Development of a gold nanoparticles based targeted delivery system

描述(由申请人提供)：靶向给药应能在副作用低到最小的情况下提高疗效。这是一个被广泛接受的概念，但由于缺乏可用的技术和验证过程，应用受到限制。生物医学纳米技术可以在这方面发挥重要作用。生物医学纳米技术是一个新兴的领域，它为推进医学科学和疾病治疗带来了无数的机会和可能性。它是一个跨越生物、化学、材料科学、工程和医学等学科的多学科领域。在纳米尺度上，由于量子尺寸效应，材料(金属、半导体等)的物理化学和生物特性与相应的块体材料有根本的不同，例如，金纳米颗粒(AuNPs)具有酒红色，而金属金是金黄色的，这种酒红色可以通过简单地控制AuNPs的大小和形状来调节为粉色、紫色或蓝色。此外，纳米粒子还具有较大的比表面积，可以负载多种诊断(如光学、放射性同位素、磁学)和治疗(如药物)。在这项应用中，我们希望利用纳米颗粒的独特性质来创建一种靶向递送系统，该系统将显示出更好的治疗效果，副作用最小甚至没有。该项目的长期目标是开发一种基于纳米颗粒的递送系统，用于靶向递送细胞毒药物，提高疗效并降低全身毒性。我们的初步研究表明，以抗EGFR抗体(C225)为靶向剂，以低剂量吉西他滨(Gem)作为金纳米结合物的靶向给药，可以增强该药物在人胰腺癌原位移植瘤模型中的抑制肿瘤生长的效果。这些结果鼓励我们深入研究纳米制造工艺，以进一步提高疗效并揭示增强药物活性的机制。因此，我们的研究集中在金纳米颗粒(AuNPs)作为载体，Gem作为细胞毒药物，C225作为靶向剂以2合1的方式结合到相同的金核上。纳米制造的有效性将首先通过测定纳米结合物对原发(PANC-1，MiaPaca2)和转移性(ASPC-1)人胰腺癌细胞株的活性进行体外测试。因此，研究这些模型的有效性将使我们能够确定这些系统在各种疾病条件下的应用(例如，原发疾病与转移性疾病、早期疾病与晚期疾病)。这些细胞的EGFR表达模式也不同，高表达的细胞(Panc-1和ASPC-1)比MiaPaca2(低表达的EGFR)摄取更多的纳米结合物。然后将在临床前胰腺癌小鼠模型上测试体内疗效。这项研究的目的是：(I)优化体外和体内靶向递送的纳米制造工艺，(Ii)确定靶向递送和非靶向递送中纳米结合物的药代动力学、生物分布和毒性，以及(Ii)确定纳米结合物在靶向递送和非靶向递送中抑制肿瘤生长、转移和提高生存率的治疗效果。胰腺癌是美国癌症死亡的第四大原因。目前，手术是唯一的选择，然而，由于表现较晚，只有10%-15%的患者愿意接受手术。这项应用的意义在于，它将研究在目前尚无有效治疗方法的情况下，使用针对胰腺癌的纳米递送系统来定向和非定向递送抗癌药物。这种有针对性的给药方式将以最小的副作用提高药物的疗效。根据我们的假设，当吉西他滨作为金纳米结合物以靶向方式给药时，将降低全身毒性，并具有更好的疗效。这项应用还将解决一些问题，以获得优化的递送载体，例如将靶向剂和药物装载到纳米颗粒、药物的生物利用度、金纳米颗粒和纳米结合物的生物兼容性和毒性。对于抗体不能靶向所有胰腺肿瘤细胞或不表达EGFR的患者，我们可能会使用其他靶点，如癌胚抗原(CEM)或糖类抗原19-9(CA-19-9)，或需要进一步确定其他靶向分子，并扩大我们的“概念验证”实验。重要的是，正如我们已经讨论的那样，EGFR在许多其他癌症如结直肠癌、非小细胞肺癌、非小细胞肺癌中过表达，吉西他滨也用于其他恶性肿瘤，如非小细胞肺癌、膀胱、乳腺等，因此，该策略不仅可用于胰腺癌的治疗，还可作为一种普遍的方法用于治疗其他一些恶性肿瘤，如结直肠癌、非小细胞肺癌、乳腺癌、卵巢癌等。

DESCRIPTION (provided by applicant): Targeted delivery of a drug should result in enhanced therapeutic efficacy with low to minimal side effects. This is a widely accepted concept, but limited in application due to lack of available technologies and process of validation. Biomedical nanotechnology can play an important role in this aspect. Biomedical nanotechnology is a burgeoning field and brings with it a myriad of opportunities and possibilities for advancing medical science and disease treatment. It is a multidisciplinary field cutting across the disciplines of biology, chemistry, materials science, engineering and medicine. At the nano scale, the physico-chemical, and biological properties of materials (metals, semiconductors, etc) differ fundamentally from their corresponding bulk counter part because of the quantum size effect e.g. gold nanoparticles (AuNPs) have wine red color whereas metallic gold is golden yellow and this wine red color can be tuned to either pink, or violet or blue by simply controlling the size and shape of AuNPs. Furthermore, nanoparticles also have large surface area to load multiple diagnostic (such as optical, radioisotope, magnetic) and therapeutic (such as drugs). In this application we want to exploit the unique properties of nanoparticles to create a targeted delivery system that will show better therapeutic efficacy with minimal to no side effects. The long-term goal of this project is to develop a nanoparticle-based delivery system for targeted delivery of cytotoxic drug with enhanced efficacy and reduced systemic toxicity. Our preliminary studies demonstrated that targeted delivery of a low dose of gemcitabine (Gem) as a gold nanoconjugate using anti-EGFR antibody (C225) as a targeting agent resulted in enhanced efficacy of the drug to inhibit the tumor growth in an orthotopic human xenograft model of pancreatic cancer. These results encouraged us to study, in depth, the nanofabrication process to improve the efficacy further and delineate the mechanism of enhanced drug activity. Therefore, our proposed studies center on gold nanoparticles (AuNPs) as a delivery vehicle, Gem as a cytotoxic drug and C225 as a targeting agent bound to the same gold core in a "2 in 1" fashion. The efficacy of nanofabrication will be tested in vitro first by determining the activity of the nanoconjugates against primary (PANC-1, MiaPaca2) and metastatic (AsPC-1) human pancreatic cancer cell lines. Therefore, studying efficacy in these models will allow us to determine the application of these systems in a wide range of disease condition (e.g. primary vs. metastatic disease, early stage vs. late stage of the disease). These cells also differ by EGFR expression pattern, cells with higher expression (PANC-1 and AsPC-1) will uptake more of the nanoconjugates than MiaPaca2 (with low EGFR expression). In vivo efficacy will then be tested in a preclinical mouse model of pancreatic cancer. The aims proposed in this study are designed to (i) optimize the nanofabrication process for targeted delivery in vitro and in vivo, (ii) to determine the pharmacokinetics, biodistribution and toxicity of the nanoconjugates in targeted vs. non-targeted delivery and (ii) to determine the therapeutic efficacy of the nanoconjugates to inhibit tumor growth, metastasis and increasing survival in targeted vs. non-targeted delivery. Pancreatic cancer is the 4th leading cause of cancer deaths in United States. Currently, surgery is the only option, however, due to late presentation only 10-15 % of the patients are amenable to surgery. The significance of this application is that it will study both targeted and non-targeted delivery of anti-cancer drugs using a nanodelivery system against pancreatic cancer where no effective therapy is currently available. Such a delivery in targeted fashion will enhance the efficacy of the drug with minimal side effects. According to our hypothesis, gemcitabine will have reduced systemic toxicity with better efficacy when delivered in a targeted fashion as a gold nanoconjugates. This application will also address a number of issues to obtain an optimized delivery vehicle such as loading of targeting agent and drug to a nanoparticle, bioavailability of the drug, biocompatibility and toxicity of gold nanoparticles and nanoconjugates. For those patients where the antibody is unsuccessful in targeting all the pancreatic tumor cells or patients that do not express EGFR, we may use other targets such as carcinoembryonic antigen (CEM) or carbohydrate antigen 19-9 (CA-19-9) or need to further identify other targeting molecules and expand our "proof of concept" experiments. Importantly, as we have already discussed that EGFR is overexpressed in a number of other cancers such as CRC, NHSC, NSCLC and gemcitabine is also used in other malignancies such as NSCLC, bladder, breast, therefore, this strategy could be used not only for the treatment of pancreatic cancers but also as a generalized approach in the treatment of a number of other malignancies such as CRC, NHLC, NSCLC, breast, ovarian, etc.