Phage Display for Improved Peptide-Based Tumor Targeting and Imaging Agents

侵袭性癌症是美国退伍军人的主要健康问题。虽然大多数癌症 是可以治疗的，与性别有关的癌症，包括乳腺癌和前列腺癌， 当在晚期和/或转移性阶段诊断时通常是致命的。到2020年，这些癌症将占 一般人群每年死亡75，400人，退伍军人的发病率明显较高 人口虽然大多数患者最初对治疗有反应，但他们经常复发并发展为侵袭性和 抗药性肿瘤更好地检测和治疗这些类型的癌症的一种有希望的方法可能是通过 开发靶向与癌症干细胞（CSC）相关的耐药性生物标志物的药物。 CSC占肿瘤的一小部分，但会驱动肿瘤的侵袭性和耐药性。有 一些经验证的CSC生物标志物，并且大多数不是癌症特异性的。我们的实验室是在新的前沿 使用噬菌体（噬菌体）展示的生物标志物和CSC鉴定。肽和抗体（Ab）片段 结合乳腺和前列腺相关的B-F糖抗原（TF）， 已经获得了结合TF并抑制免疫应答的半乳糖凝集素-3（gal-3）和CD 444 v6。这些 抗原参与细胞粘附、血管生成、免疫调节和细胞信号传导，并且被认为 代表CSC生物标志物。此外，我们证明了CD 44 v6通过TF相互作用结合gal-3， 提示该三联体可能代表探测CSC、肿瘤生长和免疫监视的新靶点。 抗体和肽两者都已在癌症成像和治疗中取得了混合成功。方面 抗体的大尺寸可导致免疫原性问题和长的分布时间，一旦放射性标记， 导致器官和组织损伤相反，肽，特别是来自噬菌体展示的肽， 通常是疏水性的，并且常常具有差的亲和力和肿瘤摄取。两种不同的策略将是 它克服了这些缺点。一种预先靶向的“点击”化学方法， 可以使用来自放射成像或放射治疗步骤的肿瘤靶向步骤。这种有针对性的方法 是基于快速和高度特异性的环加成反应，发生之间的反式环辛烯 (TCO)通过允许未标记的靶向Ab循环并结合靶点，随后，委员会注意到， 注射小分子放射性标记的探针，其具有快速的药代动力学和对药物的高亲和力。 与直接标记的Ab相比，预先靶向的Ab将导致更高的肿瘤与正常组织摄取比率。的 第二种方法是将小肽结合到纳米颗粒上用于多肽展示。康奈尔素点 （C' dots）是一种新的和良好表征的纳米颗粒平台， 和治疗，以及多路生物标记询问。C'点是二氧化硅核-壳PEG纳米颗粒， 表现出良好的血液清除率，主要通过肾脏排泄， 流通时报可调的表面化学和有利的体内药代动力学允许C'点 克服了与当前单分子癌症治疗剂相关的许多剂量限制毒性。 假设靶向gal-3/TF/CD 44 v6的Ab片段和肽与 预靶向和C 'dot多价展示将产生用于侵袭性前列腺（PCa）的治疗诊断剂， 乳腺癌（BCa）。本研究的目标是：1）放射成像和放射学 侵袭性PCa和BCa肿瘤的试剂，2）抑制肿瘤发生的探针， 肿瘤生长和转移，以及3）增加肿瘤免疫的双功能分子治疗剂 通过与gal-3和免疫检查点抑制剂的双重靶向反应。 我们的工作将对VA医疗保健产生积极影响，因为我们正在开发新的靶向检测剂 以及在体内耐受良好的癌症治疗。这项工作将有助于改善我们退伍军人的生活 与BLR&D的总体目标和妇女健康与基因组医学的特殊利益相一致。

Aggressive forms of cancer are a leading health problem for United States Veterans. While most cancers are treatable in their earliest forms, gender-related cancers including those of the breast and prostate are often fatal when diagnosed at advanced and/or metastatic stages. In 2020, these cancers will account for over 75,400 annual deaths in the general population, with morbidity rates significantly higher in the Veteran population. While most patients initially respond to treatment, they often relapse and develop aggressive and drug resistant tumors. A promising approach to better detect and treat these types of cancers may be through development of agents that target drug resistance biomarkers that associate with cancer stem cells (CSCs). CSCs make up a small proportion of a tumor but drive tumor aggressiveness and drug resistance. There are few validated CSC biomarkers, and most are not cancer-specific. Our laboratory is at the forefront of new biomarker and CSC identification using bacteriophage (phage) display. Peptides and antibody (Ab) fragments that bind the breast and prostate associated Thomsen-Friedenreich carbohydrate antigen, (TF), the lectin galectin-3 (gal-3) that binds TF and inhibits immune response, and CD444v6 have been obtained. These antigens are involved in cell adhesion, angiogenesis, immune regulation, and cell signaling, and are thought to represent CSC biomarkers. Further, we demonstrated that CD44v6 binds gal-3 through TF interaction, suggesting this triad may represent a novel target to probe CSCs, tumor growth and immune surveillance. Both Abs and peptides have been used with mixed success in cancer imaging and therapy. In terms of Abs their large size can cause immunogenic problems and long distribution times that once radiolabeled often leads to organ and tissue damage. Conversely, peptides, particularly those from phage display are usually hydrophobic and often suffer from poor affinity and tumor uptake. Two different tacks will be employed that overcome these drawbacks. A pretargeted “click” chemistry approach which decouples the tumor-targeting step from the radioimaging or radiotherapy step can be employed. This pretargeted approach is based on the rapid and highly specific cycloaddition reaction that occurs between the trans-cyclooctene (TCO) on the tumor by allowing the unlabeled targeting Ab to circulate and bind the target. Subsequently, injection of a small molecule radiolabeled probe with rapid pharmacokinetics and high affinity for the pretargeted Ab, will result in higher tumor to normal tissue uptake ratios compared to direct labeled Abs. The second tack will be to conjugate small peptides to a nanoparticle for multi-peptide display. Cornell prime dots (C' dots) are a new and well-characterized nanoparticle platform that offer both multimodal-targeted imaging and therapy, and multiplexing biomarker interrogation. C' dots are silica core-shell PEG nanoparticles that exhibit favorable clearance from the blood and are primarily excreted through the kidney overcoming extended circulation times. The tunable surface chemistries and favorable in vivo pharmacokinetics allow C' dots to overcome many of the dose limiting toxicities associated with current monomolecular cancer therapeutics. It is hypothesized that the Ab fragments and peptides that target gal-3/TF/CD44v6, coupled with pretargeting and C'dot multivalent display, will produce theranostic agents for aggressive prostate (PCa) and breast cancer (BCa). The objectives of this research are to develop: 1) radioimaging and radiotherapeutic agents of aggressive PCa and BCa tumors, 2) probes that inhibit tumorigenesis leading to reduced tumor growth and metastasis, and 3) bifunctional molecular therapeutics that increase tumor immune response through dual targeting with gal-3 and immune checkpoint inhibitors. Our work will positively impact VA healthcare in that we are developing new targeting agents for detection and treatment of cancer that will be well tolerated in vivo. This work will help improve the lives of our Veterans and coincides with BLR&D general goals and special interests of Women's Health and Genomic Medicine.