Lee Sung Hyun, Hao Yajing, Gui Tong, Dotti Gianpietro, Savoldo Barbara, Zou Fei, Kafri Tal
Gene Therapy Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
Department of Biostatistics, University of North Carolina, Chapel Hill, North Carolina, USA.
Adv Cell Gene Ther. 2022;2022. doi: 10.1155/2022/6435077. Epub 2022 May 31.
For more than a decade, genetically engineered autologous T-cells have been successfully employed as immunotherapy drugs for patients with incurable blood cancers. The active components in some of these game-changing medicines are autologous T-cells that express viral vector-delivered chimeric antigen receptors (CARs), which specifically target proteins that are preferentially expressed on cancer cells. Some of these therapeutic CAR expressing T-cells (CAR-Ts) are engineered via transduction with -retroviral vectors (-RVVs) produced in a stable producer cell line that was derived from murine PG13 packaging cells (ATCC CRL-10686). Earlier studies reported on the copackaging of murine virus-like 30S RNA (VL30) genomes with -retroviral vectors generated in murine stable packaging cells. In an earlier study, VL30 mRNA was found to enhance the metastatic potential of human melanoma cells. These findings raise biosafety concerns regarding the possibility that therapeutic CAR-Ts have been inadvertently contaminated with potentially oncogenic VL30 retrotransposons. In this study, we demonstrated the presence of infectious VL30 particles in PG13 cell-conditioned media and observed the ability of these particles to deliver transcriptionally active VL30 genomes to human cells. Notably, VL30 genomes packaged by HIV-1-based vector particles transduced naïve human cells in culture. Furthermore, we detected the transfer and expression of VL30 genomes in clinical-grade CAR-T cells generated by transduction with PG13 cell-derived -retroviral vectors. Our findings raise biosafety concerns regarding the use of murine packaging cell lines in ongoing clinical applications.
十多年来，基因工程改造的自体T细胞已成功用作治疗无法治愈的血癌患者的免疫治疗药物。其中一些具有变革性的药物的活性成分是自体T细胞，它们表达由病毒载体递送的嵌合抗原受体（CAR），这些受体特异性靶向癌细胞上优先表达的蛋白质。一些表达治疗性CAR的T细胞（CAR-T细胞）是通过用源自鼠PG13包装细胞（ATCC CRL-10686）的稳定生产细胞系产生的逆转录病毒载体（-RVV）进行转导而构建的。早期研究报道了鼠病毒样30S RNA（VL30）基因组与在鼠稳定包装细胞中产生的逆转录病毒载体的共包装情况。在一项早期研究中，发现VL30 mRNA可增强人黑色素瘤细胞的转移潜能。这些发现引发了对治疗性CAR-T细胞可能被潜在致癌的VL30逆转座子意外污染的生物安全担忧。在本研究中，我们证明了PG13细胞条件培养基中存在具有感染性的VL30颗粒，并观察到这些颗粒将转录活性VL30基因组递送至人细胞的能力。值得注意的是，由基于HIV-1的载体颗粒包装的VL30基因组在培养中可转导未接触过的人细胞。此外，我们在通过用PG13细胞衍生的逆转录病毒载体转导产生的临床级CAR-T细胞中检测到了VL30基因组的转移和表达。我们的发现引发了对在当前临床应用中使用鼠包装细胞系的生物安全担忧。
