Laboratory of Cell Therapy of Cancer, Department of Oncological Sciences, University of Torino Medical School, Institute for Cancer Research and Treatment, Provinciale 142, 10060 Candiolo, Torino, Italy.
J Biotechnol. 2011 Dec 10;156(3):218-26. doi: 10.1016/j.jbiotec.2011.09.001. Epub 2011 Sep 10.
The proteasome system restricts lentiviral transduction of stem cells. We exploited proteasome inhibition as a strategy to enhance transduction of both hematopoietic stem cells (HSC) and T lymphocytes with low dose or large-size lentiviral vectors (LV). HSC showed higher transduction efficiency if transiently exposed to proteasome inhibitor MG132 (41.8% vs 10.7%, p<0.0001). Treatment with MG132 (0.5 μM) retained its beneficial effect with 3 different LV of increasing size up to 10.9 Kb (p<0.01). We extended, for the first time, the application of proteasome inhibition to the transduction of T lymphocytes. A transient exposure to MG132 significantly improved lentiviral T-cell transduction. The mean percentage of transduced T cells progressively increased from 13.5% of untreated cells, to 21% (p=0.3), 30% (p=0.03) and 37% (p=0.01) of T lymphocytes that were pre-treated with MG132 at 0.1, 0.5 and 1 μM, respectively. MG132 did not affect viability or functionality of HSC or T cells, nor significantly increased the number of integrated vector copies. Transient proteasome inhibition appears as a new procedure to safely enhance lentiviral transduction of HSC and T lymphocytes with low viral doses. This approach could be useful in settings where the use of large size vectors may impair optimal viral production.
蛋白酶体系统限制了慢病毒对干细胞的转导。我们利用蛋白酶体抑制作为一种策略，来增强低剂量或大尺寸慢病毒载体（LV）对造血干细胞（HSC）和 T 淋巴细胞的转导效率。如果 HSC 短暂暴露于蛋白酶体抑制剂 MG132（41.8%对 10.7%，p<0.0001），则其转导效率更高。用 3 种不同大小不断增加至 10.9kb 的 LV 处理 MG132（0.5 μM），保留了其有益的效果（p<0.01）。我们首次将蛋白酶体抑制的应用扩展到 T 淋巴细胞的转导中。短暂暴露于 MG132 可显著改善慢病毒 T 细胞的转导。经 MG132 预处理的 T 细胞的转导率从未经处理细胞的 13.5%逐渐增加到 21%（p=0.3）、30%（p=0.03）和 37%（p=0.01），分别用 0.1、0.5 和 1μM 的 MG132 预处理。MG132 不影响 HSC 或 T 细胞的活力或功能，也不会显著增加整合载体拷贝数。短暂的蛋白酶体抑制似乎是一种安全增强低病毒剂量 HSC 和 T 淋巴细胞慢病毒转导的新方法。在大尺寸载体可能会损害最佳病毒产生的情况下，这种方法可能很有用。
