Disruption of Telomere Integrity and DNA Repair Machineries by KML001 Induces T Cell Senescence, Apoptosis, and Cellular Dysfunctions

Abstract

T cells in chronic viral infections are featured by premature aging with accelerated telomere erosion, but the mechanisms underlying telomere attrition remain unclear. Here, we employed human CD4 T cells treated with KML001 (a telomere-targeting drug) as a model to investigate the role of telomere integrity in remodeling T cell senescence. We demonstrated that KML001 could inhibit cell proliferation, cytokine production, and promote apoptosis via disrupting telomere integrity and DNA repair machineries. Specifically, KML001-treated T cells increased dysfunctional telomere-induced foci (TIF), DNA damage marker γH2AX, and topoisomerase cleavage complex (TOPcc) accumulation, leading to telomere attrition. Mechanistically, KML001 compromised telomere integrity by inhibiting telomeric repeat binding factor 2 (TRF2), telomerase, topoisomerase I and II alpha (Top1/2a), and ataxia telangiectasia mutated (ATM) kinase activities. Importantly, these KML001-induced telomeric DNA damage and T cell senescent phenotype and machineries recapitulated our findings in patients with clinical HCV or HIV infection in that their T cells were also senescent with short telomeres and thus more vulnerable to KML001-induced apoptosis. These results shed new insights on the T cell aging network that is critical and essential in protecting chromosomal telomeres from unwanted DNA damage and securing T cell survival during cell crisis upon genomic insult.

Inhibition of topoisomerase IIA (Top2α) induces telomeric DNA damage and T cell dysfunction during chronic viral infection

“…These results indicate that ICRF-193-induced, Top2α-mediated topological DDR involves dynamic activation and depletion of ATM. These are in line with our recent findings of ATM dynamics in healthy CD4 T cells treated with KML001 or Top1 inhibitor 8,9 and in CD4 T cells derived from patients with chronic HCV and/or HIV infection [5][6][7] .…”

“…During chronic viral infections, however, T cells are always dysregulated and often non-responsive to vaccines 1 . We and others have previously reported that T cells from chronically virusinfected individuals are prematurely aged due to accelerated telomere erosion [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17] , but the underlying mechanisms for T cell telomeric DNA damage remain unclear. Since Top2α is required to remove DNA supercoiling generated during cell proliferation, and Top2cc can become trapped during gene transcription to cause Top2cc-linked PDB due to TDP2 depletion 21,22 , we hypothesized that DNA topology in T cells may be affected during viral infections to trigger DDR as a mechanism of virus-induced immune evasion, and thus, persistent infection.…”

“…Human telomeres consist of triple guanine repeats (TTAGGG) that are sensitive to DNA damage 34,35 . We hypothesized that Top2α inhibition-mediated genomic DNA damage may affect telomeres as we have previously shown for telomere erosion in T cells derived from virus-infected subjects [5][6][7][8][9] . To determine telomeric DNA damage in Top2 inhibitor-treated T cells, we measured the number of dysfunctional telomereinduced foci (TIF, a hallmark of telomeric DNA damage 36,37 ) by examining the colocalization of 53BP1/ TRF1 (p53-binding protein 1/telomeric repeat-binding factor 1) using confocal microscopy.…”

“…4g), which are consistent with the data above showing Top2α inhibition, Top2cc accumulation, PARP1 induction, and apoptosis in these cells ( Figs. 1-4) [5][6][7][8][9] .…”

“…T cells play a critical role in control of viral infection. In studying the role of T cell dysregulation in viral persistence in humans, we and others have previously shown that chronic viral infections can cause premature T cell aging and immune senescence, as evidenced by the expression of aging markers and particularly, accumulation of DNA damage [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17] . However, the underlying mechanisms remain unclear.…”