Adaptive responses of oncogenic cells to dysfunctional telomeres

Telomerase activation is a common feature of most advanced human cancers and is postulated to restore genomic stability to a level permissive for cell viability and tumor progression. The RB/p16 INK4a and p53/p19 ARF tumor suppressor pathways are key pathways in governing cellular responses to telomere dysfunction. The Ink4a/Arf mutant background was used to study the adaptive responses of oncogenic cells to telomere dysfunction and to provide direct genetic evidence of a role for both p16 INK4a and p19 ARF in modulating the cellular and tissue phenotypes associated with telomere dysfunction. Transformed late generation mTerc-/-Ink4a/Arf-/- mouse embryonic fibroblast (MEFs) were generated to explore how telomere-based crisis relates to the evolution of cancer genomes and tumor biology. Extensive passage in culture led to genomic instability and complex karyotypes. Despite a high degree of genomic instability, these MEFs retained the capacity to form subcutaneous tumors in immunocompromised mice. However, telomere dysfunction completely abrogated lung metastases following tail-vein injection, whereas mTerc reconstitution alone conferred robust metastatic activity. Serial subcutaneous tumor formation of these late generation mTerc-/-Ink4a/Arf-/- MEFs activated telomerase-independent alternative lengthening of telomeres (ALT). Significantly, ALT did not rescue metastases, indicating in vivo functional differences between ALT and telomerase activation. In contrast to the rescue associated with p53 deficiency, Ink4a/Arf deficiency did not attenuate the degenerative phenotypes elicited by telomere dysfunction in the late generation mTerc-/- mice. Furthermore, in contrast to accelerated cancer onset and increased epithelial cancers in the p53 mutant background, late generation mTerc-/-Ink4a/Arf-/- mutant mice experienced a delayed tumor onset and maintained the stereotypical lymphoma and sarcoma spectrum. Late generation mTerc+/+Ink4a/Arf-/- tissues showed activated p53 and derivative tumor cell lines sustained frequent loss of p53 function, whereas mTerc-/-Ink4a/Arf-/- tumor cell lines retained p53. In addition, the late generation mTerc-/-Ink4a/Arf-/- tumors activated ALT, underscoring the need for adaptation despite the absence of p16 INK4a and p19 ARF . These observations highlight the importance of genetic context in dictating whether telomere dysfunction promotes or suppresses age-related degenerative conditions, the rate of initiation and type of spontaneous cancers. Lastly preliminary genomic analysis of the MEFs and tumors were performed to identify pathways pertinent to the adaptation to telomere dysfunction.