Recombinant BCG-Ag85A enhances antitumor immunity and controls melanoma progression through multimodal immune activation in mice

Melanoma is an aggressive form of skin cancer characterized by high metastatic potential and increasing incidence and mortality worldwide. Although significant advances have been achieved with immune checkpoint inhibitors and targeted therapies, effective preventive and long-term therapeutic strategies remain limited. Bacillus Calmette-Guérin (BCG), widely used in bladder cancer immunotherapy, exhibits potent immunostimulatory properties and has emerged as a promising platform for recombinant cancer vaccines. Among the candidate antigens, Antigen 85A (Ag85A), derived from Mycobacterium bovis , is notable for its strong immunogenicity and ability to induce robust Th1-mediated cellular immune responses. In this study, recombinant BCG strains overexpressing efficacy was evaluated in melanoma models in vitro and in vivo . Gene expression analysis by RT-qPCR revealed that rBCG- Ag85A induced systemic immune activation, characterized by increased Il1B and Trl4 expression in splenocytes. In tumor tissues, rBCG- Ag85A significantly upregulated genes associated with apoptosis ( Bax , Bcl2 ), oxidative stress ( Sod1 , Cat ), inflammatory and immune signaling pathways ( Tlr4 , Nfkb , Il12 , Il1b , Casp1 ), and modulation of pathways involved in cellular metabolism ( Mtor ) indicating enhanced modulation of the tumor microenvironment. Functionally, these molecular and immunological effects were associated with reduced tumor progression, slower tumor growth, and improved survival in B16F10 melanoma-bearing mice. Collectively, these findings demonstrate that rBCG- Ag85A promotes multifaceted antitumor activity through the modulation of apoptosis, inflammation and oxidative stress-related gene expression, highlighting its potential as a promising prophylactic vaccine strategy to prevent cutaneous malignant melanoma and supporting future studies aimed at elucidating the immune cell populations involved and optimizing combinatorial therapeutic approaches. Antitumor effects of recombinant BCG- Ag85A in a melanoma model. rBCG- Ag85A enhanced systemic immune activation and modulated apoptosis, oxidative stress, inflammatory and immune-related pathways in tumor tissue, resulting in reduced tumor growth and improved survival in B16F10 melanoma-bearing mice. These findings highlight rBCG-Ag85A as a promising immunotherapeutic platform for cutaneous melanoma. Created in BioRender.