3HMG-CoA reductasa inhibitor, simvastatin, inhibits cell cycle progression at the G1-S checkpoint in immortalized lymphocytes from Alzheimer´s disease patients independently of cholesterol lowering effects

Abstract

Recent work has suggested that statins may exert beneficial effects on patients suffering from Alzheimer's disease (AD). The pharmacological effects of statins extend beyond their cholesterol-lowering properties. Based on the antineoplastic and apoptotic effects of statins in several cell types, we hypothesized that statins may be able to protect neurons by controlling the regulation of cell cycle. A growing body of evidence indicates that neurodegeneration involves the activation of cell cycle machinery in postmitotic neurons. We and others have presented direct evidence to support the hypothesis that the failure of cell cycle control is not restricted to neurons in AD patients, but that it occurs in peripheral cells as well. For these reasons, we found it worthy to study the role of simvastatin on cell proliferation in immortalized lymphocytes from AD patients. We report here that simvastatin (SIM) inhibits the serum-mediated enhancement of cell proliferation in AD by blocking the events critical for G1/S transition. SIM induces a partial blockade of retinoblastoma protein phosphorylation and inhibition of cyclin E/cyclin-dependent kinase (CDK)2 activity associated with increased levels of the CDK inhibitors p21Cip1 and p27kip1. These effects of SIM on AD lymphoblasts are dependent on inhibition of the proteasome-mediated degradation of p21 and p27 proteins. The antiproliferative effect of this natural statin may provide a therapeutic approach for AD disease.

Statin therapy is a widely used treatment for hypercholesterolemia, reduces the risk of stroke, and improves cardiovascular functions (Farnier and Davignon, 1998). The statin family of drugs is competitive inhibitors of HMG-CoA reductase, the rate-limiting enzyme in the synthesis of cholesterol (Corsini et al., 1995), which converts HMG-CoA to mevalonate (MEV).

In the last decade, epidemiological, clinical, and experimental evidence has accumulated that links cholesterol to the development of AD, and recent studies showed that statin therapy might be of benefit in treating AD (Wolozin, 2004), although the efficacy of statins at slowing the cognitive decline and the progression of AD remains controversial. The link between cholesterol and AD is not surprising because the brain is the most cholesterol-rich organ, and disturbances in cholesterol homeostasis have been found associated with all major neuropathological features of AD (Shobab et al., 2005). Data from the Canadian Study of Health and Aging revealed a 74% reduced risk of developing AD in statin users compared with the total population (Rockwood et al., 2002). In other studies, a reduction in the risk of AD was observed in patients treated with statins compared with those receiving other medications typically used in cardiovascular disease (Wolozin et al., 2000), suggesting that statins in particular, rather than low cholesterol levels or lipid-lowering agents in general, are responsible for the reduction in the risk of AD.

A number of nonlipid-dependent or pleiotropic effects of statins have been reported (Takemoto and Liao, 2001). By preventing the synthesis of isoprenoid intermediates such as farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP) in the mevalonate pathway, statins may alter the subcellular localization and function of multiple proteins, including protein kinases, the subunit of trimeric G proteins, and Ras and Ras-related GTPases (Danesh et al., 2002). Pleiotropic effects of statins include anti-inflammatory properties as well as antiproliferative and proapoptotic effects (Koyuturk et al., 2004), all of these potentially relevant in treating AD. Growing evidence suggests that neuronal cell cycle regulatory failure, leading to apoptosis, may be a significant component of the AD pathogenesis (Herrup et al., 2004; Nagy, 2005). Neuronal changes supporting alteration on cell cycle control in the etiology of AD include the ectopic expression of cell cycle markers, or cytoskeletal alterations (Busser et al., 1998; Copani et al., 2001; Nagy, 2005). Moreover, it was reported that a significant number of hippocampal pyramidal and basal forebrain neurons in AD brain have undergone full or partial DNA replication (Yang et al., 2001). These events occur early in the progression of AD (Yang et al., 2003; Yang and Herrup, 2007), suggesting that cell cycle-induced death is a central mechanistic feature of the disease. There is an expanding body of evidence supporting the ability of some statins to exert direct antiproliferative and proapoptotic effects on various types of human cells (Katz, 2005). On these grounds, we have considered the possibility that the beneficial effects of statin therapy in AD could be related to their ability to interfere with cell cycle machinery. To this aim, we have investigated the effects of simvastatin, a lipophilic statin, on the distinct features of control of cell proliferation in lymphoblasts derived from late-onset AD patients. Previous work from this and other laboratories has demonstrated that cell cycle regulatory deficit is not restricted to neurons in AD; it is also observed in peripheral cells such as lymphocytes or fibroblasts (Tatebayashi et al., 1995; Nagy et al., 2002; de las Cuevas et al., 2003), thus providing a useful tool to study the involvement of cell cycle-related events in the pathogenesis of AD. A number of studies have found AD-specific changes in molecules and signaling pathways in peripheral lymphocytes that mirror changes in the brain (Eckert et al., 1998; Nagy et al., 2002; Muñoz et al., 2007). Moreover, these cells have also been used to study molecular changes in response to therapy in AD (Casademont et al., 2003; Reale et al., 2005). Conversely, Epstein-Barr virus (EBV) infection in vitro causes transformation of B cells and generates B-lymphoblastoid cell lines that resemble activated B cells (Neitzel, 1986). In fact, we have previously demonstrated identical cellular response to serum addition or withdrawal in peripheral lymphocytes or EBV-transformed lymphocytes from control and AD patients (Bartolomé et al., 2007; Muñoz et al., 2007). Taken together, these reports support a rationale for the use of peripheral cells, and in particular EBV lymphoblasts from AD patients as a model to further understand disease biology, progression, and therapeutic actions.

We report here that simvastatin selectively blocked the serum-enhanced proliferation of lymphoblasts from AD patients by regulating critical events of the G1/S transition, suggesting potential additional molecular targets for the clinical efficacy of this drug in treating AD