Poly(ε-caprolactone) Nanoparticle Tumor-Lysate Vaccination in Mice Generates Hybridoma-Derived Antibodies Enabling Breast Cancer Diagnosis and Chemotherapy Synergy.

Tumor-lysate vaccines can capture tumor heterogeneity; however, their effectiveness may be reduced by antigen instability and short antigen presentation. Here, we aimed to improve antigen protection and prolong presentation by using a slow-degrading polymeric nanocarrier and an approved adjuvant. We encapsulated breast cancer cell lysates (MCF-7 and MDA-MB-231) in poly(ε-caprolactone) (PCL) nanoparticles using a double-emulsion (w/o/w) method and co-administered them with alum. We then characterized particle size, PDI, zeta potential, morphology, and in vitro release. Next, we evaluated nitric oxide (NO), TNF-α/IL-10 responses, and cytocompatibility in J774 macrophages. Finally, we quantified serum antibody titers in Balb/c mice after six biweekly immunizations, generated hybridomas, purified IgG, and tested antibody-mediated cytotoxicity alone and together with doxorubicin. PCL nanoparticles were ~220-255 nm (PDI 0.10-0.19; ζ -2 to -3 mV) and released ~90-95% of encapsulated lysate by 800 h (~33 days). Encapsulated lysate (40 μg/mL) modestly increased NO versus control and increased further with alum ( < 0.05). TNF-α increased 7.4-9.72-fold, whereas IL-10 rose 2.82-3.11-fold. Importantly, encapsulated antigen + alum produced the highest ELISA responses after the sixth dose (6.36-fold for MCF-7 and 7.00-fold for MDA-MB-231 versus control; < 0.05). Hybridoma-derived antibody signals increased through day 42, and Protein G purification yielded up to ~395 μg and ~318 μg IgG. Purified antibodies reduced cell viability, and viability decreased further when antibodies were combined with doxorubicin (to ~31.6% in MCF-7 and ~40.3% in MDA-MB-231). Overall, sustained PCL-mediated antigen release combined with alum strengthened humoral responses to tumor lysate and enabled recovery of functional antibodies with diagnostic capture and in vitro cytotoxic activity. In future work, key mechanistic steps such as lymph-node trafficking and cross-presentation should be tested directly.