title: Enabling a robust scalable manufacturing process for therapeutic exosomes through oncogenic immortalization of human ESC-derived MSCs.
abstract: BACKGROUND
  Exosomes or secreted bi-lipid vesicles from human ESC-derived mesenchymal stem cells (hESC-MSCs) have been shown to reduce myocardial ischemia/reperfusion injury in animal models. However, as hESC-MSCs are not infinitely expansible, large scale production of these exosomes would require replenishment of hESC-MSC through derivation from hESCs and incur recurring costs for testing and validation of each new batch. Our aim was therefore to investigate if MYC immortalization of hESC-MSC would circumvent this constraint without compromising the production of therapeutically efficacious exosomes.
  
  
  METHODS
  The hESC-MSCs were transfected by lentivirus carrying a MYC gene. The transformed cells were analyzed for MYC transgene integration, transcript and protein levels, and surface markers, rate of cell cycling, telomerase activity, karyotype, genome-wide gene expression and differentiation potential. The exosomes were isolated by HPLC fractionation and tested in a mouse model of myocardial ischemia/reperfusion injury, and infarct sizes were further assessed by using Evans' blue dye injection and TTC staining.
  
  
  RESULTS
  MYC-transformed MSCs largely resembled the parental hESC-MSCs with major differences being reduced plastic adherence, faster growth, failure to senesce, increased MYC protein expression, and loss of in vitro adipogenic potential that technically rendered the transformed cells as non-MSCs. Unexpectedly, exosomes from MYC-transformed MSCs were able to reduce relative infarct size in a mouse model of myocardial ischemia/reperfusion injury indicating that the capacity for producing therapeutic exosomes was preserved.
  
  
  CONCLUSIONS
  Our results demonstrated that MYC transformation is a practical strategy in ensuring an infinite supply of cells for the production of exosomes in the milligram range as either therapeutic agents or delivery vehicles. In addition, the increased proliferative rate by MYC transformation reduces the time for cell production and thereby reduces production costs.
authors.affiliation: Institute of Medical Biology, A*STAR, 8A Biomedical Grove, 138648 Singapore.
authors.forename: Tian Sheng
authors.initials: TS
authors.lastname: Chen
authors.forename: Fatih
authors.initials: F
authors.lastname: Arslan
authors.forename: Yijun
authors.initials: Y
authors.lastname: Yin
authors.forename: Soon Sim
authors.initials: SS
authors.lastname: Tan
authors.forename: Ruenn Chai
authors.initials: RC
authors.lastname: Lai
authors.forename: Andre Boon Hwa
authors.initials: AB
authors.lastname: Choo
authors.forename: Jayanthi
authors.initials: J
authors.lastname: Padmanabhan
authors.forename: Chuen Neng
authors.initials: CN
authors.lastname: Lee
authors.forename: Dominique P V
authors.initials: DP
authors.lastname: de Kleijn
authors.forename: Sai Kiang
authors.initials: SK
authors.lastname: Lim
mesh_terms: D000818:Animals; D000954:Antigens, Surface / Q000378:metabolism; D002316:Cardiotonic Agents / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D002461:Cell Line, Transformed; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000473:pathology*; D053595:Embryonic Stem Cells / Q000473:pathology*; D055354:Exosomes / Q000378:metabolism*; D020869:Gene Expression Profiling; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000378:metabolism / Q000473:pathology*; D051379:Mice; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism
chemical_list: D000954:Antigens, Surface; D002316:Cardiotonic Agents; D016271:Proto-Oncogene Proteins c-myc