Metformin Suppresses SARS-CoV-2 in Cell Culture

Parthasarathy et al., bioRxiv, doi:10.1101/2021.11.18.469078, Nov 2021

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Metformin for COVID-19

3rd treatment shown to reduce risk in July 2020, now with p < 0.00000000001 from 105 studies.

Lower risk for mortality, ventilation, ICU, hospitalization, progression, and recovery.

No treatment is 100% effective. Protocols combine treatments.

5,700+ studies for 161 treatments. c19early.org

In Vitro study showing metformin inhibits SARS-CoV-2 in Caco2 cells. Metformin reduced viral titers by nearly 99%, and by about 90% when cells were treated prior to infection.

15 preclinical studies support the efficacy of metformin for COVID-19:

4 In Silico studies1-4

7 In Vitro studies2,5-10

4 In Vivo animal studies5,6,8,11

A systematic review and meta-analysis of 15 non-COVID-19 preclinical studies showed that metformin inhibits pulmonary inflammation and oxidative stress, minimizes lung injury, and improves survival in animal models of acute respiratory distress syndrome (ARDS) or acute lung injury (ALI)12. Metformin inhibits SARS-CoV-2 in vitro9,10, minimizes LPS-induced cytokine storm in a mouse model11, minimizes lung damage and fibrosis in a mouse model of LPS-induced ARDS8, may protect against SARS-CoV-2-induced neurological disorders7, may be beneficial via inhibitory effects on ORF3a-mediated inflammasome activation13, reduces UUO and FAN-induced kidney fibrosis8, increases mitochondrial function and decreases TGF-β-induced fibrosis, apoptosis, and inflammation markers in lung epithelial cells8, may reduce inflammation, oxidative stress, and thrombosis via regulating glucose metabolism2, attenuates spike protein S1-induced inflammatory response and α-synuclein aggregation6, may reduce COVID-19 severity and long COVID by inhibiting NETosis via suppression of protein kinase C activation14, enhances interferon responses and reduces SARS-CoV-2 infection and inflammation in diabetic models by suppressing HIF-1α signaling5, and may improve outcomes via modulation of immune responses with increased anti-inflammatory T lymphocyte gene expression and via enhanced gut microbiota diversity15.

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1. Tavares et al., Investigation of Interactions Between the Protein MPro and the Vanadium Complex VO(metf)2∙H2O: A Computational Approach for COVID-19 Treatment, Biophysica, doi:10.3390/biophysica5010004.mdpi.com, doi.org.

2. Hou et al., Metformin is a potential therapeutic for COVID-19/LUAD by regulating glucose metabolism, Scientific Reports, doi:10.1038/s41598-024-63081-0.nature.com, doi.org.

3. Agamah et al., Network-based multi-omics-disease-drug associations reveal drug repurposing candidates for COVID-19 disease phases, ScienceOpen, doi:10.58647/DRUGARXIV.PR000010.v1.scienceopen.com, doi.org.

4. Lockwood, T., Coordination chemistry suggests that independently observed benefits of metformin and Zn2+ against COVID-19 are not independent, BioMetals, doi:10.1007/s10534-024-00590-5.springer.com, doi.org.

5. Joshi et al., Severe SARS‐CoV‐2 infection in diabetes was rescued in mice supplemented with metformin and/or αKG, and patients taking metformin, via HIF1α‐IFN axis, Clinical and Translational Medicine, doi:10.1002/ctm2.70275.wiley.com, doi.org.

6. Chang et al., SARS-CoV-2 Spike Protein 1 Causes Aggregation of α-Synuclein via Microglia-Induced Inflammation and Production of Mitochondrial ROS: Potential Therapeutic Applications of Metformin, Biomedicines, doi:10.3390/biomedicines12061223.mdpi.com, doi.org.

7. Yang et al., SARS-CoV-2 infection causes dopaminergic neuron senescence, Cell Stem Cell, doi:10.1016/j.stem.2023.12.012.sciencedirect.com, doi.org.

8. Miguel et al., Enhanced fatty acid oxidation through metformin and baicalin as therapy for COVID-19 and associated inflammatory states in lung and kidney, Redox Biology, doi:10.1016/j.redox.2023.102957.sciencedirect.com, doi.org.

9. Ventura-López et al., Treatment with metformin glycinate reduces SARS-CoV-2 viral load: An in vitro model and randomized, double-blind, Phase IIb clinical trial, Biomedicine & Pharmacotherapy, doi:10.1016/j.biopha.2022.113223.sciencedirect.com, doi.org.

10. Parthasarathy et al., Metformin Suppresses SARS-CoV-2 in Cell Culture, bioRxiv, doi:10.1101/2021.11.18.469078.biorxiv.org, doi.org.

11. Taher et al., Anti‑inflammatory effect of metformin against an experimental model of LPS‑induced cytokine storm, Experimental and Therapeutic Medicine, doi:10.3892/etm.2023.12114.spandidos-publications.com, doi.org.

12. Wang et al., Effects of metformin on acute respiratory distress syndrome in preclinical studies: a systematic review and meta-analysis, Frontiers in Pharmacology, doi:10.3389/fphar.2023.1215307.frontiersin.org, doi.org.

13. Zhang et al., SARS-CoV-2 ORF3a Protein as a Therapeutic Target against COVID-19 and Long-Term Post-Infection Effects, Pathogens, doi:10.3390/pathogens13010075.mdpi.com, doi.org.

14. Monsalve et al., NETosis: A key player in autoimmunity, COVID-19, and long COVID, Journal of Translational Autoimmunity, doi:10.1016/j.jtauto.2025.100280.sciencedirect.com, doi.org.

15. Petakh et al., Metformin Alters mRNA Expression of FOXP3, RORC, and TBX21 and Modulates Gut Microbiota in COVID-19 Patients with Type 2 Diabetes, Viruses, doi:10.3390/v16020281.mdpi.com, doi.org.

Parthasarathy et al., 19 Nov 2021, preprint, 3 authors.

In Vitro studies are an important part of preclinical research, however results may be very different in vivo.

This Paper Metformin All

Metformin Suppresses SARS-CoV-2 in Cell Culture

Haripriya Parthasarathy, Dixit Tandel, Abdul Hamid Siddiqui, Krishnan H Harshan

doi:10.1101/2021.11.18.469078

People with diabetes are reported to have a higher risk of experiencing severe COVID-19 complications. Metformin, a first-line medication for type 2 diabetes, has antiviral properties. Some studies have indicated its prognostic potential in COVID-19. Here, we report that metformin significantly inhibits SARS-CoV-2 growth in cell culture models. SARS-CoV-2 infection of gut epithelial cell line, Caco2, resulted in higher phosphorylation of AMPK. Metformin reduced viral titers in the infected cells by nearly 99%, and by about 90% when cells were treated prior to infection. Metformin pre-treatment resulted in further phosphorylation of AMPK and caused a ten-fold reduction of viral titers indicating its potential in preventing naïve infections. Confirming the positive impact of AMPK activation, another AMPK activator AICAR substantially inhibited of viral titers and, AMPK inhibitor Compound C, augmented it considerably. Metformin treatment post-SARS-CoV-2 infection resulted in nearly hundred-fold reduction of viral titers, indicating that the antiviral potency of the drug is far higher in infected cells, while still being able to reduce fresh infection. Metformin displayed SARS-CoV-2 TCID50 and TCID90 at 3.5 and 8.9 mM, respectively. In conclusion, our study demonstrates that metformin is very effective in limiting the replication of SARS-CoV-2 in cell culture and thus possibly could offer double benefits to diabetic COVID-19 patients by lowering both blood glucose levels and viral load.

AMPK is activated. In this context, it is interesting to note that N protein was detected at modestly higher abundance during pharmacological activation of AMPK unlike the viral RNA and infectious titer, indicating that viral protein translation is not inhibited during the treatments. However, a significant drop in N levels during the postinfection treatment suggested an overall inhibition of viral life-stages concurrent with an overall drop of cellular activities indicated by MTT results. Inhibition of metabolic activities particularly in the infected cells upon post-infection metformin treatment indicated that metformin treatment specifically targeted the infected cells for destruction. This could be viewed as beneficial to the system fighting to clear the virus from it. Although multiple reports show AMPK as the major effector of metformin action, it is now well established that metformin exerts its affects through other pathways such as PKA and FBPase-1 mediated regulation as well (Pernicova & Korbonits, 2014) . Further study into the mechanism SARS-CoV-2 inhibition by metformin could pave the way for it to be a possible therapeutic target for COVID-patients. From a clinical perspective, our study provides some answers to the favorable prognosis of metformin-treated diabetic patients who contracted COVID-19. Institutional biosafety Institutional biosafety clearance was obtained for the experiments pertaining to SARS-CoV-2.

References

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