BackgroundSARS-CoV-2 infection involves disturbing multiple molecular pathways related to immunity and cellular functions. PIM1 is a serine/threonine-protein kinase found to be involved in the pathogenesis of several viral infections. One PIM1 substrate, Myc, was reported to interact with TMPRSS2, which is crucial for SARS-CoV-2 cell entry. PIM1 inhibitors were reported to have antiviral activity through multiple mechanisms related to immunity and proliferation. This study aimed to evaluate the antiviral activity of2-pyridone PIM1 inhibitoragainst SARS-CoV-2 and its potential role in hindering the progression of COVID-19. It also aimed to assess PIM1 inhibitor’s effect on the expression of several genes of Notch signaling and Wnt pathways. In vitro study was conducted on Vero-E6 cells infected by SARS-CoV-2 “NRC-03-nhCoV” virus. Protein–protein interaction of the study genes was assessed to evaluate their relation to cell proliferation and immunity. The effect of2-pyridone PIM1 inhibitortreatment on viral load and mRNA expression of target genes was assessed at three time points.ResultsTreatment with2-pyridone PIM1 inhibitorshowed potential antiviral activity against SARS-CoV-2 (IC50of 37.255 µg/ml), significantly lowering the viral load. Functional enrichments of the studied genes include negative regulation of growth rate, several biological processes involved in cell proliferation, and Interleukin-4 production, with interleukin-6 as a predicted functional partner. These results suggest an interplay between study genes with relation to cell proliferation and immunity. Following in vitro SARS-CoV-2 infection, Notch pathway genes, CTNNB1, SUMO1, and TDG, were found to be overexpressed compared to uninfected cells. Treatment with2-pyridone PIM1 inhibitorsignificantly lowers the expression levels of study genes, restoring Notch1 and BCL9 to the control level while decreasing Notch2 and CTNNB1 below control levels.Conclusion2-pyridone PIM1 inhibitorcould hinder cellular entry of SARS-CoV-2 and modulate several pathways implicated in immunity, suggesting a potential benefit in the development of anti-SARS-CoV-2 therapeutic approach.Supplementary InformationThe online version contains supplementary material available at 10.1186/s43141-023-00520-x.