Modulation of anti-apoptotic and survival pathways by curcumin as a strategy to induce apoptosis in cancer cells

Abstract

Apoptosis is a highly regulated mechanism by which cells undergo cell death in an active way. As one of the most challenging tasks concerning cancer is to induce apoptosis in malignant cells, researchers increasingly focus on natural products to modulate apoptotic signaling pathways. Curcumin, a natural compound isolated from the plant Curcuma longa, has chemopreventive properties, which are mainly due to its ability to arrest cell cycle and to induce apoptosis.

This article reviews the main effects of curcumin on the different apoptotic signaling pathways involved in curcumin-induced apoptosis of cancer cells, including the intrinsic and extrinsic apoptosis pathways, the NF-κB-mediated pathway as well as the PI3K/Akt signaling pathway. This review also focuses on the sensitization of cells to TRAIL-induced apoptosis after curcumin treatment and shows that curcumin enhances the capacity to induce cell death of different chemotherapeutical drugs.

Introduction

Apoptosis is an ubiquitous and highly regulated mechanism by which cells undergo programmed cell death [1]. Resistance to apoptosis is a hallmark of cancer, with both the loss of pro-apoptotic signals and the gain of anti-apoptotic mechanisms contributing to tumorigenesis [2]. Several cellular pathways cumulate in the activation of caspases and apoptosis. Overly simplified, the two main apoptosis pathways are the extrinsic and the intrinsic pathway [3]. The extrinsic pathway is initiated by the interaction between specific ligands and surface receptors [4], such as CD95/Fas/Apo1, tumor necrosis factor (TNF) receptor 1 (TNFR1), TNF receptor 2 (TNFR2) and death receptors 3–6 (DR3–6) [1], which are able to deliver a death signal from the extracellular microenvironment to the cytoplasm. Binding to the receptor induces receptor multimerization, binding of Fas-associated death domain (FADD) adapter protein, formation of the death-induced signaling complex (DISC) which recruits the initiator caspases 8 and 10 and subsequently activation of the effector caspases 3 and 7 [4].

The intrinsic pathway is activated by various stimuli, i.e. DNA damage, hypoxia, cell detachment, cellular distress and cytotoxic drugs, which act inside the cell [1]. All of these signals converge to mitochondria, where the propagation of the apoptotic signal is regulated by the Bcl-2 family members [5]. Bcl-2 and Bcl-xL exert anti-apoptotic effects, while others such as Bid, Bad and Bim are pro-apoptotic [6], [7]. An excess of pro-apoptotic over anti-apoptotic signals initiates mitochondrial outer membrane permeabilization (MOMP), which leads to the release of proteins such as cytochrome c and Smac/Diablo from the mitochondrial intermembrane space to the cytosol. Once cytochrome c is released, it binds to Apaf-1 and ATP, which then bind to pro-caspase 9 to create a protein complex known as apoptosome, which in turn activates the effector caspase 3 [8]. Smac binds to the inhibitor of apoptosis proteins (IAPs) and deactivates them, preventing the IAPs from arresting the apoptotic process and therefore allowing apoptosis to proceed.

A third apoptotic pathway, the “endoplasmic reticulum (ER) stress” pathway has recently been described [9]. ER stress, induced by the accumulation of unfolded or malfolded proteins due to hypoxia, nutrient deprivation, reduction of disulfide bonds, and over-expression of some proteins, activates various apoptotic pathways. Crosstalk between the mitochondria and ER plays an essential role in ER stress-mediated cell death. The cytochrome c-dependent apoptotic pathway is activated by ER stress [10]. On the other hand, caspase-12, which is located at the ER, is also activated by excess ER stress and results in cell death in the absence of the cytochrome c-dependent pathway [10].

The existence of various check-points in apoptosis reveals a complex balance between cell survival and cell death in cells.

Two of the main signaling pathways involved in cell survival, by inhibiting apoptotic processes, are the nuclear factor-kappa B (NF-κB) and the Akt signaling pathways. The transcription factor NF-κB is one of the most studied transcription factors in mammalian cells. Its function has been implicated in inflammation, cell proliferation, differentiation, apoptosis and cell survival. NF-κB is an ubiquitously expressed family of five proteins; p65 (RelA), p50, p52, c-Rel and RelB.

Many stimuli give survival responses to cells that are mediated by NF-κB. Indeed, overall reduction in NF-κB activity is associated with increased apoptotic index in many cell types [11]. Furthermore, NF-κB activation has been shown to inhibit p53 dependent apoptosis following expression of the oncogene AP12/MALT1 [12]. This NF-κB directed survival response is associated with increased expression of anti-apoptotic proteins. Thus, it is not surprising that NF-κB expression is deregulated in various disease states including chronic inflammation and cancer.

The phosphatidylinositol-3-kinase (PI3K) signaling pathway is also crucial for many aspects of cell growth and survival and is frequently disrupted in human cancers. The PI3Ks are a family of related enzymes that are capable of phosphorylating the 3 position hydroxyl group of the inositol ring of PI(4,5)P2, to generate PI(3,4,5)P3 [13]. Upon activation of the PI3K pathway by many growth factors (e.g. epidermal growth factor (EGF)) and regulators, PI(3,4,5)P3 is produced on the inner side of the plasma membrane and Akt binds to the phospholipids. Akt, also called PKB (Protein kinase B) or RAC, (related to protein kinase A and C), is the human homologue of the viral oncogene v-akt ([14], [15], [16] which regulates multiple targets including several apoptotic genes [17], [18]. Akt inactivates pro-apoptotic factors like Bad, which controls the release of cytochrome c[19], [20], procaspase-9 and Forkhead transcription factors (like FOXO). Akt also activates anti-apoptotic genes, including cyclic-AMP response element-binding protein (CREB) and IκB kinase (IKK) leading to NF-κB nuclear localization and the subsequent transcription of pro-survival genes, such as Bcl-xL, caspase inhibitors and c-Myb [21], [22]. Overexpression of Akt has anti-apoptotic effects in various cell types resulting in cell death resistance [23].

Curcumin, a phenolic compound isolated from the plant Curcuma longa has anti-inflammatory, antioxidant and anti-cancer activities. The anticarcinogenic properties of curcumin in animals have been demonstrated by its inhibition of tumor initiation [24] and tumor promotion [25], [26]. Although the precise mode of action of this compound is not yet elucidated, studies have shown that chemopreventive action of curcumin might be due to its ability to induce apoptosis by several pathways. The number of signaling pathways and molecular targets involved is continuously growing and consequently the picture is becoming more and more complex, not least because results often appear to be cell-type specific and dose-dependent.

In order to characterize apoptotic genes regulated by curcumin in tumor cells, Ramachandran et al. have performed a microarray study [27]. Of the 214 apoptosis-associated genes in the array, the expression of 104 genes was altered by curcumin treatment [27]. These results show that curcumin-induced apoptosis is regulated by multiple signaling pathways.