Absence of a telomere maintenance mechanism in cases of metastatic Ewing sarcoma
Telomeres are repetitive DNA structures that cap chromosome ends and protect against chromosomal fusions and genomic instability. Telomere maintenance mechanisms (TMMs) are activated during cancer development and underpin cancer cell immortality by counteracting the gradual shortening of telomeres that otherwise occurs with each cell division. Two distinct TMM have been described in human cancers: reverse transcription of telomeric DNA by the enzyme telomerase, and telomere elongation via a recombination-based mechanism known as the Alternative Lengthening of Telomeres (ALT) mechanism. While cellular immortality and telomere maintenance is broadly considered a hallmark of cancer, we and others have shown that advanced stage neuroblastoma and melanoma may develop in the absence of a TMM. Cell lines derived from these cancers exhibited a phenotype referred to as Ever Shorter Telomeres (EST) where telomeres continue to shorten over a large number of population doublings. In the current study we identified cell lines and tumour samples from patients with Ewing sarcoma (EWS) that were also negative in assays for both telomerase and ALT features (TMM-double negative). From analysis of a pan cancer panel of 976 cancer cell lines, using the qTRAP assay to quantify telomerase activity and c-circle assay for detection of ALT, EWS cell lines were found to be disproportionately represented among the cell lines that were negative in both assays (Chi-square test P<0.0001), with 4 out of 22 EWS cell lines TMM-double negative. In long-term culture, 2/4 of the TMM-double negative EWS cell lines exhibited telomere shortening, while the other two cell lines had no substantial net change in telomere length over approximately 60 population doublings. In one case telomere stabilisation was accounted for by upregulation of telomerase activity during serial passage in vitro. Among 23 EWS tumour samples, 9 were found to be TMM-double negative. Notwithstanding the potential for activation of TMM in culture, collectively the results from the EWS cell lines and tumour samples add to growing evidence that telomere maintenance is not a universal requirement for development of aggressive cancers.