Telomerase and pluripotency factors jointly regulate stemness in pancreatic cancer stem cells

To assess the role of telomerase activity and telomere length in pancreatic CSCs we used different CSC enrichment methods (CD133, ALDH, sphere formation) in primary patient-derived pancreatic cancer cells. We show that CSCs have higher telomerase activity and longer telomeres than bulk tumor cells. Inhibition of telomerase activity, using genetic knockdown or pharmacological inhibitor (BIBR1532), resulted in CSC marker depletion, abrogation of sphere formation in vitro and reduced tumorigenicity in vivo. Furthermore, we identify a positive feedback loop between stemness factors (NANOG, OCT3/4, SOX2, KLF4) and telomerase, which is essential for the self-renewal of CSCs. Disruption of the balance between telomerase activity and stemness factors eliminates CSCs via induction of DNA damage and apoptosis in primary patient-derived pancreatic cancer samples, opening future perspectives to avoid CSC-driven tumor relapse. In the present study, we demonstrate that telomerase regulation is critical for the “stemness” maintenance in pancreatic CSCs and examine the effects of telomerase inhibition as a potential treatment option of pancreatic cancer. This may significantly promote our understanding of PDAC tumor biology and may result in improved treatment for pancreatic cancer patientsThis research was funded by a Max Eder Fellowship of the German Cancer Aid (111746), a German Cancer Aid Priority Program ‘Translational Oncology’ 70112505, by a Collaborative Research Centre grant (316249678—SFB 1279) of the German Research Foundation, and by a Hector Foundation Cancer Research grant (M65.1) to P.C.H., B.S.J. is supported by a Rámon y Cajal Merit Award (RYC- 2012-12104) from the Ministerio de Economía y Competitividad, Spain and a Coordinated grant (GC16173694BARB) from the Fundación Asociación Española Contra el Cáncer (AECC). K.W. is supported by a Baustein 3.2 by Ulm University

Telomerase: The Devil Inside

High telomerase activity is detected in nearly all human cancers but most human cells are devoid of telomerase activity. There is well-documented evidence that reactivation of telomerase occurs during cellular transformation. In humans, tumors can rely in reactivation of telomerase or originate in a telomerase positive stem/progenitor cell, or rely in alternative lengthening of telomeres, a telomerase-independent telomere-length maintenance mechanism. In this review, we will focus on the telomerase positive tumors. In this context, the recent findings that telomerase reverse transcriptase (TERT) promoter mutations represent the most common non-coding mutations in human cancer have flared up the long-standing discussion whether cancer originates from telomerase positive stem cells or telomerase reactivation is a final step in cellular transformation. Here, we will discuss the pros and cons of both concepts in the context of telomere length-dependent and telomere length-independent functions of telomerase. Together, these observations may provoke a re-evaluation of telomere and telomerase based therapies, both in telomerase inhibition for cancer therapy and telomerase activation for tissue regeneration and anti-ageing strategies

IFN-γ treatment protocol for MHC-Ilo/PD-L1+ pancreatic tumor cells selectively restores their TAP-mediated presentation competence and CD8 T-cell priming potential

Background Many cancer cells express a major histocompatibility complex class I low/ programmed cell death 1 ligand 1 positive (MHC-Ilo/PD-L1+) cell surface profile. For immunotherapy, there is, thus, an urgent need to restore presentation competence of cancer cells with defects in MHC-I processing/presentation combined with immune interventions that tackle the tumor-initiated PD-L1/PD-1 signaling axis. Using pancreatic ductal adenocarcinoma cells (PDACCs) as a model, we here explored if (and how) expression/processing of tumor antigens via transporters associated with antigen processing (TAP) affects priming of CD8 T cells in PD-1/PD-L1-competent/-deficient mice.Methods We generated tumor antigen-expressing vectors, immunized TAP-competent/-deficient mice and determined de novo primed CD8 T-cell frequencies by flow cytometry. Similarly, we explored the antigenicity and PD-L1/PD-1 sensitivity of PDACCs versus interferon-γ (IFN-γ)-treated PDACCs in PD-1/PD-L1-competent/deficient mice. The IFN-γ-induced effects on gene and cell surface expression profiles were determined by microarrays and flow cytometry.Results We identified two antigens (cripto-1 and an endogenous leukemia virus-derived gp70) that were expressed in the Endoplasmic Reticulum (ER) of PDACCs and induced CD8 T-cell responses either independent (Cripto-1:Kb/Cr16-24) or dependent (gp70:Kb/p15E) on TAP by DNA immunization. IFN-γ-treatment of PDACCs in vitro upregulated MHC-I- and TAP- but also PD-L1-expression. Mechanistically, PD-L1/PD-1 signaling was superior to the reconstitution of MHC-I presentation competence, as subcutaneously transplanted IFN-γ-treated PDACCs developed tumors in C57BL/6J and PD-L1-/- but not in PD-1-/- mice. Using PDACCs, irradiated at day 3 post-IFN-γ-treatment or PD-L1 knockout PDACCs as vaccines, we could selectively bypass upregulation of PD-L1, preferentially induce TAP-dependent gp70:Kb/p15E-specific CD8 T cells associated with a weakened PD-1+ exhaustion phenotype and reject consecutively injected tumor transplants in C57BL/6J mice.Conclusions The IFN-γ-treatment protocol is attractive for cell-based immunotherapies, because it restores TAP-dependent antigen processing in cancer cells, facilitates priming of TAP-dependent effector CD8 T-cell responses without additional check point inhibitors and could be combined with genetic vaccines that complement priming of TAP-independent CD8 T cells

The promoter of human telomerase reverse transcriptase is activated during liver regeneration and hepatocyte proliferation.

Telomerase activity has not been detected in healthy human liver biopsy samples, but it is up-regulated in most human liver tumors. It is not clear whether telomerase is activated in response to acute or chronic liver injury. Telomerase activity is closely associated with expression of its catalytic subunit, telomerase reverse transcriptase (TERT). We analyzed the activity of the human TERT (hTERT) promoter during liver regeneration in vivo and hepatocyte proliferation in vitro

ATM Deficiency Generating Genomic Instability Sensitizes Pancreatic Ductal Adenocarcinoma Cells to Therapy-Induced DNA Damage

Pancreatic ductal adenocarcinomas (PDAC) harbor recurrent functional mutations of the master DNA damage response kinase ATM, which has been shown to accelerate tumorigenesis and epithelial-mesenchymal transition. To study how ATMdeficiency affects genome integrity in this setting, we evaluated the molecular and functional effects of conditional Atm deletion in a mouse model of PDAC. ATM deficiency was associated with increased mitotic defects, recurrent genomic rearrangements, and deregulated DNA integrity checkpoints, reminiscent of human PDAC. We hypothesized that altered genome integrity might allow synthetic lethality-based options for targeted therapeutic intervention. Supporting this possibility, we found that the PARP inhibitor olaparib or ATR inhibitors reduced the viability of PDAC cells in vitro and in vivo associated with a genotype-selective increase in apoptosis. Overall, our results offered a preclinical mechanistic rationale for the use of PARP and ATR inhibitors to improve treatment of ATM-mutant PDAC. (C) 2017 AACR