活pDC。最后，我们会 研究新型HS抑制剂如何影响肿瘤树突状细胞的信号转导以及肺癌的生长和免疫。 这项工作研究了改变肿瘤DC的运输和成熟的新机制 有效地重新编程内源性抗肿瘤免疫。遗传靶点验证与合理导入 糖靶向抑制剂的研究可能会指导新的治疗方法，以实现对人类肺癌的免疫根除。

Early in cancer growth, metastases and dendritic cells (DCs) traffic to draining lymph nodes (DLN). In lung cancer and other carcinomas, immune-suppression dominates a tumor microenvironment characterized by immature DCs, weak T-effector responses, proliferation of T-regulatory cells (Tregs), and over-production of immuno-suppressive cytokines. This heavily represses anti-tumor immunity. A high-impact area of new scientific discovery is immunotherapy. We find dramatic clinical responses in some advanced-stage cancer patients by blocking suppression of effector T cells. Only a fraction (<25%) of such patients, however, respond with durable remissions. Preliminary work shows that a unique class of glycans known as heparan sulfate (HS) drives subversive DC traffic and DC immaturity. While we have inhibited such DC traffic by targeting lymphatic endothelial HS, new work suggests that DC-specific HS alterations may modulate both pathologic chemokine- dependent DC traffic as well as DC maturation and function. New studies also show that these properties may favorably impact anti-tumor T cell functions, with inhibition of tumor growth and progression. Herein we target such glycans in tumor and cell-based studies, while studying immune-function and molecular mechanisms. This proposal addresses the hypothesis that targeting HS glycans on the surface of DCs in lung cancer through genetic means and novel inhibitors will inhibit DLN colonization by tolerogenic DCs and improve anti- tumor immunity. Reduced immune-tolerance and improved tumor-antigen responses by more mature DCs, with improved T cell induction will result in a novel endogenous anti-tumor state. To test this, we propose to: (1) Characterize tumor growth and anti-tumor immunity in model antigen- as well as spontaneous lung carcinoma models in mice bearing DC-glycan alterations. We will assess how a DC-targeted mutation in a key sulfating HS biosynthetic enzyme (Ndst1) affects T cell immunity in tumors and thoracic DLN of mice with orthotopic Ovalbumin-expressing Lewis lung carcinomas (LLC-Ova), including Ova-specific immunity and effects on tumor growth. Immunity and tumor growth in a KRAS transgenic mutant model will also be studied. (2) Study anti-tumor DC and T cell functions in ex-vivo preparations from lung carcinoma bearing mice with DC-specific alterations in HS biosynthesis. DC maturation, antigen presentation, and the capacity of DCs from LLC-Ova tumors grown in DC-targeted HS mutants to activate Ova-sensitized T cells will be examined, as will tumor-cytolytic capacity of CD8+ T cells isolated from DLNs of tumor-bearing mutant vs control mice. Studies will also include the effects of mutation on DLN colonization by plasmacytoid DCs and their functional capacity. (3) Assess chemokine-receptor interactions and signaling mechanisms in the setting of DC-targeted alterations in HS biosynthesis; and the effects of novel HS inhibitors on tumor growth and immunity. We will study how HS mutation affects DC-surface binding of lymphatic chemokines CCL21, CXCL12, or CX3CL1 to cognate receptors. The effect of HS mutation on DC migration and maturation signaling in response to chemokine will be assessed, including the activation of pDCs through novel TLR-dependent mechanisms. Finally, we will study how novel HS inhibitors affect signaling by tumor DCs as well as lung carcinoma growth and immunity. This work examines novel mechanisms whereby altering trafficking and maturation of tumor DCs may effectively re-program endogenous anti-tumor immunity. The genetic target validation and rational introduction of glycan-targeting inhibitors may guide new therapies to achieve immune eradication of human lung cancer.