Jorge Aguiar

The immune system is central in controlling acute hepatitis B infection and in patients resolving chronic hepatitis B (CHB). Given that 221 million (75%) of CHB patients reside in low- and middle-income countries, developing a vaccine with therapeutic properties represents a rational and cost-effective approach more than a romantic endeavor. This review systematically analyses the critical variables related to the safety, efficacy, and effectiveness of CHB treatments. HeberNasvac's experience is revisited to address the challenges and potentialities of therapeutic vaccines, as well as the current roadblocks in research and development, registration, and large-scale implementation.

HeberNasvac, a therapeutic vaccine for chronic hepatitis B, is able to safely stimulate multiple Toll-like receptors, increasing antigen presentation in vitro and in a phase II clinical trial (Profira) in elderly volunteers who were household contacts of respiratory infection patients. Thus, a new indication as a postexposure prophylaxis or early therapy for respiratory infections has been proposed. In this study, we evaluated the expression of several interferon-stimulated genes (ISGs) after mucosal administration of HeberNasvac and compared this effect with the nasal delivery of interferon alpha 2b (Nasalferon). Molecular studies of blood samples of 50 subjects from the Profira clinical trial who were locally treated with HeberNasvac or Nasalferon and concurrent untreated individuals were compared based on their relative mRNA expression of OAS1, ISG15, ISG20, STAT1, STAT3, and DRB1-HLA II genes. In most cases, the gene expression induced by HeberNasvac was similar in profile and intensity to the expression induced by Nasalferon and significantly superior to that observed in untreated controls. The immune stimulatory effect of HeberNasvac on ISGs paved the way for its future use as an innate immunity stimulator in elderly persons and immunocompromised subjects or as part of Mambisa, a nasal vaccine to prevent severe acute respiratory syndrome coronavirus 2 infection.

Nasvac (HeberNasvac^®) is a novel therapeutic vaccine for chronic hepatitis B (CHB). This product is a formulation of the core (HBcAg) and surface (HBsAg) antigens of the hepatitis B virus (HBV), administered by nasal and subcutaneous routes, in a distinctive schedule of immunizations. In the present review article, we discuss the action mechanisms of HeberNasvac, considering the immunological properties of the product and their antigens. Specifically, we discuss the capacity of HBcAg to activate different pathways of innate immunity and the signal transduction after a multi-TLR agonist effect, and we review the results of recent clinical trials and in vitro studies. Aimed at understanding the clinical results of Nasvac and other therapeutic vaccines under development, we discuss the rationale of administering a therapeutic vaccine through the nasal route and also the current alternatives to combine therapeutic vaccines and antivirals (NUCs). We also disclose potential applications of this product in novel fields of immunotherapy.

This research focused on the results of the cross-validation program related with the performance of a Cuban novel low-cost real-time quantitative polymerase chain reaction (qPCR) assay for hepatitis B virus (HBV) quantification developed by the Therapeutic Vaccine against Hepatitis B Department, Vaccines Division, Center for Genetic Engineering and Biotechnology (CIGB), Havana, Cuba. Dilution series with the plasmid standard at concentrations of 900,000 to 0.09 copies/reaction (c/r) were made for each PCR instrument. The mean cycles threshold (Ct) values and PCR efficiency were compared among the cyclers. Hepatitis B virus-positive serum samples were used for the calculation of reproducibility of the HBV assay. Biotecon Diagnostics (BCD) also ordered the oligo sequences from a second supplier and compared the PCR performance to those provided from the CIGB. All PCR cyclers were able to detect concentrations up to 0.09 c/r. However, below the concentration of 9 c/r, the variation of results increased within and between the cyclers. The PCR efficiency showed satisfying results. The overall coefficient of variation (CV) cycler values were 1.29 and 0.91% for M6 and M19 respectively. No significance was observed between the different primer suppliers. The HBV assay was performed with a good concordance between the five real-time instruments from different suppliers. The HBV assay was also performed with a high reproducibility for samples with a high and a low viral load. The HBV assay is robust against different primer suppliers.How to cite this article: Aguiar J, Silva JA, García G, Guillén G, Aguilar JC. Cross-validation Studies of a Novel Low-cost Hepatitis B Virus Quantitative Polymerase Chain Reaction System. Euroasian J Hepato-Gastroenterol 2018;8(1):38-41.

We studied the functional stability of a primer pair and the standard curve based on a plasmid carrying full-length HBV genome, from a novel low-cost real-time quantitative polymerase chain reaction (qPCR) assay. The assay was developed at the Center for Genetic Engineering and Biotechnology (CIGB) in Havana, to quantify the serum hepatitis B virus (HBV) DNA from chronic HBV-infected (CHB) patients. In-house generated oligonucleotides and plasmids were incubated at 37°C during 1 month and compared with the same materials incubated at -20, 4, and 25°C during the same time in qPCR experiments. This work shows that the oligonucleotide pair and the plasmid for the quantitative standard curve are functionally stable in severe temperature conditions during 1 month. Polymerase chain reaction amplification with both materials after its incubation 30 days at 37°C produced similar cycle threshold (CT) values and similar degree of sample quantifications compared with the same materials preserved using the conventional storage conditions at -20°C. These results are indicative of the robustness of this low-cost qPCR system for HBV DNA quantification. These results also support that this qPCR assay can be used as a low-cost technology in clinical studies to monitor the viral load changes of serum HBV DNA of CHB patients, which could be used by poor people of third world countries, where there are frequent blackouts and temperature changes that can hinder the primer and plasmid stability. Aguiar J, García G, León Y, Canales E, Silva JA, Gell O, Estrada R, Morán I, Muzio V, Guillén G, Pentón E, Aguilar JC. High Functional Stability of a Low-cost HBV DNA qPCR Primer Pair and Plasmid Standard. Euroasian J Hepato-Gastroenterol 2016;6(1):19-24.