Hybridoma-derived monoclonal antibody-conjugated carboplatin-loaded chitosan nanoparticles targeting A549 cells: an in vitro proof-of-concept study

Monoclonal antibodies (mAbs) produced against non-small cell lung cancer (NSCLC) cells have the potential to serve as targeting ligands for selective drug delivery. This study aimed to develop hybridoma-derived murine monoclonal antibodies against the A549 human lung adenocarcinoma cell line and evaluate their application in carboplatin-loaded chitosan nanoparticles (CCnp). Combining hybridoma-derived monoclonal antibodies with drug-loaded nanocarriers may enhance the selective delivery of chemotherapeutic agents to tumor cells while providing a receptor-agnostic platform for targeted therapy. Female Balb/c mice immunized with A549 (adenocarcinoma human alveolar basal epithelial) cells and fused the elicited murine splenocytes with Sp2/0 (myeloma) cells, screening the resultant hybridomas in Hypoxanthine Aminopterin Thymidine (HAT) medium. CCnp were prepared via spontaneous emulsification and conjugated with the purified mAbs (Ab-CCnp). In vitro cytotoxicity was evaluated against A549 cells using dye exclusion, sulphorhodamine B (SRB), and MTT assays; selectivity was additionally assessed via MTT on L-132 normal human lung cells. Three stable hybridoma cell lines (C4, E4, and G5) producing mAbs against A549 antigens were established, and the antibodies purified by Protein A affinity chromatography. Physicochemical characterization guided selection of the 0.25 mg/ml drug-polymer batch for antibody conjugation. Ab-CCnp showed significantly higher in vitro cytotoxicity against A549 cells than unconjugated CCnp: MTT growth inhibition increased from 48.28 ± 2.31% to 79.02 ± 2.87% at 25 µg/well. Cytotoxicity against non-target L-132 cells remained significantly lower, indicating good selectivity. Hybridoma-derived mAb-conjugated carboplatin-loaded chitosan nanoparticles demonstrated improved, in vitro cytotoxicity against A549 cells. This study provides A549-cell-line-limited proof-of-concept study, which warrants further in vivo evaluation.