title: Strategies for stabilising tetanus toxoid towards the development of a single-dose tetanus vaccine.
abstract: The stability of tetanus toxoid (TT) has been investigated for the purpose of enhancing its immunogenicity when encapsulated in TT-polylactide-co-glycolide (PLGA) microspheres. In this paper we describe our simulations of various potential inactivating events during microsphere processing and release of the antigen. These include: i) processing aqueous TT solutions in the presence of organic solvents, ii) exposing lyophilized TT to moisture, and iii) incubating vaccine with the degrading PLGA. At 37 degrees C, TT began to aggregate in solution a few hours after the addition of either methylene chloride or ethyl acetate to form a single oil-water interface. Similarly, exposure of the lyophilized vaccine to elevated humidity caused the antigen to lose solubility. Previous analysis of moisture-induced aggregates has revealed that formaldehyde, which is stored in labile linkages in the vaccine following its detoxification, is the precursor to the reactive species in the principal aggregation pathway [1]. Methods to inhibit this mechanism, such as blocking nucleophilic amino groups of TT with succinic anhydride, were verified. Succinylation of TT resulted in the incorporation of 10-fold greater antigenically active vaccine in PLGA microspheres relative to the unmodified TT following processing by double-emulsion/solvent evaporation with ethyl acetate, strongly suggesting the formaldehyde-mediated aggregation pathway also occurs during the deleterious conditions of microsphere processing. Incubation of solutions of TT in the presence of excess blank (unloaded) poly (D,L) lactide (mol. wt. 2000) microspheres led to a dramatic reduction in pH (approximately 3.2 units after one day at 45 degrees C) and simultaneous precipitation of TT. Stabilisation in the presence of the degrading polymer is likely to be the final obstacle before controlled-release preparations can be formulated to release antigenically active TT over extended time periods. Hence, mechanistic analyses as described here may be crucial for the development of effective single-dose vaccines.
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, USA.
authors.forename: S P
authors.initials: SP
authors.lastname: Schwendeman
authors.forename: H R
authors.initials: HR
authors.lastname: Costantino
authors.forename: R K
authors.initials: RK
authors.lastname: Gupta
authors.forename: M
authors.initials: M
authors.lastname: Tobio
authors.forename: A C
authors.initials: AC
authors.lastname: Chang
authors.forename: M J
authors.initials: MJ
authors.lastname: Alonso
authors.forename: G R
authors.initials: GR
authors.lastname: Siber
authors.forename: R
authors.initials: R
authors.lastname: Langer
mesh_terms: D003692:Delayed-Action Preparations; D004339:Drug Compounding; D004355:Drug Stability; D004356:Drug Storage; D005612:Freeze Drying; D006813:Humidity; D006863:Hydrogen-Ion Concentration; D007114:Immunization / Q000379:methods; D019344:Lactic Acid; D008863:Microspheres; D014677:Pharmaceutical Vehicles; D011100:Polyglycolic Acid; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011108:Polymers; D012997:Solvents / Q000494:pharmacology; D013386:Succinates; D013696:Temperature; D013745:Tetanus Toxoid / Q000008:administration & dosage / Q000737:chemistry* / Q000276:immunology
chemical_list: D003692:Delayed-Action Preparations; D014677:Pharmaceutical Vehicles; D011108:Polymers; D012997:Solvents; D013386:Succinates; D013745:Tetanus Toxoid; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011100:Polyglycolic Acid; D019344:Lactic Acid