Project 1: 5FU and Vitamin D as Neoadjuvants for Photodynamic Priming to Enhance Skin Cancer Therapy

摘要-项目1 项目1将探索光动力疗法(PDT)如何用于新生的 皮肤癌的联合治疗。非黑色素瘤皮肤癌(NMSC)，由鳞状细胞组成 癌(SCC)和基底细胞癌(BCC)是所有人类癌症中最常见的，代表着一种 巨大的医疗负担(300万美国病例/年)，某些患者亚群的致命性增加， 例如，器官移植接受者。我们以前已经证明，基于氨基酮戊酸酯(ALA)的光动力疗法是有效的 对于NMSC患者，5-氟尿嘧啶(5FU)和维生素D(维生素D)进一步改善治疗结果； 对于大型和晚期NMSC，肿瘤清扫率仍然很低。为改善这种情况，我们建议 利用光动力引发(PDP)的概念，这是光动力引发的自然结果。光动力疗法触发免疫原性 细胞死亡，增加肿瘤浸润性淋巴细胞(TIL)的流入，并诱导适应性抗肿瘤免疫 反应，使肿瘤微环境对免疫检查点抑制的影响敏感。我们的 初步数据显示，低强度PDT方案可以清除小鼠鳞状细胞癌模型中的肿瘤， 伴随着损伤相关分子模式(DAMP)的诱导和中性粒细胞的募集， 巨噬细胞和TIL，以及PD-1等免疫检查点分子表达的变化。这些 这些发现为使用PDT和免疫检查点抑制剂(ICI)的进一步组合方法奠定了基础。在……里面 目的1(临床前)我们将在小鼠鳞状细胞癌和基底细胞癌模型中评估对不同药物的治疗反应 PDT、5FU、VitD和ICI试剂的组合，并测量TIL招募的特定时间、检查点 分子表达和肿瘤清除。在目标2(临床研究)中，我们将确定其性质和时机。 光动力疗法后基底细胞癌和鳞癌患者的免疫反应。肿瘤将接受PDT(由Core指导 C剂量法测量光敏剂水平)；然后在1至14天后，肿瘤将被手术切除并 用组织学方法分析检查点分子的表达和瘤内免疫细胞的浸润 免疫核心(IS；项目3)。循环血液中的系统性T细胞激活将在合作中进行测量 这些研究的另一个分支将测试5FU或VitD对NMSC肿瘤的影响 抗肿瘤免疫反应的大小。AIM 3将进行探索性研究，以验证IS 人鳞状细胞癌体外模型中TIL募集的测定方法。与Project 3协作 核心B，3D患者来源的肿瘤免疫器官(PDIO)，由分层的SCC培养组成 与PBMCs(淋巴细胞)混合后将测试PDT后不同免疫细胞的激活情况 核心B开发的尖端高光谱光学细胞术技术可以对多种免疫进行成像 标记同时出现。相关性和影响：总体而言，该项目将通过提供 为结合光动力激发、5FU、维生素D和ICI免疫疗法来改进的新方法奠定了基础 疑难和晚期NMSC的治疗结果。

ABSTRACT – Project 1 Project 1 will explore how photodynamic therapy (PDT) can be used for immunological priming in new combination treatments for skin cancer. Nonmelanoma skin cancers (NMSC), comprising squamous cell carcinoma (SCC) and basal cell carcinoma (BCC), are the most common of all human cancers, representing a significant healthcare burden (>3 million U.S. cases/year), with increased lethality for certain patient subsets, e.g. organ transplant recipients. We showed previously that aminolevulinate (ALA)-based PDT can be effective for NMSC patients, and that 5-fluorouracil (5FU) and Vitamin D (VitD) further improve treatment outcomes; yet tumor clearance rates remain poor for large and advanced NMSC. To ameliorate this situation, we propose to exploit the concept of photodynamic priming (PDP), a natural consequence of PDT. PDT triggers immunogenic cell death, enhances influx of tumor infiltrating lymphocytes (TILs), and induces an adaptive anti-tumor immune response, sensitizing the tumor microenvironment to the effects of immune checkpoint inhibition. Our preliminary data show that a low-intensity PDT regimen can clear tumors in a murine SCC model, accompanied by induction of damage-associated molecular patterns (DAMPs) and recruitment of neutrophils, macrophages and TILs, along with altered expression of immune checkpoint molecules such as PD-1. These findings set the stage for a further combination approach using PDT and immune checkpoint inhibitors (ICI). In Aim 1 (preclinical) we will evaluate in murine SCC and BCC models the therapeutic response to various combinations of PDT, 5FU, VitD, and ICI agents, and measure specific timing of TIL recruitment, checkpoint molecule expression, and tumor clearance. In Aim 2 (clinical studies) we will determine the nature and timing of immune responses in BCC and SCC patients, post-PDT. Tumors will be subjected to PDT (guided by Core C dosimetry to measure photosensitizer levels); then after 1-to-14 days the tumor will be surgically excised and analyzed for checkpoint molecule expression and intratumoral immune cell infiltration using the histological immunoscore (IS; Project 3). Systemic T-cell activation in circulating blood will be measured in collaboration with Project 2. Another arm of these studies will test effects of 5FU or VitD pretreatment of NMSC tumors upon the magnitude of anti-tumoral immune responses. Aim 3 will feature exploratory studies to validate the IS approach for measuring TIL recruitment in human SCC tumor models in vitro. In collaboration with Project 3 and Core B, 3D patient-derived tumor-immune organoids (PDIOs), consisting of stratified SCC cultures admixed with PBMCs (lymphocytes) will be tested for differential immune cell activation following PDT using cutting-edge hyperspectral optical cytometry techniques developed in Core B that can image multiple immune markers simultaneously. Relevance and Impact: Overall, this Project will benefit patients by providing the basis for a new approach that combines photodynamic priming, 5FU, VitD, and ICI immunotherapy to improve treatment outcomes for difficult and advanced cases of NMSC.