Elsevier

Vaccine

Volume 18, Issue 23, 22 May 2000, Pages 2482-2493
Vaccine

In vivo antibody response and in vitro CTL activation induced by selected measles vaccine candidates, prepared with purified Quil A components

https://doi.org/10.1016/S0264-410X(00)00026-8Get rights and content

Abstract

Semipurified Quil A and purified Quil A were used to prepare well-characterized subunit vaccine candidates against measles. Variation in the relative amounts of the measles virus (MV) fusion (F) protein, Quil A-components and lipids did not influence induction of antibody responses in mice, but had a pronounced effect on the capacity to induce cytotoxic T cell (CTL) activity of a CD8+ MV F-protein specific human T cell clone in vitro. A characteristic MV iscom preparation based on the combined use of HPLC-purified Quil A-components QA-3 and QA-22 (QA-3/22) efficiently induced CTL activity in vitro. Comparable results were obtained by mixing β-propiolactone inactivated MV with iscom-matrix QA-3/22 or free QA-22. On the basis of the data presented it was concluded that these three preparations are interesting MV vaccine candidates for further evaluation in pre-clinical experiments in a primate model.

Introduction

Measles is still a major cause of serious disease and mortality in developing countries, annually causing the deaths of more than 1 million children [1]. The limited ability of the currently used live attenuated measles vaccines to induce protective immunity in the presence of maternally derived measles virus (MV) neutralizing antibodies is considered a major stumbling-block in the control of measles in developing countries [2], [3], [4], [5], [6]. Therefore, the development of a vaccine that induces protective immunity in the presence of maternally derived MV specific antibodies, would be a major step forward for the eradication of measles, as is currently envisaged by the World Health Organization (WHO).

The specific immune response after MV infection against most of the MV proteins has been the subject of many studies in humans, non-human primates, and rodent models [7]. Still, the individual contribution to protective immunity of the MV-specific humoral and cell mediated immune responses, against each of these proteins, is largely elusive. However, the combined induction of long lasting virus neutralizing (VN) antibodies and CD4+ as well as CD8+ memory T cells warrants solid protection against measles [8].

The exploitation of new generations of vaccines and vaccination strategies may contribute to the successful control and eventual eradication of measles, especially in very young infants with maternal derived MV-specific VN antibodies. Among the currently considered candidate vaccines, recombinant pox-viruses [9], [10], recombinant plasmid DNA [11], [12], [13] and immune-stimulating complex (iscom) based vaccines [9] have attracted major attention. The iscom formulation proved to be of particular interest since it has been shown to induce both strong VN antibody and cell-mediated immune responses. In the presence of pre-existing VN antibodies, adequate protection could be induced with a MV candidate iscom vaccine in a macaque model [9]. The latter studies have all been carried out with iscoms prepared with semipurified Quil A using ‘classical’ centrifugation methods [14]. However, iscom based measles vaccines may only be considered for use in infants, if well-defined non-toxic purified Quil A components would be used. Crude Quil A, as was initially used for iscom preparation has been extensively analyzed by semi-preparative reversed phase high-pressure liquid chromatography (HPLC). Some purified fractions have been characterized in terms of sugar composition, and hemolytic and adjuvant activity [15], [16], [17].

Here we present a study in which we have analyzed five purified Quil A fractions (QA-3; QA-17; QA-20; QA-22 and QA-23) for their capacity to — either alone or in combination — allow the formation of iscoms using various preparation methods. The resulting iscom preparations were characterized with regard to the MV fusion (F)- and hemagglutinin (H) protein, Quil A components and lipid content, and antigenic properties. In addition, we compared the immunogenic properties (antibody and T cell responses) of these preparations in vitro and in vivo.

Section snippets

Virus

Vero cells grown in a 40 l bioreactor on microcarriers (Cytodex 1, Pharmacia, Uppsala, Sweden) were infected with plaque-purified MV (Edmonston strain) at a multiplicity of infection (m.o.i.) of 0.2. After 64 h, culture supernatant was harvested and concentrated using a 100 Kd 0.92 m2 cross-flow filter (Centracette, Filtron Technology Corporation, Massachusetts, USA). Further purification and concentration of virus were achieved by ultra-centrifugation (SW28 rotor 50′ 18.000 rpm) through a 20%

Iscom formation with MV components

Using semi-purified Quil A and purified MV, both the use of D. and C. yielded characteristic iscom-like structures which could not be distinguished morphologically. The MV–iscoms D./QA-3/22 (ratio 1:4) are shown in Fig. 1 as an example. The mean particle size found for the characteristic iscom-like structures was 47±5.8 nm (ranging from 39 to 56 nm) when measured by EM and DLS, which is within the range previously published for these structures [14]. We subsequently studied whether the five

Discussion

In the present paper we have shown that selected Quil A based measles vaccine candidates induced stronger and qualitatively different MV-specific antibody responses in mice than BPL-inactivated MV without Quil A-components. Furthermore, we showed that only preparations containing Quil A components induced in vitro cytolytic activity of a F-protein specific CTL clone.

A series of HPLC-purified Quil A components was used to study their individual contribution to the formation of the characteristic

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