CLINICAL TRANSLATION OF 19F MRI TO VISUALIZE CANCER IMMUNOTHERAPEUTIC CELLS

描述（由申请人提供）：我们建议翻译一种新的细胞MRI技术，以可视化结肠直肠癌（CRC）细胞治疗药物的贩运。每年约有15万例结直肠癌新病例，其中6000例在匹兹堡大学癌症研究所（UPCI）接受治疗。伴有可切除肝转移的结直肠癌患者是一组整体预后不良且5年总生存率在20-35%之间的独特组合患者，尽管仅存在少量残留疾病。免疫治疗干预可能对这类患者有效。目前，用于结直肠癌和其他癌症类型的一种主要的基于细胞的治疗方法是使用树突状细胞（dc）作为活疫苗，将肿瘤特异性抗原递送到淋巴结，从而在患者体内诱导内源性杀伤T细胞（即细胞毒性T细胞或ctl）。迄今为止，治疗性细胞转移后无法在体内观察到细胞是癌症免疫治疗发展的最大瓶颈之一。目前基于DC的疫苗临床结果差异很大的一个常见原因是DC对淋巴结的递送不理想。最近，阿伦斯博士的实验室展示了一项引人注目的技术，通过核磁共振成像可以看到移植的细胞。在这种方法中，用一种新型的全氟聚醚（PFPE）纳米颗粒组合物在体外标记细胞，然后将其引入患者体内。随后使用19F MRI监测体内细胞迁移。这种方法的主要优点是，19F图像对标记细胞具有极强的选择性，没有来自宿主组织的背景信号。此外，在感兴趣的区域精确的细胞定量是可能的。数据表明，PFPE纳米颗粒在细胞内具有高度的生物相容性。迄今为止，已经在啮齿动物模型中进行了几项体内成像研究。一种临床级的PFPE纳米颗粒试剂最近由Celsense公司配制和制造。Celsense目前正在对该试剂进行强制性的体外和动物安全性研究，以供最终用于人类。拟议的研究将使我们能够同时推进人体细胞治疗的体内跟踪技术，同时为正在进行的CRC免疫治疗临床研究提供见解。UPCI的Kalinski和Bartlett。提出的项目有三个具体目标：（1a）建立治疗细胞的PFPE标记方案。我们将开发最佳的组织培养方案PFPE标记免疫治疗dc。（1b）确定PFPE标记后治疗细胞的体外特性。我们将严格评估PFPE标记在体外诱导DC功能和表型改变的程度。（2a,b）在啮齿动物模型中研究标记dc的细胞运输和PFPE清除。（3a）在3T临床MRI系统上实施敏感的19F MRI/MRS方法。我们将优化脉冲序列，并评估最小可检测细胞数使用幻影研究。（3b）追踪CRC患者免疫治疗细胞的初步研究。我们将修改现有的UPCI CRC治疗方案（UPCI 05-063; BB-IND 13,234），纳入转移细胞的成像。在一小群结直肠癌患者（n=6）中，我们将成像基于dc的疫苗向淋巴结的迁移。我们将遵循传统的皮内给药途径和一种新的淋巴内给药途径，以加速DC向淋巴结的递送，并保护DC的加强剂量不被先前诱导的ctl消除。提出的转化研究是评估19F MRI/ mrs细胞跟踪临床应用潜力的第一次努力。此外，我们将获得对改善转移性结直肠癌和其他形式癌症的免疫治疗策略至关重要的初步数据。匹兹堡地区在癌症免疫治疗和成像创新方面具有卓越的实力。我们从该地区组织了一支强大的研究小组来进行这个项目。公共卫生相关性：在本提案中，我们将使用先进的磁共振成像（MRI）技术来可视化结肠直肠癌细胞免疫疗法的贩运。细胞免疫疗法通常应用于晚期癌症，现有的治疗方法疗效有限，细胞免疫疗法是使用专门的细胞杀死癌细胞或产生对疾病的免疫力。免疫疗法有望成为继手术、化疗和放疗之后的第四种癌症治疗策略。目前，接种后无法看到体内细胞是癌症免疫治疗发展的最大瓶颈之一。该提案将采用现代成像工具来解决这一迫切需求，同时为正在进行的结直肠癌免疫治疗临床研究提供见解。

DESCRIPTION (provided by applicant): We propose to translate a novel cellular MRI technology to visualize the trafficking of cellular therapeutics against colorectal cancer (CRC). CRC accounts for approximately 150,000 new cases annually, and 6,000 of these are treated every year at the University of Pittsburgh Cancer Institute (UPCI). CRC patients with resectable liver metastases constitute a group of patients with a unique combination of an overall poor prognosis and a 5-year overall survival rate between 20-35%, despite the presence of only minimal residual disease. Immunotherapeutic interventions may potentially be effective in this patient population. Currently, a major type of cell-based therapy used for CRC and other cancer types uses dendritic cells (DCs) as live vaccines that deliver tumor-specific antigens to the lymph nodes thereby inducing endogenous killer T cells (i.e., cytotoxic T cells or CTLs) in patients. To date, the inability to see the cells in vivo following therapeutic cell transfer is one of the greatest bottlenecks in the development of cancer immunotherapy. A commonly cited reason for the highly varied clinical outcomes of current DC-based vaccines is sub-optimal DC delivery to the lymph nodes. Recently, the laboratory of Dr. Ahrens has demonstrated a compelling technology that makes transplanted cells visible via MRI. In this approach, cells are labeled ex vivo with a novel perfluoropolyether (PFPE) nanoparticle composition and then introduced into the patient. Cell migration is subsequently monitored in vivo using 19F MRI. The key advantage of this approach is that the 19F images are extremely selective for the labeled cells, with no background signal from the host's tissues. Furthermore, accurate cell quantification in regions of interest is possible. Data show that PFPE nanoparticles are highly biocompatible inside cells. To date, several in vivo imaging studies have been conducted in rodent models. A clinical-grade version of the enabling PFPE nanoparticle reagent has recently been formulated and manufactured by Celsense, Inc. Celsense is currently performing mandatory in vitro and animal safety studies of this reagent for eventual human use. The proposed studies will allow us to simultaneously advance the technology of in vivo tracking of cell therapy in humans, while giving insights into ongoing clinical studies of immunotherapy in CRC, directed by Drs. Kalinski and Bartlett at the UPCI. The proposed project has three Specific Aims: (1a) Establish PFPE labeling protocols for therapeutic cells. We will develop optimal tissue culture protocols for PFPE labeling of immunotherapeutic DCs. (1b) Determine in vitro characteristics of therapeutic cells following labeling with PFPE. We will rigorously evaluate the degree to which PFPE labeling induces any alterations in DC function and phenotype in vitro. (2a,b) study cell trafficking and PFPE clearance of labeled DCs in a rodent model. (3a) Implementation of sensitive 19F MRI/MRS methods on a 3T clinical MRI system. We will optimize pulse sequences and evaluate the minimum detectable cell numbers using phantom studies. (3b) Pilot study tracking immunotherapeutic cells in CRC patients. We will amend the existing UPCI CRC therapeutic protocol (UPCI 05-063; BB-IND 13,234) to include imaging of transferred cells. In a small cohort of CRC patients (n=6), we will image the migration of DC-based vaccines to the lymph nodes. We will follow their administration by the traditional intradermal route and a novel intralymphatic route developed to accelerate the DC delivery to the lymph nodes and to protect booster doses of DCs from elimination by the previously-induced CTLs. The proposed translational studies are the first effort to evaluate the potential of 19F MRI/MRS-based cell tracking for clinical use. Additionally, we will garner preliminary data critically important for improving the immunotherapeutic strategies in metastatic CRC and other forms of cancer. The Pittsburgh region has exceptional strength in cancer immunotherapy and imaging innovations. We have organized a potent research team from the region to conduct this project. PUBLIC HEALTH RELEVANCE: In this proposal we will use advanced magnetic resonance imaging (MRI) technology to visualize the trafficking of cellular immunotherapy against colorectal cancer. Most often applied to advanced cancer, where existing therapies have limited efficacy, cellular immunotherapy is the administration of specialized cells that kill cancer cells or produce immunity to the disease. Immunotherapy is positioned to become a fourth strategy in cancer treatment supplementing surgery, chemotherapy, and radiation. Currently, the inability to see cells in the body following inoculation is one of the greatest bottlenecks in the development of cancer immunotherapy. This proposal will adapt modern imaging tools to address this urgent need, while giving insights into ongoing clinical studies of immunotherapy in colorectal cancer.