Kashani-Sabet Mohammed, Liu Yong, Fong Sylvia, Desprez Pierre-Yves, Liu Shuqing, Tu Guanghuan, Nosrati Mehdi, Handumrongkul Chakkrapong, Liggitt Denny, Thor Ann D, Debs Robert J
Auerback Melanoma Research Laboratory, Cutaneous Oncology Program, University of California at San Francisco Cancer Center and Department of Dermatology, University of California, San Francisco, CA 94115, USA.
Proc Natl Acad Sci U S A. 2002 Mar 19;99(6):3878-83. doi: 10.1073/pnas.002025599. Epub 2002 Mar 12.
To date, functional genomic studies have been confined to either cell-based assays or germline mutations, using transgenic or knockout animals. However, these approaches are often unable either to recapitulate complex biologic phenotypes, such as tumor metastasis, or to identify the specific genes and functional pathways that produce serious diseases in adult animals. Although the transcription factor NF-kappaB transactivates many metastasis-related genes in cells, the precise genes and functional-pathways through which NF-kappaB regulates metastasis in tumor-bearing hosts are poorly understood. Here, we show that the systemic delivery of plasmid-based ribozymes targeting NF-kappaB in adult, tumor-bearing mice suppressed NF-kappaB expression in metastatic melanoma cells, as well as in normal cell types, and significantly reduced metastatic spread. Plasmid-based ribozymes suppressed target-gene expression with sequence specificity not achievable by using synthetic oligonucleotide-based approaches. NF-kappaB seemed to regulate tumor metastasis through invasion-related, rather than angiogenesis-, cell-cycle- or apoptosis-related pathways in tumor-bearing mice. Furthermore, ribozymes targeting either of the NF-kappaB-regulated genes, integrin beta(3) or PECAM-1 (a ligand-receptor pair linked to cell adhesion), reduced tumor metastasis at a level comparable to NF-kappaB. These studies demonstrate the utility of gene targeting by means of systemic, plasmid-based ribozymes to dissect out the functional genomics of complex biologic phenotypes, including tumor metastasis.
迄今为止，功能基因组学研究一直局限于基于细胞的分析方法或种系突变，采用转基因或基因敲除动物。然而，这些方法往往既无法重现复杂的生物学表型，如肿瘤转移，也无法识别在成年动物中引发严重疾病的特定基因和功能途径。尽管转录因子核因子-κB（NF-κB）在细胞中可反式激活许多与转移相关的基因，但对于NF-κB在荷瘤宿主中调节转移的确切基因和功能途径仍知之甚少。在此，我们表明，在成年荷瘤小鼠中系统性递送靶向NF-κB的基于质粒的核酶，可抑制转移性黑色素瘤细胞以及正常细胞类型中NF-κB的表达，并显著减少转移扩散。基于质粒的核酶以序列特异性抑制靶基因表达，这是使用基于合成寡核苷酸的方法无法实现的。在荷瘤小鼠中，NF-κB似乎通过与侵袭相关而非与血管生成、细胞周期或凋亡相关的途径来调节肿瘤转移。此外，靶向NF-κB调控基因整合素β3或血小板内皮细胞黏附分子-1（PECAM-1，一种与细胞黏附相关的配体-受体对）的核酶，可在与NF-κB相当的水平上减少肿瘤转移。这些研究证明了通过系统性的、基于质粒的核酶进行基因靶向以剖析包括肿瘤转移在内的复杂生物学表型的功能基因组学的实用性。
