Adoptive Immunotherapy after Haploidentical Stem Cell Transplantation with T Cells Allodepleted by Photodynamic Purging
Authors: Perruccio Katia , Topini Fabiana , Tosti Antonella , Carotti Alessandra , Aloisi Teresa , Aversa Franco , Martelli Massimo F , Velardi Andrea 
Published: April 2020

Abstract
After haploidentical stem cell transplantation, immune recovery is slow due to decaying thymic function and extensive T-cell depletion of the graft which is needed to prevent Graft-versus-Host Disease (GvHD). Consequently, infectious related mortality is about 30–40%. To address this problem, we investigated the efficacy of adoptive immunotherapy after photodynamic purging of alloreactive T cells (ATIR, Kiadis Pharma, Amsterdam, The Netherlands) in preventing GvHD and improving immune reconstitution. The 4,5-dibromorhodamine methyl ester (TH9402) is a photosensitizer structurally similar to rhodamine. When donor T cells are activated with allogeneic cells, they retain TH9402 which becomes highly cytotoxic upon activation with visible light. MLR-activated donor T cells are exposed to a fluorescent-light scanning device to eliminate alloreactive T cells. We designed a protocol which provided 3,260 ± 450 (mean ± SD)-fold allodepletion, full retention of T-regulatory cells, and preservation of pathogen- and leukaemia-specific T-cell responses (against Aspergillus, Candida, Cytomegalovirus (CMV), Adenovirus (ADV), Herpes Simplex Virus (HSV), Varicella Zoster Virus (VZV), Toxoplasma antigens; and against leukaemia cells and leukaemia antigens (WT1 and Ca125)). Optimized protocol conditions are:
MLR cell concentration: 3–5 × 10e6/ml; MLR duration: 24 hours; TH9402 concentration: 5 μM; TH9402 incorporation, as measured by median fluorescence index (MFI): 20,000 – 25,000; energy delivery: 0.1 J/cm2. Here we present the preliminary results of a clinical trial. Escalating doses of photodynamically allodeleted donor T cells, i.e., 1.25 × 10e5/Kg, 2.5 × 10e5/Kg, 5 × 10e5/Kg, 1 × 10e6/Kg and 1.25 × 10e6/Kg, were infused into groups of haploidentical transplant recipients.
Only 1 patient developed grade III aGvHD at the 1 × 10e6/Kg cell dose and responded to immune suppressive treatment. Immune assessment analyses revealed that infusion of cell doses equal or greater than 5 × 10e5/Kg are associated with significant reconstitution of T-cell counts and appearance of pathogen-specific T-cell responses. One month after infusion, CD4+ and CD8+ T cells were 124 ± 54/cmm and 327 ± 42/cmm (versus 11 ± 4/cmm and 8 ± 4/cmm respectively, in patients receiving T-cell doses below 5 × 10e5/Kg, P = 0.0007). Aspergillus, Candida, CMV, ADV, HSV, VZV, Toxoplasma-specific CD4+ and CD8+ T-cell responses had recovered to frequencies within the normal ranges while they were absent in patients who received T cell doses under 5 × 10e5/Kg (P = 0.0002). In conclusion, this study demonstrates the feasibility, safety and preliminary indications of efficacy of adoptive immunotherapy after photodynamic purging of alloreactive T cells in recipients of haploidentical stem cell transplantation. A larger study will evaluate the impact of these T-cell infusions on transplant related mortality and disease free survival.