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

 描述(由申请人提供)： 癌症是美国退伍军人的主要健康问题。虽然大多数癌症最早的形式是可以治疗的，但与性别有关的癌症，包括卵巢癌、乳腺癌和 前列腺癌在诊断为晚期和/或转移性前列腺癌时往往是致命的。据估计，2015年，这些癌症每年将在普通人群中造成超过8万人的死亡，其中退伍军人的发病率要高得多。许多患者最初对治疗有反应，但往往会复发。复发可能是由于肿瘤的异质性，各种信号通路的失调，异常的凝集素-碳水化合物，或侵袭性和耐药性肿瘤的发展。最好的方法是识别和治疗这些癌症，可能是通过实施更个人化的诊断和/或治疗方法，其中使用针对癌细胞上的多个生物标记物的药物。然而，令人惊讶的是，很少有肿瘤相关抗原或生物标记物被证明与癌症类型或分期有关。我们的实验室以前曾利用噬菌体展示技术来识别生物标志物，并分离识别卵巢癌、乳腺癌和前列腺癌的多肽靶向分子。针对ErbB-2和ErbB-3受体的多肽、碳水化合物(Thomsen-Friedenreich抗原，Tf)、凝集素(Galectin-3，GAL-3)和CD44剪接变异体CD44v6已经被获得。这些生物标志物参与细胞黏附、血管生成和信号转导。Tf在90%以上的腺癌中表达，GAL-3是侵袭性和转移潜能的标志，ErbB-2是侵袭性转移的标志，ErbB-3常与不良预后和治疗耐药相关，CD44v6与侵袭性癌症起始细胞(CIC)相关。在体内检测此类生物标记物的能力 将有助于区分肿瘤进展的不同阶段，并可能有助于监测对治疗的反应。肿瘤生物标记物的靶向和成像通常依赖于放射性标记抗体(Abs)的使用。不幸的是，单抗非常大，经常会引起免疫原性相关的毒性和耐药性。小肽在人类中耐受性良好，但肿瘤摄取率低，不稳定，肾脏滞留较高。展示在小支架或单链抗体片段(ScFv)背景下的多肽具有高亲和力、稳定性和良好的体内药代动力学，为开发有效的肿瘤靶向药物提供了理想的平台。该噬菌体展示筛选出的Gal-3/Tf、ErbB-2/ErbB-3和CD44v6多肽可以被工程化为高亲和力、稳定、靶向性的支架，用于卵巢癌、乳腺癌和前列腺癌的治疗的成像和响应。这些研究的目标是将ErbB-2/ErbB-3和Gal-3/Tf靶向多肽和新发现的CD44v6靶向多肽开发成个性化的癌症检测或治疗试剂。这项研究的目的是：1)从功能上表征这些多肽的阻断作用 体外和人乳腺癌、卵巢癌或前列腺癌内的信号通路和增殖；2)开发新一代用于放射成像/治疗的多肽：i)合成逆转录和多抗原肽结构，ii)工程多肽 3)利用荧光成像技术同时靶向多个生物标志物，以简化癌症检测探针的开发；以及4)利用64Cu(或68Ga)标记的支架进行体内生物分布和正电子发射断层扫描(PET)成像，以评估AS对体内放射诊断和/或治疗药物的响应。我们的工作将对医疗保健产生积极影响，因为我们正在开发用于检测和治疗癌症的靶向药物。这些基于多肽的药物将在体内获得良好的耐受性，并有助于改善退伍军人的生活。这项研究与BLR&D对妇女健康和基因组医学的特殊兴趣不谋而合。

 DESCRIPTION (provided by applicant): Cancer is a leading health problem for United States Veterans. While most cancers are treatable in their earliest forms, gender-related cancers including those of the ovary, breast, and prostate are often fatal when diagnosed at advanced and/or metastatic stages. It is estimated that in 2015, these cancers will account for over 80,000 annual deaths in the general population, with morbidity rates significantly higher in the Veteran population. Many patients initially respon to treatment, but often relapse. Relapse may be due to the heterogeneous nature of the tumor, dysregulation of various signaling pathways, aberrant lectin- carbohydrates, or development of aggressive and drug resistant tumors. The best was to identify and treat these cancers may be through implementation of a more personally tailored diagnostic and/or therapeutic approach in which agents that target multiple biomarkers on cancer cells are employed. However, surprisingly few tumor-associated antigens or biomarkers have been shown to associate with cancer type or stage. Our laboratory has previously utilized bacteriophage (phage) display technology to identify biomarkers and isolate peptide-based targeting molecules that recognize ovarian, breast, and prostate carcinomas. Peptides that target the ErbB-2 and ErbB-3 receptors, carbohydrates (Thomsen-Friedenreich antigen, TF), lectins (galectin- 3, gal-3), and the CD44 splice variant CD44v6, have been obtained. These biomarkers are involved in cell adhesion, angiogenesis, and signaling. TF is expressed in over 90% of all adenocarcinomas, gal-3 is a marker of invasiveness and metastatic potential, ErbB-2 is a marker of aggressive metastasis, ErbB-3 often associates with poor prognosis and treatment resistance, and CD44v6 associates with aggressive cancer initiating (stem) cells (CIC). The ability to detect such biomarkers in vivo will help discern various stages of tumor progression and may help monitor response to therapy. The targeting and imaging of cancer biomarkers has generally relied on the use of radiolabeled antibodies (Abs). Unfortunately, Abs are very large and often cause immunogenic related toxicity and drug resistance. Small peptides are well tolerated in humans but suffer from low tumor uptake, instability, and high kidney retention. Peptides displayed in the context of small scaffolds or single chain Ab fragments (scFv) may provide ideal platforms for development of efficacious tumor targeting agents due to their high affinity, stability, and favorable in vivo pharmacokinetics. It is hypothesized that the phage display selected gal-3/TF, ErbB- 2/ErbB-3, and CD44v6 peptides can be engineered into high affinity, stable, targeting scaffolds to be used in the imaging and response to therapy of ovary, breast, and prostate cancer. The goal of these studies is to develop ErbB-2/ErbB-3 and gal-3/TF-targeting peptides and newly identified CD44v6-targeting peptides into personalized cancer detection or therapy agents. The objectives of this proposed research are to: 1) functionally characterize the peptides for the interruption of signaling pathways and proliferation in vitro and within human breast, ovary, or prostate carcinomas; 2) develop new generation of peptides tailored to radioimaging/therapy applications by: i) synthesis of retro-inverso and multiple antigen peptide constructs, ii) engineering peptides into novel scFv scaffolds, and iii) generating phage display scFv libraries for affinity maturation 3) simultaneous target multiple biomarkers using fluorescence imaging for streamlining the development of cancer detection probes, and 4) perform in vivo biodistribution and positron emission tomography (PET) imaging with 64Cu (or 68Ga)-labeled scaffolds to evaluate as in vivo radiodiagnostic and/or response to therapy agents. Our work will positively impact healthcare in that we are developing targeted agents for the detection and treatment of cancer. The peptide-based agents will be well tolerated in vivo and help improve the lives of our Veterans. This research coincides with BLR&D special interests of Women's Health and Genomic Medicine.