Thermostability of Freeze‐Dried Plant‐Made VLP‐Based Vaccines

Freeze-drying or lyophilisation is a technique commonly used for pharmaceutical products, in which stability is required to be preserved beyond 4°C temperature. Although lyophilisation is a well‐established method, designing a sustainable process for a new product is still to a large extent subjected to empirical practice and often requires trial and error approach. Moreover, even successful lyophilisation of the product may not assure its good long‐term storage stability, and progressive decrease in activity may still be encountered. In the past decades, numerous studies have been conducted in the area of protein instability and preservation during lyophilisation and long‐term storage. Many critical issues have been identified with regard to physical and chemical instability of proteins in the solid

Assembly and Characterization of HBc Derived Virus-like Particles with Magnetic Core

Core-virus like particles (VLPs) assembly is a kinetically complex cascade of interactions between viral proteins, nanoparticle’s surface and an ionic environment. Despite many in silico simulations regarding this process, there is still a lack of experimental data. The main goal of this study was to investigate the capsid protein of hepatitis B virus (HBc) assembly into virus-like particles with superparamagnetic iron oxide nanoparticles (SPIONs) as a magnetic core in relation to their characteristics. The native form of HBc was obtained via agroinfection of Nicotiana benthamiana with pEAQ-HBc plasmid. SPIONs of diameter of 15 nm were synthesized and functionalized with two ligands, providing variety in ζ-potential and hydrodynamic diameter. The antigenic potential of the assembled core-VLPs was assessed with enzyme-linked immunosorbent assay (ELISA). Morphology of SPIONs and core-VLPs was evaluated via transmission electron microscopy (TEM). The most successful core-VLPs assembly was obtained for SPIONs functionalized with dihexadecyl phosphate (DHP) at SPIONs/HBc ratio of 0.2/0.05 mg/mL. ELISA results indicate significant decrease of antigenicity concomitant with core-VLPs assembly. In summary, this study provides an experimental assessment of the crucial parameters guiding SPION-HBc VLPs assembly and evaluates the antigenicity of the obtained structures

Plant expression, lyophilisation and storage of HBV medium and large surface antigens for a prototype oral vaccine formulation

Current immunisation programmes against hepatitis B virus (HBV) increasingly often involve novel tri-component vaccines containing—together with the small (S-HBsAg)—also medium and large surface antigens of HBV (M- and L-HBsAg). Plants producing all HBsAg proteins can be a source of components for a potential oral ‘triple’ anti-HBV vaccine. The objective of the presented research was to study the potential of M/L-HBsAg expression in leaf tissue and conditions of its processing for a prototype oral vaccine. Tobacco and lettuce carrying M- or L-HBsAg genes and resistant to the herbicide glufosinate were engineered and integration of the transgenes was verified by PCR and Southern hybridizations. M- and L-HBsAg expression was confirmed by Western blot and assayed by ELISA at the level of micrograms per g of fresh weight. The antigens displayed a common S domain and characteristic domains preS2 and preS1 and were assembled into virus-like particles (VLPs). Leaf tissues containing M- and L-HBsAg were lyophilised to produce a starting material of an orally administered vaccine formula. The antigens were distinctly sensitive to freeze-drying conditions and storage temperature, in the aspect of stability of S and preS domains and formation of multimeric particles. Efficiency of lyophilisation and storage depended also on the initial antigen content in plant tissue, yet M-HBsAg appeared to be approximately 1.5–2 times more stable than L-HBsAg. The results of the study provide indications concerning the preparation of two other constituents, next to S-HBsAg, for a plant-derived prototype oral tri-component vaccine against hepatitis B

The Twenty-Year Story of a Plant-Based Vaccine Against Hepatitis B: Stagnation or Promising Prospects?

Hepatitis B persists as a common human disease despite effective vaccines having been employed for almost 30 years. Plants were considered as alternative sources of vaccines, to be mainly orally administered. Despite 20-year attempts, no real anti-HBV plant-based vaccine has been developed. Immunization trials, based on ingestion of raw plant tissue and conjugated with injection or exclusively oral administration of lyophilized tissue, were either impractical or insufficient due to oral tolerance acquisition. Plant-produced purified HBV antigens were highly immunogenic when injected, but their yields were initially insufficient for practical purposes. However, knowledge and technology have progressed, hence new plant-derived anti-HBV vaccines can be proposed today. All HBV antigens can be efficiently produced in stable or transient expression systems. Processing of injection vaccines has been developed and needs only to be successfully completed. Purified antigens can be used for injection in an equivalent manner to the present commercial vaccines. Although oral vaccines require improvement, plant tissue, lyophilized or extracted and converted into tablets, etc., may serve as a boosting vaccine. Preliminary data indicate also that both vaccines can be combined in an effective parenteral-oral immunization procedure. A partial substitution of injection vaccines with oral formulations still offers good prospects for economically viable and efficacious anti-HBV plant-based vaccines

Interaction of silicone hydrogel contact lenses with lipids – a chronological review

Silicone hydrogel (SiHy) contact lenses are a common form of correction of refractive errors prescribed by eye care professionals around the world. SiHy lenses perform in a complex environment, which is the surface of the eye and the tear film. Therefore, they are exposed to various factors, such as lipid deposits. The aims of this paper are to review available scientific reports on the study of SiHy lens interactions with lipids and search for further research objectives. A total of 57 publications were identified and reviewed, from 2003 to 2020. In general, SiHy lenses are more likely to accumulate lipid deposits than traditional hydrogel lenses, although there are significant differences between SiHy lens materials that may result from different methods used in the studies. The review includes studies on various aspects of interactions between lenses and lipids, such as those concerning the effectiveness of lipids removal from lenses by care solutions. The conclusion points out future research directions, such as measurements of lipid diffusion in SiHy lens' matrices

Analiza opinii polskich specjalistów ochrony wzroku dotyczących osadów na soczewkach kontaktowych

Soft contact lenses are used worldwide to correct ametropia. This medical device should be fitted and systematically evaluated on the patient eye by eye care professionals. The examination should include an evaluation of lens deposits. The widespread use of silicone hydrogel (Si-Hy) lenses and their tendency to build up lipid deposits indicate that eye care professionals should pay special attention to this kind of deposits. This paper aims to analyze the opinions of Polish ECPs regarding deposits on contact lenses, particularly regarding lipids and Si-Hy lenses. Data were collected from 103 Polish eye care professionals through an online survey. Respondents reported that lipid deposits are most often present on Si-Hy lenses, and protein deposits occur most often on hydrogel lenses. Almost all of them declare that they evaluate deposits on contact lenses on follow-up visits, which concludes that this is an essential part of the lens selection process for them. Respondents believe that different lenses and lens care solutions vary in the context of their interaction with lipid deposits. These parameters should be considered at the time of product selection. Eye care professionals expect descriptions of lenses’ resistance to lipid deposits, and care solution effectiveness in reducing lipid deposits.Miękkie soczewki kontaktowe służące do korekcji ametropii są powszechnie używane na całym świecie. Jako wyrób medyczny powinny być dopasowywane i systematycznie oceniane na oku pacjenta przez specjalistów ochrony wzroku. Badanie kontrolne powinno obejmować ocenę osadów na soczewkach. Powszechne stosowanie soczewek silikonowo-hydrożelowych (Si-Hy) i ich tendencja do odkładania się złogów lipidowych wskazują, że specjaliści ochrony wzroku powinni zwracać szczególną uwagę na ten rodzaj osadów. Celem niniejszej pracy jest analiza opinii polskich specjalistów ochrony wzroku w zakresie osadów na soczewkach kontaktowych, ze szczególnym uwzględnieniem lipidów i soczewek Si-Hy. Dane zebrano od 103 polskich specjalistów ochrony wzroku poprzez ankietę internetową. Respondenci wskazali, że osady lipidowe najczęściej występują na soczewkach Si-Hy, a osady białkowe na soczewkach hydrożelowych. Niemal wszyscy ankietowani deklarują, że oceniają osady na soczewkach kontaktowych podczas wizyt kontrolnych, co pozwala wnioskować, iż jest to dla nich istotna część procesu doboru soczewek. Respondenci uważają, że poszczególne soczewki i płyny do ich pielęgnacji różnią się w kontekście ich interakcji z osadami lipidowymi. Parametry te powinny być brane pod uwagę w momencie wyboru produktu dla pacjenta. Specjaliści ochrony wzroku oczekują opisów odporności soczewek na powstawanie osadów lipidowych oraz skuteczności płynów pielęgnacyjnych w redukcji osadów lipidowych

Plant-Based Vaccines in Combat against Coronavirus Diseases

Coronavirus (CoV) diseases, including Middle East Respiratory Syndrome (MERS) and Severe Acute Respiratory Syndrome (SARS) have gained in importance worldwide, especially with the current COVID-19 pandemic caused by SARS-CoV-2. Due to the huge global demand, various types of vaccines have been developed, such as more traditional attenuated or inactivated viruses, subunit and VLP-based vaccines, as well as novel DNA and RNA vaccines. Nonetheless, emerging new COVID-19 variants are necessitating continuous research on vaccines, including these produced in plants, either via stable expression in transgenic or transplastomic plants or transient expression using viral vectors or agroinfection. Plant systems provide low cost, high scalability, safety and capacity to produce multimeric or glycosylated proteins. To date, from among CoVs antigens, spike and capsid proteins have been produced in plants, mostly using transient expression systems, at the additional advantage of rapid production. Immunogenicity of plant-produced CoVs proteins was positively evaluated after injection of purified antigens. However, this review indicates that plant-produced CoVs proteins or their carrier-fused immunodominant epitopes can be potentially applied also as mucosal vaccines, either after purification to be administered to particular membranes (nasal, bronchus mucosa) associated with the respiratory system, or as oral vaccines obtained from partly processed plant tissue