Research Institute for Children, Children's Hospital, New Orleans, LA 70118, USA.
Cancer Gene Ther. 2012 Dec;19(12):828-38. doi: 10.1038/cgt.2012.66. Epub 2012 Oct 19.
The INSM1 gene encodes a transcriptional repressor that is exclusively expressed in neuronal and neuroendocrine tissue during embryonic development that is re-activated in neuroendocrine tumors. Using the 1.7 kbp INSM1 promoter, an adenoviral HSV thymidine kinase gene therapy was tested for the treatment of neuroendocrine tumors. An unforeseen interference on the INSM1 promoter specificity from the adenoviral genome was observed. Attempts were made to protect the INSM1 promoter from the influence of essential adenoviral sequences and to further enhance the tissue specificity of the INSM1 promoter region. Using the chicken β-globin HS4 insulator sequence, we eliminated off-target tissue expression from the Ad-INSM1 promoter-luciferase2 constructs in vivo. In addition, inclusion of two copies of the mouse nicotinic acetylcholine receptor (n(AchR)) neuronal-restrictive silencer element (NRSE) reduced nonspecific activation of the INSM1 promoter both in vitro and in vivo. Further, inclusion of both the HS4 insulator with the n(AchR) 2 × NRSE modification showed a two log increase in luciferase activity measured from the NCI-H1155 xenograft tumors compared with the original adenovirus construct. The alterations increase the therapeutic potential of adenoviral INSM1 promoter-driven suicide gene therapy for the treatment of a variety of neuroendocrine tumors.
INSM1 基因编码一种转录抑制剂，在胚胎发育过程中仅在神经元和神经内分泌组织中表达，在神经内分泌肿瘤中重新激活。使用 1.7 kbp INSM1 启动子，测试了腺病毒 HSV 胸苷激酶基因治疗神经内分泌肿瘤的效果。观察到腺病毒基因组对 INSM1 启动子特异性的意外干扰。试图保护 INSM1 启动子免受必需腺病毒序列的影响，并进一步增强 INSM1 启动子区域的组织特异性。使用鸡β-珠蛋白 HS4 绝缘子序列，我们消除了体内 Ad-INSM1 启动子-荧光素酶 2 构建体的非靶向组织表达。此外，包含两个拷贝的小鼠烟碱型乙酰胆碱受体（n（AchR））神经元限制性沉默元件（NRSE）可减少 INSM1 启动子在体外和体内的非特异性激活。此外，包含 HS4 绝缘子和 n（AchR）2×NRSE 修饰可使 NCI-H1155 异种移植肿瘤中测量的荧光素酶活性增加两个对数级，与原始腺病毒构建体相比。这些改变增加了腺病毒 INSM1 启动子驱动自杀基因治疗治疗各种神经内分泌肿瘤的治疗潜力。
