Analysis and validation of critical signatures and immune cell infiltration characteristics in doxorubicin-induced cardiotoxicity by integrating bioinformatics and machine learning

Abstract

Doxorubicin-induced cardiotoxicity (DIC) is a severe side reaction in cancer chemotherapy that greatly impacts the well-being of cancer patients. Currently, there is still an insufficiency of effective and reliable biomarkers in the field of clinical practice for the early detection of doxorubicin-induced cardiotoxicity. We obtained high-throughput sequencing data from the Gene Expression Omnibus (GEO) database and performed data analysis and visualization using R software, GO, KEGG and Cytoscape. Machine learning methods and weighted gene coexpression network (WGCNA) were used to identify key genes for diagnostic model construction. Receiver operating characteristic (ROC) analysis and a nomogram were used to assess their diagnostic values. A multiregulatory network was built to reveal the possible regulatory relationships of critical signatures. Cell-cype identification by estimating relative subsets of RNA transcript (CIBERSORT) analysis was used to investigate differential immune cell infiltration. Additionally, a doxorubicin-induced cardiotoxicity cell model was constructed to investigate the relationship between the identified genes and doxorubicin-induced cardiotoxicity. Finally, among the 3713 differentially expressed genes, three hub genes ( CSGALNACT1, ZNF296 and FANCB) were identified. A nomogram and ROC curves based on three hub genes showed excellent diagnostic predictive performance. The regulatory network analysis showed that the TFs CREB1, EP300, FLI1, FOXA1, MAX, and MAZ modulated CSGALNACT1, ZNF296 and FANCB. An analysis of immune cell infiltration indicated that many immune cells (activated NK cells, M0 macrophages, activated dendritic cells and neutrophils) might be related to the progression of DOX-induced cardiotoxicity. Furthermore, there may be various degrees of correlation between the three critical signatures and immune cells. RT‒qPCR in vitro demonstrated that the mRNA expression of CSGALNACT1 and ZNF296 was significantly upregulated, while FANCB was significantly downregulated in doxorubicin-treated cardiomyocytes. Our results suggested that the differential expression of CSGALNACT1, ZNF296 and FANCB is associated with cardiotoxicity and is involved in immune cell infiltration in doxorubicin-induced cardiotoxicity. They might be potential biomarkers for the early occurrence of doxorubicin-induced cardiotoxicity.