ROLE OF NICOTINE IN LUNG CANCER

1 – INTRODUCTION

Tobacco use in cancer patients is associated with increased cancer treatment failure and decreased survival. Tobacco use is also associated with a more advanced stage at diagnosis, younger age at cancer presentation, decreased compliance to cancer treatment, decreased quality of life, increased risk of treatment toxicity, increased risk of developing second primary cancers, increased surgical risk, increased recurrence, and increased risk of cancer-related and non-cancer-related mortality. Nicotine is one of over 7,000 compounds in tobacco smoke and nicotine is the principal chemical associated with addiction. Nicotine is the reason that people cannot stop smoking, but it is not the cause of lung cancer. There has been a steady stream of recent studies demonstrating effects of nicotine in in vitro systems. The results of several studies have shown among other effects increased cell proliferation, inhibition of apoptosis, stimulation of cancer cell growth, and enhancement or inhibition of angiogenesis. Furthermore, nicotine can decrease the biologic effectiveness of conventional cancer treatments such as chemotherapy and radiotherapy. Nicotine may be a carcinogen, tumor promoter, or co-carcinogen, and it has been suggested to be the ‘‘estrogen of lung cancer.’’ Common mechanisms appear to involve activation of nicotinic acetylcholine receptors (nAchR), and also β-adrenergic receptors, leading to downstream activation of parallel signal transduction pathways that facilitate tumor progression and resistance to treatment. Data suggest that nicotine may be an important mechanism by which tobacco promotes tumor development, progression, and resistance to cancer treatment. Several studies found specific protein that could be inactivated using siRNA, in order to develop a new treatment for lung cancer.

Nicotine and lung cancer, 2013
Lung carcinogenesis by tobacco smoke, 2012

2 – NICOTINE

2.1 – Substance

Nicotine is an alkaloid named after the tobacco plant Nicotiana tabacum, which in turn is named after Jean Nicot. It is also present in lower quantities in tomato, potato, eggplant, green pepper and in the leaves of the coca plant. Pure nicotine is a clear liquid with a characteristic odor, whereas it turns brown on exposure to air. It is a strong base.

2.2 – Absorption

Nicotine absorption can occur through the oral cavity, skin, lung, urinary bladder, and gastrointestinal tract. The absorption of nicotine through the oral mucosa has been shown to be the principal route of absorption for smokers who do not inhale. The principal route of nicotine absorption in smokers who inhale is through the alveoli of the lung. When tobacco smoke reaches the small airways and alveoli of the lung, the nicotine is rapidly absorbed. Absorption through the alveoli is also dependent on the nicotine concentration in the smoke. Plasma nicotine levels depend on the type of cigarette smoked. Absorption of nicotine through the buccal mucosa is poor whereas the absorption through the lung is rapid. Nevertheless, absorption of nicotine through the skin is important during replacement therapies.

2.3 – Metabolism

During the tobacco curing and smoking process, nicotine can be converted NNK, NNAL and NNN through nitrosation. After entering the body, nicotine is rapidly absorbed and distributed throughout the body. Approximately 75% of nicotine is metabolized to cotinine by cytochrome P450. Further metabolism of cotinine involves additional hydroxylation primarily to 3-OH cotinine, glucuronidation, and excretion primarily in urine. The remaining nicotine can be converted to other metabolites such as nicotine-N-oxide. Glucuronidated nicotine and its metabolites are also excreted in the urine.

2.4 – Nicotinic Acetylcholine Receptors (nAchR)

The diverse functional properties of nicotine are due to its agonistic interaction with various subtypes of nicotinic acetylcholine receptors. nAChR consist of homo- or hetero-pentamer composed of the various subunits that are arranged symmetrically around an axis perpendicular to the membrane, thus delineating the ionic pore. The presence of β2 or β4 subunits in the receptor pentamer seems correlated with the respective high and low affinity for nicotine. All nAChR subunits share homologous structure with a large extracellular domain, four transmembrane regions (M1–M4) structured in α-helice, and a short extracellular C-terminal tail. As an allosteric receptor, nAChR may undergo rapid conformational transitions from a resting basal state to an active or desensitized state. Application of nicotine provokes first the stabilization of the receptor in a high-affinity open state followed by a progressive stabilization of a closed desensitized state. Normal human bronchial epithelial cells express α3-, α4-, α5-, and α7- subunits of nAChR that form channels modulating Ca++ metabolism and regulating cell adhesion and motility. Long-term exposure to millimolar concentrations of nicotine resulted in a steady increase of [Ca++], which may lead to cell damage.

Multiple roles of nicotine on cell proliferation and inhibition of apoptosis: Implications on lung carcinogenesis, 2008
Nicotine and lung cancer, 2013

3 – LUNG CANCER

3.1 – Antiapoptotic effect

Mcl-1, a major antiapoptotic protein of the Bcl2 family, is extensively expressed in both small cell and non–small cell lung cancer cells. Mcl-1 can be stimulated by multiple growth factors including interleukins, vascular endothelial growth factor (VEGF), α-IFN, and epidermal growth factor (EGF). Nicotine, a major component of cigarette smoke, can mimic growth factors to simulate Mcl-1 phosphorylation. Nicotine activates ERK1/2 through the upstream β-adrenergic receptors. ERK1 and ERK2 are physiologic Mcl-1 kinases that can phosphorylate Mcl-1 at T163 site. Nicotine not only stimulates phosphorylation and activation of ERK1/2 but also facilitates the phosphorylated, active ERK1/2 to colocalize with Mcl-1 in cytoplasm. The T163 site should be a critical target for nicotine to positively regulate the antiapoptotic function of Mcl-1. Nicotine not only markedly prolongs the half-life of Mcl-1, but also upregulates its expression level in various human lung cancer cells.

Multiple roles of nicotine on cell proliferation and inhibition of apoptosis: Implications on lung carcinogenesis, 2008
Nicotine Enhances the Antiapoptotic Function of Mcl-1 through Phosphorylation, 2009

3.2 – Proliferation

Nicotine promotes lung cancer proliferation via the α7-nicotinic acetylcholine receptor (α7-nAChR) subtype. It increases the levels of α7-nAChR mRNA and α7-nAChR transcription in human SCC-L cell lines and SCC-L tumors, which facilitates tumor growth and progression. The α7-nAChR has been shown to cause direct activation of inactivation of signaling kinases and phosphatases. The α7-nAChR promoter has several binding sites for Sp1. Studies have demonstrated that the Sp1 protein can directly associate with the GATA family of transcription factors to regulate gene expression. The transcription factor Sp1 can interact directly with GATA4 and GATA6 in a signal-dependent manner in several experimental systems to regulate gene expression. Nicotine induces the binding of GATA4 or GATA6 to Sp1 on the α7-nAChR promoter. This process increased α7-nAChR transcription and expression in SCC-Ls. Nicotine activates one of the best-characterized signaling pathways that promotes cellular survival, the PI3K/Akt pathway. Akt increased phosphorylation of multiple downstream components that control cellular cell cycle.

Nicotine Induces the Up-regulation of the α7-Nicotinic Receptor (α7-nAChR) in Human Squamous Cell Lung Cancer Cells via the Sp1/GATA Protein Pathway, 2013
Activated Cholinergic Signaling Provides a Target in Squamous Cell Lung Carcinoma, 2008

h3. 3.3 – Angiogenesis

In order to grow beyond a critical size, tumors must recruit endothelial cells from the surrounding stroma to form their own endogenous microcirculation. Nicotine, asides from its psychoactive and addictive effects, can promote tumor growth and angiogenesis in lung cancer. Hypoxia-inducible factor-1α (HIF-1α) and VEGF are overexpressed in human lung cancers. Nicotine promotes HIF-1α protein accumulation and VEGF expression in human lung cells by activating various downstream nAChR-mediated signaling pathways which include the influx of Ca2+ and activation of calmodulin, PKC and c-Src. VEGF has been recognized as one of the principal initiators of tumor angiogenesis. VEGF expression is regulated by external factors, of which hypoxia is the best-characterized mediator of VEGF secretion, whereas HIF-1 protein is stabilized and bound to the hypoxia-responsive elements on VEGF promoter, thus leading to the transcriptional activation of the VEGF gene. HIF-1α contributes to the up-regulation of VEGF expression in tumor angiogenesis. The endothelial nAChR mediates an angiogenic pathway that is interdependent with growth factor mediated pathways. Endothelial nAChRs modulate blood vessel formation and remodeling, and mediate the effect of nicotine (or endogenous acetylcholine) on angiogenesis. α7-nAChR is upregulated during proliferation of subconfluent endothelial cells, or by hypoxia. Nicotine stimulates endothelial cell proliferation, and promotes the synthesis of growth factors and autocoids (NO, endothelin, prostacyclin) that may have angiogenic effects, increases the expression of matrix metalloproteinases that facilitate migration of vascular cells through the extracellular matrix, potentiates endothelial-monocyte interactions that contribute to arteriogenesis and increases the incorporation of endothelial progenitor cells into newly forming vessels.

Nicotine Induces Hypoxia-Inducible Factor-1α Expression in Human Lung Cancer Cells via Nicotinic Acetylcholine Receptor–Mediated Signaling Pathways,2007
Angiogenesis and the role of the endothelial nicotinic acetylcholine receptor,2007

3.4 – Migration and invasion

From primary tumor to secondary growth, cancer cells must invade the surrounding tissues, penetrate vessels, and travel to other sites where they arrest and resume growth. During the metastatic cascade, tumor cells disrupt many physical barriers formed by epithelial and endothelial basement membranes. Active cell motility is essential during intravasation and extravasation. Cigarette smoke constituents not only contribute to tumorigenesis but also may increase the spread of cancer in the body. NNK is formed by nitrosation of nicotine and has been identified as the most potent carcinogen. NNK, an important component in cigarette smoke, may also promote tumor metastasis by regulating cell motility. It can induce activation of a functionally interdependent protein kinase cascade, including c-Src, PKC and FAK (localized in cytoplasm), in association with increased migration and invasion of human lung cancer cells. Treatment of cells with α7 nAChR specific inhibitor α-BTX blocks NNK-stimulated activation of c-Src, PKC and FAK and suppresses cell migration and invasion. NNK-induced cell migration and invasion may occur through activation of the signal transduction pathway involving α7 nAChR/c-Src/PKC/FAK in lung cancer cells. c-Src, that is a PKC upstream kinase directly interacts with FAK and this association facilitates activation of both c-Src and FAK. PKC is a multigene family consisting of at least 11 distinct lipid-regulated protein-serine/threonine kinases that promotes tumor-cell migration in a mechanism by regulating localization of cytoskeletal proteins and phosphorylation of focal adhesion kinase (FAK). FAK plays a critical role in tumor invasion and metastasis because FAK binds to the cytoplasmic domain of β1 integrin, and subsequently binds to the SH2 domain of c-Src. This molecular complex facilitates activation of the c-Src/FAK signaling cascade and is critical in many cytoskeletal functions, as adhesions and cell-cell contacts.

Multiple roles of nicotine on cell proliferation and inhibition of apoptosis: Implications on lung carcinogenesis, 2008
NNK promotes migration and invasion of lung cancer cells through activation of c-Src/PKCι/FAK loop, 2013