• Asian Pacific Journal of Cancer Prevention
• /
• v.16 no.9
• /
• pp.3843-3847
• /
• 2015

Background: Cervical cancer is the second most common cause of cancer related death of women. Persistent HPV infection, especially with high-risk types such as HPV16 and HPV18, has been identified to be the primary cause of cervical cancer. E6 and E7 are the major oncoproteins of high-risk HPVs, which are expressed exclusively in HPV infected tissues, and thereby represent ideal therapeutic targets for immunotherapy of cervical cancer. Materials and Methods: In this work, we used recombinant adenovirus expressing coden-optimized HPV16 E6 and E7 fusion protein (Ad-ofE6E7) to prime dendritic cells (DC-ofE6E7), to investigate the ability of primed DC vaccine in eliciting antitumor immunity in vitro and vivo. Results: Our results indicated that DC-ofE6E7 vaccine co-culturing with splenocytes could strongly induce a tumor-specific cytotoxic T lymphocyte (CTL) response and kill the TC-1 cells effectively in vitro. Moreover, DC-ofE6E7 vaccine induced protective immunity against the challenge of TC-1 cancer cells in vivo. Conclusions: The results suggested that the HPV16 ofE6E7 primed DC vaccine has potential application for cervical cancer immunotherapy.

• Korean Journal of Veterinary Research
• /
• v.51 no.2
• /
• pp.151-158
• /
• 2011

The study evaluated whether a transgenic carrot vaccine could induce a K88-specific immune response in sows and whether the resultant maternal antibody could protect piglets against enterotoxigenic Escherichia coli (ETEC) K88ac infection. Sows (n = 15) selected randomly from a farm in Korea were assigned to three groups (n = 5 per group: control [untreated]), group A (orally inoculated with a nontransgenic and transgenic carrot vaccines at 2 and 4 weeks ante partum, respectively), and group B (conventionally vaccinated according to the manufacturer's instructions). After 7 days of lactation, 5 piglets selected randomly from each group were challenged with $1{\times}10^{10}$ colony forming units/mL ETEC K88ac. Group C had the lowest mean fecal consistency score on post-challenge days 1 and 7. Histiologically, On post-challenge day 7, group C showed an increased duodenum and ileum villus:crypt ratio, compared to group A in the duodenum, with group B displaying the highest ratio. Groups B and C had more increased villus width than group A in the jejunum. Group C displayed the greatest increase in villus width in the ileum. The colostrums and serum from groups B and C displayed higher concentrations of IgA and IgG against ETEC K88, compared to group A. Based on the results, it was concluded that the transgenic carrot vaccine in sow per oral may have an effect on preventing piglet diarrhea as good as commercial recombinant vaccine.

• Tuberculosis and Respiratory Diseases
• /
• v.57 no.2
• /
• pp.125-131
• /
• 2004

Tuberculosis (TB) remains an enormous global health problem, and a new vaccine against TB more potent than the current inadequate BCG vaccine is urgently needed. We constructed three recombinant Mycobacterium bovis BCG (rBCG) strains over-expressing antigen (Ag) 85A, Ag85B, or both of M. tuberculosis using their own promoter and secretory sequence, or hsp60 promoter. SDS-PAGE analysis of rBCG proteins showed overexpression of Ag85A and Ag85B proteins in higher level than of those in their parental strain of BCG. In addition, rBCG(rBCG/B.FA) over-expressing Ag85A and Ag85B induced strong IFN-${\gamma}$ production in splenocytes. However, there was no significant difference in protective efficacy between rBCG and their parental BCG strain. In this study, therefore, rBCG over-expressing Ag85A, Ag85B, or both failed to show enhanced protection against M. tuberculosis infection in a mouse model.

• Journal of Microbiology and Biotechnology
• /
• v.27 no.9
• /
• pp.1701-1710
• /
• 2017

Classical swine fever virus (CSFV) is the etiologic agent of classical swine fever, a highly contagious disease that causes significant economic losses to the swine industry. The lapinized C-strain, a widely used vaccine strain against CSFV, has low growth efficiency in cell culture, which limits the productivity in the vaccine industry. In this study, a recombinant virus derived from C-strain was constructed and subjected to continuous passaging in PK-15 cells with the goal of acquiring a high progeny virus yield. A cell-adapted virus variant, RecCpp80, had nearly 1,000-fold higher titer than its parent C-strain but lost the ability to induce fever in rabbits. Sequence analysis of cell-adapted RecC variants indicated that at least six nucleotide changes were fixed in RecCpp80. Further adaption of RecCpp80 variant in swine testicle cells led to a higher virus yield without additional mutations. Introduction of each of these residues into the wild-type RecC backbone showed that one mutation, M979R (T3310G), located in the C-terminal region of E2 might be closely related to the cell-adapted phenotype. Rabbit inoculation revealed that $RecCpp40_{+10}$ failed to induce fever in rabbits, whereas $RecCpp80_{+10}$ caused a fever response similar to the commercial C-strain vaccine. In conclusion, the C-strain can be adapted to cell culture by introducing specific mutations in its E2 protein. The mutations in RecCpp80 that led to the loss of fever response in rabbits require further investigation. Continuous passaging of the C-strain-based recombinant viruses in PK-15 cells could enhance its in vitro adaption. The non-synonymous mutations at 3310 and 3531 might play major roles in the enhanced capacity of general virus reproduction. Such findings may help design a modified C-strain for improved productivity of commercial vaccines at reduced production cost.

• Journal of Microbiology and Biotechnology
• /
• v.29 no.2
• /
• pp.330-338
• /
• 2019

Chronic infection with intracellular Brucella abortus (B. abortus) in livestock remains as a major problem worldwide. Thus, the search for an ideal vaccine is still ongoing. In this study, we evaluated the protective efficacy of a combination of B. abortus recombinant proteins; superoxide dismutase (rSodC), riboflavin synthase subunit beta (rRibH), nucleoside diphosphate kinase (rNdk), 50S ribosomal protein (rL7/L12) and malate dehydrogenase (rMDH), cloned and expressed into a pMal vector system and $DH5{\alpha}$, respectively, and further purified and applied intraperitoneally into BALB/c mice. After first immunization and two boosters, mice were infected intraperitoneally (IP) with $5{\times}10^4CFU$ of virulent B. abortus 544. Spleens were harvested and bacterial loads were evaluated at two weeks post-infection. Results revealed that this combination showed significant reduction in bacterial colonization in the spleen with a log protection unit of 1.31, which is comparable to the average protection conferred by the widely used live attenuated vaccine RB51. Cytokine analysis exhibited enhancement of cell-mediated immune response as IFN-${\gamma}$ is significantly elevated while IL-10, which is considered beneficial to the pathogen's survival, was reduced compared to control group. Furthermore, both titers of IgG1 and IgG2a were significantly elevated at three and four-week time points from first immunization. In summary, our in vivo data revealed that vaccination with a combination of five different proteins conferred a heightened host response to Brucella infection through cell-mediated immunity which is desirable in the control of intracellular pathogens. Thus, this combination might be considered for further improvement as a potential candidate vaccine against Brucella infection.

• Journal of Microbiology and Biotechnology
• /
• v.17 no.6
• /
• pp.879-890
• /
• 2007

The identification of tumor antigens is essential for the development of anticancer therapeutic vaccines and clinical diagnosis of cancer. SEREX (serological analysis of recombinant cDNA expression libraries) has been used to identify such tumor antigens by screening sera of patients with cDNA expression libraries. SEREX-defined antigens provide markers for the diagnosis of cancers. Potential diagnostic values of these SEREX-defined antigens have been evaluated. SEREX is also a powerful method for the development of anticancer therapeutics. The development of anticancer vaccines requires that tumor antigens can elicit antigen-specific antibodies or T lymphocytes. More than 2,000 antigens have been discovered by SEFEX. Peptides derived from some of these antigens have been evaluated in clinical trials. This review provides information on the application of SEREX for identification of tumor-associated antigens (TAA) for the development of cancer diagnostics and anticancer therapeutics.

• Journal of Microbiology and Biotechnology
• /
• v.21 no.12
• /
• pp.1264-1269
• /
• 2011

Interleukin-2 (IL-2) is a vital cytokine secreted by activated T lymphocytes, and plays an important role in the regulation of cellular functions and immunity of animals. In this study, the recombinant duck IL-2 (rduIL-2) was secretory expressed in Pichia pastoris (P. pastoris). The recombinant P. pastoris strain was cultured in shake flasks and then scaled up in a 5.0-l bioreactor. The result showed that the maximal fresh-cell-weight of 594.1 g/l and the maximal $OD_{600}$ of 408 were achieved in the bioreactor. The rduIL-2 was purified by two steps of purification procedures, and approximately 311 mg of rduIL-2/L fermentation supernatant was obtained. SDS-PAGE showed that the purified rduIL-2 constituted a homogeneous band of ~16 kDa or ~14 kDa corresponding to the glycosylated or non-glycosylated duIL-2 protein in size, respectively. The bioactivity of rduIL-2 was determined by lymphocyte proliferation assay. The result indicated that the rduIL-2 greatly promoted the proliferation of ConA-stimulated lymphocytes in vitro. The P. pastoris expression system described here could provide promising, inexpensive, and large-scale production of the rduIL-2, which lays the foundation for development of novel immunoadjuvants to enhance both the immunity of ducks against various infectious pathogens and vaccine efficacy.

• Journal of Microbiology and Biotechnology
• /
• v.8 no.4
• /
• pp.361-368
• /
• 1998

The E2 protein of HCV (hepatitis C virus) is thought to have a potential role in the development of subunit vaccines and diagnostics. To express it by the insect cell/baculovirus expression (Bacu) system, we constructed a recombinant Autographa californica nuclear polyhedrosis virus (AcIL3E2), determined the most appropriate expression conditions in terms of host cell line and culture medium, and characterized the expressed HCV E2 protein. A culture system using Trichoplusia ni BTI-TN5Bl-4 cells and SF 900IISFM medium expressed a relatively high level of HCV E2 protein. It was revealed that its glycosylation properties and subcellular localization were almost the same as the ones in the mammalian cell expression system previously reported, suggesting the recombinant HCV E2 protein derived from our Bacu system can be utilized for development of a subunit vaccine and diagnostics. Interestingly, HCV E2 protein was not degraded at all even at 43 h post-heat shock in the heat shock-induced necrotic cells, probably due to its integration into the microsomal membrane, indicating that heat shock can be employed to purify HCV E2 protein.

• Archives of Pharmacal Research
• /
• v.21 no.5
• /
• pp.543-548
• /
• 1998

We examined the immunological properties of the recombinant hepatitis B surface antigen (r-HBsAg) which was expressed in mammalian cell (C127). The cross-immunity of r-HBsAg and plasma-derived hepatitis B surface antigen (p-HBsAg) were tested using Western blotting and ELISA with guinea pig polyclonal antibody and naturally infected human-derived antibody and the both antigens show the same results in their response pattern and intensity, which indicate they have a good cross-immunity. from the measurement of $ED_{50}$ after formalin- or heat-inactivation, both r-HBsAg and p-HBsAg and p-HBsAg showed $ED_{50}$ of 0.2-0.3 in formalin-inactivaton, while r-HBsAg was 0.05-0.09 and p-HBsAg was 0.03-0.07 in heat-inactivation, which means heat-inactivation method is 3-4 times superior in immunogenicity. In the immunopersistency test performed in guinea pig for the period of 3 months with two different adjuvants, antibody titer was 34.2 with muramyl dipeptide adjuvant, which was 1.8 times greater than the antibody titer of 18.9 with $AIPO_{4}$ adjuvant. the mutagenicity of r-HBsAg has the same cross-immunity with p-HBsAg, and heat-inactivation method and muramyl dipeptide adjuvant allow development of r-HBsAg vaccine with excellent immunogenicity.

• Parasites, Hosts and Diseases
• /
• v.62 no.1
• /
• pp.75-84
• /
• 2024

Ancylostoma ceylanicum is a zoonotic soil-derived nematode that parasitizes the intestines of humans and animals (dogs and cats), leading to malnutrition and iron-deficiency anemia. Helminth parasites secrete calreticulin (CRT), which regulates or blocks the host's immune response. However, no data on A. ceylanicum calreticulin (Ace-CRT) are available. We investigated the biological function of recombinant Ace-CRT (rAce-CRT). rAce-CRT showed reliable antigenicity and stimulated the proliferation of mouse splenocytes and canine peripheral blood mononuclear cells. Quantitative reverse-transcription PCR assays revealed that rAce-CRT primarily promoted the expression of T helper 2 cytokines, particularly IL-13, in canine peripheral blood lymphocytes. rAce-CRT inhibited complement-mediated sheep erythrocyte hemolysis in vitro. Our findings indicate that Ace-CRT plays an immunomodulatory role and may be a promising candidate molecule for a hookworm vaccine.