Department of Bioengineering, University of California San Diego, La Jolla, CA, USA.
Department of NanoEngineering, University of California San Diego, La Jolla, CA, USA.
Methods Mol Biol. 2024;2720:221-245. doi: 10.1007/978-1-0716-3469-1_16.
Melt processing is an emerging production method to efficiently encapsulate proteins into polymeric devices for sustained release. In the context of vaccines, melt processing is well-suited to develop vaccine delivery devices that are stable outside the cold chain and can generate protective immunity from a single dose. We have demonstrated the compatibility of bacteriophage Qβ virus-like particles (VLPs) with hot-melt extrusion (HME) and have leveraged this technology to develop a single-dose vaccine candidate for vaccination against human papillomavirus (HPV). Here, we detail the methods for chemically conjugating an HPV peptide epitope from the L2 minor capsid protein to Qβ VLPs to generate HPV-Qβ particles. We outline techniques used to characterize HPV-Qβ particles, and we elaborate on the process to encapsulate HPV-Qβ into biodegradable poly(lactic-co-glycolic acid) (PLGA) implants and discuss methods for the materials characterization of the HPV-Qβ/polymer melts. The methods described could be adapted to other disease targets, i.e., by conjugation of a different peptide epitope, or transferred to other VLP systems suited for conjugation, immune response, or stability during processing. Such VLPs are ideally suited for use in HME, a mature, scalable, continuous, and solvent-free process which can be adapted to mold devices, therefore allowing the processing of the melts into various geometries, such as subcutaneous implants, or self-administrable microneedle patches.
熔融加工是一种新兴的生产方法，可有效地将蛋白质封装到聚合物设备中，以实现持续释放。在疫苗方面，熔融加工非常适合开发疫苗输送装置，这些装置在冷链外稳定，并且可以通过单次剂量产生保护免疫。我们已经证明了噬菌体 Qβ 病毒样颗粒（VLPs）与热熔挤出（HME）的兼容性，并利用这项技术开发了一种用于 HPV 疫苗接种的单剂量候选疫苗。在这里，我们详细介绍了将 HPV 衣壳蛋白 L2 次要衣壳蛋白的 HPV 肽表位化学偶联到 Qβ VLPs 上以生成 HPV-Qβ 颗粒的方法。我们概述了用于表征 HPV-Qβ 颗粒的技术，并详细介绍了将 HPV-Qβ 封装到可生物降解的聚（乳酸-共-羟基乙酸）（PLGA）植入物中的过程，并讨论了 HPV-Qβ/聚合物熔体的材料特性的方法。描述的方法可以适应其他疾病靶标，即通过偶联不同的肽表位，或转移到其他适合偶联、免疫反应或加工过程中稳定性的 VLP 系统。此类 VLP 非常适合用于 HME，这是一种成熟、可扩展、连续且无溶剂的工艺，可以适应模具设备，因此可以将熔体加工成各种形状，例如皮下植入物或可自我管理的微针贴片。
