Impact of adjuvants on CD4(+) T cell and B cell responses to a protein antigen vaccine: Results from a phase II, randomized, multicenter trial.

Immunogenicity and safety of different adjuvants combined with a model antigen (HBsAg) were compared. Healthy HBV-naïve adults were randomized to receive HBs adjuvanted with alum or Adjuvant Systems AS01B, AS01E, AS03A or AS04 at Days 0 and 30. Different frequencies of HBs-specific CD4+ T cells 14days post dose 2 but similar polyfunctionality profiles were induced by the different adjuvants with frequencies significantly higher in the AS01B and AS01E groups than in the other groups. Antibody concentrations 30days post-dose 2 were significantly higher in AS01B, AS01E and AS03A than in other groups. Limited correlations were observed between HBs-specific CD4+ T cell and antibody responses. Injection site pain was the most common solicited local symptom and was more frequent in AS groups than in alum group. Different adjuvants formulated with the same antigen induced different adaptive immune responses and reactogenicity patterns in healthy naïve adults. The results summary for this study (GSK study number 112115 - NCT# NCT00805389) is available on the GSK Clinical Study Register and can be accessed at www.gsk-clinicalstudyregister.com.

An adjuvanted polyprotein HIV-1 vaccine induces polyfunctional cross-reactive CD4+ T cell responses in seronegative volunteers.

BACKGROUND: This phase I/II partially blinded, randomized, dose-ranging study assessed the safety and immunogenicity of a novel human immunodeficiency virus type 1 (HIV-1) vaccine candidate consisting of a recombinant fusion protein (F4) containing 4 HIV-1 clade B antigens (Gag p24, Pol reverse transcriptase, Nef, and Gag p17) adjuvanted with AS01 in HIV-seronegative volunteers. Methods. Two doses of the recombinant F4 protein (10, 30, or 90 µg/dose), adjuvanted with AS01 or reconstituted with water for injection, were administered 1 month apart to 180 healthy volunteers aged 18-40 years. F4-specific CD4(+) T cell responses were measured using intracellular cytokine staining after in vitro stimulation by overlapping peptide pools covering the 4 individual antigens. Results. Reactogenicity was higher during the 7-day period after each vaccine dose in the adjuvanted than in the nonadjuvanted groups. In the adjuvanted groups, the overall immune response rate was high after the second vaccine dose, with highest responder rates seen in the 10-µg F4/AS01 group (100% to 3 HIV-1 antigens and 80% to all 4 HIV-1 antigens). High and long-lasting CD4(+) T cell frequencies were observed (up to a median value of 1.2% F4-specific CD4(+) T cells at day 44), with strongest responses directed against reverse transcriptase. Antigen-specific CD4(+) T cells exhibited a polyfunctional phenotype, expressing at least CD40 ligand and interleukin 2, often in combination with tumor necrosis factor α and/or interferon γ. Vaccine-induced CD4(+) T cell responses were broadly cross-reactive to all 4 antigens derived from HIV-1 clades A and C. Conclusions. These results support further clinical investigation of this HIV-1 vaccine candidate both in a prophylactic setting (alone, in conjunction with an envelope-based antigen or in combination with other vaccine approaches in a heterologous prime-boost regimen) and as a potentially disease-modifying therapeutic vaccine in HIV-1-infected subjects. CLINICAL TRIALS REGISTRATION: NCT00434512.

Synthetic studies on cyclobuta[a]indanes: stereocontrolled access to C9-substituted derivatives.

We report the preparation of novel ethyl cyclobuta[a]indane derivatives of pharmacological interest. The synthesis of compounds 5 was used as a model to study stereocontrolled access to C9-substituted cyclobutane from the corresponding cyclobutanone 1. Progress was made on two complementary aspects: (1) catalytic hydrogenation from the appropriate cyclobutene precursors; and (2) delivery of the C9 substituent through an intramolecular process. The level of diastereoselectivity obtained through hydrogenation of cyclobutene depends both on the metal used as catalyst and the nature of the functional group proximal to the double bond. The intramolecular delivery approach was diastereospecific, provided that the spiro-intermediate 7 does not ring open during the stereoinduction step. The latter route should allow preparation of quaternary carbon at C9 not available through hydrogenation.

Glycosylation engineering of spinosyn analogues containing an L-olivose moiety.

Biosynthetic genes encoding proteins involved in the first steps of deoxyhexose biosynthesis from D-glucose-1-phosphate were expressed in Saccharopolyspora erythraea. The resulting mutant was able to accumulate and utilise TDP-L-olivose. Co-expression of the spinosyn glycosyl transferase SpnP in the resulting mutant endowed upon it the ability to biotransform exogenously added spinosyn aglycones to yield novel spinosyn analogues.

Comparison of the immunogenicity and safety of Cervarix and Gardasil human papillomavirus (HPV) cervical cancer vaccines in healthy women aged 18-45 years.

This observer-blind study compared the prophylactic human papillomavirus (HPV) vaccines, Cervarix (GlaxoSmithKline) and Gardasil (Merck), by assessing immunogenicity and safety through one month after completion of the three-dose vaccination course. Women (n = 1106) were stratified by age (18-26, 27-35, 36-45 years) and randomized (1:1) to receive Cervarix (Months 0, 1, 6) or Gardasil (Months 0, 2, 6). At Month 7 after first vaccination, all women in the according-to-protocol cohort who were seronegative/DNA negative before vaccination for the HPV type analyzed had seroconverted for HPV-16 and HPV-18 serum neutralizing antibodies, as measured by pseudovirion-based neutralization assay (PBNA), except for two women aged 27-35 years in the Gardasil group who did not seroconvert for HPV-18 (98%). Geometric mean titers of serum neutralizing antibodies ranged from 2.3-4.8-fold higher for HPV-16 and 6.8-9.1-fold higher for HPV-18 after vaccination with Cervarix compared with Gardasil, across all age strata. In the total vaccinated cohort (all women who received at least one vaccine dose, regardless of their serological and DNA status prior to vaccination), Cervarix induced significantly higher serum neutralizing antibody titers in all age strata (p < 0.0001). Positivity rates for anti-HPV-16 and -18 neutralizing antibodies in cervicovaginal secretions and circulating HPV-16 and -18 specific memory B-cell frequencies were also higher after vaccination with Cervarix compared with Gardasil. Both vaccines were generally well tolerated. The incidence of unsolicited adverse events was comparable between vaccinated groups. The incidence of solicited symptoms was generally higher after Cervarix, injection site reactions being most common. However, compliance rates with the three-dose schedules were similarly high (>or= 84%) for both vaccines. Although the importance of differences in magnitude of immune response between these vaccines is unknown, they may represent determinants of duration of protection against HPV-16/18. Long-term studies evaluating duration of efficacy after vaccination are needed for both vaccines.

Molecular characterization of macbecin as an Hsp90 inhibitor.

Macbecin compares favorably to geldanamycin as an Hsp90 inhibitor, being more soluble, stable, more potently inhibiting ATPase activity (IC50 = 2 microM) and binding with higher affinity (Kd = 0.24 microM). Structural studies reveal significant differences in their Hsp90 binding characteristics, and macbecin-induced tumor cell growth inhibition is accompanied by characteristic degradation of Hsp90 client proteins. Macbecin significantly reduced tumor growth rates (minimum T/C: 32%) in a DU145 murine xenograft. Macbecin thus represents an attractive lead for further optimization.

Engineered biosynthesis of hybrid macrolide polyketides containing D-angolosamine and D-mycaminose moieties.

The glycosylation of natural product scaffolds with highly modified deoxysugars is often essential for their biological activity, being responsible for specific contacts to molecular targets and significantly affecting their pharmacokinetic properties. In order to provide tools for the targeted alteration of natural product glycosylation patterns, significant strides have been made to understand the biosynthesis of activated deoxysugars and their transfer. We report here efforts towards the production of plasmid-borne biosynthetic gene cassettes capable of producing TDP-activated forms of D-mycaminose, D-angolosamine and D-desosamine. We additionally describe the transfer of these deoxysugars to macrolide aglycones using the glycosyl transferases EryCIII, TylMII and AngMII, which display usefully broad substrate tolerance.

Biosynthesis of the angiogenesis inhibitor borrelidin: directed biosynthesis of novel analogues.

We report the directed biosynthesis of borrelidin analogues and their selective anti-proliferative activity against human cancer cell lines.

Vaccine Adjuvant Systems containing monophosphoryl lipid A and QS-21 induce strong humoral and cellular immune responses against hepatitis B surface antigen which persist for at least 4 years after vaccination.

BACKGROUND: Recombinant hepatitis B surface antigen (HBsAg) was used as a model antigen to evaluate persistence of cellular and humoral immune responses when formulated with three different Adjuvant Systems containing 3-O-desacyl-4'-monophosphoryl lipid A (MPL) and QS-21, in an oil-in-water emulsion (AS02B and AS02V), or with liposomes (AS01B). METHODS: This is an open, 4-year follow-up of a previous randomised, double-blind study. Healthy subjects aged 18-40 years received three vaccine doses on a month 0, 1, 10 schedule and were initially followed for 18 months. A total of 93 subjects (AS02B: n=30; AS02V: n=28; AS01B: n=35) were enrolled in this follow-up and had an additional blood sample taken at Year 4 (NCT02153320). The primary endpoint was the frequency of HBsAg-specific CD4(+) and CD8(+) T-cells expressing cytokines upon short-term in vitro stimulation of peripheral blood mononuclear cells with HBsAg-derived peptides. Secondary endpoints were anti-HBs antibody titres and frequency of HBsAg-specific memory B-cells. RESULTS: A strong and persistent specific CD4(+) T-cell response was observed at Year 4 in all groups. HBsAg-specific CD4(+) T-cells expressed mainly CD40L and IL-2, and to a lesser extent TNF-α and IFN-γ. HBsAg-specific CD8(+) T-cells were not detected in any group. A high, persistent HBsAg-specific humoral immune response was observed in all groups, with all subjects seroprotected (antibody titre ≥10mIU/mL) at Year 4. The geometric mean antibody titre at Year 4 was above 100,000mIU/mL in all groups. A strong memory B-cell response was observed post-dose 2, which tended to increase post-dose 3 and persisted at Year 4 in all groups. CONCLUSION: The MPL/QS-21/HBsAg vaccine formulations induced persistent immune responses up to 4 years after first vaccination. These Adjuvant Systems offer potential for combination with recombinant, synthetic or highly purified subunit vaccines, particularly for vaccination against challenging diseases, or in specific populations, although additional studies are needed.

A phase II, randomized, safety and immunogenicity study of a re-derived, live-attenuated dengue virus vaccine in healthy adults.

Two formulations of a new live tetravalent dengue virus (DENV) vaccine produced using re-derived master seeds from a precursor vaccine and that same precursor vaccine as a control were compared in a placebo-controlled, randomized, observer-blind, phase II trial of 86 healthy adults. Two vaccine doses were administered 6 months apart; a third dose was offered to a subset. Symptoms and signs of dengue-like illness reported after vaccination were mild to moderate, transient, and occurred with similar frequency among recipients of the new DENV vaccine and placebo, except for rash. Neither dengue nor vaccine-related serious adverse events were reported. The first DENV vaccine dose was moderately immunogenic; the second dose increased the potency and breadth of the neutralizing antibody response. Tetravalent response rates to the new formulations were 60% and 66.7% in unprimed subjects. A third dose did not increase tetravalent antibody rates. The new DENV vaccine candidates merit additional evaluation.

Separation of anti-angiogenic and cytotoxic activities of borrelidin by modification at the C17 side chain.

A set of novel borrelidin analogues have been prepared by precursor-directed biosynthesis. Structure-activity relationship analysis suggests that steric structural arrangement within the C17 side chain is important for differentiating cytotoxic and anti-angiogenic activities. A C17-cyclobutyl analogue 3 was found to have markedly increased selectivity for in vitro angiogenesis inhibition over cytotoxicity and is therefore potentially useful as an anticancer agent.

Rapamycin biosynthesis: Elucidation of gene product function.

The function of gene products involved in the biosynthesis of the clinically important polyketide rapamycin were elucidated by biotransformation and gene complementation.

Yardenone A and B: new cytotoxic triterpenes from the Indian Ocean sponge Axinella cf. bidderi.

Two new polyepoxysqualene-derived triterpenes, yardenone A (1) and B (2), together with the known yardenone (3) and sodwanone A (4), have been isolated from the marine sponge Axinella cf. bidderi from Yemen's Socotra Island in the Indian Ocean. The structures were elucidated using spectroscopic data. The relative stereochemistry was established by the analysis of ROESY spectra as well as coupling constants and molecular modeling. Furthermore, the absolute configuration of 1 was confirmed by the advanced Mosher's method. The cytotoxicity of these compounds was evaluated against a NSCLC cell line.