Clinical grade adjuvants to mature CD141+ DCs for immunotherapy.

Blanca Alegria,Carlos Alfaro

doi:10.31083/j.fbe1401002

Abstract

Stimulation of dendritic cells (DC) is considered critical in cancer immunotherapy. BATF-3-dependent subsets, that express in humans CD141 (BDCA-3), promote CD8 T-cell cross-priming against tumor antigens. Here, we evaluate two clinical-grade stimuli for peripheral blood CD141+ myeloid dendritic cells (mDCs), a rare DC subset that is currently being explored for use in immunotherapy. In contrast to routine evaluation methods, which focus on predefined maturation markers on the surface or factors released from the activated cells, we applied an unbiased transcriptome-based method using both RNA-sequencing (RNA-seq) and microarrays. Specifically, we analyzed the mRNA of CD141+ mDCs from five human donors upon activation with two clinical-grade adjuvants, Hiltonol (poly-ICLC, a TLR3 ligand) and protamine RNA (pRNA, a TLR7/8 ligand), and compared these samples to unstimulated counterparts. Both methods, RNA-seq, and microarray showed that Hiltonol and pRNA lead to almost identical changes in the transcriptome of CD141+ mDCs. A gene ontology (GO) term analysis suggested that these changes were mainly related to activation and maturation pathways, including induction of type I IFN and IL-12 transcription, while pathways related to adverse effects or cell damage were not strongly affected. The combination of both reagents in the DC cultures gave a very similar result as compared to either stimulus alone, suggesting no synergistic effect. Furthermore, our analysis demonstrates that microarray and RNA-seq analysis gave similar conclusions about the activation status of these cells. Importantly, microarray analyses instead of the advantages of RNA sequencing may still be suitable for studying the activation of rare cell types that are minimally represented or in very low frequency in the organism. Together, our results indicate that both stimuli are potent clinical grade adjuvants with comparable effects to mature CD141+ mDCs in short-term cultures to be used in immunotherapy.

Highlights

Dendritic cells (DCs) are professional antigenpresenting cells (APCs) and are found in most tissues of the human body [1]
Our results indicate that both stimuli are potent clinical grade adjuvants with comparable effects to mature CD141+ Myeloid dendritic cells (mDC) in short-term cultures to be used in immunotherapy
Our analyses revealed no strong signs of toxic effects of either stimulus on this DC subset, since no gene ontology (GO) terms and genes related to adverse responses, e.g., the activation of pathways related to nonsense-mediated decay were upregulated or changed upon stimulation [29]

Summary

Introduction

Dendritic cells (DCs) are professional antigenpresenting cells (APCs) and are found in most tissues of the human body [1]. CD141+ mDCs show a high expression of TLR3, have been characterized as high IFN-λ and IL-12 producers, and are highly efficient cross-presenters to CD8 T cells, raising interest in them as a target for DC-based immunotherapies [3,4,5,6,7,8,9,10]. Due to their low frequencies in the human blood, CD141+ mDCs have not been included in clinical trials so far and have not been studied as extensively as the more abundant CD1c+ mDCs and pDCs

Methods

Results

Conclusion