Existing approaches for the integration and expression of genes of interest in a desired human cellular context are marred by the safety concerns related to either the random nature of viral-mediated integration or unpredictable pattern of gene expression in currently employed targeted genomic integration sites. Disadvantages of these methods lead to their limited use in clinical practice, thus encouraging future research in identifying novel human genomic sites that allow for predictable and safe expression of genes of interest. We conducted a bioinformatic search followed by experimental validation of novel genomic sites and identified two that demonstrated stable expression of integrated reporter and therapeutic genes without detrimental changes to cellular transcriptome. The cell-type agnostic criteria used in our bioinformatic search suggest wide-scale applicability of our sites for engineering of a diverse range of tissues for therapeutic as well as enhancement purposes, including modified T-cells for cancer therapy and engineered skin to ameliorate inherited diseases and aging. Additionally, the stable and robust levels of gene expression from identified sites allow for their use in industry-scale biomanufacturing of desired proteins in human cells.

中文翻译：

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发现和验证用于基因和细胞疗法的新型人类基因组安全港

在所需的人类细胞环境中整合和表达目的基因的现有方法受到与目前介导的靶向基因组整合位点中病毒介导的整合的随机性或基因表达的不可预测模式有关的安全性的关注。这些方法的缺点导致它们在临床实践中的使用受到限制，因此鼓励了未来的研究来确定新的人类基因组位点，以允许可预测和安全地表达感兴趣的基因。我们进行了生物信息学搜索，然后对新型基因组位点进行了实验验证，并确定了两个能够证明整合的报告基因和治疗基因稳定表达而对细胞转录组无不利影响的基因。在我们的生物信息学搜索中使用的细胞类型不可知标准表明，我们的网站可广泛用于各种组织工程的治疗和增强用途，包括用于癌症治疗的改良T细胞和用于改善遗传性疾病的经工程改造的皮肤和老化。另外，来自已鉴定位点的基因表达的稳定和鲁棒水平允许其用于人细胞中所需蛋白质的工业规模生物制造。