Screening and improving the safe provision of mesenchymal stem cells in regenerative medicine: an in vitro study

Abstract

A number of studies have suggested that mesenchymal stem cells (MSCs) may undergo genetic alterations and spontaneous malignant transformation to form tumour cells, or at least can become contaminated with other cell types following extended periods in culture. Possible transformation or contamination of MSCs during cell culture expansion prior to their use in transplantation therapies is a risk, which should be taken into account. There is a continued need for the development of improved tools for monitoring safety and release criteria of cells intended for cell-based therapies. In order to help address these risks, this thesis aimed to develop improved safety measures in MSC-based cell therapies. Initially, this was investigated through the use of microscopic imaging and image analysis platforms to screen, characterise and distinguish between cultures of non-transformed MSCs and MSC-derived tumour cells, i.e. the osteosarcoma cell lines, SAOS2 and MG63, as well as cells derived from a chondrosarcoma. High content screening (HCS) and live-cell imaging and analysis platforms were used to enable these experiments. Phenotypic features that distinguished the normal versus malignant cell types were identified, including immunoreactivity for the proliferation-associated Ki67 antigen and pluripotency marker Oct4, as well as significant differences in nuclear morphology. These findings help inform release criteria for therapeutic MSCs. To further potentially improve the safety of MSC-based therapies, research was also performed to address the possibility that tumour cells in MSC cultures might remain undetected, thereby still providing a risk in MSC transplantations. Novel combinatorial regimes of potential anti-tumour drugs, i.e. bezafibrate, medroxyprogesterone, and valproic acid (termed V-BAP) were tested in vitro for their effects on MSCs versus SAOS2 and MG63 cells. At determined concentrations, these drugs were shown to significantly inhibit the growth of the osteosarcoma cells, but had little effect on MSCs. Thus, this thesis has made inroads into improved safety of MSC-based therapies by (i) demonstrating the application of imaging-based cell screening platforms to help characterise MSC cultures intended for cell transplantation, and (ii) identifying a novel drug regime that selectively targets osteosarcoma cells whilst having little effect on MSCs. The findings on V-BAP also have application in anti-tumour treatments for osteosarcoma.