Homologous recombination (HR), a mechanism to accurately repair DNA in normal cells, is deregulated in cancer. Elevated/deregulated HR is implicated in genomic instability and telomere maintenance, which are critical lifelines of cancer cells. We have previously shown that HR activity is elevated and significantly contributes to genomic instability in Barrett’s esophageal adenocarcinoma (BAC). The purpose of this study was to evaluate therapeutic potential of HR inhibition, alone and in combination with telomerase inhibition, in BAC. We demonstrate that telomerase inhibition in BAC cells increases HR activity, RAD51 expression, and association of RAD51 to telomeres. Suppression of HR leads to shorter telomeres as well as markedly reduced genomic instability in BAC cells over time. Combination of HR suppression (whether transgenic or chemical) with telomerase inhibition, causes a significant increase in telomere attrition and apoptotic death in all BAC cell lines tested, relative to either treatment alone. A subset of treated cells also stain positive for β-galactosidase, indicating senescence. The combined treatment is also associated with decline in S-phase and a strong G2/M arrest, indicating massive telomere attrition. In a subcutaneous tumor model, the combined treatment resulted in the smallest tumors, which were even smaller (P=0.001) than those that resulted from either treatment alone. Even the tumors removed from these mice had significantly reduced telomeres and evidence of apoptosis. We therefore conclude that although telomeres are elongated by telomerase, elevated RAD51/HR assist in their maintenance/stabilization in BAC cells. Telomerase inhibitor prevents telomere elongation but induces RAD51/HR, which contributes to telomere maintenance/stabilization and prevention of apoptosis, reducing the efficacy of treatment. Combining HR inhibition with telomerase renders telomeres more vulnerable to degradation and significantly increases/expedites their attrition, leading to apoptosis. We therefore demonstrate that a therapy targeting HR and telomerase has the potential to prevent both tumor growth and genomic evolution in BAC.
This is a preview of subscription content
Subscribe to Journal
Get full journal access for 1 year
only $2.38 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Get time limited or full article access on ReadCube.
All prices are NET prices.
Barrett’s esophageal adenocarcinoma
Allshire RC, Gosden JR, Cross SH, Cranston G, Rout D, Sugawara N et al. Telomeric repeat from T. thermophila cross hybridizes with human telomeres. Nature 1988; 332: 656–659.
de Lange T, Shiue L, Myers RM, Cox DR, Naylor SL, Killery AM et al. Structure and variability of human chromosome ends. Mol Cell Biol 1990; 10: 518–527.
Moyzis RK, Buckingham JM, Cram LS, Dani M, Deaven LL, Jones MD et al. A highly conserved repetitive DNA sequence, (TTAGGG)n, present at the telomeres of human chromosomes. Proc Natl Acad Sci USA 1988; 85: 6622–6626.
Griffith JD, Comeau L, Rosenfield S, Stansel RM, Bianchi A, Moss H et al. Mammalian telomeres end in a large duplex loop. Cell 1999; 97: 503–514.
van Steensel B, Smogorzewska A, de Lange T . TRF2 protects human telomeres from end-to-end fusions. Cell 1998; 92: 401–413.
Day JP, Marder BA, Morgan WF . Telomeres and their possible role in chromosome stabilization. Environ Mol Mutagen 1993; 22: 245–249.
Blasco MA, Lee HW, Hande MP, Samper E, Lansdorp PM, DePinho RA et al. Telomere shortening and tumor formation by mouse cells lacking telomerase RNA [see comments]. Cell 1997; 91: 25–34.
Collins K . Mammalian telomeres and telomerase. Curr Opin Cell Biol 2000; 12: 378–383.
Ducray C, Pommier JP, Martins L, Boussin FD, Sabatier L . Telomere dynamics, end-to-end fusions and telomerase activation during the human fibroblast immortalization process. Oncogene 1999; 18: 4211–4223.
McEachern MJ, Krauskopf A, Blackburn EH . Telomeres and their control. Annu Rev Genet 2000; 34: 331–358.
Shammas MA, Rao MY . Purification of diseased cells from Barrett's esophagus and related lesions by laser capture microdissection. Methods Mol Biol 2011; 755: 181–187.
Harley CB, Futcher AB, Greider CW . Telomeres shorten during ageing of human fibroblasts. Nature 1990; 345: 458–460.
Shammas MA, Koley H, Beer DG, Li C, Goyal RK, Munshi NC . Growth arrest, apoptosis, and telomere shortening of Barrett's-associated adenocarcinoma cells by a telomerase inhibitor. Gastroenterology 2004a; 126: 1337–1346.
Shammas MA, Liu X, Gavory G, Raney KD, Balasubramanian S, Shmookler Reis RJ . Targeting the single-strand G-rich overhang of telomeres with PNA inhibits cell growth and induces apoptosis of human immortal cells. Exp Cell Res 2004b; 295: 204–214.
Shammas MA, Koley H, Batchu RB, Bertheau RC, Protopopov A, Munshi NC et al. Telomerase inhibition by siRNA causes senescence and apoptosis in Barrett's adenocarcinoma cells: mechanism and therapeutic potential. Mol Cancer 2005; 4: 24.
Shammas MA, Koley H, Bertheau RC, Neri P, Fulciniti M, Tassone P et al. Telomerase inhibitor GRN163L inhibits myeloma cell growth in vitro and in vivo. Leukemia 2008a; 22: 1410–1418.
Shammas MA, Qazi A, Batchu RB, Bertheau RC, Wong JY, Rao MY et al. Telomere maintenance in laser capture microdissection-purified Barrett’s adenocarcinoma cells and effect of telomerase inhibition in vivo. Clin Cancer Res 2008b; 14: 4971–4980.
Harley CB . Telomere loss: mitotic clock or genetic time bomb? Mutat Res 1991; 256: 271–282.
Blackburn EH, Greider CW, Henderson E, Lee MS, Shampay J, Shippen-Lentz D . Recognition and elongation of telomeres by telomerase. Genome 1989; 31: 553–560.
Shay JW, Wright WE . The reactivation of telomerase activity in cancer progression. Trends Genet 1996; 12: 129–131.
Shay JW . Telomerase in human development and cancer. J Cell Physiol 1997; 173: 266–270.
Shay JW, Bacchetti S . A survey of telomerase activity in human cancer. Eur J Cancer 1997; 33: 787–791.
Engelhardt M, Martens UM . The implication of telomerase activity and telomere stability for replicative aging and cellular immortality (Review). Oncol Rep 1998; 5: 1043–1052.
Wai LK . Telomeres, telomerase, and tumorigenesis—a review. Med Gen Med 2004; 6: 19.
Counter CM, Avilion AA, LeFeuvre CE, Stewart NG, Greider CW, Harley CB et al. Telomere shortening associated with chromosome instability is arrested in immortal cells which express telomerase activity. Embo J 1992; 11: 1921–1929.
Kim NW, Piatyszek MA, Prowse KR, Harley CB, West MD, Ho PL et al. Specific association of human telomerase activity with immortal cells and cancer [see comments]. Science 1994; 266: 2011–2015.
Lord RV, Salonga D, Danenberg KD, Peters JH, DeMeester TR, Park JM et al. Telomerase reverse transcriptase expression is increased early in the Barrett's metaplasia, dysplasia, adenocarcinoma sequence. J Gastrointest Surg 2000; 4: 135–142.
Bechter OE, Eisterer W, Pall G, Hilbe W, Kuhr T, Thaler J . Telomere length and telomerase activity predict survival in patients with B cell chronic lymphocytic leukemia. Cancer Res 1998; 58: 4918–4922.
Shammas MA, Shmookler Reis RJ, Li C, Koley H, Hurley LH, Anderson KC et al. Telomerase inhibition and cell growth arrest following telomestatin treatment in multiple myeloma. Clin Cancer Res 2003; 10: 770–776.
Fu W, Begley JG, Killen MW, Mattson MP . Anti-apoptotic role of telomerase in pheochromocytoma cells. J Biol Chem 1999; 274: 7264–7271.
Nakajima A, Tauchi T, Sashida G, Sumi M, Abe K, Yamamoto K et al. Telomerase inhibition enhances apoptosis in human acute leukemia cells: possibility of antitelomerase therapy. Leukemia 2003; 17: 560–567.
Seimiya H, Oh-hara T, Suzuki T, Naasani I, Shimazaki T, Tsuchiya K et al. Telomere shortening and growth inhibition of human cancer cells by novel synthetic telomerase inhibitors MST-312, MST-295, and MST-1991. Mol Cancer Ther 2002; 1: 657–665.
Sumi M, Tauchi T, Sashida G, Nakajima A, Gotoh A, Shin-Ya K et al. A G-quadruplex-interactive agent, telomestatin (SOT-095), induces telomere shortening with apoptosis and enhances chemosensitivity in acute myeloid leukemia. Int J Oncol 2004; 24: 1481–1487.
Tauchi T, Shin-Ya K, Sashida G, Sumi M, Nakajima A, Shimamoto T et al. Activity of a novel G-quadruplex-interactive telomerase inhibitor, telomestatin (SOT-095), against human leukemia cells: involvement of ATM-dependent DNA damage response pathways. Oncogene 2003; 22: 5338–5347.
Shammas MA, Reis RJ, Li C, Koley H, Hurley LH, Anderson KC et al. Telomerase inhibition and cell growth arrest after telomestatin treatment in multiple myeloma. Clin Cancer Res 2004c; 10: 770–776.
Akiyama M, Hideshima T, Shammas MA, Hayashi T, Hamasaki M, Tai YT et al. Effects of oligonucleotide N3'—>P5' thio-phosphoramidate (GRN163) targeting telomerase RNA in human multiple myeloma cells. Cancer Res 2003; 63: 6187–6194.
Shammas MA, Simmons CG, Corey DR, Reis RJ . Telomerase inhibition by peptide nucleic acids reverses ‘immortality’ of transformed human cells. Oncogene 1999; 18: 6191–6200.
Bryan TM, Englezou A, Gupta J, Bacchetti S, Reddel RR . Telomere elongation in immortal human cells without detectable telomerase activity. Embo J 1995; 14: 4240–4248.
Opitz OG, Suliman Y, Hahn WC, Harada H, Blum HE, Rustgi AK . Cyclin D1 overexpression and p53 inactivation immortalize primary oral keratinocytes by a telomerase-independent mechanism. J Clin Invest 2001; 108: 725–732.
Dunham MA, Neumann AA, Fasching CL, Reddel RR . Telomere maintenance by recombination in human cells.[see comment]. Nat Genet 2000a; 26: 447–450.
Cerone MA, Londono-Vallejo JA, Bacchetti S . Telomere maintenance by telomerase and by recombination can coexist in human cells. Hum Mol Genet 2001; 10: 1945–1952.
Pal J, Bertheau R, Buon L, Qazi A, Batchu RB, Bandyopadhyay S et al. Genomic evolution in Barrett's adenocarcinoma cells: critical roles of elevated hsRAD51, homologous recombination and Alu sequences in the genome. Oncogene 2011; 30: 3585–3598.
Shammas MA, Shmookler Reis RJ, Koley H, Batchu RB, Li C, Munshi NC . Dysfunctional homologous recombination mediates genomic instability and progression in myeloma. Blood 2009; 113: 2290–2297.
Pierce AJ, Johnson RD, Thompson LH, Jasin M . XRCC3 promotes homology-directed repair of DNA damage in mammalian cells. Genes Dev 1999; 13: 2633–2638.
Munshi NC, Hideshima T, Carrasco D, Shammas M, Auclair D, Davies F et al. Identification of genes modulated in multiple myeloma using genetically identical twin samples. Blood 2004; 103: 1799–1806.
Shammas MA, Shmookler Reis RJ, Akiyama M, Koley H, Chauhan D, Hideshima T et al. Telomerase inhibition and cell growth arrest by G-quadruplex interactive agent in multiple myeloma. Mol Cancer Ther 2003; 2: 825–833.
Ganapathipillai SS, Medova M, Aebersold DM, Manley PW, Berthou S, Streit B et al. Coupling of mutated Met variants to DNA repair via Abl and Rad51. Cancer Res 2008; 68: 5769–5777.
Wallweber G, Gryaznov S, Pongracz K, Pruzan R . Interaction of human telomerase with its primer substrate. Biochemistry 2003; 42: 589–600.
Dunham MA, Neumann AA, Fasching CL, Reddel RR . Telomere maintenance by recombination in human cells. Nat Genet 2000b; 26: 447–450.
Henson JD, Neumann AA, Yeager TR, Reddel RR . Alternative lengthening of telomeres in mammalian cells. Oncogene 2002; 21: 598–610.
Morrish TA, Greider CW . Short telomeres initiate telomere recombination in primary and tumor cells. PLoS Genet 2009; 5: e1000357.
IJpma AS, Greider CW . Short telomeres induce a DNA damage response in Saccharomyces cerevisiae. Mol Biol Cell 2003; 14: 987–1001.
Badie S, Escandell JM, Bouwman P, Carlos AR, Thanasoula M, Gallardo MM et al. BRCA2 acts as a RAD51 loader to facilitate telomere replication and capping. Nat Struct Mol Biol 2010; 17: 1461–1469.
Abdallah P, Luciano P, Runge KW, Lisby M, Geli V, Gilson E et al. A two-step model for senescence triggered by a single critically short telomere. Nat Cell Biol 2009; 11: 988–993.
Ogunwobi OO, Beales IL . Statins inhibit proliferation and induce apoptosis in Barrett’s esophageal adenocarcinoma cells. Am J Gastroenterol 2008; 103: 825–837.
Pal J, Fulciniti M, Nanjappa P, Buon L, Tai YT, Tassone P et al. Targeting PI3K and RAD51 in Barrett's adenocarcinoma: impact on DNA damage checkpoints, expression profile and tumor growth. Cancer Genomics Proteomics 2012; 9: 55–66.
Aggarwal S, Taneja N, Lin L, Orringer MB, Rehemtulla A, Beer DG . Indomethacin-induced apoptosis in esophageal adenocarcinoma cells involves upregulation of Bax and translocation of mitochondrial cytochrome C independent of COX-2 expression. Neoplasia 2000; 2: 346–356.
Shi J, Shi Y, Waehrens LN, Rasmussen JT, Heegaard CW, Gilbert GE . Lactadherin detects early phosphatidylserine exposure on immortalized leukemia cells undergoing programmed cell death. Cytometry A 2006; 69: 1193–1201.
Shammas MA, Neri P, Koley H, Batchu RB, Bertheau RC, Munshi V et al. Specific killing of multiple myeloma cells by (-)-epigallocatechin-3-gallate extracted from green tea: biologic activity and therapeutic implications. Blood 2006; 108: 2804–2810.
Cawthon RM . Telomere measurement by quantitative PCR. Nucleic Acids Res 2002; 30: e47.
Dimri GP, Lee X, Basile G, Acosta M, Scott G, Roskelley C et al. A biomarker that identifies senescent human cells in culture and in aging skin in vivo. Proc Natl Acad Sci USA 1995; 92: 9363–9367.
We are grateful to Dr David G Beer, Department of Surgery, University of Michigan, Ann Arbor, MI, USA, for providing FLO-1 cells. This work was supported in part by grants from National Cancer Institute R01CA125711 to MAS, from the Department of Veterans Affairs Merit Review Awards (to NCM) and from the National Institutes of Health Grants RO1-1375555, P50-100007 and PO1-78378 to NCM.
The authors declare no conflict of interest.
Supplementary Information accompanies this paper on the Oncogene website
About this article
Cite this article
Lu, R., Pal, J., Buon, L. et al. Targeting homologous recombination and telomerase in Barrett’s adenocarcinoma: impact on telomere maintenance, genomic instability and tumor growth. Oncogene 33, 1495–1505 (2014). https://doi.org/10.1038/onc.2013.103
- homologous recombination
- Barrett’s adenocarcinoma
A pan-cancer landscape of telomeric content shows that RAD21 and HGF alterations are associated with longer telomeres
Genome Medicine (2022)
Medical Oncology (2022)
Blood Cancer Journal (2021)
Integrated genomics and comprehensive validation reveal drivers of genomic evolution in esophageal adenocarcinoma
Communications Biology (2021)
Radiation Oncology (2019)