Detection of alternative lengthening of telomeres mechanism on tumor sections

Abstract

The vast majority of adult cancer cells achieve cellular immortality by activating a telomere maintenance mechanism (TMM). While this is mostly achieved by the de-silencing of hTERT telomerase gene expression, an alternative homologous recombination-based and telomerase-independent mechanism, known as ALT (Alternative Lengthening of Telomeres), is frequently activated in a subset of tumors, including paediatric cancers. Being absent from normal cells, the ALT mechanism offers interesting perspectives for new targeted cancer therapies. To date, however, the development of better translationally applicable tools for ALT detection in tumor sections is still needed. Here, using a newly derived ALT-positive cancer cell mouse xenograft model, we extensively examined how the previously known ALT markers could be used as reliable tools for ALT diagnosis in tumor sections. We found that, together with the detection of ultra-bright telomeric signals (UBS), an ALT hallmark, native telomeric FISH, that detects single-stranded C-rich telomeric DNA, provides a very sensitive and robust tool for ALT diagnosis in tissues. We applied these assays to paediatric tumor samples and readily identified three ALT-positive tumors for which the TMM was confirmed by the gold-standard C-circle amplification assay. Although the latter offers a robust assay for ALT detection in the context of research laboratories, it is more difficult to set up in histopathological laboratories and could therefore be conveniently replaced by the combination of UBS detection and native telomeric FISH.

Orphan nuclear receptors-induced ALT-associated PML bodies are targets for ALT inhibition

“…In addition, we examined whether ALT activity is abolished upon As 2 O 3 administration. We utilized the native FISH assay to detect single-stranded C-rich telomeric DNA (ssTeloC), such as C-circles, which are features of ALT activity ( 19 , 54 , 55 ). Our findings demonstrate that As 2 O 3 treatment led to a reduction of ssTeloC in SaOS2 xenografts (Figure 7D, E ), suggesting a loss of ALT activity.…”

ALT-FISH quantifies alternative lengthening of telomeres activity by imaging of single-stranded repeats

“…The application of ALT-FISH to tumor tissue provides a new approach to resolve intra- and inter-tumor heterogeneity in cancer entities with high ALT prevalence as for example pediatric glioblastoma, neuroblastoma, leiomyosarcoma and pancreatic neuroendocrine tumors ( 23 , 24 , 50 ). The usefulness of native telomere FISH against C-rich repeats for assessing the ALT status was very recently also shown for mouse xenograft models and FFPE material from other tumor entities ( 51 ). Our results from leiomyosarcoma and neuroblastoma demonstrate the use of ALT-FISH for resolving TMM heterogeneity in a spatial context with high sensitivity.…”