Hans Peter Merkle

Tetanus toxoid and synthetic malaria antigen containing poly(lactide)/poly(lactide-co-glycolide) microspheres: importance of polymer degradation and antigen release for immune response

Abstract The importance of in vitro degradation of poly(lactide)/poly(lactide-co-glycolide) (PLA/PLGA) microspheres and of the concomitant in vitro release of a natural and a synthetic antigen for eliciting immune response was studied in mice. A variety of PLAs and PLGAs differing in molecular weight ( M w of 14–130 kDa) and in polymer composition (lactic/glycolic acid ratio of 100:00, 75:25, and 50:50) were examined for their in vitro degradation, which ranged from approximately 4 to 20 weeks. Three specific polymers were then selected for microencapsulation of the two antigens tetanus toxoid (TT) and a weakly immunogenic synthetic branched malaria peptide (P30B2). The in vitro release data showed that antigen delivery correlates fairly well with polymer degradation giving rise to a distinct burst release during the first 24 h and an additional release pulse towards the end of polymer degradation. After single subcutaneous administration in mice, long lasting high antibody titers were obtained with the antigen containing microspheres, as compared to TT adsorbed on alum or to P30B2 in Incomplete Freunds Adjuvant. Moreover, the immune responses induced by microspheres were clearly influenced by the antigen release kinetics, the polymer type and the size of the microspheres. The results demonstrate the immunopotentiating properties of the biodegradable microspheres and their potential to elicit long-lasting immune responses after single administration when tailoring in vitro release characteristics and particle size.

MHC class I- and class II-restricted processing and presentation of microencapsulated antigens

Macrophages were found of having a strong capacity of phagocytosing small size microcapsules (MS) and presenting microencapsulated antigens to either CD4+ and CD8- T cells. The class I-restricted presentation of microencapsulated tetanus toxoid by macrophages requires an intracellular processing which might follow the phagosome-to-cytosol route to enter the classical MHC class I presentation pathway. In contrast, presentation of microencapsulated cytotoxic peptide PbCS252-260 to specific CD8+ T cells has been observed with different APC and is not blocked by cytochalasin D, suggesting that peptide released from MS may directly bind to MHC class I molecules on the cell surface. In the case of MHC class II-restricted T cells, prefixation or treatment of macrophages with chloroquine, brefeldin A and cycloheximide inhibits the presentation of microencapsulated and soluble tetanus toxoid. These findings illustrate the capacity of microencapsulated antigens to enter different presentation pathways and should facilitate the development of subunit vaccines.

Competitive adsorption of serum proteins at microparticles affects phagocytosis by dendritic cells

Serum protein adsorption to the surface of particulate synthetic drug carrier systems has a major influence on their uptake by phagocytes. The influence of alpha2-human serum glycoprotein (alpha2GP) on the phagocytosis of various surface modified microparticles was studied in dendritic cells (DC) and was compared with a potent opsonin, IgG, and a dysopsonin, human serum albumin (HSA). The microparticles were administered to DC before and after the incubation with alpha2GP, IgG and HSA in single, binary or ternary protein systems and in whole blood serum. Phagocytosis of microparticles was vastly affected by the surface character of the microparticles themselves and by the adsorption of the proteins. Poly-L-lysine (PLL)-modified microparticles were under all conditions internalized with highest efficiency which is suggested to be mediated by their positive surface charge. The adsorption of commonly phagocytosis promoting proteins reduced the uptake of PLL-modified particles and is explained by compensation of the positive surface charge by the adsorbed negatively charged proteins. In all other particle types tested, freshly adsorbed alpha2GP was found to exhibit a strong phagocytosis promoting activity which was comparable to that of adsorbed IgG. Interestingly, this opsonic activity was lost already 2 h after adsorption to the particle surface. Protein adsorption from binary and ternary protein systems and from whole blood serum occurred in a competitive manner. Significant inhibition of phagocytosis was observed, even when HSA was combined with strong opsonins such as alpha2GP or IgG or in mixtures of all three proteins, indicating the importance of studying the influence of protein adsorption in protein mixtures.

Membrane modulated dissolution of oral drug delivery systems

A kinetic model analogous to two chemical reactions occurring in series describes the dissolution behavior of tablets and pellets coated with either microporous or semipermeable membranes. Microporous membranes permit free water influx, but moderated core dissolution and dialytically controlled solution efflux. Semipermeable membranes allow osmotically limited water influx, moderated core dissolution, but relatively unimpeded solution efflux through a release orifice. An explanation is offered which illustrates why the systems perform similarly despite major differences in the membrane structure and need for a release orifice. Applications to literature examples and experimental evidence demonstrate the suitability of the model.

Intraoral Peptide Absorption

Among the different sites of administration for proteinaceous drugs, the intraoral route of delivery has recently gained considerable interest. In spite of its apparent disadvantages such as limited absorptive surface and moderate mucosal permeability, this route of administration might be of benefit especially for small- or medium-size peptide drugs. Potential advantages for intraoral delivery are the avoidance of gastrointestinal or liver first-pass effect, the feasibility of locally controlled absorption enhancement, the ease of administration and removal of an administered device, and the sustainment of drug action.