Synthesis, Evaluation, and Effect of Anti-CD38 Single-chain Immunotoxin on HL-60 acute myeloid Leukemia cell Line

Acute myeloid leukemia (AML) remains a highly aggressive hematologic malignancy with poor prognosis, especially in elderly patients. Targeted immunotherapies have emerged as promising alternatives to conventional chemotherapy and monoclonal antibodies, which are often limited by poor pharmacokinetics and off-target effects. In this study, a novel immunotoxin was designed by fusing a single-chain antibody (scFv) specific for CD38 with a low-immunogenic mutant form of bouganin, a plant-derived ribosome-inactivating protein.

The recombinant constructs were expressed in E. coli, purified using affinity chromatography, and validated by SDS-PAGE and Western blot analysis.

Functional assays demonstrated that the immunotoxin significantly reduced viability and induced apoptosis in CD38-positive HL-60 cells compared to both untreated controls and single-chain antibody treatments. Flow cytometry confirmed increased apoptotic cell death, while gene expression analysis revealed upregulation of the pro-apoptotic Bax gene alongside downregulation of anti-apoptotic BCL2 and STAT3, indicating activation of apoptosis-related pathways.

Collectively, these findings highlight the superior cytotoxicity and specificity of the CD38-targeted immunotoxin, suggesting its potential as an effective therapeutic strategy against AML.