Abstract
DDX3 RNA helicase is intensively studied as a therapeutic target due to participation in the replication of some viruses and involvement in cancer progression. Here we used transcriptome analysis to estimate the primary response of hepatocytes to different levels of RNAi-mediated knockdown of DDX3 RNA helicase both in vitro and in vivo. We found that a strong reduction of DDX3 protein (>85%) led to similar changes in vitro and in vivo—deregulation of the cell cycle and Wnt and cadherin pathways. Also, we observed the appearance of dead hepatocytes in the healthy liver and a decrease of cell viability in vitro after prolonged treatment. However, more modest downregulation of the DDX3 protein (60–65%) showed discordant results in vitro and in vivo—similar changes in vitro as in the case of strong knockdown and a different phenotype in vivo. These results demonstrate that the level of DDX3 protein can dramatically influence the cell phenotype in vivo and the decrease of DDX3, for more than 85% leads to cell death in normal tissues, which should be taken into account during the drug development of DDX3 inhibitors.
Highlights
Recent progress in the consolidation of systems biology, high-throughput methods and bioinformatics has significantly promoted drug development
We found the activation of caspase-3 after the administration of lipid nanoparticles (LNPs)-siRNA #5 for 13 days, which correlates with the hepatocyte death detected by morphological study (Figure 2B)
The same approach was used in vivo: we analyzed the transcriptome of the murine liver at days 6 and 13 after DDX3 KD by LNP-siRNA #5 and
Summary
Recent progress in the consolidation of systems biology, high-throughput methods and bioinformatics has significantly promoted drug development. Multiple omics techniques can provide exhaustive data on the changes of RNA, proteins, lipids and metabolites both in vitro and in vivo Among these methods, transcriptome analysis increases our ability to evaluate the primary response of the cell to any external or internal signal both in vitro and in vivo, providing additional valuable information to common cellular assays and long-term animal studies. More modest downregulation of DDX3 protein (60–65%) resulted in discordant results between gene expression in vitro and in vivo; in vitro data for both siRNAs were similar, while in vivo, the phenotype was different in comparison to data obtained for the strong reduction of DDX3 protein These results demonstrate that the amount of DDX3 protein can dramatically influence the phenotype in vivo, which should be taken into account during drug development
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