In silicon desinging of RANKL-targeting vaccine for protection of osteoporosis based on the epitope of Denosumab

Abstract

<p>Background: Life quality of osteoporosis patients is affected significantly due to the severely complications of fracture and pain. RANKL, indicated as the key mediator of osteoporosis, plays a pathogenic role of osteoclasts induction. To target this program, two medications, bisphosphonate and Denosumab, were developed and achieved remarkable advantages in clinics. Unfortunately, fracture-related side-effects always emerge unavoidably, after either long-term administration of bisphosphonates or Denosumab withdrawing. To address these challenges, vaccine-based approach has been adopted to achieve sustainable protection through induction and maintenance of effective antibodies in mild level over decades. Methods: A Denosumab binding peptide was firstly identified as the basic component of vaccine. This peptide was then fused with diphtheria toxin T domain, a widely used adjuvant protein. Its capabilities to penetrate the autologous tolerance and induce the immune responses was then demonstrated with in-silicon evaluation. Finally, the efficacy of the DR3 vaccine was assessed through immunization on the human RANKL transgenic mice model of osteoporosis. Results: The DTT-RANKL_(220–245)3 vaccine, termed as DR3, were predicted as highly antigenic and non-allergenicity. This molecule was comprised of 46.5 % of helix, 8.5 % strand and 45.1 % coil, the optimized Z-value of the tertiary structure was 6.39, and the favored area in the Ramachandran plot was 96.1 % after refinement. Molecular docking showed a tight binding of DR3 vaccine to TLR2 (−9.2 kcal/mol) and TLR4 (−9.5 kcal/mol). In addition, the immune stimulation indicated robust responses post administration of DR3 vaccine, including high level production of of antibodies and cytokines, activated T and B lymphocytes, and the long-last immune memory. In agree with the simulation, vaccinated mice generated high titers anti-hRANKL antibodies and elevated levels of IL-4 and IL-10 at 7th week post immunization. Conclusion: DR3 vaccine was aroused to benefit the prevention and treatment of osteoporosis, and other bone-resorptive diseases potentially. <br></p>