Selective Killing of Activated T Cells by 5-Aminolevulinic Acid Mediated Photodynamic Effect: Potential Improvement of Extracorporeal Photopheresis.

Sagar Darvekar,Mouldy Sioud,Andrius Kleinauskas,Qian Peng,Trond Stokke,Petras Juzenas,Morten Oksvold,Eidi Christensen,Toril Holien

doi:10.3390/cancers12020377

Abstract

Extracorporeal photopheresis (ECP), a modality that exposes isolated leukocytes to the photosensitizer 8-methoxypsoralen (8-MOP) and ultraviolet-A (UV-A) light, is used to treat conditions such as cutaneous T-cell lymphoma and graft-versus-host disease. However, the current procedure of ECP has limited selectivity and efficiency; and produces only partial response in the majority of treated patients. Additionally, the treatment is expensive and time-consuming, so the improvement for this modality is needed. In this study, we used the concept of photodynamic therapy (PDT) with 5-aminolevulinic acid (ALA), a precursor of an endogenously synthesized photosensitizer protoporphyrin IX (PpIX) in combination with blue light to explore the possibility of targeting activated human blood T cells ex vivo. With various T-cell activation protocols, a high ALA-induced PpIX production took place in activated CD3+, CD4+CD25+, and CD8+ T cell populations with their subsequent killing after blue light exposure. By contrast, resting T cells were much less damaged by the treatment. The selective and effective killing effect on the activated cells was also seen after co-cultivating activated and resting T cells. Under our clinically relevant experimental conditions, ALA-PDT killed activated T cells more selectively and efficiently than 8-MOP/UV-A. Monocyte-derived dendritic cells (DCs) were not affected by the treatment. Incubation of ALA-PDT damaged T cells with autologous DCs induced a downregulation of the co-stimulatory molecules CD80/CD86 and also upregulation of interleukin 10 (IL-10) and indoleamine 2,3-dioxygenase expression, two immunosuppressive factors that may account for the generation of tolerogenic DCs. Overall, the data support the potential use of ALA-PDT strategy for improving ECP by selective and effective killing of activated T cells and induction of immune tolerance.

Highlights

Extracorporeal photopheresis (ECP) is a cell-based immuno-modulatory treatment modality that exposes isolated leukocytes ex vivo to the photosensitizer 8-methoxypsoralen (8-MOP) followed by ultraviolet A (UV-A) radiation before infusion back to the patient in vivo
This was consistent with an increase in mitotic cells in the activated peripheral blood mononuclear cells (PBMCs) (Figure 1B)
The CD3+, CD4+, CD8+, and CD25+ T cell subsets in the activated PBMCs demonstrated increased Ki-67 staining, indicating cell proliferation as compared to resting PBMCs (Figure 1C). This was evident in the CD4+ and CD25+ T cell subsets. The sizes of both cells and their mitochondria were increased in the activated PBMCs, as confirmed by fluorescent microscopy with Rhodamine 123 staining (Figure A2)

Summary

Introduction

Extracorporeal photopheresis (ECP) is a cell-based immuno-modulatory treatment modality that exposes isolated leukocytes ex vivo to the photosensitizer 8-methoxypsoralen (8-MOP) followed by ultraviolet A (UV-A) radiation before infusion back to the patient in vivo. ECP was officially approved three decades ago for the treatment of cutaneous T-cell lymphoma (CTCL) [1]. Since this treatment has been applied for a number of T-lymphocyte (T cell) mediated diseases, including graft-versus-host disease (GvHD), organ transplant rejection, and certain autoimmune diseases [2,3,4]. The exact mechanism by which ECP exerts its immune modulatory effects is not fully understood. These tolerogenic DCs can induce regulatory T cells (Tregs) for immune tolerance [6,7,8]

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Discussion

Conclusion