Preclinical safety assessment of recombinant botulinum vaccine A/B (rBV A/B).

A recombinant botulinum vaccine (rBV A/B) is being developed to protect adults 18-55 years of age from fatal botulism caused by inhalational intoxication with botulinum neurotoxin complex (BoNT) serotype A, subtype A1 (BoNT/A1) and BoNT serotype B, subtype B1 (BoNT/B1). Fundamental to the advanced development process is an initial demonstration of product safety in animals. A comprehensive series of studies was conducted to evaluate the general toxicity, neurobehavioral toxicity and local reactogenicity of the rBV A/B vaccine prior to first use in humans. Toxicity was evaluated in CD-1 mice vaccinated with control material and three dosages of rBV A/B with or without Alhydrogel(®) by intramuscular (IM) injection on Study Days 0, 28, 56 and 70 in a volume of 100µL. Total immunizing protein given in each dose was either 0, 2, 4 or 8 µg/animal. Local reactogenicity was evaluated in mice at the dosages given and in New Zealand white (NZW) rabbits using the same injection volume (0.5 mL) and formulations (10, 20 and 40 g/mL total antigen with 0.2% (w/v) Alhydrogel(®)) intended for human use. The rBV A/B vaccine produced no apparent systemic or neurobehavioral toxicity and only transient mild inflammation at the injection site. Together these results indicated a favorable safety profile for rBV A/B and supported its use in a Phase 1 clinical trial.

[Evaluation of immunobiological activity of Francisella tularensis C-complex preparations as promising component of subunit vaccines].

Data on influence of Francisella tularensis C-complex preparations on formation of immunity against tularemia are presented. Study of cellular immunity characteristics as well as dynamics of antibody response was carried out on white mice and guinea pigs models. Absence of toxicity, pyrogenicity, and negative effects on immunocompetent cells in combination with protective activity points to possibility of use the C-complex as a component of a subunit vaccine.

Technology evaluation: ChimeriVax-DEN, Acambis/Aventis.

Acambis, in collaboration with Aventis Pasteur, is developing a chimeric vaccine based on a recombinant yellow fever vaccine for the potential prevention of dengue virus infection. The vaccine is undergoing phase I clinical trials.

Pre-clinical evaluation of the malaria vaccine candidate P. falciparum MSP1(42) formulated with novel adjuvants or with alum.

We compared the safety and immunogenicity of the recombinant Plasmodium falciparum MSP1(42) antigen formulated with four novel adjuvant systems (AS01B, AS02A, AS05 and AS08) to alum in rhesus monkeys. All five formulations of MSP1(42) were safe and immunogenic. Whereas, all MSP1(42) formulations tested generated high stimulation indices for lymphocyte proliferation (ranging from 27 to 50), the AS02A and AS01B formulations induced the highest levels of specific anti-MSP1(42) antibody. ELISPOT assays showed that the AS02A and AS01B vaccine formulations-induced different cytokine response profiles. Using the ratio of IFN-gamma/IL-5 secreting cells as the metric, the AS01B formulation induced a strong Th1 response, whereas the AS02A formulation induced a balanced Th1/Th2 response. The IFN-gamma response generated by AS02A and AS01B formulations persisted at least 24 weeks after final vaccination. The notable difference in Th1/Th2 polarization induced by the AS02A and AS01B formulations warrants comparative clinical testing.

Safety and immunogenicity of rts,s+trap malaria vaccine, formulated in the as02a adjuvant system, in infant rhesus monkeys.

Malaria vaccine RTS,S combined with thrombospondin-related anonymous protein (TRAP) and formulated with AS02A (RTS,S+TRAP/AS02A) is safe and immunogenic in adult humans and rhesus monkeys (Macaca mulatta). Here, RTS,S+TRAP/AS02A was administered on a 0-, 1-, and 3-month schedule to three cohorts of infant monkeys, along with adult comparators. Cohort 1 evaluated 1/5, 1/2, and full adult doses, with the first dose administration at one month of age; cohort 2 monkeys received full adult doses, with the first dose administration at one versus three months of age; and, cohort 3 compared infants gestated in mothers with or without previous RTS,S/AS02A immunization. Immunization site reactogenicity was mild. Some infants, including the phosphate-buffered saline only recipient, developed transient iron-deficiency anemia, which is considered a result of repeated phlebotomies. All RTS,S+TRAP/AS02A regimens induced vigorous antibody responses that persisted through 12 weeks after the last vaccine dose. Modest lymphoproliferative and ELISPOT (interferon-gamma and interleukin-5) responses, particularly to TRAP, approximated adult comparators. RTS,S+TRAP/AS02A was safe and well tolerated. Vigorous antibody production and modest, selective cell-mediated immune responses suggest that RTS,S+TRAP/AS02A may be immunogenic in human infants.

[The efficacy and outlook for the study of live vaccines for the prevention of melioidosis]. / Effektivnost' i perspektivy issledovaniia zhivykh vaktsin dlia profilaktiki melioidoza.

The effect of immunization with Burkholderia pseudomallei, (Pur- and Ts), heterologous vaccines and the recombinant culture of Francisella tularensis RM2, carrying a plasmid with fragments of B. pseudomallei chromosome, was studied on four species of experimental animals, essentially differing by their sensitivity to melioidosis. B. pseudomallei mutants formed the statistically significant level of protection in subcutaneously challenged animals, moderately sensitive to melioidosis, but were not effective when tested, under the same conditions, in animals, highly sensitive to melioidosis. The effect produced by the experimental vaccines under study in animals of all species, subjected to aerogenic challenge, was leveled. The study showed good prospects for the use of tularemia vaccine with a view to create heterologous immunity to melioidosis and the possibility of its use as the basis of bivalent gene engineering vaccine.

[The outer membranes of Vibrio cholerae as a potential component in a chemical vaccine]. / Naruzhnye membrany kholernogo vibriona kak potentsial'nyi komponent khimicheskoi vaktsiny.

The results of the study of the preparation of V. cholerae eltor membrane, obtained by the lysis and inactivation of microbial cells with urea and the subsequent differential centrifugation and nuclease treatment. As revealed in this study, the outer membrane preparation, when introduced parenterally and orally to mice, induced pronounced immunity to experimental cholera infection and the production of vibriocidal antibodies in high titers. The treatment of V. cholerae eltor membranes with trypsin led to further increase of the immunogenic potency of the preparation. The protective action of V. cholerae eltor outer membranes considerably exceeded the protective effect of currently used whole-cell eltor vaccine. This opens prospects for using the above-mentioned preparation for the improvement of chemical vaccine as a component ensuring the formation of antibacterial immunity.

[A chemical typhoid vaccine containing a surface-antigen complex including the K-antigen]. / Khimicheskaia briushnotifoznaia vaktsina, soderzhashchaia kompleks poverkhnostnykh antigenov, vkliuchaiushchii K-antigen.

A new chemical typhoid vaccine containing the complex of surface Vi- and K-antigens has been obtained and experimentally studied. The vaccine has been found to possess high immunogenic and antigenic potency. Typhoid complex vaccine containing K-antigen stimulates the development of anti-infection and postvaccinal complex.

A phase I study of HGP-30, a 30 amino acid subunit of the human immunodeficiency virus (HIV) p17 synthetic peptide analogue sub-unit vaccine in seronegative subjects.

HGP-30-KLH vaccine in alum at doses of 10, 25, 50, and 100 micrograms/kg administered intramuscularly at weeks 0, 4, and 10 appear well-tolerated clinically. Local pain at the injection site, appears to be the main clinical toxicity. Laboratory parameters are not affected by administration of the vaccine candidate except for perhaps mild urinalysis abnormalities at the highest dose. This vaccine candidate has no apparent immunotoxicity and does not appear to affect lymphocyte populations or T-cell functional studies. Low levels and transient antibodies develop in a minority of subjects early after immunization with the vaccine candidate. These responses were observed in the lowest dose range. Higher doses, and longer follow-up will be needed to confirm this observation. T-cell proliferative responses to KLH and KLH-HGP-30 are consistent and may not be dose dependent, but the proliferative responses are variable and more data need to be accumulated. Preliminary, there appears to be an HGP-30-induced CTL response of HGP-30-coated EBV-transformed autologous B cell lines. This study was approved under an IND for the California Department of Health Services' Food and Drug Branch. They have provided excellent support and regulatory guidelines for this project. Future work will extend and confirm these initial observations.

[An oral chemical vaccine from the hypertoxigenic strains of the causative agent of cholera KM-76 Inaba and KM-68 Ogawa]. / Oral'naia khimicheskaia vaktsina iz gipertoksigennykh shtammov KM-76 Inaba i KM-68 Ogava vozbuditel'a kholery.

The material on the development of chemical vaccine, prepared from two newly formed strains (KM-76 Inaba and KM-68 Ogawa) and intended for oral administration, is presented. The conditions for the submerged cultivation of these strains have been established, which makes it possible to increase the production of choleragen 8- to 10-fold and O-antigen 3- to 4-fold in comparison with V. cholerae natural strain 569B. The maximum accumulation of neuraminidase, protease, phospholipase, along with choleragen, has been registered in the logarithmic phase and that of O-antigen, in the stationary phase of growth. The use of strains KM-76 and KM-68 has led to the fourfold increase of the specific activity of the main immunogens, thus permitting the respective increase of the yield of the oral vaccine without changes in its high capacity for the formation of specific antibodies and its low residual toxigenicity.

Synthesis and immunologic properties in mice of vaccines composed of Staphylococcus aureus type 5 and type 8 capsular polysaccharides conjugated to Pseudomonas aeruginosa exotoxin A.

Epidemiological, serological and in vitro phagocytosis experiments provide evidence that the newly discovered type 5 and type 8 capsular polysaccharides (CPs) are both virulence factors and protective antigens for bacteremia caused by Staphylococcus aureus. Neither type 5 nor type 8 CP elicited serum antibodies when injected into mice. These two CPs were bound to Pseudomonas aeruginosa exotoxin A (ETA) to form conjugates by using the synthetic scheme devised for the CP (Vi) of Salmonella typhi and of pneumococcus type 12F (A. Fattom, W. F. Vann, S. C. Szu, A. Sutton, X. Li, D. Bryla, G. Schiffman, J. B. Robbins, and R. Schneerson, Infect. Immun. 56:2292-2298, 1988; S. C. Szu, A. L. Stone, J. D. Robbins, R. Schneerson, and J. B. Robbins, J. Exp. Med. 166:1510-1524, 1987). Both S. aureus CP-ETA conjugates elicited a rise in CP antibodies. As components of conjugates, both S. aureus CPs acquired T-cell-dependent properties, as shown by their ability to respond to carrier priming and to stimulate booster responses. The conjugate-induced antibodies facilitated type-specific opsonization of S. aureus by human polymorphonuclear leukocytes. The conjugates also induced ETA antibodies which neutralized the native toxin in vitro. Clinical studies of these two conjugates for active or passive immunization of patients at risk for S. aureus bacteremia are planned.