Mechanism of Action of PLX-R18, a Placental-Derived Cellular Therapy for the Treatment of Radiation-Induced Bone Marrow Failure
Authors: Ofir Racheli , Pinzur Lena , Levent Akyüz , Aberman Zami , Gorodetsky Raphael , Volk Hans Dieter 
Published: October 2019

Abstract
Bone marrow (BM) failure occurs in individuals who fail to produce sufficient red blood cells, white blood cells, or platelets. This may be a result of damage to hematopoietic stem cells by a congenital defect or by exposure to a noxious substance or factor.
PLX-R18 are 3D-expanded placenta-derived cells, with biological properties including a profound capacity to protect and regenerate bone marrow. The cells secrete a broad array of cytokines including G-CSF, IL-6, MCP-1, MCP-3 and GRO that contribute to the reconstitution of the hematopoietic and immune systems.
To assess their therapeutic potential, PLX-RAD cells were administered to C3H/HeN male mice intramuscularly, one and five days following 7.7 Gy total body irradiation. Body weight and animal survival were monitored for 3 weeks, then the animals were euthanized for BM and blood analysis. In the PLX-R18-treated group, 10 out of 11 (91%) animals survived, compared to only 4 out of 9 in the vehicle treated group (44%), (P < 0.05). Weight reduction after irradiation was also improved in the PLX-RAD treated mice where the average weight loss was less than 10% compared to almost 20% for vehicle-treated controls. Consistent with the improved survival, cell counts of all the three hematopoietic lineages were significantly increased in the BM and blood of PLX-R18 treated mice as compared to the control animals, and attained close to normal levels. Analysis of plasma in the irradiated treated animals detected the presence of critical, PLX-R18-derived (human) cytokines as well as alterations in the equivalent murine cytokines, suggesting a direct role of PLX-R18 secreted cytokines in animal survival. Interestingly, human cytokines were detected only after irradiation and were not present in the plasma of sham non-irradiated animals treated with PLX-R18. This may imply that the active in vivo cytokine secretion by PLX-R18 is a response to signals from the environment in the irradiated animals. Already by day 4-6 after irradiation, the number of colony forming progenitors in the BM of PLX-R18 treated animals was significantly higher than in control animals suggesting that earlier regeneration of BM induced by PLX-R18 secreted cytokines results in improved blood counts and increased survival. Studies in vitro demonstrated that PLX-R18-derived conditioned medium induced the formation of all colony types in a methyl cellulose colony formation assay as well as a 3.1 fold-increase in the number of migrating cells in a BM migration assay compared to the SDF-1-supplemented positive control. These results, in vivo and in vitro, reveal key clues to the resolution of the underlying mechanism of action.
Administration of the novel cell product PLX-R18 markedly improved survival and recovery of the three hematopoietic blood lineages after radiation induced BM failure indicating potential as a highly effective therapy for general radiation-induced BM damage, or in a nuclear disaster scenario. Further clinical studies are planned.

Disclosures
Ofir: Pluristem ltd: Employment. Pinzur:Pluristem ltd: Employment. Aberman:Pluristem ltd: Employment. Gorodetsky:Pluristem ltd: Consultancy. Volk:Pluristem ltd: Consultancy.