Canola oilseed- and Escherichia coli- derived hepatitis C virus (HCV) core proteins adjuvanted with oil bodies, induced robust Th1-oriented immune responses in immunized mice.

Induction of broad Th1 cellular immune responses and cytokines is crucial characteristics for vaccines against intracellular infections such as hepatitis C virus (HCV). Plants (especially oilseed tissues) and plant-immunomodulators (like oil bodies) offer cost-effective and scalable possibilities for the production of immunologically relevant and safe vaccine antigens and adjuvants, respectively. Herein, we provide data of the murine immunization by transgenic canola oilseed-derived HCV core protein (HCVcp) soluble extract (TSE) and Escherichia coli- derived rHCVcp in combination with Canola oil bodies (oil) compared to that of the Freund's (FA) adjuvant. Mice immunized by TSE+ oil developed both strong humeral (IgG) and Th1-biased cellular responses, manifested by high levels of IFN-γ and lower IgG1/IgG2a ratio and IL-4 secretion. Results of the intracellular cytokine staining indicated that TSE+ oil immunization in mice triggered both CD4+ and CD8+ T cells to release IFN-γ, while CD4+ cells were mostly triggered when FA was used. Analyses by qRT-PCR indicated that a combination of rHCVcp/TSE with oil body induced high levels of IL-10 cytokines compared to that of the FA adjuvant. These characteristics are important properties for the design of an HCV vaccine candidate and indicate the potential of Canola-derived antigen and oil bodies in addressing these concerns.

Pre-clinical evaluation of a quadrivalent HCV VLP vaccine in pigs following microneedle delivery.

The introduction of directly acting antiviral agents (DAAs) has produced significant improvements in the ability to cure chronic hepatitis C infection. However, with over 2% of the world's population infected with HCV, complications arising from the development of cirrhosis of the liver, chronic hepatitis C infection remains the leading indication for liver transplantation. Several modelling studies have indicated that DAAs alone will not be sufficient to eliminate HCV, but if combined with an effective vaccine this regimen would provide a significant advance towards achieving this critical World Health Organisation goal. We have previously generated a genotype 1a, 1b, 2a, 3a HCV virus like particle (VLP) quadrivalent vaccine. The HCV VLPs contain the core and envelope proteins (E1 and E2) of HCV and the vaccine has been shown to produce broad humoral and T cell immune responses following vaccination of mice. In this report we further advanced this work by investigating vaccine responses in a large animal model. We demonstrate that intradermal microneedle vaccination of pigs with our quadrivalent HCV VLP based vaccine produces long-lived multi-genotype specific and neutralizing antibody (NAb) responses together with strong T cell and granzyme B responses and normal Th1 and Th2 cytokine responses. These responses were achieved without the addition of adjuvant. Our study demonstrates that our vaccine is able to produce broad immune responses in a large animal that, next to primates, is the closest animal model to humans. Our results are important as they show that the vaccine can produce robust immune responses in a large animal model before progressing the vaccine to human trials.

Management of hepatocellular carcinoma.

PURPOSE: Hepatocellular carcinoma (HCC), a common cause for cancer-related death, is increasing worldwide. Over the past decade, survival and quality of life of HCC patients have significantly improved due to better prevention strategies, early diagnosis, and improved treatment options. We performed this narrative review to synthesize current status on the HCC management. METHODS: Literature search for publications especially over the last decade, which has changed the paradigm on the management of HCC. RESULTS: Hepatitis B vaccination and treatment of chronic hepatitis B and C are important measures for HCC prevention. Screening and surveillance for HCC using ultrasonogram and alpha-fetoprotein estimation are directed toward cirrhotics and hepatitis B patients at high risk of HCC. If detected at an early stage, curative treatments for HCC can be used such as tumor resection, ablation and liver transplantation. HCC patients without curative options are managed by loco-regional therapies and systemic chemotherapy. Loco-regional treatments include trans-arterial chemoembolization, radioembolization and combinations of loco-regional plus systemic therapies. Currently, sorafenib is the only FDA-approved systemic therapy and newer better chemotherapeutic agents are being investigated. Palliative care for terminally ill patients with metastatic disease and/or poor functional status focusses on comfort care and symptom control. CONCLUSIONS: In spite of significant advancement in HCC management, its incidence continues to rise. There remains an urgent need to continue refining understanding of HCC and develop strategies to increase utilization of the available preventive measures and curative treatment modalities for HCC.

Hepatitis C virus hypervariable region 1 variants presented on hepatitis B virus capsid-like particles induce cross-neutralizing antibodies.

Hepatitis C virus (HCV) infection is still a serious global health burden. Despite improved therapeutic options, a preventative vaccine would be desirable especially in undeveloped countries. Traditionally, highly conserved epitopes are targets for antibody-based prophylactic vaccines. In HCV-infected patients, however, neutralizing antibodies are primarily directed against hypervariable region I (HVRI) in the envelope protein E2. HVRI is the most variable region of HCV, and this heterogeneity contributes to viral persistence and has thus far prevented the development of an effective HVRI-based vaccine. The primary goal of an antibody-based HCV vaccine should therefore be the induction of cross-reactive HVRI antibodies. In this study we approached this problem by presenting selected cross-reactive HVRI variants in a highly symmetric repeated array on capsid-like particles (CLPs). SplitCore CLPs, a novel particulate antigen presentation system derived from the HBV core protein, were used to deliberately manipulate the orientation of HVRI and therefore enable the presentation of conserved parts of HVRI. These HVRI-CLPs induced high titers of cross-reactive antibodies, including neutralizing antibodies. The combination of only four HVRI CLPs was sufficient to induce antibodies cross-reactive with 81 of 326 (24.8%) naturally occurring HVRI peptides. Most importantly, HVRI CLPs with AS03 as an adjuvant induced antibodies with a 10-fold increase in neutralizing capability. These antibodies were able to neutralize infectious HCVcc isolates and 4 of 19 (21%) patient-derived HCVpp isolates. Taken together, these results demonstrate that the induction of at least partially cross-neutralizing antibodies is possible. This approach might be useful for the development of a prophylactic HCV vaccine and should also be adaptable to other highly variable viruses.

Alginate coated chitosan nanoparticles are an effective subcutaneous adjuvant for hepatitis B surface antigen.

We recently described a delivery system that is composed of a chitosan core to which the hepatitis B surface antigen (HBsAg) was adsorbed and subsequently coated with sodium alginate. In this present work, alginate coated chitosan nanoparticles were evaluated as a subcutaneous adjuvant for HBsAg. HBsAg loaded, alginate coated or uncoated chitosan nanoparticles, associated or not with CpGODN were subcutaneously administered to mice and several immunological parameters were evaluated. A high anti-HBsAg IgG titer (2271+/-120 mIU/ml), with the majority of antibodies being of Th2 type, was observed within group I, vaccinated with HBsAg loaded onto coated nanoparticles. However, regarding cellular immune response, no significant differences were observed for antigen-specific splenocyte proliferation or for the secretion of IFN-gamma and IL-4, when compared to the control group. The co-delivery of antigen-loaded nanoparticles in the presence of the immunopotentiator, CpG ODN 1826, resulted in an increase of anti-HBsAg IgG titers that was not statistically different from the first group; however, an increase of the IgG2a/IgG1 ratio from 0.1 to 1.0 and an increase (p<0.01) of the IFN-gamma production by the splenocytes stimulated with the HBV antigen was observed. The enhancement of the immune response observed with the antigen-loaded nanoparticles demonstrated that chitosan is a promising platform for parenteral HBsAg delivery and, when co-administered with the CpG ODN, resulted in a mixed Th1/Th2 type immune response.

Optimization strategies for DNA vaccines.

DNA immunization is a relatively new vaccination strategy that involves the direct introduction into the host of plasmid DNA encoding the desired antigen. The DNA enters host cells and results in immune responses following in vivo expression of the antigen. Although DNA-based immunization works well in animal models for the induction of both humoral and cell-mediated immune responses, its success in humans has been limited. This paper discusses different approaches that have attempted to optimize DNA vaccines, and presents results evaluating some of these approaches in mice.

Reactogenicity and immunogenicity of inactivated hepatitis A vaccines.

Two inactivated hepatitis A virus (HAV) candidate vaccine strain were tested, derived from strains CLF and HM175. Neither vaccine increased liver enzymes levels and reactogenicity was similar to that observed with other alum-absorbed products. Antibody responses were dose-dependent and protection against HAV can be presumed to last for at least three years. All persons receiving 720 ELISA units (El.U) of the CLF vaccine seroconverted after one dose. For the HM175 vaccine, anti-HAV persisted until month 12 after injections at months 0 and 1, suggesting that the third dose of vaccine could be given at any time from month 6 to 12. A double dose of HM175 vaccine (1440 El.U) given as a single bolus resulted in 100% seroconversion by day 14 with a geometric mean anti-HAV level of 121 mIU/ml. This implies that rapid protection can be induced using large doses of inactivated HAV vaccine should time constraints dictate such an approach.

A modified immunisation schedule for the hepatitis B vaccine Recombivax HB suitable for mass vaccination in childhood.

Following the demonstration of a fully satisfactory immunogenic activity of a hepatitis B vaccination protocol consisting of three doses of Hevac B Pasteur vaccine given at 3,5 and 11 months of age, it was possible to administer this vaccine at the same times as the vaccinations for diphtheria, tetanus and polio which are mandatory in Italy at those ages. We have also shown that both another plasma-derived vaccine, H-B-VAX (MSD), as well as the DNA-recombinant Engerix B (SK&F) are highly immunogenic when given at the same times as the mandatory childhood vaccinations. In this paper we demonstrate that the same schedule can be used for another hepatitis B vaccine prepared by a DNA-recombinant technique, Recombivax HB (MSD) recently introduced in Italy. In fact two doses of this vaccine, the first given at three months of age and the second two months later, resulted in a 100% seroconversion rate and a mean anti-HBs titre of 440 mUI/ml. Although the date are incomplete since the third dose will be given at 11 months of age, we conclude that this hepatitis B vaccine can also be used in the mass vaccination campaigns of infants in Italy, the first of which was initiated in January 1987 in an hyperendemic area near Naples (HBsAg prevalence about 14%). We underline that this mass vaccination campaign is the first in Europe.

[Eradication of hepatitis B in Portugal. A challenge within reach of the National Health Service]. / Erradicação da hepatite B em Portugal. Um desafio ao alcance do Serviço Nacional de Saúde.

The aim of this work is to evaluate a major epidemiological problem--human hepatitis B virus (HBV) infection--in the world and in Portugal--reinforcing WHO's view that it is a worldwide health problem. Portugal is considered a risk area for HBV infection, with 30% of the population having had contact with the virus and 1 to 5% of chronic carriers (100 to 150,000 persons). Deficient information, education and sanitary conditions aggravate the national situation, urging for official measures that may, in future, lead to a status of controlled infection (less than 0.1% chronic carriers). We propose an educative and informative campaign, the fulfillment of sanitary measures legally controlled as well as the central support for vaccination against HBV. Some of the fundamental points for a legal document that institute a national program for HBV infection control are proposed. At present, cost-benefit relation seems to be favourable--but an adequate plan considering scientific and technical criteria is needed and essential to guarantee that benefits will outweigh investment in terms of Public Health. Recombinant vaccines, obtained by genetic engineering, are referred as the preferential type of vaccine to choose.

Demonstration of mother-to-infant transmission of hepatitis B virus by means of polymerase chain reaction.

To investigate the failure of vaccines to prevent mother-to-infant transmission of hepatitis B virus (HBV), serum, cord blood, and colostrum samples from eleven mothers, known to be carriers of hepatitis B surface antigen, and their infants were examined by means of a highly sensitive polymerase chain reaction (PCR) method. HBV-specific DNA was detected in ten maternal serum samples, eight samples of colostral whey, eight samples of colostral cells, and one cord blood sample. Four infants of mothers with HBV-DNA-positive colostrum showed low responsiveness to hepatitis B vaccine. The infant whose cord blood was positive for HBV DNA showed low responsiveness to hepatitis B vaccine and subsequently became an HBV carrier. These results suggest the need for further study to evaluate whether breastfeeding is advisable for HBV carriers.

Trials of intradermal hepatitis B vaccines in Gambian children.

Three trials of intradermal hepatitis B virus (HBV) vaccines were made in Gambian children. In the first trial HBV vaccine (1 microgram) was given to neonates in the same syringe with BCG followed by two further does of 1.0 microgram of intradermal HBV vaccine. The trial was a failure, for 19 of 32 subjects had an HBV surface antibody response of less than 10 m.i.u./ml. In the second trial in young children two different regimes were used: two doses of 2 micrograms HBV vaccine were given intradermally after a 20 micrograms intramuscular dose or three doses of 2 micrograms were given intradermally. In both cases geometric mean antibody responses were significantly lower than in the control group who were given 20 micrograms HBV intramuscularly followed by two 10 micrograms doses intramuscularly. Vaccine failures, defined as the presence of HBV surface antigen or core antibody or absence of surface antibody, were also significantly higher in the intradermal groups. In the third trial 4 micrograms of vaccine were given intradermally to 20 young children with a multiple orifice head fired by a jet gun: all had a good HBV surface antibody response of greater than 100 m.i.u./ml of serum.

[Clinical and immunological studies of an experimental series of vaccines against hepatitis B]. / Klinicheskie i immunologicheskie issledovaniia éksperimental'nykh serii vaktsiny protiv gepatita B.

Two experimental lots of hepatitis B vaccine were prepared by purification of HBsAg from human plasma, inactivation at 100 degrees C for 2 min and at 37 degrees C for 72 hours with formalin in a concentration of 1:4000. The former lot comprises purified HBsAg (40 mg/ml) adsorbed with 0.32 mg % Al(OH)3, the latter consists of purified HBsAg stabilized with 0.1% human albumin and adsorbed with 1 mg% Al(OH)3. The immune response after vaccination with the first lot was observed in 54.54% of the vaccinees. The second vaccine after 3 administrations of 80 micrograms/ml each produced a very good primary and a very good secondary immune response. Both vaccines are nonreactogenic and well tolerated by the vaccinees.