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Antagomir is chemically-modified single-strand miRNA inhibitor functioning by blocking miRNA regulation of target gene expression efficiently. They are synthesized to reduce the ability of endogenous miRNAs to silence target mRNA transcripts. They can down-regulate the corresponding endogenous miRNAs by either local or systemic injection into the animals.
Creative Biogene's microDOWN™ miRNA antagomir is single-strand miRNA inhibitor carrying the chemically modifications functioning by blocking miRNA regulation of target gene expression efficiently. The strand of the antagomir has 2 phosphorothioates at the 5' end, 4 phosphorothioates, 1 cholesterol group at the 3' end, and full-length nucleotide 2'-methoxy modification. Stability of microDOWN™ miRNA antagomir appears to be significantly higher than miRNA inhibitors. It exhibits enhanced cellular uptake, stability and regulatory activity and is recommended for miRNA functional studies in vitro and in vivo.
Figure 1. Mechanism of microDOWN™ miRNA antagomir.
Figure 2. qRT-PCR and western blot assays show the mRNA (A) and protein (B) expressions of TOB1 after transfection with antagomir-NC and antagomir-25.
Our microDOWN™ miRNA antagomir has unparalleled advantages
Resuspension Protocol
Cell Assay
Recommended amounts of microDOWN™ antagomir for your cell assay.
| miRNA | Cell | Antagomir | References |
| miR-122 | Huh-7 WT, CNS3 | 50 nM | Shan Y, et al. Gastroenterology. 2007 |
| miR-206 | HeLa, MDA-231, MCF-7 | 20-50 nM | Brian D, et al., Mol Ther. 2007 |
| miR-27a, miR-451 | A2780 | 100 nM | Zhu H, et al. Biochem Pharmacol. 2008 |
| miR-126, miR-126 | LNCaP | 10~30 nM | Alla Musiyenko, et al. J Mol Med. 2008 |
| miR-184, miR-205 | HeLa, HEKs | 1 μM | Jia Yu, et al. PNAS. 2008 |
| miR-221, miR-222 | Me665/1 | 50~250 nM | Federica et al. Cancer Res. 2008 |
| miR-196b | bone marrow cells | 100 nM | Relja Popovic, et al. Blood. 2009 |
| miR-221, miR-222 | hepatocellular carcinoma-derived | 100 μM | Pascal Pineau, et al. PNAS. 2010 |
| miR-214 | MC3T3-E1, hFOB | 200 μM | Xiaogang Wang, et al. Nat Med. 2012 |
| miR-214 | MC3T3-E1 | 200 nM | Wang X, et al. Nat Med. 2013 |
In Vivo Delivery
| miRNA | Animal | Delivery | Doses | Volume | Injection Times | Detection | References |
| miR-122 miR-192 | 6-week-old mice | tail-vein injections | 80 mg per kg body weight | 0.2 ml per injection | 1 to 3 consecutive days | 24 h after the last injection | Krützfeldt J, et al. Nature. 2005 |
| miR-16 | 6-week-old mice | Intracerebral injection | ~800 ng per mouse | 0.2 ml per injection | a single injection | 72 h after injection | Jan Krützfeldt, et al., Nucleic Acids Research. 2007 |
| miR-133 | 8-week-old mice | minipump implantation | 80 mg per kg body weight | N/A | a single infusion | 1 month after minipump implantation | Alessandra Care, et al. Nature Medicine. 2007 |
| miR-17-5p | 6-week-old nude mice | intratumorally injection | 0.2 mg per mouse | 0.1 ml | three times per week for two weeks | 2 weeks after the first injection | Laura Fontana, et al. PLoS ONE. 2008 |
| miR- 21 | 10~12 weeks old mice | jugular vein catheter | 80 mg per kg body weight | N/A | injected daily for three days | 3 weeks after the first injection | Thomas Thum, et al. Nature. 2008 |
| miR-221 miR-222 | 6-week-old mice | intratumorally injection | 1 ug of each antagomir | 40 μl | at day 0, 5 and 9, for a injections per tumor | N/A | Neri Mercatelli, et al. PLoS ONE. 2008 |
| miR-145 | rat | minipump implantation | totally 960 pmol each rat | 48 μl | N/A | N/A | Ashuthosh Dharap, et al. J. Cereb. Blood Flow Metab. 2009 |
| miR-2861 | 6-week-old female mice | tail vein injection | 80 mg/kg body weight | 0.2 ml per injection | days 1 to 3 for 3 consecutive weeks | 4 days, 3 weeks, and 6 weeks after the last injection | Li H, Xie H, et al. J Clin Invest. 2009 |
| miR-92 | mice | intravenous injection | 8 mg per kg of body weight | 0.2 ml per injection | at days 0, 2, 4, 7, 9 of the left coronary artery | 14 days | Bonauer A, et al. Science. 2009 |
| miR-151-5p | nude mice | transplantion | 2x106 cells transfected by antagomir | N/A | 5 weeks after injection | Jie Ding, et al. Nature cell biology. 2010 | |
| miR-125b | 6~8 weeks old nude mice | subcutaneously injection | 2x105 cells transfected by antagomir | N/A | 4 weeks after injection | LinHui Liang, et al. Hepatology. 2010 | |
| miR-335 | 4-week-old nude mice | Intratumorally injection | 4 mg/ml | 50 μl | every two days for 2 weeks | 2 weeks after the first injection | Minfeng Shu, et al. Molecular Cancer. 2011 |
Case Study 1
Myotonic dystrophy type 1 (DM1) is a rare neuromuscular disease caused by expansion of unstable CTG repeats in the noncoding region of the DMPK gene. CUG expansion in mutant DMPK transcripts sequesters MBNL1 protein into ribonuclear foci. Antisense oligonucleotides targeting miR-218 were previously shown to enhance MBNL1 expression and rescue the phenotype in disease models. Here, researchers provide preclinical characterization of the antagomiR-218 molecule using HSALR mouse models and patient-derived myotubes. In HSALR, antagomiR-218 reached 40-60 pM 2 weeks after injection, rescued molecular and functional phenotypes in a dose- and time-dependent manner, and showed a favorable toxicity profile after a single subcutaneous injection. Furthermore, miR-218 is significantly overexpressed in DM1 muscle samples and disease models and thus may contribute to the disease phenotype through MBNL1- and 2-independent pathways. These results highlight the therapeutic potential of miR-218 blocking oligonucleotides as potential treatments for DM1.
Figure 3. Duration of antagomiR-218 therapeutic effects upon single subcutaneous injection. (AntagomiR-218 and scramble control were synthesized by Creative Biogene.) (Cerro-Herreros E, et al., 2021)
Case Study 2
Defects in the epithelial tight junction (TJ) barrier contribute to the development of disease-associated intestinal inflammation. Interleukin 1 beta (IL1B) increases intestinal permeability in mice. Studies here show that IL1B mRNA and MIR200C-3p levels are elevated in intestinal tissue from ulcerative colitis patients and colitis mice, thereby reducing the expression of occludin by enterocytes and increasing TJ permeability. antagomiR-200C prevents the reduction of occludin in intestinal epithelial cells and intestinal tissue in mice with colitis, thereby maintaining the TJ barrier.
Figure 4. Effects of mouse DSS-induced colitis on the cooperation between IL-1β, miR-200c3p, and mouse intestinal permeability. Effects of oral administration of DSS for 7 days on IL-1β mRNA expression (A), miR-200c-3p expression (B), and occludin mRNA expression (C). The colonic perfusion of DSS mice to analyze the effect of antagomiR-200c on DSS-induced increase in IL-1β mRNA expression (D), increase in miR-200c-3p expression (E), decrease in occludin mRNA expression (F) and increase in dextran 10kD flux (G). Effect of antagomiR-200c gavage on body weight of DSS mice (H), histological observation of the effect of antagomiR-200c on mouse colon, (I). (AntagomiR-200c and non-specific antagomiR were designed by Creative biogene) (Rawat M, et al., 2020)
Case Study 3
Myotonic dystrophy type 1 (DM1) is a chronic, debilitating rare genetic disorder that originates from the expansion of noncoding CTG repeats in the dystrophia myotonica protein kinase (DMPK) gene. This expansion is pathogenic when DMPK transcripts contain 50 or more repeats due to sequestration in the myoblading-like (MBNL) protein family. Here, researchers demonstrate the therapeutic effects of antagomiR-23b administered as a treatment in human skeletal actin long repeat (HSALR) mice over time. Subcutaneous administration of antagomiR-23b upregulates MBNL1 protein expression and rescues splicing alterations, grip strength, and myotonia in a dose-dependent manner with long-lasting effects.
Figure 5. Comparative Study between Subcutaneous and Intravenous Delivery of AntagomiRs in Mice. (AntagomiR against miR-23b and the SC control was purchased from Creative Biogene.) (Cerro-Herreros E, et al., 2020)
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