The use of chitosan for cancer vaccine delivery

壳聚糖溶液增强粒-巨噬细胞的免疫佐剂特性 集落刺激因子（GM-CSF）。免疫调节细胞因子的持续局部递送是 正在研究其增强疫苗和抗肿瘤反应的能力， 临床和临床前。本研究评价了壳聚糖，一种生物相容性 多糖，以（1）控制细胞因子GM-CSF的传播，和（2）增强 GM-CSF的免疫佐剂性质。虽然细胞因子先前已经在细胞内递送， 基于脂质的佐剂和其他媒介物，这些不具有临床安全性特征，或 壳聚糖的独特性能。我们发现，壳聚糖溶液保持了可测量的贮库 重组GM-CSF（rGM-CSF）在皮下注射部位给药长达9天。在 相反，当在生理盐水载体中递送时，在1224小时内未检测到rGM-CSF。此外，委员会认为， 单次皮下注射20 g rGM-CSF壳聚糖溶液 （壳聚糖/rGM-CSF（20 g））瞬时扩增淋巴结达4.6倍， 使MHC II类表达细胞和树突状细胞的数量增加7.4倍， 6.8倍。这些增加明显大于当 rGM-CSF以标准临床前剂量和时间表，即每天4次，在盐水中施用 S.C.注射20 g。此外，来自注射了 壳聚糖/rGM-CSF（20 μ g）诱导更大的同种异体T细胞增殖， 表明增强的抗原提呈能力，比来自小鼠的淋巴结细胞注射 单独使用rGM-CSF。最后，在疫苗接种实验中，壳聚糖/rGM-CSF上级优于 壳聚糖或rGM-CSF单独增强抗原特异性CD 4+细胞的诱导 增殖、肽特异性CD 8+五聚体染色和细胞毒性T细胞裂解。总之， 壳聚糖/rGM-CSF在树突状细胞募集方面优于标准rGM-CSF施用， 抗原呈递和疫苗增强。我们得出结论，壳聚糖溶液是一种 有前途的交付平台的持续，局部交付的rGM-CSF。在一项相关研究中， 壳聚糖溶液增强了皮下注射的体液和细胞介导的免疫应答 预防针开发安全的新型佐剂是必要的， 新的和/或现有的疫苗。壳聚糖是一种无毒、生物相容、可生物降解、天然 从甲壳类动物和昆虫的外骨骼中提取的多糖。壳聚糖 生物降解性、免疫活性和高粘度使其成为作为 用于肠胃外疫苗接种的贮库/佐剂。为此，我们探索了壳聚糖溶液作为 用于用模型蛋白抗原皮下接种小鼠的佐剂。我们发现 壳聚糖增强抗原特异性抗体滴度超过5倍， CD 4+增殖超过6倍。抗体滴度大幅增加， 迟发型超敏反应（DTH）反应表明，壳聚糖诱导体液和 细胞介导的免疫反应。与传统疫苗佐剂相比，壳聚糖 与不完全弗氏佐剂（IFA）相当，优于氢氧化铝（上级）。 机制研究表明，壳聚糖表现出至少两个特性， 使其发挥免疫佐剂的作用。首先，粘稠的壳聚糖溶液产生了 抗原库更具体地，当在盐水中递送时， 8小时后保留在注射部位。然而，超过60%的蛋白质抗原 在注射部位保持7天。其次，壳聚糖诱导了一种 引流淋巴结中一过性67%细胞扩增。扩张在14岁到19岁之间达到顶峰， 注射壳聚糖后21天，随着多糖的降解而减少。这些 机制研究以及疫苗应答的增强表明， 壳聚糖是一种有前途且安全的胃肠外疫苗输送平台。

Chitosan solution enhances the immunoadjuvant properties of granulocyte-macrophage colony-stimulating factor (GM-CSF). Sustained, local delivery of immunomodulatory cytokines is under investigation for its ability to enhance vaccine and anti-tumor responses both clinically and preclinically. This study evaluates the ability of chitosan, a biocompatible polysaccharide, to (1) control the dissemination of a cytokine, GM-CSF, and (2) enhance the immunoadjuvant properties of GM-CSF. While cytokines have previously been delivered in lipid-based adjuvants and other vehicles, these do not have the clinical safety profile or unique properties of chitosan. We found that chitosan solution maintained a measurable depot of recombinant GM-CSF (rGM-CSF) at a subcutaneous injection site for up to 9 days. In contrast, when delivered in a saline vehicle, rGM-CSF was undetectable in 1224 h. Furthermore, a single s.c. injection of 20 μg rGM-CSF in chitosan solution (chitosan/rGM-CSF(20 μg)) transiently expanded lymph nodes up to 4.6-fold and increased the number of MHC class II expressing cells and dendritic cells by 7.4-fold and 6.8-fold, respectively. These increases were significantly greater than those measured when rGM-CSF was administered in saline at the standard preclinical dose and schedule, i.e. 4 daily s.c. injections of 20 μg. Furthermore, lymph node cells from mice injected with chitosan/rGM-CSF(20 μg) induced greater allogeneic T cell proliferation, indicating enhanced antigen presenting capability, than lymph node cells from mice injected with rGM-CSF alone. Finally, in vaccination experiments, chitosan/rGM-CSF was superior to either chitosan or rGM-CSF alone in enhancing the induction of antigen-specific CD4+ proliferation, peptide-specific CD8+ pentamer staining and cytotoxic T cell lysis. Altogether, chitosan/rGM-CSF outperformed standard rGM-CSF administrations in dendritic cell recruitment, antigen presentation and vaccine enhancement. We conclude that chitosan solution is a promising delivery platform for the sustained, local delivery of rGM-CSF. In a related study, chitosan solution enhances both humoral and cell-mediated immune responses to subcutaneous vaccination. The development of safe, novel adjuvants is necessary to maximize the efficacy of new and/or available vaccines. Chitosan is a non-toxic, biocompatible, biodegradable, natural polysaccharide derived from the exoskeletons of crustaceans and insects. Chitosans biodegradability, immunological activity and high viscosity make it an excellent candidate as a depot/adjuvant for parenteral vaccination. To this end, we explored chitosan solution as an adjuvant for subcutaneous vaccination of mice with a model protein antigen. We found that chitosan enhanced antigen-specific antibody titers over five-fold and antigen-specific splenic CD4+ proliferation over six-fold. Strong increases in antibody titers together with robust delayed-type hypersensitivity (DTH) responses revealed that chitosan induced both humoral and cell-mediated immune responses. When compared with traditional vaccine adjuvants, chitosan was equipotent to incomplete Freunds adjuvant (IFA) and superior to aluminum hydroxide. Mechanistic studies revealed that chitosan exhibited at least two characteristics that may allow it to function as an immune adjuvant. First, the viscous chitosan solution created an antigen depot. More specifically, less than 9% of a protein antigen, when delivered in saline, remained at the injection site after 8 h. However, more than 60% of a protein antigen delivered in chitosan remained at the injection site for 7 days. Second, chitosan induced a transient 67% cellular expansion in draining lymph nodes. The expansion peaked between 14 and 21 days after chitosan injection and diminished as the polysaccharide was degraded. These mechanistic studies, taken together with the enhancement of a vaccine response, demonstrate that chitosan is a promising and safe platform for parenteral vaccine delivery.