Abstract
BackgroundIn osteosarcoma (OS) and most Ewing sarcoma (EWS) patients, the primary tumor originates in the bone. Although tumor resection surgery is commonly used to treat these diseases, it frequently leaves massive bone defects that are particularly difficult to be treated. Due to the therapeutic potential of mesenchymal stem cells (MSCs), OS and EWS patients could benefit from an autologous MSCs-based bone reconstruction. However, safety concerns regarding the in vitro expansion of bone marrow-derived MSCs have been raised. To investigate the possible oncogenic potential of MSCs from OS or EWS patients (MSC-SAR) after expansion, this study focused on a biosafety assessment of MSC-SAR obtained after short- and long-term cultivation compared with MSCs from healthy donors (MSC-CTRL).MethodsWe initially characterized the morphology, immunophenotype, and differentiation multipotency of isolated MSC-SAR. MSC-SAR and MSC-CTRL were subsequently expanded under identical culture conditions. Cells at the early (P3/P4) and late (P10) passages were collected for the in vitro analyses including: sequencing of genes frequently mutated in OS and EWS, evaluation of telomerase activity, assessment of the gene expression profile and activity of major cancer pathways, cytogenetic analysis on synchronous MSCs, and molecular karyotyping using a comparative genomic hybridization (CGH) array.ResultsMSC-SAR displayed comparable morphology, immunophenotype, proliferation rate, differentiation potential, and telomerase activity to MSC-CTRL. Both cell types displayed signs of senescence in the late stages of culture with no relevant changes in cancer gene expression. However, cytogenetic analysis detected chromosomal anomalies in the early and late stages of MSC-SAR and MSC-CTRL after culture.ConclusionsOur results demonstrated that the in vitro expansion of MSCs does not influence or favor malignant transformation since MSC-SAR were not more prone than MSC-CTRL to deleterious changes during culture. However, the presence of chromosomal aberrations supports rigorous phenotypic, functional and genetic evaluation of the biosafety of MSCs, which is important for clinical applications.
Highlights
In osteosarcoma (OS) and most Ewing sarcoma (EWS) patients, the primary tumor originates in the bone
Characterization of mesenchymal stem cells (MSCs) To characterize MSCs isolated from the bone marrow (BM) of sarcoma patients and controls, we evaluated cell morphology, expression of typical surface markers and differentiation potential into mesodermal lineage cells
Despite the considerable variability in proliferation kinetics, cumulative population doubling (CPD) analysis revealed that the MSCs from BM sarcoma patients (MSC-SAR) proliferation rate from P3 to P10 was comparable to MSC-CTRL (Figure 2)
Summary
In osteosarcoma (OS) and most Ewing sarcoma (EWS) patients, the primary tumor originates in the bone. Bone sarcomas are classified genetically into two categories: osteosarcoma (OS) is characterized by complex karyotypes indicative of severe genetic and chromosomal instability [2], while Ewing’s sarcoma (EWS) is characterized by the presence of tumor-specific translocations. The most common mutation associated with EWS involves a translocation of chromosomes 22 and 11 (t (11;22)), which fuses a portion of the EWSR1 gene with a portion of the FLI1 gene to create a EWS/FLI-1 fusion. This is a non-inheritable somatic mutation acquired only in tumor cells during a person’s lifetime [5,6]
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