Bevec D, Dobrovnik M, Hauber J, Böhnlein E
Sandoz Research Institute, Vienna, Austria.
Proc Natl Acad Sci U S A. 1992 Oct 15;89(20):9870-4. doi: 10.1073/pnas.89.20.9870.
Human immunodeficiency virus type 1 (HIV-1) is the causative agent of the acquired immunodeficiency syndrome (AIDS). Currently, no satisfactory treatment for this viral disease is available. Somatic gene therapy has been proposed as an alternative to conventional therapies. Several antiviral gene therapy approaches including ribozymes, antisense inhibition, and RNA-decoy strategies, as well as dominant-negative mutants of HIV-1 proteins (Gag, Tat, and Rev) have been suggested. To prove the concept of trans-dominant inhibition of HIV-1 replication, we transduced CEM cells with a retroviral vector encoding a dominant-negative rev gene. Amplification of integrase-specific proviral sequences from high molecular weight DNA indicated successful HIV-1 human T-lymphotropic virus type IIIB (HTLV-IIIB) infection of all cells. In contrast to CEM cells and CEM cells expressing the rev wild-type (wt) gene, infection of two CEM-RevM10 clones with HIV-1 did not result in the release of significant levels of p24 Gag antigen as measured by antigen capture assay, indicating a block in HIV-1 replication due to the presence of the trans-dominant Rev protein. Furthermore, the parental CEM cells as well as CEM cells expressing the Rev wt protein were effectively killed in the course of the HIV-1 infection, whereas all CEM cells expressing the RevM10 protein were unaffected in their growth rate.
人类免疫缺陷病毒1型(HIV-1)是获得性免疫缺陷综合征(AIDS)的病原体。目前,尚无针对这种病毒性疾病的令人满意的治疗方法。体细胞基因治疗已被提议作为传统疗法的替代方法。已经提出了几种抗病毒基因治疗方法,包括核酶、反义抑制和RNA诱饵策略,以及HIV-1蛋白(Gag、Tat和Rev)的显性负突变体。为了证明对HIV-1复制进行反式显性抑制的概念,我们用编码显性负rev基因的逆转录病毒载体转导CEM细胞。从高分子量DNA中扩增整合酶特异性前病毒序列表明所有细胞均成功感染了HIV-1人类嗜T淋巴细胞病毒IIIB型(HTLV-IIIB)。与CEM细胞和表达rev野生型(wt)基因的CEM细胞相比,用HIV-1感染两个CEM-RevM10克隆并未导致通过抗原捕获测定法测得的显著水平的p24 Gag抗原释放,这表明由于反式显性Rev蛋白的存在导致HIV-1复制受阻。此外,亲代CEM细胞以及表达Rev wt蛋白的CEM细胞在HIV-1感染过程中被有效杀死,而所有表达RevM10蛋白的CEM细胞的生长速率均未受影响。