A novel synthetic adjuvant enhances dendritic cell function

Abstract

The lipid core peptide (LCP) is a novel, synthetic, self-adjuvanted vaccine delivery system that neatly incorporates the adjuvant, carrier and antigenic peptides of a vaccine into a single molecular entity. This system has been previously shown to efficiently deliver vaccines and induce immunity. Because adjuvants target sentinels of the immune response, such as dendritic cells (DCs), that are widely distributed throughout the body to initiate specific immune responses, we investigated the effects of the adjuvant on DCs. Here we show that LCP targets vaccines to DCs and induces their activation.

Figure 1

A schematic representation of the synthetic lipid core peptide construct.

Figure 2

Flow-cytometry profiles of murine splenic dendritic cells (DCs) from lipid core peptide (LCP)-treated mice. (a) Multiple cohorts of mice were administered with either the synthetic compound LCP or phosphate-buffered saline (PBS) and CD11c⁺ DC isolated after 3, 7 and 11 days. DCs were labelled for major histocompatibility complex (MHC) class II, CD80, CD86 and CD11b expression and analysed by flow cytometry. (b) Multiple cohorts of mice were treated with LCP alone, LCP with ovalbumin (OVA) or OVA alone and DC isolated from naïve or treated mice after 7 days. The DCs were labelled for expression of MHC class II and DC subpopulation markers, CD8 and CD4, for flow cytometry analysis. The error bars shown are the standard error of the mean (SEM) and the data shown are an example of one from two experiments with three or four mice in each cohort.

Figure 3

Function of murine splenic dendritic cells (DCs) from lipid core peptide (LCP)-treated mice. Splenic DCs were isolated from mice 7 days after the administration of ovalbumin (OVA), LCP + OVA, LCP alone or phosphate-buffered saline (PBS), and co-cultured for 4 days with unlabelled or carboxyfluorescein succinimidyl ester (CFSE) labelled, naïve CD4⁺ T cells. (a) Examples of flow cytometry profiles showing the proliferation frequency of the CFSE-labelled T cells. (b) Bar chart showing the proliferation frequency of the CFSE-labelled T cells from cohorts of mice, in replicate experiments. The error bars shown are the standard error of the mean (SEM) and the data shown are an example of one from two experiments with three or four mice in each cohort.

Figure 4

Interleukin (IL)-12 secretion by murine splenic dendritic cells (DCs) from lipid core peptide (LCP)-treated mice. Splenic DCs were isolated from mice 7 days after the administration of ovalbumin (OVA), LCP + OVA, LCP alone or phosphate-buffered saline (PBS), and were re-stimulated in vitro with either LPS or CpG. The numbers of DCs secreting IL-12 were then assessed by enzyme-linked immunosorbent spot-forming cell assay (ELISPOT). The error bars shown are the standard error of the mean (SEM) and the data shown are an example of one from two experiments with three or four mice in each cohort.

Figure 5

The lipid core peptide (LCP) construct is able to stimulate Toll-like receptor 2 (TLR2) activity. Cells lines expressing TLR2 were transfected with a plasmid carrying the luciferase gene under control of the nuclear factor (NF)-κB promoter, or mock transfected and then treated with the LCP construct or with a positive control (PC), Pam2Cys. Luceferase activity was then measured to assess TLR2 signalling PC, positive control.