Targeted Prodrug Therapy of Liver Cancers

描述(申请人提供)：肝细胞癌和结直肠癌是世界上最常见和最致命的恶性肿瘤之一。在美国，结直肠癌是与癌症相关的死亡的第二大原因，主要是由于肝脏转移。5-氟尿嘧啶(5-FU)仍然是治疗包括不可切除的肝转移癌在内的肝癌的主要联合化疗方案。最近的研究表明，与全身化疗相比，以5-FU为基础的区域直接肝灌注化疗可提高结直肠癌患者的有效率和生存率。然而，这种投放系统技术复杂、侵入性强、风险大、成本高。本申请是一种用于改进肝癌和转移的化疗的专利递送系统。它利用由疟疾环子孢子(CS)蛋白组成的重组融合蛋白，CS蛋白是一种肝细胞特异性靶向配体，连接到细菌胞嘧啶脱氨酶(CD)，CD是一种自杀基因产物，催化其前体药物5-氟胞嘧啶(5-FC)产生5-FU。我们已经证明了CD-CS融合蛋白可以以细胞类型特异性的方式内化。更重要的是，内化的蛋白质至少在四周内稳定，并在给药前药5-FC时发挥旁观者细胞杀伤作用。这种持久的稳定性可以归因于这样一个事实，即内化的融合蛋白被包裹在特定的隔室(S)，这些隔室不受细胞质降解机制的影响。为了进一步开发和商业化该系统，我们提出了以下两个具体目标：(1)构建CD-CS的缺失突变体和修饰版本，以提高融合蛋白的产量、稳定性和活性；(2)优化修饰的CD-CS蛋白的生产，并确定其在产品生产和未来临床试验的放大准备中的有效性。这种新型的前药靶向治疗有效、技术简单、非侵入性和成本效益高，应该具有巨大的商业潜力。

DESCRIPTION (provided by applicant): Hepatocellular carcinoma (HCC) and colorectal cancer (CRC) are among the most common and deadly malignancy worldwide. CRC is the second leading cause of cancer-related death in the USA, mostly due to metastases to the liver. 5-fluorouracil (5-FU) remains the mainstay of combination chemotherapy for liver cancers including non-resectable liver metastases. Recent studies demonstrated that regional 5-FU-based chemotherapy by directly hepatic infusion showed improved response rates and survival for CRC patients as compared with those with systemic treatment. However, this delivery system is technically complicated, highly invasive, risky and costly. The present application is a patented delivery system for improved chemotherapy of liver cancer and metastasis. It utilizes a recombinant fusion protein consisted of the malarial circumsporozoite (CS) protein, a hepatocyte-specific targeting ligand, linked to the bacterial cytosine deaminase (CD), a suicidal gene product which catalyzes the production of 5-FU from its prodrug 5- fluorocytosine (5-FC). We have demonstrated that the CD-CS fusion protein can be internalized in a cell type-specific manner. More importantly, the internalized protein is stable for at least four weeks and exerts bystander cell-killing effects upon the administration of the prodrug 5-FC. The prolonged stability can be attributed to the fact that the internalized fusion protein is entrapped in the particular compartment(s) that are free from the cytoplasmic degradation machinery. To further develop and commercialize this system, we propose the following 2 specific aims: (1) to construct deletion mutants and modified versions of CD-CS for increasing yield, stability and activity of the fusion protein; and (2) to optimize the production of the modified CD-CS protein and to determine their effectiveness in scale-up preparations for product manufacturing and future clinical trials. This novel prodrug targeting therapy is effective, technically simple, non-invasive and cost effective, which should have enormous commercial potential.