The Role of Shelterin Complex and Post-Translational Non-Enzymatic Modification in Telomere Maintenance in Chronic Myeloid Leukemia

Abstract The introduction of tyrosine kinase inhibitors (TKI) revolutionized therapy of chronic myeloid leukemia (CML) and gives long-lasting remission in majority of patients. However, a significant proportion of patients develops resistance and are at risk of progression. Genomic instability is one of the key features associated with CML progression and as in majority of human malignancies, reduction in telomere length characterize transition from chronic phase (CML-CP) to the blastic phase (CML-BP). Telomere maintenance depends on telomerase nucleoprotein complex consisting of reverse transcriptase TERT, RNA template TERC and structural proteins, such as dyskerin (DKC1), but also on telomere-associated proteins including but not limited to shelterin complex(TRF1, TRF2, RAP1, TIN2, TPP1, POT1) and other telomeric-associated proteins (such as TEP1 and Tankyrase). The role of shelterin complex and its post-translational non-enzymatic modifications in BCR/ABL1-mediated genomic instability in CML progression and resistance to TKIs is not fully elucidated, thus we decided to examine the possible role of different components of shelterin complex and its glycation and carbonylation in promoting genomic instability and CML progression. We employed CML CD34+ primary cells isolated from peripheral blood leukocytes or bone marrow of CML patients at different stages of disease: CML-CP, CML-BP and CML-TKIres (TKI-resistant cells). Blood samples were taken after informed consent. The length of telomeres was determined by Southern blotting and PNA-FISH technique. Enzymatic activity of telomerase, as well as protein glycation was measured immunoenzymatically, while expression of subunits of telomerase and shelterin complexes was examined by RT-qPCR and by Western Blot. We confirmed that reduction in telomere length was positively correlated with CML progression. However, in samples from CD34+ CML-TKIres patients in comparison to CML-CP patients, an increase in telomere length was observed (Fig 1). In addition, the negative correlation between mean telomere length and expression of BCR-ABL1 in CML samples was observed. This suggests involvement of alternative telomere lengthening mechanisms, since dynamic changes in telomere length were neither associated with enzymatic activity of telomerase nor with gene copy number/expression of TERT/TERC. Therefore we postulate that telomere dynamics in CML may have a bi-phasic scenario. We found that the expression of RAP1, DKC1 and Tankyrase1 was increased in CML-BP as compared to CML-CP. Moreover, RAP1 is involved in increased metabolism, which is associated with elevated glycolytic and oxidative rates leading to post-translational non-enzymatic modification of proteins and damage of cellular proteostasis which may indirectly influence telomere length in CML. We found that telomere shortening was associated with accumulation of post-translational non-enzymatic modification ofselected proteins of shelterin complex, namely glycation and carbonylation. In conclusion, we postulate that aberrant expression of members of the shelterin complex such as RAP1 may be responsible for aberrant telomere maintenance mechanisms in CML cells and may play role in genomic instability associated with CML progression as well as the clonal selection and resistance to TKIs. In addition, the changes in the level of protein glycation may influence telomere length in CML. Disclosures No relevant conflicts of interest to declare.