The motivation behind this paper comes from the need to better understand the molecular mechanisms and HPI driving Mpox pathogenesis, especially given its growing threat to global health ¹. While previous research has looked into various aspects of Mpox biology, there is still a lack of comprehensive insights into host-virus interactions at the systems biology level ^2,3. This study fills that gap by combining Weighted Gene Co-expression Network Analysis (WGCNA) ^4,5 with HPI analysis ^6,7, offering a novel approach to studying Mpox ^2,3. WGCNA identifies key gene modules and regulatory hubs impacted by Mpox infection, while HPI analysis reveals how viral proteins interact with host components to alter cellular processes ⁸. Together, these methods provide a systems-level view of how Mpox reshapes host molecular networks, focusing on the transcriptional and signaling regulators involved in immune evasion and viral persistence ⁹. Our findings not only deepen the understanding of Mpox immunopathogenesis but also identify potential therapeutic targets, such as kinase inhibitors, hormone-based therapies, and agents that influence PI3K/AKT and STAT3 signaling. This combined approach is a significant contribution to Mpox research, bridging molecular virology with computational biology to develop targeted antiviral strategies.

Inspiration

The Mpox outbreak of 2022, which resulted in over 94,000 confirmed cases worldwide, highlighted the urgent need for effective antiviral treatments ^10,11. With limited therapeutic options and the virus's ability to rapidly evolve, there was a clear gap in our preparedness for emerging infectious diseases. In this context, drug repurposing became a promising strategy to identify potential treatments using existing medications. Our work is driven by this need, using advanced bioinformatics tools to pinpoint critical molecular targets and pathways involved in the infection and immune response. By uncovering these targets, we aim to identify new drug candidates and strengthen our ability to respond quickly to future outbreaks, ultimately contributing to better global health preparedness.

Advancements and Scientific Significance

In this in silico study, we constructed a HPI network by utilizing experimentally validated protein-protein interaction data from publicly available repositories such as STRING, BioGRID, and IntAct, alongside virus-host interaction databases specific to orthopoxviruses. This allowed us to develop a high-confidence HPI network encompassing 1,120 interactions between 76 Mpox proteins and 314 human proteins. Centrality analysis of the network revealed hub proteins, including EGFR, TRAF6, and CASP8, which appear to play significant roles in immune regulation. Specifically, EGFR was found to be involved in viral entry and immune suppression, while TRAF6 and CASP8 were linked to inflammatory responses and apoptosis pathways. These findings highlight potential targets for immunomodulatory therapies aimed at mitigating the effects of Mpox infection.

Drug Target Identification

we predicted 11 kinases, including JAK1, TYK2, and MAPK1, as potential regulatory targets involved in Mpox pathogenesis. Additionally, we identified 15 transcription factors, such as IRF7, STAT1, and NFKB1, which play a critical role in driving antiviral gene expression. These transcription factors are essential for regulating immune responses during infection. To further explore therapeutic avenues, we employed a drug repurposing strategy by mapping druggable targets to existing FDA-approved compounds using the DrugBank and LINCS databases. Several candidate drugs were proposed, including Ruxolitinib, a JAK1/2 inhibitor, to suppress hyperinflammatory responses, Ribavirin and Favipiravir as antiviral agents targeting the viral replication machinery, and Baricitinib as an immunomodulator to mitigate cytokine storms. These findings offer promising therapeutic options for the treatment of Mpox, particularly in reducing inflammation and controlling viral replication.

The Future

This study paves the way for future research by integrating multi-omics data to provide a deeper understanding of Mpox pathogenesis. Expanding beyond transcriptomics, it includes proteomics and epigenomics, offering a more comprehensive view of how the virus interacts with the host. Additionally, refining HPI predictions by using CRISPR-Cas9 screening and experimental validation of the identified targets will further confirm the key molecular players involved in the disease ¹². In terms of therapeutic development, investigating combination therapies that target both immune response regulators and viral entry pathways could improve treatment outcomes ¹³. Furthermore, the development of immunomodulatory vaccines, which incorporate antiviral cytokine inducers, offers a novel approach to enhance the immune response against Mpox. To ensure these therapeutic strategies translate into clinical practice, conducting in vivo studies and clinical trials for the identified drug candidates is essential ^14,15. Lastly, establishing real-time monitoring systems powered by computational models will help prepare for future outbreaks and enable more effective responses. These approaches collectively aim to advance our understanding of Mpox and lead to the development of targeted therapies and vaccines.

Summary

By combining HPI and WGCNA, we uncovered key molecular targets and druggable pathways in Mpox infection. This integrative approach identified potential antiviral agents, including kinase inhibitors, IFN pathway activators, and immune regulators. Our study highlights a data-driven strategy for drug repurposing, paving the way for innovative therapeutic solutions against future viral outbreaks.

About Authors

Saber Imani is an independent Assistant Professor at Shulan International Medical College, Zhejiang Shuren University, Hangzhou, China. He leads a research lab focused on advancing the field of mRNA-based cancer vaccines and translational oncology. His work primarily revolves around understanding the molecular mechanisms of cancer and viral diseases, with a particular emphasis on developing innovative therapeutic strategies. Through his leadership, his lab is making significant contributions to the development of effective treatments and vaccines, to address global health challenges. Sargol Aminnezhad is a postdoctoral researcher at Tarbiat Modares University, where she leads a team dedicated to advancing AI applications in medicine, including innovative projects like skin cancer diagnostic tools and insulin management apps. Holding a Ph.D. in Cellular and Molecular Biology and extensive postdoctoral experience in cancer genomics and systems biology, her research encompasses bioinformatics, vaccine design, drug discovery, and biomarker identification. She is committed to integrating interdisciplinary approaches to advance immunology, microbiome studies, and precision medicine, leveraging AI-driven technologies to tackle healthcare challenges and enhance patient outcomes. Mazaher Maghsoudloo holds a Ph.D. in Bioinformatics from Tehran University, where he graduated in 2020. With a background in Computer Science, he combines computational expertise with a deep understanding of biological systems. Since 2023, he has been serving as a Research Associate at Southwest Medical University, working in the Key Laboratory of Epigenetics and Oncology within the Research Center for Preclinical Medicine. His research focuses on cancer biology, particularly on biomarker detection and the reconstruction of disease networks, with a special emphasis on cancer-related applications.

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