Transductionally Redirected /Transcriptionally Restricte

腺病毒载体是癌症基因治疗方案最受欢迎的载体之一。然而，腺病毒通过柯萨奇和腺病毒受体感染细胞，后者通常在正常细胞上比在肿瘤细胞上更广泛地表达。我们利用全身注射CMVfLuc后的生物发光成像来监测腺病毒通过sCAREGF转导的“去靶向”和“重定向”。SCAR-EGF是一种双特异性重定向分子，含有与表皮生长因子相连的CAR的可溶性部分。非侵入性光学成像显示，系统注射[Ad.CMVfLuc][SCAR-EGF]复合体降低了感染肝脏的能力，但可以感染EGF受体阳性的异种移植物。COX-2在大多数正常组织中不表达，但在许多肿瘤中过表达。我们使用了非侵入性的 光学成像技术检测瘤内注射Ad.COX2fLuc(AN)后Flc的表达 表达COX-2启动子荧光素酶的腺病毒)。肝脏不表达。相反，在COX-2阳性的肿瘤中有表达。结直肠癌(CRC)是癌症死亡的主要原因。结直肠癌常转移至肝脏。结直肠癌肝转移常同时过度表达癌胚抗原(CEA)和COX-2。我们将结合我们的经验--包括转导“去靶向”和“重定向”，以及限制COX-2基因。 表达-开发治疗结直肠癌肝转移的有效方法。我们将建立异种结直肠癌肝转移模型。我们将创建Ad.COX2.fLucTK.COX2，这是一种腺病毒，表达包含萤火虫荧光素酶和HSV1-胸苷激酶(FLucTK)的融合蛋白。COX-2启动子和COX-2mRNA3‘非翻译区共同调控了结直肠癌肿瘤限制性fLucTK的表达。转导肝脏非靶向和Ad.COX2.fLuTK.COX2转导重定向针对结直肠癌移植瘤转移将通过SCAR-f-αCEA优化，SCAR-f-αCEA是一种包含可溶性CAR、噬菌体T4纤维蛋白三聚结构域和单链抗CEA抗体的重组分子。联合SCAR-f-αCEA转导肝靶向、转导肿瘤重定向和COX-2限制 在结直肠癌肝转移中的表达应优化fLucTK的表达。HSV1-TK/更昔洛韦治疗应能有效根除结直肠癌转移，副作用最小。我们将使用生物发光成像来监测限制性的fLucTK表达，使用Renilla荧光素酶标记的异种移植的生物发光成像和124I-antiCEA微体/microPET成像来监测HSV1-TK/GCV治疗期间的肿瘤负荷，并使用FDG和Flt/microPET来监测早期治疗反应。我们的目标是在人类疾病的临床模型中优化这一治疗方案来治疗结直肠癌肝转移。

Adenovirus vectors are among the most popular vehicles for cancer gene therapy protocols. However, adenovirus infect cells via the Coxsackie and Adenovirus Receptor, which is often more extensively expressed on normal cells then on tumors. We utilized bioluminescent imaging following systemic Ad.CMVfLuc administration to monitor adenovirus transductional "untargeting" and "retargeting" by sCAREGF. sCAR-EGF is a bi-specific retargeting molecule containing the soluble portion of CAR linked to epidermal growth factor. Noninvasive optical imaging demonstrates that systemically injected [Ad.CMVfLuc] [sCAR-EGF] complexes have reduced ability to infect liver, but can infect EGF receptor-positive xenografts. COX-2 is not expressed in most normal tissues, but is overexpressed in many tumors. We used noninvasive optical imaging to examine fLuc expression following intratumoral injection of Ad.COX2fLuc (an adenovirus expressing luciferase from the COX-2 promoter). No expression occurs in liver. In contrast, expression occurs in COX-2 positive tumors. Colorectal cancer (CRC) is a leading cause of cancer death. CRC frequently metastasizes to liver. CRC liver metastases often over-express both carcinoembryonic antigen (CEA) and COX-2. We will combine our experience - both with transductional "untargeting" and "retargeting", and with restricted COX-2 gene expression - to develop effective therapies for CRC hepatic metastases. We will establish xenograft CRC models of liver metastasis. We will create Ad.COX2.fLucTK.COX2, an adenovirus that expresses a fusion protein containing both firefly luciferase and HSV1-thymidine kinase (fLucTK). CRC tumor-restricted fLucTK expression will result from regulation by both the COX-2 promoter and the COX-2 mRNA 3' untranslated region. Transductional liver untargeting and Ad.COX2.fLuTK.COX2 transductional retargeting to CRC xenograft metastases will be optimized with sCAR-f-alphaCEA, a recombinant molecule containing soluble CAR, the phage T4 fibritin trimerization domain and a single-chain anti-CEA antibody. Combined sCAR-f-alphaCEA transductional liver untargeting, transductional tumor retargeting and COX-2 restricted expression should optimize fLucTK expression in CRC hepatic metastases. HSV1-TK/ganciclovir therapy should then effectively eradicate CRC metastases, with minimal side effects. We will employ bioluminescent imaging to monitor restricted fLucTK expression, bioluminescent imaging from xenografts marked with Renilla luciferase and 124I-antiCEA minibody/microPET imaging to monitor tumor burden during HSV1- TK/GCV therapy, and FDG and FLT/microPET to monitor early therapeutic responses. Our goal is to optimize this therapeutic protocol for CRC hepatic metastases, in apreclinical model of human disease.