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Background OTUB1 is a deubiquitin enzyme that plays important role in cancer-related events. Prompted by the lack of information about OTUB1's potential as a breast cancer molecular marker, this study is conducted to evaluate the significance of OTUB1 as a diagnostic tool based on its expression in MCF-7, as well as to validate its potential as a therapeutic target using network-based analysis. Methods Relative expression of OTUB1 in MCF-7 and MCF-10A was compared using bioinformatics analysis on microarray datasets obtained from Gene Expression Omnibus and validated by qPCR experiment. Protein-protein interaction (PPI) network was constructed using Cytoscape ver3.6.0 to analyse the topological position of OTUB1 in cancer and differentially expressed genes (DEGs) neighbourhoods. Results Microarray datasets analysis showed that OTUB1 expression value is significantly higher in MCF-7 in seven out of 16 probes with log(FC) value ranging from 0.24 to 1.43. This result is consistent with qPCR analysis, with qPCR showing a larger log(FC) value of 2.44–4.95. Despite a significant difference in some microarray probes and qPCR experiment, the fold change value is too small to ensure sensitivity in breast cancer diagnosis. Meanwhile, PPI network analysis revealed that OTUB1 is the immediate neighbour for 31 cancer proteins and DEGs ESR1 and BIRC3, a strategic topological position for perturbing cancer interactome. Conclusion Taken together, our findings suggest that OTUB1 is an unfavourable candidate for breast cancer diagnosis marker, but it could be a very promising molecular target for breast cancer therapeutic intervention.