Carl J. Burke

Safety, immunogenicity and efficacy in healthy infants of G1 and G2 human reassortant rotavirus vaccine in a new stabilizer/buffer liquid formulation.

Background. A refrigerator-stable rotavirus (RV) vaccine that withstands gastric acid is anticipated to permit more widespread use of RV vaccine. Objective. We investigated for the first time in infants an oral, liquid formulation of G1 and G2 human bovine reassortant rotavirus vaccine (HRRV) with a new stabilizer/buffer (S/B) containing sucrose, sodium phosphate and sodium citrate. Methods. During 1997 through 1998, 731 healthy infants ∼2 to 4 months of age were enrolled at 19 US sites to receive 3 HRRV or placebo doses ∼6 to 8 weeks apart in a partially double blinded study. Infants were randomized to: (1) HRRV with no S/B but with prefeeding; (2) HRRV plus 1 of 3 different concentrations/volumes of S/B; or (3) placebo. Results. No serious vaccine-related adverse experiences or intussusception cases were reported. No statistically significant differences were observed between vaccine and placebo recipients for fever (≥38.1°C) 0 to 7 days after any dose, irritability, vomiting or diarrhea incidence 0 to 42 days after any dose. Vaccine virus shedding among vaccine recipients was uncommon. Among S/B vaccine groups, proportions of infants with a ≥3-fold titer rise from baseline to Postdose 3 for G1 serum-neutralizing antibody (SNA), G2 SNA, WC3 SNA, serum anti-RV IgA, serum anti-RV IgG and stool anti-RV IgA were generally similar to those of the prefed non-S/B group. Conclusions. HRRV with a new S/B was generally well-tolerated; immunogenicity was generally similar to the prefed non-S/B group. No intussusception cases were reported, but the small sample size precluded a definitive conclusion. A large international clinical study is under way to address safety and efficacy of an S/B formulation of a pentavalent version of HRRV.

Preformulation studies as an essential guide to formulation development and manufacture of protein pharmaceuticals

Preformulation generally refers to a process in which a bulk drug material is characterized to a sufficient extent that it can be converted to a pharmaceutically acceptable drug substance. In the case of conventional, small-molecule (nonmacromolecular) pharmaceuticals, this procedure can usually be accomplished to a high degree of scientific rigor. For example, the atomic level structures of these compounds are usually known from a combination of X-ray crystallography, nuclear magnetic resonance (NMR), and mass spectrometry. Furthermore, the sensitivity of modern high-performance chromatography systems coupled to mass spectrometric detectors often permits altered forms of these molecules to be simply detected and identified. Unfortunately, the increased complexity of proteins, arising from a combination of their large size and higher order forms (secondary, tertiary, quaternary) of structure does not yet permit the more straightforward type of analysis utilized for small molecules to be simply applied. Nevertheless, it is still possible to treat proteins primarily as chemical (rather than biological) entities through an understanding of their structure and stability and use of a more complex array of lower resolution methodologies.

Large-scale purification and characterization of recombinant tick anticoagulant peptide

Recombinant tick anticoagulant peptide (r-TAP), a potent and specific inhibitor of blood coagulation factor Xa, was purified to greater than 99% homogeneity at the multi-gram scale. Genetically engineered yeast secreted 200-250 mg/l of the heterologous protein into the medium. Cells were separated from broth by diafiltration and purification was done by two chromatographic steps, both conducive to operation on a large scale. Analysis of the purified protein by several methods indicated that it was greater than 99% homogeneous and no incompletely processed or truncated proteins were detected. Physico-chemical characterization data of r-TAP show that it exists as a monomer in solution and no evidence of post-translational modification was observed. The purified protein was fully active in inhibiting human coagulation factor Xa.