Recombination-deletion between homologous cassettes in retrovirus is suppressed via a strategy of degenerate codon substitution.
Recombination-deletion between homologous cassettes in retrovirus is suppressed via a strategy of degenerate codon substitution.
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DOI：
10.1038/mtm.2014.22
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发表时间：
2014-07-09
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基因治疗方案中的转导和表达程序可以最佳地转移多于单个基因以校正缺陷和/或将新功能传递到受体细胞或生物体。这可以通过用两个（或更多个）载体转导来实现，或者更有效地在单个载体中实现。偶尔，共表达同源基因或具有共享同源性区域的嵌合蛋白可能是有用的。逆转录病毒科包括用于基因转移的显性载体系统（例如，γ-逆转录病毒和慢病毒）并且能够进行这种多基因表达。然而，已知当结构域在病毒基因组内复制时，这些相同的病毒具有有效的重组-缺失。这个问题可以通过诉诸双向量策略（双链双向量）来避免，但代价是成本、便利性和效率。采用嵌合抗原受体系统作为一个例子，我们证实，在一个单一的γ-逆转录病毒载体（双链单载体）的同源结构域的两个基因的共表达导致重复序列之间的重组删除，切除相当于一个嵌合抗原受体。在这里，我们表明，在双链单载体格式的简并密码子取代策略有效地抑制了载体内缺失的损失与救援的平衡基因的共表达，通过最大限度地减少重复结构域之间的序列同源性，并保留最终的蛋白质序列。
Transduction and expression procedures in gene therapy protocols may optimally transfer more than a single gene to correct a defect and/or transmit new functions to recipient cells or organisms. This may be accomplished by transduction with two (or more) vectors, or, more efficiently, in a single vector. Occasionally, it may be useful to coexpress homologous genes or chimeric proteins with regions of shared homology. Retroviridae include the dominant vector systems for gene transfer (e.g., gamma-retro and lentiviruses) and are capable of such multigene expression. However, these same viruses are known for efficient recombination–deletion when domains are duplicated within the viral genome. This problem can be averted by resorting to two-vector strategies (two-chain two-vector), but at a penalty to cost, convenience, and efficiency. Employing a chimeric antigen receptor system as an example, we confirm that coexpression of two genes with homologous domains in a single gamma-retroviral vector (two-chain single-vector) leads to recombination–deletion between repeated sequences, excising the equivalent of one of the chimeric antigen receptors. Here, we show that a degenerate codon substitution strategy in the two-chain single-vector format efficiently suppressed intravector deletional loss with rescue of balanced gene coexpression by minimizing sequence homology between repeated domains and preserving the final protein sequence.
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DOI：
10.1158/1078-0432.ccr-07-4910
发表时间：
2008-12-15
期刊：
Clinical cancer research : an official journal of the American Association for Cancer Research
影响因子：
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作者：
Emtage PC;Lo AS;Gomes EM;Liu DL;Gonzalo-Daganzo RM;Junghans RP
通讯作者：
Junghans RP
影响因子：
46.9
作者：
Maher, J;Brentjens, RJ;Sadelain, M
通讯作者：
Sadelain, M
影响因子：
8.7
作者：
Jensen MC;Riddell SR
通讯作者：
Riddell SR
影响因子：
56.9
作者：
Sun, T;Patoine, C;Walsh, CA
通讯作者：
Walsh, CA
影响因子：
64.5
作者：
JUNGHANS, RP;BOONE, LR;SKALKA, AM
通讯作者：
SKALKA, AM