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Targeted efficacy of BCG Hsp70-anti-CD123 immunoconjugate in childhood acute myeloid leukemia
Xiao-Ling Li1, Wenting Pei1, Chun-Lei Liu2
1Department of Pediatrics, Children's Hospital Affiliated to Shandong University (Jinan Children's Hospital), Huaiyin District, Jinan, Shandong, China.
Background:
Relapsed/refractory acute myeloid leukemia (AML) in children remains challenging due to leukemia stem cells (LSCs) and poor immunogenicity. This study aimed to construct a novel immunoconjugate combining BCG heat shock protein 70 (HSP70) with an anti-CD123 monoclonal antibody (MAb) and evaluate its targeted anti-leukemic efficacy and mechanisms.
Methods:
The conjugate (HSP70-MAb) was synthesized via SPDP crosslinker and characterized by ESI-MS and UV spectrophotometry. Antigen-binding activity was assessed by flow cytometry. In vitro, autologous T-cell proliferation (CFSE staining), IFN-γ secretion (ELISA), and cytotoxic activity (LDH release) were assessed. In vivo, the anti-leukemic effect was evaluated in a MOLM13 xenograft nude mouse model.
Results:
The HSP70-MAb conjugate showed high purity (96.2 ± 1.5)% and near 1:1 molar conjugation efficiency, retaining antigen-binding activity. In vitro, HSP70-MAb significantly enhanced autologous T-lymphocyte proliferation and IFN-γ secretion, and markedly increased CTL-mediated cytotoxicity against CD123⁺ AML cells in an E:T ratio-dependent manner. In vivo, HSP70-MAb significantly reduced tumor volume, strongly stimulated MOLM13-specific Th1 cell generation, and induced coagulative necrosis with abundant CD8⁺ T lymphocyte infiltration in tumor tissues.
Conclusion:
The BCG HSP70/anti-CD123 immunoconjugate exerts potent targeted anti-leukemic effects by enhancing immunogenicity and T-cell immunity, representing a promising therapeutic candidate for childhood AML, especially relapsed/refractory cases.
Impact:
We constructed an HSP70/anti-CD123 MAb conjugate retaining binding/immune activity, exerting potent targeted anti-leukemic effects via enhanced immunogenicity and T-cell immunity in vitro/in vivo. Targeting LSCs/MRD, it outperforms single agents, filling gaps in pediatric AML CD123 + LSC targeted immune enhancement. This conjugate offers a new immunotherapy, laying preclinical foundations for LSC/MRD-clearing anti-AML drugs. It establishes an HSP70-mAb coupling method, providing new ideas for other hematologic/solid tumors. The high-quality conjugate shows strong anti-tumor effects with no noted toxicity, accelerating its pediatric oncology translational application.
