A novel TLR4 binding protein, 40S ribosomal protein S3, has potential utility as an adjuvant in a dendritic cell-based vaccine

Hyun Jin Park,Yeong-Min Park,Tae Heung Kang,In Duk Jung,Sung Eun Lee,Manh-Cuong Vo,Young Seob Kim,Jung Hwa Park,Hee Dong Han,Je-Jung Lee,Gun-Young Jang

doi:10.1186/s40425-019-0539-7

Hyun Jin Park, Yeong-Min Park + Show 9 more

Open Access

https://doi.org/10.1186/s40425-019-0539-7

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Abstract

BackgroundDendritic cells (DCs) are professional antigen presenting cells (APCs), which can activate antigen-specific CD8+ T cell immunity, resulting in tumor clearance. Immature DCs are usually stimulated by various adjuvants through their immune receptors. Among them, Toll-like receptor 4 (TLR4) has an important role in activating DCs to cause their maturation. In fact, TLR4 is well-known to induce innate and adaptive immune responses against various external microbial or internal damage associated molecular patterns (DAMP). LPS is widely regarded as a strong stimulator of TLR4 signaling. However, LPS is inappropriate for use in humans since it is an endotoxin. Unfortunately, other TLR4 ligands such as HMGB1 or heat shock proteins have weak adjuvant effects. Therefore, there is a need to identify novel, biocompatible, strong, TLR4 ligands.Methods40S ribosomal protein S3 (RPS3) was screened through pull-down assay using TLR4. BMDCs from wild type (WT) and TLR4 knock-out mice were treated by RPS3 to identify the activation and maturation of DCs. T cell generation including memory T cells, tumor prevention, and treatment experiments were performed with BMDCs based vaccination. Also, human DCs originated from patients were treated by RPS3 to confirm the activation and maturation of DCs.ResultsIn this study, we identified 40S ribosomal protein S3 (RPS3) through a pull-down assay using a variety of human cancer cell-derived proteins that could bind to TLR4. RPS3 was released from tumor cells following treatment with an anticancer drug, and it was shown that the released RPS3 binds to TLR4. Recombinant RPS3 induced maturation and activation of DCs, and following pulsing with tumor specific antigens, these DCs could be used as a vaccine to significantly increase tumor specific CD8+IFN-γ+ T cells, and provide both tumor prevention and tumor treatment effects. The effect of RPS3 on DC maturation and its utility as a vaccine were shown to be dependent on TLR4 using TLR4 knockout mice.ConclusionsThis study therefore proved that human cancer cell-derived RPS3, a novel TLR4 ligand, has great potential as an adjuvant in tumor-specific antigen DC-based vaccines.

Highlights

Dendritic cells (DCs) are professional antigen presenting cells (APCs), which can activate antigenspecific CD8+ T cell immunity, resulting in tumor clearance
Release of ribosomal protein S3 (RPS3) from tumor cells and the binding of recombinant RPS3 from E. coli to recombinant Toll-like receptor 4 (TLR4) To identify protein candidates in human cancer cells that can associate with TLR4, we screened human cancer cells using a luciferase assay and three cancer cell lines were selected in which NF-kB activity could be observed (Additional File 2: Figure S1)
Among the various ribosomal protein families that were found to bind to TLR4, ribosomal protein S3 (RPS3) was selected for use in our experiments because it had the greatest effects when used to treat BMDCs

Summary

Introduction

Dendritic cells (DCs) are professional antigen presenting cells (APCs), which can activate antigenspecific CD8+ T cell immunity, resulting in tumor clearance. TLRs recognize various ligands (e.g. DNA, RNA, proteins), and activate intracellular signaling pathways (e.g., MAPK, NF-кB) and produce pro-inflammatory cytokines in antigen presenting cells (APCs), e.g., dendritic cell (DCs) [5,6,7]. DC-based cancer vaccines have been produced, and of the different cancer vaccines available for use in humans (antigen/adjuvant vaccines, anti-idiotype vaccines, DNA vaccines, tumor cell vaccines) are the most preferred [12,13,14]. In order to generate DC-based cancer vaccines, an adjuvant that binds to TLRs that can induce maturation and activation of DCs is needed. It is well known that LPS, a TLR4 ligand, has a significant effect on inducing maturation and activation of DCs through the MyD88 and TRIF cell signaling pathways [17]. We need to find a new adjuvant that can bind to TLR4 to effectively induce DC maturation

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