The association of statins and taxanes: an efficient combination trigger of cancer cell apoptosis

Abstract

Background: Cancer cell killing might be achieved by the combined use of available drugs. Statins are major anti-hypercholesterolemia drugs, which also trigger apoptosis of many cancer cell types, while docetaxel is a potent microtubule-stabilising agent.

Methods: Here, we looked at the combined effects of lovastatin and docetaxel in cancer cells.

Results: Whole transcriptome microarrays in HGT-1 gastric cancer cells demonstrated that lovastatin strongly suppressed expression of genes involved in cell division, while docetaxel had very little transcriptional effects. Both drugs triggered apoptosis, and their combination was more than additive. A marked rise in the cell-cycle inhibitor p21, together with reduction of aurora kinases A and B, cyclins B1 and D1 proteins was induced by lovastatin alone or in combination with docetaxel. The drug treatments induced the proteolytic cleavage of procaspase-3, a drop of the anti-apoptotic Mcl-1 protein, Poly-ADP-Ribose Polymerase and Bax. Strikingly, docetaxel-resistant HGT-1 cell derivatives overexpressing the MDR-1 gene were much more sensitive to lovastatin than docetaxel-sensitive cells.

Conclusion: These results suggest that the association of lovastatin and docetaxel, or lovastatin alone, shows promise as plausible anticancer strategies, either as a direct therapeutic approach or following acquired P-glycoprotein-dependent resistance.

Figure 1

Apoptosis induction by lovastatin and docetaxel in HGT-1 gastric cancer cells. HGT-1 cells were treated with 12.5 μM lovastatin (L12.5) or with 5 or 10 nM docetaxel (D5 or D10) alone or in combination for 48 h. Apoptosis was determined by Hoechst 33342 staining. Values are means ±s.d. (n=6). ^***Compared with control, ^###compared with docetaxel treatment, ^†††compared with lovastatin treatment (P<0.001, Student's t-test).

Figure 2

Effect of lovastatin and docetaxel on lipid synthesis gene expression levels. HGT-1 cells were treated with 12.5 μM lovastatin (L12.5) or with 5 or 10 nM docetaxel (D5 or D10) alone or in combination for 48 h. HMG-CoA reductase (HMG-CoA red), LDL receptor (LDL-R), fatty acid synthase (FAS), FPP synthase (FPPS) (A) and SREBP-1 and SREBP-2 (B) mRNA levels were analysed by real-time RT–PCR (see Materials and Methods). Relative mRNA levels were normalised to P0 mRNA levels. Values are means±s.d. (n=4). ^*Compared with control and ^#compared with docetaxel treatment. One symbol: P<0.05, two symbols: P<0.01, three symbols: P<0.001 (Student's t-test).

Figure 3

Effect of lovastatin and docetaxel on caspase-3, Poly-ADP-Ribose Polymerase (A), Mcl-1 and Bax protein levels (B). HGT-1 cells were treated with 12.5 μM lovastatin (L12.5) or with 5 or 10 nM docetaxel (D5 or D10) alone or in combination for 48 h. Protein levels were analysed by western blotting. Hsc70 was used as a loading control. The results are representative of three experiments with similar results.

Figure 4

Effect of lovastatin and docetaxel on p21 and p27 expression. HGT-1 cells were treated with 12.5 μM lovastatin (L12.5) or with 5 or 10 nM docetaxel (D5 or D10) alone or in combination for 48 h. (A) p21 and p27 mRNA levels were analysed by real-time RT–PCR. Relative mRNA levels were normalised to GAPDH mRNA levels. Values are means±s.d. (n=4). ^*Compared with control, ^#compared with docetaxel treatment, †compared with lovastatin treatment. One symbol: P<0.05, two symbols: P<0.01, three symbols: P<0.001 (Student's t-test). (B) p21 and p27 protein levels were analysed by western blotting. Hsc70 was used as a loading control. The western blotting analyses are representative of three experiments with similar results.

Figure 5

Effect of lovastatin and docetaxel on expression of genes involved in the initiation and progression of mitosis, cytokinesis, and MAP kinases signalling pathway. HGT-1 cells were treated with 12.5 μM lovastatin (L12.5) or with 5 or 10 nM docetaxel (D5 or D10) alone or in combination for 48 h. Cyclin D1, cyclin B1, aurora kinase A (A), aurora kinase B and survivin (B) mRNA levels were analysed by real-time RT–PCR. Relative mRNA levels were normalised to P0 mRNA levels. Values are means±s.d. (n=4). ^*Compared with control, #compared with docetaxel treatment, ^†compared with lovastatin treatment. One symbol: P<0.05, two symbols: P<0.01, three symbols: P<0.001 (Student's t-test). (C) Cyclin B1, cyclin D1, aurora kinase A, aurora kinase B, and survivin protein levels were analysed by western blotting. Hsc70 was used as a loading control. (D) phospho-JNK, phospho-MEK1/2, ERK1/2, phospho-ERK1/2, and p38 protein levels were analysed by western blotting. Hsc70 was used as a loading control. The western blotting analyses are representative of three experiments with similar results.

Figure 6

Comparison of the effect of lovastatin in HGT-1 and HGT-1-D5 cells. HGT-1 and HGT-1-D5 cells were treated with 0, 2.5, or 5 μM lovastatin for 48 or 72 h (A). Apoptosis was determined by Hoechst 33342 staining. Values are means±s.d (n=3). ^*P<0.1 or ^**P<0.01 for HGT-1-D5 cells compared with HGT-1 cells (Student's t-test). (B) HGT-1 and HGT-1-D5 cells were treated with 2.5 μM lovastatin for 72 h. Protein levels were analysed by western blotting. Hsc70 was used as a loading control. The western blotting analyses are representative of three experiments with similar results.