| As the "booster" of the vaccine,the vaccine adjuvant can enhance the immune response of the body to the antigen after being injected into the body in combination with antigens.The development of human vaccine adjuvant has been slow.Since aluminum adjuvant was approved for use in human vaccines in 1926,only six vaccine adjuvants have been approved worldwide in the past century.The research and development of vaccine adjuvants are mainly faced with issues such as higher safety requirements urgent requirements for synergistic enhancement of immune response,limited source of raw materials,and difficulty in process amplification.Most enterprises and institutions in China still focus on following the lead in research and development of vaccine adjuvants and vaccines,lacking independent innovation capabilities and core competitiveness.Based on the above background,this thesis focuses on the application research of chiral poly(lactic acid)(PLA)nanoadjuvant.First,we synthesized PLA with different chiralities(PLLA and PDLA)by ring-opening polymerization of lactide,and prepared PLA nanoparticles with different chiralities by nano-precipitation method.Suitable physical and chemical properties,good biological safety,efficient endocytosis efficiency by antigen presenting cells(APC),and excellent immuneactivating ability confirmed the potential of different chiral PLA nanoparticles as nanoadjuvant.Furthermore,we co-assembled amino-terminated PLA(PLA-NH2)and model antigen ovalbumin(OVA)to obtain a spherical chiral nanovaccine(PLAOVA)with uniform particle size of about 80 nm.Further characterization through flow cytometry and confocal laser microscopy showed that the PLA-OVA nanovaccine could be effectively internalized by DC cells,and the internalization efficiency of D-type chiral PLA nanovaccine was higher than that of L-type chiral nanovaccine.48 h after subcutaneous injection,the nanovaccine could migrate to lymph nodes,promote dendritic cell(DC)maturation,improve antigen presenting efficiency,and activate T lymphocytes,while enhancing humoral immune response.Through comparison,we found that PDLA nanoadjuvant has stronger immune activation function than PLLA.Meanwhile,we confirmed that PDLA-OVA had a stronger ability to enhance specific antibodies and activate complement.In the mouse melanoma prevention model,PLA-OVA nanovaccine could effectively prolong the survival period of immunized mice.On the 41st day after tumor inoculation,the survival rates of mice in the PLLA-OVA and PDLA-OVA groups were still 40%and 60%,respectively.In the mouse melanoma treatment model,the tumor inhibition rate(TIR)of PLLA-OVA and PDLA-OVA groups was 49.1%and 65.3%respectively on the 20th day after tumor inoculation.In three models of mouse prostate cancer,mouse orthotopic breast cancer,and mouse colon cancer,the prominent advantages of PDLA-NH2 as an adjuvant of nanovaccine and the good effectiveness and universality of the prepared PDLA tumor nanovaccine were also confirmed again.Next,we further verified the safety of PLA-OVA nanovaccine.The results showed that the pH of different chiral PLA-OVA nanovaccines was between 6.2 and 7.1,while the osmotic pressure was between 280 and 320 mOsm/kg,and the hemolysis rate was also lower than 2%.Finally,after confirming the safety and efficacy of chiral PLA nanoadjuvants,we summarized the overall research and development ideas of PLA nanoadjuvant and vaccine,and promoted the process of establishing the preparation process of nanoadjuvant and vaccine,and establishing the screening and technical platform of nanoadjuvant and vaccine with cooperation companies.At present,based on the previous research,the first draft of the quality standard for PLA nanoadjuvant products has been completed.At the same time,we’ve also made necessary analysis and summary of intellectual property and technology reserves.In conclusion,the chiral PLA materials we prepared,especially PDLA,showed good safety and high effectiveness as a new vaccine adjuvant.Next,we will conduct more in-depth clinical transformation research on PLA nanoadjuvant,with a view to improving the independent innovation ability and core competitiveness of China’s vaccine industry,and contributing to the goal of changing from a "large vaccine country"to a "powerful vaccine country". |
