Differential modulation of paclitaxel-mediated apoptosis by p21Waf1 and p27Kip1

Abstract

The impact of the cyclin dependent kinase (CDK) inhibitors p21 Waf1 and p27 Kip1 on paclitaxel-mediated cytotoxicity was investigated in RKO human colon adenocarcinoma cells with the ecdysone-inducible expression of p21 Waf1 or p27 Kip1 . Ectopic expression of p27 Kip1 arrested cells at G1 phase, whereas p21 Waf1 expression arrested cells at G1 and G2. Expression of p21 Waf1 after paclitaxel treatment produced much greater resistance to paclitaxel than did expression of p27 Kip1 . We attributed this di erence to the additional block at G2 induced by p21 Waf1 , which prevented cells from entering M phase and becoming paclitaxel susceptible. Expression of p21 Waf1 inhibited p34cdc2 activity and markedly reduced paclitaxel-mediated mitotic arrest, from 87.5 to 23%. In contrast, p27 Kip1 expression also inhibited p34cdc2 but reduced mitotic arrest only slightly, from 87.4 to 74.5%. We concluded that the G2 block produced by p21 Waf1 , but not by p27 Kip1 , contributed to their unequal modulation of sensitivity to paclitaxel-mediated apoptosis in RKO cells, and there is no causal relationship between paclitaxelmediated cytotoxicity and elevation of p34cdc2 activity.

Novel high-affinity PPARγ agonist alone and in combination with paclitaxel inhibits human anaplastic thyroid carcinoma tumor growth via p21WAF1/CIP1

“…The loss of normal p53 function (p53 transcriptionally upregulates p21 WAF1/CIP1 ) in fibroblast cells conferred sensitization to paclitaxel by increasing G 2 -M arrest and apoptosis (Wahl et al, 1996). This was also shown in sarcoma and breast adenocarcinoma cell lines (Barboule et al, 1997;Li et al, 1999;Schmidt et al, 2000). p21 WAF1/CIP1 has also been implicated in desensitizing cells to other chemotherapeutic agents such as cisplatin and nitrogen mustard (Fan et al, 1997).…”

BRCA1 and GADD45 mediated G2/M cell cycle arrest in response to antimicrotubule agents

“…We therefore postulated that the observed cell cycle arrest may represent activation of a mitotic rather than a G2 checkpoint. To test this hypothesis we distinguished the G2 population from the M population by FACS analysis of formaldehyde-®xed cells stained with Mithramycin A. Formaldehyde ®xation alters the chromatin structure of DNA in such a way that mitotic nuclei are di erentially stained using the¯uorescent dye Mithramycin A thereby di erentiating between the G2 and M populations (Schmidt et al, 2000). The FACS data indicated a 32% increase in the mitotic population when cells were induced to express BRCA1 in the presence of Taxol, supporting our hypothesis that BRCA1 was also inducing a mitotic arrest in response to Taxol treatment in these cells (Figure 2b).…”

“…G2 and mitotic population were separated by staining with Mithramycin A, as previously described (Schmidt et al, 2000). In all cases FACS analysis was carried out using an EPICS ELITE¯ow cytometer (Coulter).…”

Agonists of an ecdysone-inducible mammalian expression system inhibit Fas Ligand- and TRAIL-induced apoptosis in the human colon carcinoma cell line RKO

“…As an additional advantage, ecdysteroids are neither toxic nor teratogenic, and they should not affect mammalian physiology, 3 although a recent publication reported effects of muristerone A and ponasterone A on cytokine signaling in mammalian cells. 5 We used an established system of ecdysone-inducible p27 overexpression in the human colon carcinoma cell line RKO 6 to analyze the functional relationship between death receptormediated apoptosis and the cell cycle. During the course of these experiments, we noticed that the substances used to induce p27 expression (namely muristerone A, ponasterone A and GSt-E) exhibited a strong antiapoptotic effect independent of the ecdysone-inducible system.…”