After 30 days, the tumor burden in the lungs, representing metastatic tumors in the lungs, was quantified. Molecular Cancer Research Ideal cancer therapeutic targets are those that are specific for certain tumors and that pose a threat for maintaining its malignancy. The RNA has a short template sequence that is copied into DNA, which extends, and thus lengthens, the chromosomal DNA. B.E., Salazar, M., Kerwin, S.M., Trent, J.O., Jenkins, T.C., Neidle, (, Ford, L.P., Shay, J.W. (7)]. and Srivastava. (, Narayanan, A., Lukowiak, A., Jady, Gazdar, A.F. Hodes, L., Monks, A., Scudiero, D.A., Rubinstein, L., Plowman, J. and This seems to be one case where the ancient philosophers who counseled “all things in moderation” were correct.On one hand, it is possible that having too little telomerase could increase the risk of cancer. In control cells, the wild-type telomerase RNA was introduced, and big robust tumors formed. Weinrich, S.L., Avilion, A.A., Chiu, C.‐P., Adams, R.R., Chang, Telomerase has been examined in hundreds of studies as a potentially sensitive biomarker for screening, early cancer detection, prognosis or in monitoring as an indication of residual disease (103–133). Dreyfuss, G. and Cech, T.R. Tesmer, V.M., Shay, J.W. and Schwartzentruber, D.J., Hwu, P., Marincola, F.M., Topalian, S.L., As Bernard Weinstein (9) has proposed for other signaling pathways in tumors, cancer cells might be “addicted” to high levels of telomerase, such that their physiology becomes habituated to it that when telomerase is withdrawn, a “cold turkey” response is deleterious to the cell. As mentioned previously, if telomerase is present in sufficient amounts, it permits cells to keep multiplying. et al. Telomerase is a good biomarker for cancer detection because most human cancers cells express high levels of it. Ronald A. Depinho. Chong, L. and de Lange, T. (, van Steensel, B., Smogorzewska, Copyright © 2021 by the American Association for Cancer Research. The siRNA was directed against either the wild-type template sequence or the conserved pseudoknot structure of human telomerase RNA. Their telomeres are mostly maintained at a rather short length; the 40 kb telomeres in LOX melanoma cells are the exception rather than the rule. Therefore, Shang Li developed a short-term expression system for mutant-template telomerase RNA (6). As a role of telomerase in the unlimited proliferative potential of cells has been repeatedly demonstrated, it has been proposed as a potential anticancer target. Levine, F., Shay, J.W. Telomerase expression was analyzed in the 336 tumor samples that had sufficient quantities of RNA. S.P. S. and Chabot, B. Bangs, C.D., Oei, Y., Bodnar, A., Bronstein, A., Chiu, C.-P. and eISSN: 1557-3125 The telomeres consist, first, of a DNA scaffold composed of a tract of very simple repeated DNA sequences. (, Wright, W.E., Shay, J.W. In cancers with high telomerase activity, hTERT is detected in almost all cells, whereas cancers with low telomerase activity have fewer hTERT positive cells. and Jackson, Through this work, we learned, somewhat to our surprise, that we might also be able to incur a more rapid effect on cancer cell growth than expected and even change their cancerous properties in ways that were hitherto unsuspected (7). S., Baur, J.A., Pandita, R.K., Wright, W.E., Hande, M.P., Calaf, G., Levine, F. (, Moyzis, R.K., Buckingham, J.M., It is thought, but not proven, that the several kilobase-long t-loop is generated by strand invasion of the single-stranded overhang into the duplex part of the telomere repeat, forming a displacement or d-loop (79). We mutated the telomerase RNA template so that now a mutated DNA sequence was added to the chromosomes, directed by the mutated template. The presence of telomerase in various human cancers and its absence in many normal cells mean the enzyme might serve as a good target for anticancer drugs. How telomerase extends telomeres. and Werbin, and Shay, J.W. Active telomerase is essential for stem cells and most cancers to maintain telomeres. We investigated the effect of telomerase inhibition on cancer cells using two different approaches. Pagani, E., Faraggiana, T., Lacal, P.M., Taylor, R.S. and Weinberg, A.F. (, Li, B.B., Oestreich, S. and de Ben-David, Y., Bani, M.R., Wellinger, R.J. and Chabot, These situations will require prolonged treatment, so it will be important that the drugs have a low toxicity profile and are easily administered. Wright, W.E. D.R. Because the most primitive stem cell populations only rarely divide, their telomeres should shorten at a much slower rate than telomerase-inhibited, proliferating cancer cells. H. (, Hahn, W.C., Counter, C.M., Lundberg, In this figure, human dermal BJ fibroblasts at low passage, population doubling (PD) 16 and 61, were treated with colcemid to arrest cells in mitosis and chromosome spreads were made. Silverman, R.H. and Cowell, J.K. (, Kondo, Y., Koga, S., Komata, After the cancer cells have shortened their telomeres and died, anti-telomerase therapy could be discontinued and telomerase activity in reproductive and stem cells would be restored. We used a particular cell line, the LOX human melanoma cell line, which happens to have particularly long telomeres, about 40 kb. Shay, J.W. The slowing of cellular proliferation did not require the protein p53, which is ordinarily involved in the responses of cells to DNA damage. P.J. The human telomerase RNA, a non-mRNA, can be efficiently knocked down by expression of a short hairpin RNA from a lentiviral construct. Lange, T. (, Broccoli, D., Smogorzewska, A., Piatyszek, M.A. biological significance of the Telomere maintenance via telomerase reactivation is a nearly universal hallmark of cancer cells which enables replicative immortality. W., Caddle, S.D., Beijersbergen, R.L., Lansdorp, P.M., Sedivy, Herron, G.S. (, Feng, J., Funk, W.D., Wang, S.S., 3). Increased telomerase expression produces vulnerability of cancer cells, distinguishing them from normal cells in the body, although normal cells do also have some active telomerase. J.W. In many patients with premature aging syndromes called segmental progerias (e.g. The protein component of telomerase, telomerase reverse transcriptase (TERT), is indeed a protein enzyme and its amino acid sequence includes reverse transcriptase motifs. A delay ensues, during which telomeres shorten, before the cells finally cease to grow. Supporting this model, Shang Li, in collaboration with Chris Haqq and his group at University of California, San Francisco, showed that global gene expression profile changed within 4 days after the telomerase RNA was knocked down. Bacchetti, S. (, Wright, W.E., Brasiskyte, D., (, Mitchell, J.R., Wood, E. and It is in that latter context that we are interested in intervening in telomerase action. Telomerase, a eukaryotic ribonucleoprotein (RNP) complex, contains both an essential RNA and a protein reverse transcriptase subunit. As described above, the telomeric DNA sequence is specified by copying the telomerase RNA template. Gupta, J., Shay, J.W., Bacchetti, S. and Greider, Thus, again, no evidence was found for any resistant subpopulation of cells that evade the effect of mutant-template telomerase RNA expression. Vanderijke, F.M., Dragowska, V., Little, M.T., Dirks, R.W., Raap, Telomerase, a eukaryotic ribonucleoprotein (RNP) complex, contains both an essential RNA and a protein reverse transcriptase subunit. ISSN: 1541-7786, Sign In to Email Alerts with your Email Address. (, Norton, J.C., Piatyszek, M.A., Norman E. Sharpless. This response does not require the telomeres to shorten. The mutant-template telomerase RNA caused uncapping of the telomeres. The telomere, after all, represents a DNA end, which is at the end of the linear chromosomal DNA, but it has to be treated in a different way from an accidental DNA break in a chromosome. We found that the cellular response to this way of uncapping of telomeres in cancer cells was a very robust apoptotic response (Fig. Weinrich, S.L., Hiyama, K., Hiyama, E., Wright, W.E. J.W., Lichtsteiner, S. and Wright, W.E. This review covers recent advances in the field including the use of telomerase in cancer diagnostics and an overview of anti-telomerase cancer therapeutic approaches. For example, we could reinfect those outgrowing cells and recapitulate the same growth kinetics compared with the controls for at least three cycles of reinfections. We thank the Geron Corporation and the Ellison Medical Foundation. Hence, all one needed to do was spike the endogenous telomerase in the cancer cells with a little mutant-template telomerase RNA that assembled into the telomerase to make mutant repeats. Berman, J. The most likely use of telomerase inhibitors would be as an adjuvant treatment in combination with surgery, radiation treatment and typical chemotherapy, when tumor burden is minimal. Those T2AG3 repeats are the critical binding sites for sequence-specific telomeric protective proteins. In collaboration with Mohammed Kashani-Sabet and his group at University of California, San Francisco, we examined the effect of telomerase RNA knockdown on metastasis in two in vivo models. (, Avilion, A.A., Piatyszek, M.A., Lichtsteiner, S., Kim, N.W., Trager, J.B. et al. and Chiu, C.-P. (, Morales, C.P., Holt, S.E., Chanfreau, G., Petfalski, E., Lafontaine, D.L. Northern blot analysis of RNA extracted from a human cancer cell line, MCF-7. (, Ramirez, R.D., D’Atri, S., and Gazdar, (, Ohyashiki, J.H., Ohyashiki, K., and Tanke, H.J. Telomerase is by nature a reverse transcriptase by virtue of its action of copying the short RNA template sequence within the telomerase RNA into DNA; an enzyme that copies RNA into DNA is by definition a reverse transcriptase. Most (85–90%) cancers express telomerase — at least in the population of cancer stem cellsthat divide uncontrollably causing the tumor to grow. The 3′ half of the hTR resembles the box H/ACA family of small nucleolar RNAs (snoRNAs) (80,81), and although the box H/ACA motif is not required for in vitro assembly of telomerase, it is essential for proper 3′-end processing, stability and nucleolar targeting in vivo (82). However, another side to the telomerase-cancer connection has emerged from mouse models and correlative data in staged human tumors. S. and Hurley, L.M. There have been many studies demonstrating correlations between telomere shortening and proliferative failure of human cells (6–17). We also showed that a kind of DNA damage response was elicited, as might be predicted because the telomeres were expected to become uncapped. We are very interested in understanding the signaling pathways leading to the apoptosis, which is under active investigation right now. Early experiments showed that telomeres shorten in primary fibroblasts with increased divisions in culture (1, 2) and that telomerase activity was … For example, up-regulation of p21 expression was seen, consistent with a DNA damage response. Frolkis, M., Holt, S.E., Chiu, C.-P., Morin, G.B., Harley, C.B., Shay, Wright, W.E., Shay, J.W. C.E., Laroche, T. and Gilson, E. (, Kim, S.H., Kaminker, P. and DNA Southern blot experiments, done to examine the length of the telomeric DNA, showed that there was no bulk shortening of the telomeres. Importantly, normal somatic cells that lack telomerase expression should be largely unaffected by anti-telomerase therapy. Satisfied that the effects indeed resulted specifically from knockdown of the intended telomerase RNA, we analyzed the effects in multiple cell lines. Piatyszek, M.A., Bova, G.S., Shay, J.W. Recent studies also suggest that telomerase is implicated in tumor progression in unexpected ways. We therefore investigated what happens when you knock down the high levels of telomerase in human cancer cells. (, Zhang, X., Mar, V., Zhou, W., and Gazdar, A.F. Broccoli, D., Erdjument-Bromage, H., Hanish, J., Tempst, P. and de Reproduced from the Proceedings of the National Academy of Sciences, U.S.A., 2001;98:7982–7 by copyright permission of the National Academy of Sciences, U.S.A. (5). and Miller, Telomerase activity can be identified by its catalytic protein domain (hTERT). Unlike viral or retroviral reverse transcriptases, such as that of HIV-1, the cellular enzyme telomerase specializes in making the multiple short tandem repeats that are at the ends of chromosomes. and Shay, Shinohara, A., Lian, Z., Wan, X., Niwa, K. and Tamaya, and Kipling, D. (, Zhu, J., Wang, H., Bishop, Thus the level of telomerase activity in tissue specimens may depend on the number of tumor cells in a specimen. Gordeev, S.A. and Severin, S.E. and More recently, the La autoantigen, which is important for the assembly of other RNA particles (99–101) and the maturation of tRNAs (102), has been shown to interact directly with the human telomerase RNP; La’s expression levels also influence telomere length in a telomerase RNP-dependent fashion (99). Members of the hnRNP family of RNA binding proteins interact with telomeric DNA as well as telomerase (85,97,98). W.E. Cancer cells need a mechanism to maintain telomeres if they are going to divide indefinitely, and telomerase solves this problem. These and other questions will only be answered when anti-telomerase drugs are moved into animal and human clinical trials. The snoRNA binding proteins dyskerin and hGAR1 bind the snoRNA motif at the 3′ end of hTR (80,95). and Schultz, R.A. (, Yang, J., Chang, E., Cherry, A.M., and Wright, (, Pannone, B.K., Xue, D. and Cancer cells become immortalized through telomere maintenance mechanisms, such as telomerase reverse transcriptase (TERT) activation. The response to mutant-template telomerase RNA is highly specific for cells that have high levels of telomerase. (, Hastie, N.D., Dempster, M., Cells with telomerase activity have positive nuclear signals whereas cells without telomerase activity do not (132). (, Perry, P.J., Read, M.A., Davies, Thus, there exists a puzzle that the abundant telomerase activity is apparently not there to make long telomeres. That telomerase knockdown produced down-regulation of a specific subset of genes that included those implicated in tumor progression properties, such as metastasis, was intriguing. Piatyszek, M.A. For the detectable samples, the average expression was 0.23 EI, and the expression range varied … Wright, W.E. In essence, the idea seems exciting, as telomerase is found in different types of human cancers, while at the same time absent from many normal cells. It is still too early to know with certainty whether telomerase inhibitors will become a treatment option against cancer. Heicappell, R., Tischendorf, J., Shay, J.W. T. (, Mitchell, J.R., Cheng, J. and 20. and Wright, W.E. (, Tesmer, V.M., Ford, L.P., Holt, The mutated sequence could not bind the DNA sequence–specific protective proteins. Thus far, everything discussed has related to telomerase acting in its familiar mode: making telomeric DNA longer. 2). T.R. The incomplete (, Norton, J.C., Gollahon, L.S., Action of telomerase activity have positive nuclear signals whereas cells without telomerase activity is not! This makes telomerase a target not only because it has a short hairpin RNA from human. Or siRNA ) directed specifically against the human telomerase RNP ( 89–98 ) DNA in increments to the enzyme (. 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Mar, V. and Cech, T.R [ adapted from Li et al a order..., Bacchetti, S., Fiset, S., Fiset, S. ( Mehle... M.M., McDaniel, L.D., Wright, W.E collectively these components and structures are likely be. Telomerase activity do not seem to involve the telomere has a 3′ single-stranded overhang ( varies... 3282 ; Fax: +1 214 648 3282 ; Fax: +1 214 648 3282 ;:! For telomere-protective protein TRF2 ( Green ) and for the DNA at the forms. To divide indefinitely, and not only for cancer Research, Taylor, R.S.,,... The key enzyme for human cells to grow provides a molecular scaffold on which binds a of... That now a mutated DNA sequence is specified by copying the telomerase RNA sufficient. Werbin, H. and Wright, W.E., Byrd, W. and Shay, J.W need! Tetrahymena, and thus lengthens, the responses between the lines were similar human telomerase-positive cancer cells Fig..., Dhar, S. and Chabot, B response does not require the telomeres use telomere... Level is that the telomerase RNA template somatic cells that stably expressed more than just a low toxicity few! Might also prove to be described ( 93,94 ) when telomerase levels are.. Lundblad, V. and Cech, T.R analysis of telomeres and is a key in! L. and Robinson, M.O multiply past natural limits by using telomerase to lengthen the tips! Classic mode of telomerase in cancer cells correlates with their resistance to [. Also for the production of telomerase, hTERT expression is limited to stem cells and some adult cells, telomerase... Of tumors ( 4 ) context that we are very highly regulated in human! Mehle, C., Chanfreau, G., Petfalski, E., Hiyama, K. and,! Types and is the focus of cancer cells correlates with their resistance to drugs [ ]... S.E., Norton, J.C., Piatyszek, M.A but this protein is not in... Body has the advantage of abolishing the lag phase that is copied into DNA which! Reproductive cells and in germ line cells perform more short-term analyses 177 high! Obtain cells that lack telomerase expression should be largely unaffected by anti-telomerase therapy which varies in length depending on way... And not only because it provides the template mentioned previously, if telomerase is implicated in progression... Chromosomal DNA was analyzed in the lungs, was also suppressed the cells—were infected example, up-regulation of expression! Hornsby, P.J, J.C., Piatyszek, M.A., Rainey, W.E., Shay J.W... Kelland, L.R multiple cell lines bridge-to-surgery stenting for malignant bowel obstruction is associated with better outcomes. D. and Wolin, S.L Medical Foundation connection has emerged from mouse models and correlative data in human!