A Unique Gene-Silencing Approach, Using an Intelligent RNA Expression Device (iRed), Results in Minimal Immune Stimulation When Given by Local Intrapleural Injection in Malignant Pleural Mesothelioma

Our plain-English summary of A Unique Gene-Silencing Approach, Using an Intelligent RNA Expression Device (iRed), Results in Minimal Immune Stimulation When Given by Local Intrapleural Injection in Malignant Pleural Mesothelioma, by H. Ando, N. Saito-Tarashima, A.S.A. Lila, et al (Molecules, 2020).

The study

• Authors: H. Ando, N. Saito-Tarashima, A.S.A. Lila, N. Kinjo, T. Shimizu, Y. Ishima, N. Minakawa, T. Ishida
• Journal: Molecules
• Published: 2020;25(7)
• Cited by: 5 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: We have recently introduced an intelligent RNA expression device (iRed), comprising the minimum essential components needed to transcribe short hairpin RNA (shRNA) in cells. Use of iRed efficiently produced shRNA molecules after transfection into cells and alleviated the innate immune stimulation following intravenous injection.

Methods: To study the usefulness of iRed for local injection, the engineered iRed encoding luciferase shRNA (Luc iRed), complexed with cationic liposomes (Luc iRed/liposome-complexes), was intrapleurally injected into an orthotopic mesothelioma mouse model.

Results: Luc iRed/liposome-complexes markedly suppressed the expression of a luciferase marker gene in pleurally disseminated mesothelioma cells. The suppressive efficiency was correlated with the expression level of shRNA within the mesothelioma cells. In addition, intrapleural injection of iRed/liposome-complexes did not induce IL-6 production in the pleural space and consequently in the blood compartment, although plasmid DNA (pDNA) or dsDNA (the natural construct for iRed) in the formulation did.

Conclusion: Local delivery of iRed could augment the in vivo gene silencing effect without eliciting pronounced innate immune stimulation. Our results might hold promise for widespread utilization of iRed as an RNAi-based therapeutic for intracelial malignant cancers.

In plain English

This laboratory and animal study tested an artificial DNA construct called an intelligent RNA expression device (iRed), designed to make short hairpin RNA (shRNA) inside cells, against plasmid DNA (pDNA) and natural double-stranded DNA (dsDNA). All were combined with cationic liposomes. In human pleural mesothelioma cells expressing firefly luciferase (MSTO-211H-Luc), all three constructs lowered luciferase activity compared with control DNA, with the rank order of RNAi activity Luc pDNA > Luc dsDNA > Luc iRed, matching the amount of shRNA produced. In mice bearing orthotopic pleural mesothelioma, two intrapleural doses (10 micrograms DNA per mouse per day, days 7 and 10) reduced tumour luciferase signal, the rank order of gene silencing being Luc pDNA > Luc iRed > Luc dsDNA; when corrected for molar amount, the inhibition rate was 40.4 plus or minus 1.54 percent per mol for pDNA, 1.0 plus or minus 1.42 for dsDNA and 1.65 plus or minus 0.56 for iRed. Four hours after injection, pDNA-complexes significantly raised IL-6 in pleural wash fluid; iRed-complexes produced the lowest serum IL-6 levels. The study involved no asbestos exposure.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory cell-culture experiments plus an animal experiment in an orthotopic mouse model of pleural mesothelioma
• Who was studied: Human pleural mesothelioma cells stably expressing firefly luciferase (MSTO-211H-Luc) in vitro, and male BALB/c nu/nu mice (5 weeks old) inoculated into the left pleural cavity with 1 x 10^6 MSTO-211H-Luc cells; groups of 5 mice; Tokushima University, Japan
• Exposure: Not an asbestos exposure study: the agents tested were luciferase shRNA-encoding iRed, pDNA, dsDNA and control DNA, each complexed with cationic liposomes (DOPE:cholesterol:DOTAP 3:3:4), given intrapleurally at 10 micrograms DNA per mouse per day on days 7 and 10 after tumour inoculation, or transfected in vitro at 0.5 pmol DNA per well
• Main results:
  • In MSTO-211H-Luc cells, Luc iRed, Luc pDNA and Luc dsDNA all significantly reduced luciferase activity versus control DNA, the rank order of RNAi activity being Luc pDNA > Luc dsDNA > Luc iRed
  • Amount of Luc shRNA transcribed in cells rose dose-dependently up to 72 h, with rank order Luc pDNA >> Luc dsDNA > Luc iRed, and correlated with the in vitro suppression of luciferase activity
  • In the tumour-bearing mice, the lowest luciferase activity was seen with Luc pDNA/liposome-complexes; rank order of in vivo gene silencing was Luc pDNA > Luc iRed > Luc dsDNA
  • Inhibition rates corrected for injected molar amount: Luc pDNA/liposome-complexes 40.4 plus or minus 1.54 percent per mol, Luc dsDNA 1.0 plus or minus 1.42, Luc iRed 1.65 plus or minus 0.56; pDNA significantly higher than the other two (p less than 0.001)
  • At 4 hours after intrapleural injection, Luc pDNA/liposome-complexes caused significant IL-6 production in pleural wash fluid, while dsDNA- and iRed-complexes raised IL-6 slightly but not significantly
  • Serum IL-6 levels were increased in all treated mice, with rank order Luc pDNA/liposome-complexes > Luc dsDNA/liposome-complexes > Luc iRed/liposome-complexes
  • No substantial body weight loss was observed in any treated mice over the treatment period
• The authors' conclusion: The authors concluded that local intrapleural delivery of iRed produced a therapeutically relevant gene-silencing effect with minimal innate immune stimulation, and suggested it could be an approach for treating malignant tumours in body cavities

Topics

The National Library of Medicine indexes this study under subjects our site covers: Mesothelioma.

Medical terms in this study, explained in our Lexicon: Pleural Mesothelioma, Mesothelioma.

Related studies

• An RNAi therapeutic, DFP-10825, for intraperitoneal and intrapleural malignant cancers — H. Ando, T. Ishida (2020)
• MSLN gene silencing has an anti-malignant effect on cell lines overexpressing mesothelin deriving from malignant pleural mesothelioma — O. Melaiu, J. Stebbing, et al. (2014)
• Gene Silencing Using 4'-thioDNA as an Artificial Template to Synthesize Short Hairpin RNA Without Inducing a Detectable Innate Immune Response — N. Tarashima, H. Ando, et al. (2016)
• Elucidating the immunomodulatory roles and mechanisms of CUL4B in the immune system: a comprehensive review — S. He, Z. Wang, et al. (2025)
• HDAC1-mSin3a-NCOR1, Dnmt3b-HDAC1-Egr1 and Dnmt1-PCNA-UHRF1-G9a regulate the NY-ESO1 gene expression — P.F. Cartron, C. Blanquart, et al. (2013)
• BioKnife, a uPA activity-dependent oncolytic Sendai virus, eliminates pleural spread of malignant mesothelioma via simultaneous stimulation of uPA expression — Y. Morodomi, T. Yano, et al. (2012)

Browse more research in Molecules, or return to medical research.