Effect of Bleomycin Hydrolase Expression in Tumor Tissue on the Therapeutic Effectiveness of Electrochemotherapy

Background: Electrochemotherapy (ECT) enhances the intracellular delivery of chemotherapeutic agents, most notably bleomycin, through electroporation. Despite high response rates, tumor sensitivity to ECT varies, highlighting the need for predictive biomarkers. Bleomycin hydrolase (BLMH), an enzyme that metabolically inactivates bleomycin, may influence treatment outcomes. This study investigated the relationship between BLMH expression and bleomycin-based ECT effectiveness. Methods: BLMH expression was evaluated at the mRNA and protein levels in six murine tumor cell lines and their corresponding syngeneic tumors using qPCR, immunofluorescence, and immunohistochemistry. Correlations between BLMH expression and tumor response to ECT were assessed both in vitro and in vivo. Results: BLMH expression varied significantly among tumor models, without consistent patterns across cancer types. In vitro, BLMH mRNA levels strongly correlated with IC30 values for bleomycin (R = 0.74), while the correlation weakened at IC50 doses, suggesting enzyme saturation. In vivo, BLMH expression levels moderately correlated with complete tumor response rates following ECT (R = 0.50). Differences between in vitro and in vivo expression highlighted the role of the tumor microenvironment. Conclusions: High BLMH expression reduces tumor sensitivity to bleomycin-based ECT, supporting its role as a predictive biomarker. Measuring BLMH levels may help stratify patients and personalize ECT application; however, it is not the sole factor for response prediction. Future studies in clinical tumor samples are warranted to evaluate its predictive value and to develop integrated biomarker models.