Transductionally Redirected /Transcriptionally Restricte

腺病毒载体是癌症基因治疗方案中最受欢迎的载体之一。然而，腺病毒通过科萨基和腺病毒受体感染细胞，其通常在正常细胞上比在肿瘤上更广泛地表达。我们在全身性Ad.CMVfLuc施用后利用生物发光成像来监测腺病毒转导通过sCAREGF的“非靶向”和“重靶向”。sCAR-EGF是一种双特异性重靶向分子，含有与表皮生长因子连接的CAR可溶性部分。非侵入性光学成像表明，全身注射的[Ad.CMVfLuc] [sCAR-EGF]复合物感染肝脏的能力降低，但可以感染EGF受体阳性异种移植物。考克斯-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 光学成像检查肿瘤内注射Ad.COX2fLuc后fLuc的表达（一种免疫抑制剂）。 从考克斯-2启动子表达荧光素酶的腺病毒）。在肝脏中无表达。相反，表达发生在考克斯-2阳性肿瘤中。结直肠癌（CRC）是癌症死亡的主要原因。CRC经常转移到肝脏。CRC肝转移瘤通常过表达癌胚抗原（CEA）和考克斯-2。我们将联合收割机结合我们的经验--转导“非靶向”和“重靶向”，以及限制性考克斯-2基因， 表达-为CRC肝转移开发有效的疗法。我们将建立大肠癌肝转移的异种移植模型。我们将创建Ad.COX2.fLucTK.COX2，这是一种表达包含萤火虫荧光素酶和HSV 1-胸苷激酶（fLucTK）的融合蛋白的腺病毒。CRC肿瘤限制性fLucTK表达将由考克斯-2启动子和考克斯-2 mRNA 3'非翻译区的调节引起。将用sCAR-f-alphaCEA（一种含有可溶性CAR、噬菌体T4纤维蛋白三聚化结构域和单链抗CEA抗体的重组分子）优化转导肝脏非靶向和Ad.COX2.fLuTK.COX2转导重靶向CRC异种移植物转移。联合sCAR-f-alphaCEA转导肝脏非靶向、转导肿瘤重靶向和考克斯-2限制 表达应该优化CRC肝转移中fLucTK的表达。HSV 1-TK/更昔洛韦治疗应有效根除CRC转移，副作用最小。我们将采用生物发光成像监测限制性fLucTK表达，采用海肾荧光素酶标记的异种移植物的生物发光成像和124 I-抗CEA微抗体/microPET成像监测HSV 1- TK/GCV治疗期间的肿瘤负荷，并采用FDG和FLT/microPET监测早期治疗反应。我们的目标是在人类疾病的临床前模型中优化这种治疗方案用于CRC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.