Production and Immunogenicity Assessment of LTp50: An Escherichia coli-Made Chimeric Antigen Targeting S1- and S2-Epitopes from the SARS-CoV-2/BA.5 Spike Protein.

Subunit vaccines stand as a leading approach to expanding the current portfolio of vaccines to fight against COVID-19, seeking not only to lower costs but to achieve long-term immunity against variants of concern and have the main attributes that could overcome the limitations of the current vaccines. Herein a chimeric protein targeting S1 and S2 epitopes, called LTp50, was designed as a convenient approach to induce humoral responses against SARS-CoV-2. LTp50 was produced in recombinant Escherichia coli using a conventional pET vector, recovering the expected antigen in the insoluble fraction. LTp50 was purified by chromatography (purity > 90%). The solubilization and refolding stages helped to obtain a stable protein amenable for vaccine formulation. LTp50 was adsorbed onto alum, resulting in a stable formulation whose immunogenic properties were assessed in BALB/c mice. Significant humoral responses against the S protein (BA.5 variant) were detected in mice subjected to three subcutaneous doses (10 µg) of the LTp50/alum formulation. This study opens the path for the vaccine formulation optimization using additional adjuvants to advance in the development of a highly effective anti-COVID-19 vaccine directed against the antigenic regions of the S protein, which are less prone to mutations.

Expression and immunogenicity assessment of a plant-made immunogen targeting the cytotoxic T-lymphocyte associated antigen-4: a possible approach for cancer immunotherapy.

Cancer immunotherapy is a promising intervention to fight against this global health problem. In particular targeting immune checkpoints, such as cytotoxic T-lymphocyte associated antigen-4 (CTLA-4) and programmed-death protein 1 (PD-1), by specific monoclonal antibodies is a current treatment for many malignances. A possible innovation in this field is based on the induction of humoral responses in the host by suppressing the effects of such immune checkpoints and as consequence favoring the activation of cellular immunity against the tumor cells. In this study, chimeric protein comprising the B subunit of Escherichia coli heat-labile enterotoxin as carrier and the extracellular domain of CTLA-4 (LTB-CTLA4) was produced in Nicotiana benthamiana by transient expression. The recombinant protein was accumulated up to 1.29 µg/g of leaves fresh weight on 4 day-post-infiltration. The integrity of the plant-made LTB-CTLA4 antigen was confirmed by western blot analysis and ELISA. Immunogenicity of the plant-made LTB-CTLA4 was assessed in BALB/c mice and the results showed that humoral responses were induced against both the LTB and CTLA-4 moieties. The plant-made LTB-CTLA4 stands as a promising candidate for the design of advanced protection studies against cancer in murine models.

Evaluation of acute and chronic exposure to aflatoxin B1 in indigenous women of the Huasteca Potosina, Mexico.

Aflatoxin B1 (AFB1) is one of the most studied mycotoxins due to its high occurrence in food and its hepatotoxic, immunosuppressive, carcinogenic, childhood growth, genotoxic, mutagenic, and teratogenic effects in humans and animals. Exposure to AFB1 is reported to be both, acute and chronic; the main exposure pathway to AFB1 is through the intake of contaminated food. In Mexico, although the reports of several studies addressing the problem of aflatoxins in maize and other foods, the evidence has been centered on exposure to AFB1 and to the quantification of the Aflatoxins themselves, but there is null evidence about genotoxic effects of aflatoxins in vulnerable populations. Therefore, this study focused on assessing chronic AFB1 exposure through the AFB1-lys biomarker adduct and acute exposure through total AFB1-DNA adducts in women from a rural indigenous community in the Huasteca Potosina (Mexico). A hundred percent of the studied population presented total AFB1-DNA and AFB1-lys adducts in concentrations of 1.08 (0.48-1.34) µmol of adduct/mol of DNA and 2.33 (1.08-102.6) pg/mg of albumin respectively (median (min-max)). Thus, continuous monitoring and important changes in regulations are desired and recommended. The results in this study provide enough evidence to support the toxic effects that the exposure to AFB1 represents, as well as the population risk that it poses, and in the same sense, the current need to create an intervention program that directly influences the control of the sources of exposure in order to reduce it.

Expression in algae of a chimeric protein carrying several epitopes from tumor associated antigens.

Immunotherapies for cancer treatment constitute promising avenues to fight this global health issue. Algae can be used as both biofactories and delivery vehicles of vaccines; having low cost, fast growth, enhanced safety, and adjuvant effects as advantages. In the present study a multiepitope protein, called BCB, was designed as an attractive approach to develop new cancer immunotherapies. The BCB protein targets epitopes from the following tumor-associated antigens: human epidermal growth factor receptor-2 (HER2), mucin-like glycoprotein 1 (MUC1), Wilms' tumor antigen (WT1), and mammaglobin. Moreover, the BCB protein is based on the B subunit of the heat labile E. coli enterotoxin as immunogenic carrier to brake tolerance against self-antigens. A synthetic BCB-coding gene was obtained and expressed in Schizochytrium sp. using the Algevir system. The BCB protein was successfully expressed in transformed algae at levels up to 637 µg/g fresh weight, retaining the GM1-binding activity. The algae-made BCB showed reactivity towards an anti-serum against the tumor cell line 4T1; evidencing its antigenicity. Moreover the immunogenicity was evidenced in mice immunized with BCB, which developed serum IgG antibodies reacting against the 4T1 lysate. This study constitutes the first step in the development of innovative algae-based vaccines against cancer.

Mesoporous Silicon Particles Favor the Induction of Long-Lived Humoral Responses in Mice to a Peptide-Based Vaccine.

Vaccinology faces the challenge of developing improved immunization approaches that are able to induce long-term immunity with the desired Th profile according to the pathology. In this context, new vehicles for efficient antigen delivery that exert adjuvant effects play a critical role in addressing this goal. Herein, mesoporous silicon particles (PSiP) were assessed as carriers for a peptide-based vaccine targeting the receptor for advanced glycation end products (RAGE), which is a relevant receptor in Alzheimer´s disease and other diseases. A RAGE peptide was adsorbed onto PSiP (PSiP vaccine) and administered to BALB/c mice, leading to immune responses that were similar in magnitude to those induced by the soluble peptide. However, the response induced by PSiP lasted for a significantly longer period when compared with the behavior of the group immunized with the peptide alone. Therefore, PSiP are proposed as carriers to enhance immune memory, which is critical in vaccination. This study opens interesting perspectives related to the application of PSiP in vaccinology.

Plant-Made Vaccines in the Fight Against Cancer.

Immunotherapies constitute an important trend in developing new cancer treatments, and several promising candidates are under evaluation in clinical trials. Plants have entered the fight against cancer because they constitute low-cost and efficient hosts for biopharmaceutical production that can also serve as oral delivery vehicles. This review is focused on the knowledge gained through the development of anticancer plant-made vaccines reported thus far, and highlights the potential of this technology - its success being reflected in the number of candidates that are close to market. Future prospects for anticancer plant-made vaccines are also identified.