Microarray and Biochemical Analysis of Lovastatin-induced Apoptosis of Squamous Cell Carcinomas

Abstract

We recently identified 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, the rate-limiting enzyme of the mevalonate pathway, as a potential therapeutic target of the head and neck squamous cell carcinomas (HNSCC) and cervical carcinomas (CC). The products of this complex biochemical pathway, including de novo cholesterol, are vital for a variety of key cellular functions affecting membrane integrity, cell signaling, protein synthesis, and cell cycle progression. Lovastatin, a specific inhibitor of HMG-CoA reductase, induces a pronounced apoptotic response in a specific subset of tumor types, including HNSCC and CC. The mediators of this response are not well established. Identification of differentially expressed genes represents a feasible approach to delineate these mediators as lovastatin has the potential to modulate transcription indirectly by perturbing levels of sterols and other mevalonate metabolites. Expression analysis following treatment of the HNSCC cell lines SCC9 or SCC25 with 10 microM lovastatin for 1 day showed that less than 2% (9 cDNAs) of the 588 cDNAs on this microarray were affected in both cell lines. These included diazepam-binding inhibitor/acyl-CoA-binding protein, the activated transcription factor 4 and rhoA. Because the biosynthesis of mevalonate leads to its incorporation into more than a dozen classes of end products, their role in lovastatin-induced apoptosis was also evaluated. Addition of the metabolites of all the major branches of the mevalonate pathway indicated that only the nonsterol moiety, geranylgeranyl pyrophosphate (GGPP), significantly inhibited the apoptotic effects of lovastatin in HNSCC and CC cells. Because rhoA requires GGPP for its function, this links the microarray and biochemical data and identifies rhoA as a potential mediator of the anticancer properties of lovastatin. Our data suggest that the depletion of nonsterol mevalonate metabolites, particularly GGPP, can be potential mediators of lovastatin-induced apoptosis of HNSCC and CC cells.

Determinants of sensitivity to lovastatin-induced apoptosis in multiple myeloma

“…As isopentenyl PPi is also depleted by statin exposure, it would be unavailable to the statin-treated cell (49,50). These results, as well as the ability of GGTI-298 to induce apoptosis comparably with statins in sensitive tumor lines, build on previous reports (1,11,16,41) to clearly indicate the importance of protein geranylgeranylation in statininduced apoptosis.…”

“…We and others have shown that of these downstream end-products, GGPP most consistently reverses statin-induced apoptosis in sensitive tumor cells (1,11,16,41). To evaluate the mechanism of lovastatininduced apoptosis in MM, four sensitive MM cell lines (KMS11, OPM2, H929 and OCI MY7) were screened by 3-4,5-dimethylthiazolyl-2,2,5-diphenyl tetrazolium bromide assay, as we have previously conducted (41).…”

“…Of the known isoprenylated proteins, Rho and Ras GTPases are critical in promoting survival, proliferation, transformation, cell cycle control, and the regulation of cytoskeleton formation (9). Indeed, it has recently been shown that statin exposure leads to the mislocalization of several Ras and Rho GTPases from the membrane (10 -15) and that the expression of many of these GTPases has also been shown to be up-regulated in response to statin exposure (14,16,17). These observations have been made in a wide range of cell types, presumably as part of a feedback response to replace functionally depleted isoprenylated proteins in statin-treated cells.…”

The HMG-CoA reductase inhibitor simvastatin overcomes cell adhesion–mediated drug resistance in multiple myeloma by geranylgeranylation of Rho protein and activation of Rho kinase

“…26 Mevastatin, for example, inhibits bone resorption and induces osteoclast apoptosis in vitro by inhibition of protein prenylation. 27 Furthermore, statins have been shown to induce apoptosis in melanoma, 28 thyroid cancer, [29][30] colon carcinoma, 32,33 squamous cell carcinoma, 34 breast cancer, 35 acute myeloid leukemia, 36 malignant lymphoma, 37 and multiple myeloma cells 14,38 in vitro. In our experiments, statins have been used in much lower concentrations than in most in vitro studies published with complete reversal of CAM-DR, suggesting realistically achievable doses in humans.…”

Statin use and cancer risk in the General Practice Research Database

“…Rodent studies indicate that statins are carcinogenic (Newman and Hulley, 1996). Other evidence, however, suggests that statins may inhibit the growth of cancer cells by triggering apoptosis (Muller et al, 1998;Dimitroulakos et al, 2002;Wong et al, 2002), inhibiting angiogenesis (Weis et al, 2002;Park et al, 2002), or impairing the metastatic process (Alonso et al, 1998;Kusama et al, 2002). We undertook a matched case -control study of the relation between statin use and cancer risk.…”