Chondrosarcoma is the second most frequent sarcoma occurring primary in bone. More than 80% of total chondrosarcoma (CS) cases are represented by conventional, central chondrosarcoma. 3D “in vitro” models could help to design alternative treatment options for this rare bone cancer that is resistant to conventional chemo- and radiotherapy. In this article the authors created a multicellular tumour spheroid (MCTS) model using the HEMC-SS cell line, which is not representative of CS as it is derived from an extraskeletal myxoid chondrosarcoma (EMC). EMC almost exclusively occurs in the soft tissues, and is classified as a tumour of uncertain differentiation in the WHO 2013 classification instead of a chondro-osseous tumor, as the designation as “chondrosarcoma” is considered a misnomer in the field. Thus, extraskeletal myxoid chondrosarcoma and chondrosarcoma of bone are not related ¹.
Primary central CS arise centrally within the medullar cavity of any bone derived from enchondral ossification and represent 20% of all malignant bone tumours. They are defined by the formation of hyaline cartilage, presence of mucoid matrix as well as cartilaginous markers such as collagen II. The higher grade variants often contain areas of high cell density, myxoid or mucoid material and are much more likely to metastasize ¹. They are considered highly resistant to radio- and chemotherapy. For instance, the 10 year survival for grade III chondrosarcoma is merely 29% ². Mutations in isocitrate dehydrogenase (IDH), an enzyme involved in the tricarboxylic acid cycle, have been found in 38-86% of conventional central chondrosarcomas and this is thus far the only common genetic abnormality detected in these tumours ³.
Extraskeletal myxoid chondrosarcoma differs from conventional chondrosarcoma by site of occurrence, expression of differentiation markers and genetic characteristics. EMC was originally classified as chondrosarcoma due to positivity for S100, resemblance to chondroblasts when using electron microscopy and focal cartilage formation. On the contrary, expression of cartilaginous differentiation markers such as collagen II and aggrecan was found to be absent in 86% of EMCs and S100 expression was very focal or absent ⁴. Well-formed hyaline cartilage was also only found in a minority of EMCs ⁵. Numerous ultrastructural and genetic studies of EMC have uncovered neuroendocrine differentiation markers such as class III B-tubulin and microtubule-associated protein 2, which disputes a chondrocytic origin of this malignancy ^6–8. Due to an overall lack of convincing evidence of a cartilaginous differentiation, EMC is now considered a unique entity of uncertain differentiation ¹. Furthermore, most EMCs contain a characteristic translocational juxtaposition of NR4A3 on chromosome 9 with EWSR1 on chromosome 22, t(9;22) which separates them further from the typical chondrosarcoma genotype ⁹.
In the article, a 3D model was created in order to more sufficiently represent the tumour microenvironment, as it has been suggested that this plays a key role in chemo- and radiotherapeutic resistance in cartilage tumours ². EMC is decidedly resistant to conventional chemotherapy and has a high rate of reported local and distant recurrence ranging from 37 to 48% ¹. Although the model is not representative of CS the authors have created a much needed 3D model of EMC. There is a large requirement for better pre-clinical models in this field ⁹. Overall, the methods chosen to characterise and test the model were well-suited and the article demonstrates a much needed step forward toward the development of 3D cell models of rare tumours.
There are similarities in results expected for CS compared to EMC with regards to the characterisation assays used. For example, a significant increase in proteoglycan quantity would be observed in a myxoid sarcoma, as it is the type of glycosaminoglycans (GAGs) that would differ ¹⁰. Additionally, expression of collagen II by a fraction of these tumours has been previously shown, despite their overall lack of chondrogenic phenotype ⁴. The penetration of doxorubicin into 3D multicellular chondrosarcoma pellets of diameter of around ~1mm has been formerly investigated ¹¹. Conversely, it was determined that Doxorubicin penetrated the larger masses fully, albeit over a longer chemotherapeutic incubation period. Possibly, the resulting difference in penetration could also be due to difference in sarcoma type, as EMC is also strongly resistant to cytotoxic chemotherapy ¹².
There are several recognized chondrosarcoma cell lines available for use. For example, CH2879 (conventional, grade III), OUMS27 (conventional, grade III), JJ012 (conventional, grade II) and SW1353 (conventional, grade II) are well established and some newly derived lines such as CH3573 (conventional, grade II) and L835 (conventional, grade III) could be used ^13–18. There have been recent attempts to model central conventional chondrosarcoma in 3D scaffolds using chondrosarcoma cell lines. Recently, a 3D cell culture model of chondrosarcoma using the CH2879, JJ012 and SW1353 cell lines was characterised and its response to chemotherapeutics was compared to its 2D counterparts as well as xenografts ¹⁹. The SW1353 cell line was also used previously in a collagen-based 3D model in order to observe cell survival post-irradiation ²⁰. A previously mentioned study utilized 3D pellet culture of several chondrosarcoma cell lines in order to observe restoration of chemosensitivity compared to 2D ¹¹. Additionally, Truong and colleagues demonstrated 3D culture of primary cells isolated from fresh chondrosarcoma tissue, where cell suspensions were extracted directly from biopsies and cultured within a collagen scaffold ²¹.
In conclusion, the authors have not presented a model of chondrosarcoma. They did, however, present a well characterised model of extraskeletal myxoid chondrosarcoma, which is also an important tumour to investigate. This model could contribute towards elucidation of the molecular mechanisms contributing to EMC tumourigenesis and chemoresistance.

Ieva Palubeckaite and Judith V.M.G. Bovée, MD, PhD
 
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