Pilot scale production of the vaccine adjuvant Proteoliposome derived Cochleates (AFCo1) from Neisseria meningitidis serogroup B.

The use of new adjuvants in vaccine formulations is a subject of current research. Only few parenteral adjuvants have been licensed. We have developed a mucosal and parenteral adjuvant known as AFCo1 (Adjuvant Finlay Cochleate 1, derived from proteoliposomes of N. meningitidis B) using a dialysis procedure to produce them on lab scale. The immunogenicity of the AFCo1 produced by dialysis has been already evaluated, but it was necessary to demonstrate the feasibility of a larger-scale manufacturing process. Therefore, we used a crossflow diafiltration system (CFS) that allows easy scale up to obtain large batches in an aseptic environment. The aim of this work was to produce AFCo1 on pilot scale, while conserving the adjuvant properties. The proteoliposomes (raw material) were resuspended in a buffer containing sodium deoxycholate and were transformed into AFCo1 under the action of a calcium forming buffer. The detergent was removed from the protein solution by diafiltration to a constant volume. In this CFS, we used a hollow fiber cartridge from Amicon (polysulfona cartridge of 10 kDa porosity, 1mm channel diameter of fiber and 0.45 m² area of filtration), allowing production of a batch of up to 20 L. AFCo1 were successfully produced by tangential filtration to pilot scale. The batch passed preliminary stability tests. Nasal immunization of BALB/c mice, induced specific saliva IgA and serum IgG. The induction of Th1 responses were demonstrated by the induction of IgG2a, IFNγ and not IL-5. The adjuvant action over Neisseria (self) antigens and with co-administered (heterologous) antigens such as ovalbumin and a synthetic peptide from haemolytic Streptococcus B was also demonstrated.

Mucosal immunization using proteoliposome and cochleate structures from Neisseria meningitidis serogroup B induce mucosal and systemic responses.

Most pathogens either invade the body or establish infection in mucosal tissues and represent an enormous challenge for vaccine development by the absence of good mucosal adjuvants. A proteoliposome-derived adjuvant from Neisseria meningitidis serogroup B (AFPL1, Adjuvant Finlay Proteoliposome 1) and its derived cochleate form (Co, AFCo1) contain multiple pathogen-associated molecular patterns as immunopotentiators, and can also serve as delivery systems to elicit a Th1-type immune response. The present studies demonstrate the ability of AFPL1and AFCo1 to induce mucosal and systemic immune responses by different mucosal immunizations routes and significant adjuvant activity for antibody responses of both structures: a microparticle and a nanoparticle with a heterologous antigen. Therefore, we used female mice immunized by intragastric, intravaginal, intranasal or intramuscular routes with both structures alone or incorporated with ovalbumin (OVA). High levels of specific IgG antibody were detected in all sera and in vaginal washes, but specific IgA antibody in external secretions was only detected in mucosally immunized mice. Furthermore, antigen specific IgG1 and IgG2a isotypes were all induced. AFPL1 and AFCo1 are capable of inducing IFN-gamma responses, and chemokine secretions, like MIP-1alpha and MIP-1beta. However, AFCo1 is a better alternative to induce immune responses at mucosal level. Even when we use a heterologous antigen, the AFCo1 response was better than with AFPL1 in inducing mucosal and systemic immune responses. These results support the use of AFCo1 as a potent Th1 inducing adjuvant particularly suitable for mucosal immunization.

[Skin tests, serum specific IgE and total IgE in the diagnosis of patients with perennial allergic rhinitis]. / Pruebas cutáneas, IgE sérica específica e IgE total en el diagnóstico de pacientes con rinitis alérgica perenne.

BACKGROUND: Skin tests are the most used diagnostic method of allergic rhinitis, which, in addition to identify specific allergen, can determine the relative sensitivity of one patient to the allergen. OBJECTIVE: To assess the association between skin test reactivity and total and specific serum IgE levels on the diagnosis of patients with perennial allergic rhinitis. MATERIAL AND METHODS: We measured the response to skin test reactivity and total and specific serum IgE levels in 69 patients with perennial allergic rhinitis. RESULTS: The skin test reactivity showed responses to: Dermatophagoides pt in 62 patients (90%), house dust in 57 (83%), cat in 41 (59%), dog in 23 (33%), and Lolium p in 16 (23%). The mean level of total IgE was of 378 UI (19 to 4,036) and that of specific IgE was of 39.2 UI (0.2 to 98.6). Total IgE > 200 UI was observed in fifty two patients (75%), which was significantly lesser than the frequency of specific IgE > 0.35 UI (at least for an allergen), which was 94% (p < 0.05, Z). At least two tests of skin reactivity were positive for 90% of the patients. This frequency was similar to the 95% for the specific IgE but different to the 75% for the total IgE (p < 0.05, X2). The correlation between the results of the skin test reactivity and the specific seric IgE showed Spearman r from 0.23 to 0.35 (p < 0.05). The correlation between the total IgE and the specific IgE showed a Spearman r of 0.08 (p < 0.05) and between the total IgE and the skin test reactivity an r of 0.15 (p < 0.05). CONCLUSION: In the diagnosis of perennial allergic rhinitis, the results of the skin test reactivity and the specific serum IgE are correlated but these two results are non consistent with the results of the total serum IgE.