2597: Assessment of engraftment and safety of a new tolerance inducing therapy of human cord blood derived ex-vivo created di-chimeric cells in NOD SCID mouse model

Abstract

2597: Assessment of engraftment and safety of a new tolerance inducing therapy of human cord blood derived ex-vivo created di-chimeric cells in NOD SCID mouse model Joanna Cwykiel, MSc, Natalia Filipek, BSc, Malgorzata Cyran, Can Emre Bas, Erzsebet Szilagyi, Ewa Bryndza Tfaily, Medhat Askar, MD, PhD, and Maria Siemionow University of Illinois at Chicago, Chicago, IL, USA; Cleveland Clinic, Cleveland, OH, USA Background The aim of this study was to evaluate the phenotype, engraftment and safety of ex-vivo fused human cord blood derived di-chimeric cell therapy (DCC) in the NOD SCID mouse model. Methods A total of 36 fusions of human umbilical cord blood (UCB) mononuclear clls were performed UCB from 2 unrelated donors were separately stained with PKH26 and PKH67 dyes Fused with polyethylene glycol, double (PKH26/PKH67) stained DCC were sorted and subjected to the in vitro evaluations (15 fusions): lymphocytotoxicity (LCT) test, PCR-rSSOP, STR-PCR, viability, apoptosis, colony forming unit (CFU) assay and COMET assay DCC (3–5 £ 10 cells) from 18 fusions (n D 6/Group) were delivered: Group 1: intraosseous, Group 2: intramuscular, and Group 3: subcutaneous to the NOD SCID recipient mice Control mice in Groups 4, 5, and 6 (n D 6) received 3 £ 10UCB utilizing the same 3 delivery sites Mice were observed daily for 90 days for changes in weight, activity, posture and hair loss Moreover, mice were evaluated bi-weekly by palpation and at 90 days by magnetic resonance imaging (MRI) The presence of DCC in the blood was determined by complete blood count (CBC) and flow cytometry (FC) The migratory pathways of DCC, peripheral blood, bone marrow and lymphoid organs were assessed at 3 months after delivery using immunofluorescent staining Furthermore, H&E staining was performed to assess harvested tissues for tumor growth. Results The presence of HLA class I and II from both UCB donors was confirmed by LCT, PCR-rSSOP, and STR COMET assay showed no damage to the DNA of DCC following fusion procedure Migratory properties of DCC were confirmed at 24 hours after cell delivery Human derived cells (CD45C, CD19C, HLA class I and CD4C) were detected at a level up to 2% in the peripheral blood as tested by CBC and flow cytometry at 90 days following delivery No DCC derived tumorlike growth was observed. Conclusions Phenotype, engraftment and safety of DCC were characterized The unique concept of supportive therapy of DCC introducing cells presenting phenotype characteristics of both transplant donor and recipient is a new promising approach for tolerance induction and prolonging VCA survival. CONTACT Joanna Cwykiel, MSc jcwykiel@uic.edu © 2016 Joanna Cwykiel, Natalia Filipek, Malgorzata Cyran, Can Emre Bas, Erzsebet Szilagyi, Ewa Bryndza Tfaily, Medhat Askar, and Maria Siemionow. Published with license by Taylor & Francis. This is an Open Access article distributed under the terms of the Creative Commons Attribution-Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. The moral rights of the named author(s) have been asserted. VASCULARIZED COMPOSITE ALLOTRANSPLANTATION 2016, VOL. 3, NOS. 1–2, 56 http://dx.doi.org/10.1080/23723505.2016.1234262