Revolutionizing gene editing techniques in medicine
siRNAs, saRNAs, miRNAs, CRISPR/Cas system
DOI:
https://doi.org/10.58445/rars.2194Keywords:
Gene therapy, RNA interference, Gene editing, small interfering RNA, small activating RNA, microRNA, CRISPR/CasAbstract
Gene therapy is a new modality of medical treatment that can treat or prevent diseases by directly modifying gene expression. This review focuses on four prominent mechanisms: small interfering RNAs (siRNAs), small activating RNAs (saRNAs), micro RNAs (miRNAs), and the CRISPR/Cas system. siRNA and miRNA silence gene expression by either degrading mRNA or repressing protein translation, respectively, while saRNA promotes gene expression through targeting promoter regions. CRISPR/Cas precisely edits genes through insertion, deletion, or correction of specific sequences. Although these mechanisms show significant therapeutic potential, challenges such as delivery barriers and off-target effects remain as obstacles. This review synthesizes findings from peer-reviewed articles found using Google Scholar, while focusing on research from the past 20 years. While exceptions were made for a few older articles due to their importance to the paper, their credibility was assessed through cross-referencing with reliable medical sources. Preliminary findings highlight the capability of siRNA and miRNA in silencing genes associated with various diseases, the potential of saRNA in activating therapeutic targets, and the precision of CRISPR/Cas in correcting genetic mutations. By providing a comprehensive evaluation of four innovative gene therapy approaches via their mechanisms of action, advantages, challenges, and therapeutic applications, this review highlights the need for continued innovation to overcome current limitations and revolutionize medicine.
References
B. E. Jaski, M. L. Jessup, D. M. Mancini, T. P. Cappola, D. F. Pauly, B. Greenberg, K. Borrow, H. Dittrich, K. M. Zsebo, R. J. Hajjar. Calcium upregulation by percutaneous administration of gene therapy in cardiac disease (CUPID Trial), a first-in-human phase 1/2 clinical trial. Journal of Cardiac Failure 15, 171-181 (2009).
W. Filipowicz, J. Paszkowski. Gene silencing. Brenner’s Encyclopedia of Genetics (Second Edition), 221-222 (2013).
S. M. Elbashir, J. Harborth, W. Lendeckel, A. Yalcin, K. Weber, T. Tuschl. Duplexes of 21-nucleotide RNAs mediated RNA interference in cultured mammalian cells. Nature 411, 494-498 (2001).
N. Agrawal, P. V. N. Dasaradhi, A. Mohmmed, P. Malhotra, R. K. Bhatnagar, S. K. Mukherjee. RNA interference: biology, mechanism, and applications. Microbiology and Molecular Biology Reviews 67, 657-686 (2003).
K. Gavrilov, W. M. Saltzman. Therapeutic siRNA: principles, challenges, and strategies. The Yale Journal of Biology and Medicine 85, 187-200 (2012).
A. Kwok, N. Raulf, N. Habib. Developing small activating RNA as a therapeutic: current challenges and promises. Therapeutic Delivery 10, 151-164 (2019).
F. Wahid, A. Shehzad, T. Khan, Y. Y. Kim. MicroRNAs: synthesis, mechanism, function, and recent clinical trials. Biochemica Et Biophysica Acta (BBA) - Molecular Cell Research 1803, 1231-1243 (2010).
M. Jie, T. Feng, W. Huang, M. Zhang, Y. Feng, H. Jiang, Z. Wen. Subcellular localization of miRNAs and implications in cellular homeostasis. Genes 12 (2021).
M. Segal, F. J. Slack. Challenges identifying efficacious miRNA therapeutics for cancer. Expert Opinion on Drug Discovery 15, 987 (2020).
N. Gohil, G. Bhattacharjee, N. L. Lam, S. D. Perli, V. Singh. CRISPR-Cas systems: challenges and future prospects. Progress in Molecular Biology and Translational Science 180, 141-151 (2021).
Y. Xu, Z. Li. CRISPR-Cas systems: overview, innovations and applications in human disease research and gene therapy. Computational and Structural Biotechnology 18, 2401-2415 (2020).
A. J. Hamilton, D. C. Baulcombe. A species of small antisense RNA in posttranscriptional gene silencing in plants. Science 286, 950-952 (1999).
T. Tuschl, P. D. Zamore, R. Lehmann, D. P. Bartel, P. A. Sharp. Targeted mRNA degradation by double-stranded RNA in vitro. Genes & Development 13, 3191-3197 (1999).
S. M. Elbashir, W. Lendeckel, T. Tuschl. RNA interference is mediated by 21- and 22- nucleotide RNAs. Genes & Development 15, 188-200 (2001).
P. D. Zamore, T. Tuschl, P. A. Sharp, D. P. Bartel. RNAi: double-stranded RNA directs the ATP-dependent cleavage of mRNA at 21 to 23 nucleotide intervals. Cell 101, 25-33 (2000).
B. L. Bass. Double-stranded RNA as a template for gene silencing. Cell 101, 235-238 (2000).
C. Lipardi, Q. Wei, B. M. Paterson. RNAi as random degradative PCR: siRNA primers convert mRNA into dsRNAs that are degraded to generate new siRNAs. Cell 107, 297-307 (2001).
T. Sijen, J. Fleenor, F. Simmer, K. L. Thijssen, S. Parrish, L. Timmons, R. H. Plasterk, A. Fire. On the role of RNA amplification in dsRNA-triggerend gene silencing. Cell 107, 465-476 (2001).
K. Ciechanowska, M. Pokornowska, A. Kurzyńska-Kokorniak. Genetic insight into the domain structure and functions of dicer-type ribonucleases. International Journal of Molecular Sciences 22, 616 (2021).
L. Gherardini, G. Bardi, M. Gennaro, T. Pizzorusso. Novel siRNA delivery strategy: a new “strand” in CNS translational medicine?. Cellular and Molecular Life Sciences 71, 1-20 (2013).
H. Kim, K. Cho, S. K. Lee, G. W. Kim. Apoptosis signal-regulating kinase 1 (Ask1) targeted small interfering RNA on ischemic neuronal cell death. Brain Research 1412, 73-78 (2011).
P. Cowled, R. Fitridge. Pathophysiology of reperfusion injury. Mechanisms of Vascular Disease: A Reference Book for Vascular Specialists (2011).
H. Yamaguchi, H. Wang. CHOP is involved in endoplasmic reticulum stress-induced apoptosis by enhancing DR5 expression in human carcinoma cells. Mechanisms of Signal Transduction 279, 45495-45502 (2004).
Z. He, R. P. Ostrowski, X. Sun, Q. Ma, B. Huang, Y. Zhan, J. H. Zhang. CHOP silencing reduces acute brain injury in the rat model of subarachnoid hemorrhage. American Heart Association Journals 43 (2011).
K. T. Al-Jamal, L. Gherardini, G. Bardi, A. Nunes, C. Guo, C. Bussy, M. A. Herrero, A. Bianco, M. Prato, K. Kostarelos, T. Pizzorusso. Functional motor recovery from brain ischemic insult by carbon nanotube-mediated siRNA silencing. Proceedings of the National Academy of Sciences 108, 10952-10957 (2011).
B. Tizon, S. Sahoo, H. Yu, S. Gauthier, A. R. Kumar, P. Mohan, M. Figliola, M. Pawlik, A. Grubb, Y. Uchiyama, U. Bandyopadhyay, A. M. Cuervo, R. A. Nixon, E. Levy. Induction of autophagy by Cystatin C: a mechanism that protects murine primary cortical neurons and neuronal cell lines. PLoS ONE 5, 9819 (2010).
A. L. Jackson, S. R. Bartz, J. Schelter, S. V. Kobayashi, J. Burchard, M. Mao, B. Li, G. Caveat, P. S. Linsley. Expression profiling reveals off-target gene regulation by RNAi. Nature Biotechnology 21, 636-637 (2003).
V. Portnoy, S. H. S. Lin, K. H. Li, A. Burlingame, Z. Hu, H. Li, L. Li. saRNA-guided Ago2 targets the RITA complex to promoters to stimulate transcription. Cell Research 26, 320-335 (2016).
X. Zhao, J. Voutila, S. Ghobrial, N. A. Habib, V. Reebye. Treatment of liver cancer by C/EBPA saRNA. RNA Activation 983, 189-194 (2017).
V. Reebye, P. Sætrom, P. J. Mintz, K. Huang, P. Swiderski, L. Peng, C. Liu, X. Liu, S. Lindkær-Jensen, D. Zacharoulis, N. Kostomitsopoulos, N. Kasahara, J. P. Nicholls, L. R. Jiao, M. Pai, D. R. Spalding, M. Mizandari, T. Chikovani, M. M. Emara, A. Haoudi, D. A. Tomalia, J. J. Rossi, N. A. Habib. Novel RNA oligonucleotie improves liver function and inhibits liver carcinogenesis in vivo. Hepatology 59, 216-227 (2013).
K. Yamamoto, K. Tateishi, Y. Kudo, T. Sato, S. Yamamoto, K. Miyabayashi, K. Matsusaka, Y. Asaoka, H. Ijichi, Y. Hirata, M. Otsuka, Y. Nakai, H. Isayama, T. Ikenoue, M. Kurokawa, M. Fukayama, N. Kokudo, M. Omata, K. Koike. Loss of histone demethylase KDM6B enhances aggressiveness of pancreatic cancer through downregulation of C/EBPα. Carcinogenesis 35, 2404-2414 (2014).
R. F. Place, J. Wang, E. J. Noonan, R. Meyers, M. Manoharan, K. Charisse, R. Duncan, V. Huang, X. Wang, L. Li. Formulation of small activating RNA into lipidoid nanoparticles inhibits xenograft prostate tumor growth by inducing p21 expression. Moleculary Therapy 1 (2012).
J. Voutila, V. Reebye, T. C. Roberts, P. Protopapa, P. Andrikakou, D. C. Blakey, R. Habib, H. Huber, P. Sætrom, J. J. Rossi, N. A. Habib. Development and mechanism of small activating RNA targeting CEBPA, a novel therapeutic in clinical trials for liver cancer. Molecular Therapy 25, 2705-2714 (2017).
C. P. Petersen, M. Bordeleau, J. Pelletier, P. A. Sharp. Short RNAs repress translation after initiation in mammalian cells. Molecular Cell 21, 533-542 (2006).
Y. Funakoshi, Y. Doi, N. Hosoda, N. Uchida, M. Osawa, I. Shimada, M. Tsujimoto, T. Suzuki, T. Katada, S. Hoshino. Mechanism of mRNA deadenylation: evidence for a molecular interplay between translation termination factor eRF3 and mRNA deadenylases. Genes and Development 21, 3135-3148 (2007).
Z. A. Syeda, S. S. S. Langden, C. Munkhzul, M. Lee, S. J. Song. Regulatory mechanism of MicroRNA expression in cancer. International Journal of Molecular Sciences 21, 1723 (2020).
G. Reid, S. C. Kao, N. Pavlakis, H. Brahmbhatt, J. MacDiarmid, S. Clarke, M. Boyer, N. Van Zandwijk. Clinical development of TargomiRs, a miRNA mimic-based treatment for patients with recurrent thoracic cancer. Epigenomics 8, 1079-1085 (2016).
Cleveland Clinic. Pleural Mesothelioma. https://my.clevelandclinic.org/health/diseases/15044-pleural-mesothelioma (2022).
M. Asmamaw, B. Zawdie. Mechanism and applications of CRISPR/Cas-9-mediated genome editing. Biologics: Targets and Therapy 15, 353-361 (2021).
F. Hille, E. Charpentier. CRISPR-Cas: biology mechanisms and relevance. Philosophical Transactions of the Royal Society of Biological Sciences 371 (2016).
A. Mir, A. Edraki, J. Lee, E. J. Sontheimer. Type II-C CRISPR-Cas9 biology, mechanism, and application. ACS Chemical Biology 13 (2017).
I. Gostimskaya. CRISPR-Cas9: a history of its discovery and ethical considerations of its use in genome editing. Biochemistry Moscow 87, 777-788 (2022).
E. V. Hillary, S. A. Caesar. A review on the mechanism and applications of CRISPR/Cas9/Cas12/Cas13/Cas14 proteins utilized for genome engineering. Molecular Biotechnology 65, 311-325 (2023).
A. Sharma, J. Boelens, M. Cancio, J. S. Hankins, P. Bhad, M. Azizy, A. Lewandowski, X. Zhao, S. Chitnis, R. Peddinti, Y. Zheng, N. Kapoor, F. Ciceri, T. Maclachlan, Y. Yang, Y Liu, J. Yuan, U. Naumann, V. W. C. Yu, S. C. Stevenson, S. D. Vita, J. L. Labelle. CRISPR-Cas9 editing of the HBG1 and HBG2 promoters to treat sickle cell disease. The New England Journal of Medicine 389 (2023).
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