Genetic analysis of cellular senescence

Vojta, Patrick J.; Barrett, J. Carl

doi:10.1016/0304-419x(95)00002-w

Cited by 55 publications

(60 citation statements)

References 138 publications

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“…Predictive mathematical simulations indicate that after inhibition of telomerase a relatively long period of time corresponding to about 20-30 cell divisions would be necessary for telomerase inhibitors to induce a dramatic telomere shortening and chromosomal instability (Vojta and Barret, 1995). Such a number of cell divisions is theoretically sufficient to increase the tumour mass by 1000 times in patients and is certainly sufficient to kill the host Harley and Villeponteau, 1995).…”

Section: Discussionmentioning

confidence: 99%

A human breast cancer model for the study of telomerase inhibitors based on a new biotinylated-primer extension assay

Raymond¹,

Sun²,

Izbicka³

et al. 1999

Br J Cancer

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SummaryTelomerase is an RNA-dependent polymerase that synthesizes telomeric DNA (TTAGGG) n repeats. The overall goal of our work was to establish human cancer models that can be used to design clinical trials with telomerase inhibitors. The objectives of this study were (1) to set up a human breast cancer system that allows evaluation of the effects of telomerase inhibitors in cultured cells using a non-amplified telomerase assay and (2) to test this system using two drugs (cisplatin and TMPyP 4 ) that affect the telomerase expression in breast cancer cells in culture. We first compared the telomerase activity in a variety of human breast cancer cell lines to that of other tumour types using a new biotinylated-primer extension assay. Our method, based on a non-amplified primer extension assay shows the direct incorporation of 32 Plabelled nucleotides induced by telomerase on human telomeric primers. The 32 P-dGTP labelled telomerase-extended 5′-biotinylated (TTAGGG) 3 primer can subsequently be separated using streptavidin-coated magnetic beads. As compared to other non-amplified method, we showed that this procedure improved the characterization and the quantification of the banding pattern resulting from telomerase extension by reducing the radioactive background. Using this method, we observed that telomerase activity varies markedly in a panel of 39 human cancer cell lines. For example, MCF7 breast cancer cells in culture showed intermediate telomerase activity corresponding to 33.8 ± 3.4% of that of the HeLa cells (reference cell line). Similarly, the telomere length varied with each cell line (average: 6.24 ± 6.16). No correlation between the level of telomerase and telomere length was observed, suggesting that a high processivity is not required to maintain telomeres and that, in some cell lines, another mechanism of telomere elongation can maintain telomere length. From this study, we selected MCF7 and MX1 models that showed reproducible telomerase activity and a relatively limited telomere length for the testing of potential telomere-telomerase interacting agents. Using cisplatin and a new porphyrin-derived compound TMPyP 4 , we showed that our model was able to detect a down-regulation of the telomerase activity in MCF7 cells in culture and in a human MX1 tumour xenografts. Based on these results, a breast cancer model for evaluating telomerase and telomere interactive agents is proposed.

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Section: Discussionmentioning

confidence: 99%

A human breast cancer model for the study of telomerase inhibitors based on a new biotinylated-primer extension assay

Raymond¹,

Sun²,

Izbicka³

et al. 1999

Br J Cancer

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“…At the end of their lifespan, normal cells enter into a permanent growth-arrest known as cellular senescence (reviewed in Votja and Barrett, 1995). In human cells, telomere attrition is directly implicated in triggering senescence (see for example Bodnar et al, 1998;Kiyono et al, 1998); but in rodent cells, senescence is independent of telomere shortening (Blasco et al, 1997;Carman et al, 1998;Russo et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

Murine fibroblasts lacking p21 undergo senescence and are resistant to transformation by oncogenic Ras

1999

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The cell-cycle inhibitor p21 is upregulated during senescence and upon induction of senescence-like arrest by oncogenic Ras. We have used primary ®broblasts derived from p21-null mice to evaluate the role of p21 in these processes. We ®nd that primary p21 7/7 cells enter senescence and have a lifespan similar to wild-type cells. Upon immortalization, most wild-type and p21 7/7 cultures acquire alterations in either p53 or p16 INK4a , further indicating that p21-de®ciency is not su cient by itself to allow immortalization. Primary p21 7/7 cells, like wild-type cells, respond to oncogenic Ras by accumulating p53 and p16 INK4a , and by decreasing their proliferation rate. In agreement with this, p21 7/7 cells are refractory to neoplasic transformation by oncogenic Ras when compared to p53 7/7 cells. We conclude that, in murine ®broblasts, p21 is not essential neither for senescence nor for preventing neoplasic transformation by oncogenic Ras.

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“…In human cells, for instance, an immortalizing oncogene like SV40 large T antigen extends the replicative potential of infected ®broblasts past their Hay¯ick limit (Hay¯ick and Moorhead, 1961). Telomerase is only expressed in post-crisis cells (Meyerson et al, 1997), however, and the emergence of immortal clones is a rare event occurring with an extremely low probability (reviewed in Vojta and Barrett, 1995). Activation of telomerase in isolated Figure 2 Induction of telomerase activity by MC29 and MC29 td10H in quail myoblasts.…”

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confidence: 99%

Induction of telomerase activity in v-myc-transformed avian cells

Falchetti¹,

Falcone²,

D’Ambrosio³

et al. 1999

Oncogene

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Telomerase activity is detectable in the majority of tumors or immortalized cell lines, but is repressed in most normal human somatic cells. It is generally assumed that reactivation of telomerase prevents the erosion of chromosome ends which occurs in cycling cells and, hence, hinders cellular replicative senescence. Here, we show that the expression of v-Myc oncoprotein by retroviral infection of telomerase-negative embryonal quail myoblasts and chicken neuroretina cells is su cient for reactivating telomerase activity, earlier than telomere shortening could occur. Furthermore, the use of a conditional v-Myc-estrogen receptor protein (v-MycER) causes estrogen-dependent expression of detectable levels of telomerase activity in recently infected chick embryo ®broblasts and neuroretina cells. We conclude that the high levels of telomerase activity in v-Myc-expressing avian cells are not the mere consequence of transformation or of a di erentiative block, since v-Src tyrosine kinase, which prevents terminal di erentiation and promotes cell transformation, fails to induce telomerase activity.

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Genetic analysis of cellular senescence

Cited by 55 publications

References 138 publications

A human breast cancer model for the study of telomerase inhibitors based on a new biotinylated-primer extension assay

A human breast cancer model for the study of telomerase inhibitors based on a new biotinylated-primer extension assay

Murine fibroblasts lacking p21 undergo senescence and are resistant to transformation by oncogenic Ras

Induction of telomerase activity in v-myc-transformed avian cells

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