Elsevier

Toxicology in Vitro

Volume 27, Issue 8, December 2013, Pages 2256-2263
Toxicology in Vitro

Nicotine increases survival in human colon cancer cells treated with chemotherapeutic drugs

https://doi.org/10.1016/j.tiv.2013.09.020Get rights and content

Highlights

  • Nicotine increases proliferation and inhibits apoptosis in human colon carcinoma cells treated with 5-FU and CPT.

  • Nicotine effects are mediated through the interaction with the α7-nAChR.

  • Nicotine addition to chemotherapeutic agents inhibits apoptosis through ERK and AKT pathways.

Abstract

Cigarette smoking is implicated in the development of colon cancer. Furthermore, nicotine increases cell proliferation and inhibits apoptosis through α7-nicotinic acetylcholine receptor (α7-nAChR) activation in human colon carcinoma cells. An open issue is whether nicotine interfere with colorectal cancer pharmacological treatment, by inhibiting drug-mediated apoptosis. To assess this hypothesis, we evaluated nicotine effect on Caco-2 and HCT-8 colon cancer cells, treated with 5-Fluorouracil (5-FU) and Camptothecin (CPT), chemotherapeutics commonly utilized as adjuvant treatment of colon cancer. Nicotine decreased anti-proliferative and pro-apoptotic effects exerted by chemotherapeutics on both cell lines. These effects partially reverted by exposure to α-bungarotoxin (α-BTX), an inhibitor of α7-nAChR. Nicotine addition to Caco-2 and HCT-8, treated with 5-FU or CPT, decreased the cleavage of substrate of caspase 3 and 7, poly-ADP-ribose polymerase (PARP). Moreover, P-ERK/ERK ratio was modified by nicotine addition to 5-FU and CPT treated cells in an opposite manner. However, when co-administrating PD98059, an ERK phosphorylation inhibitor, an increased apoptosis was observed. In Caco-2 and HCT-8 nicotine reverted 5-FU and CPT apoptotic effects through AKT phosphorylation, as demonstrated by apoptotic increase in presence of LY294002, an AKT phosphorylation inhibitor. Nicotine interfered with colorectal cancer pharmacological treatment in vitro by inhibiting apoptosis induced by chemotherapeutic drugs. Nicotine anti-apoptotic effects were exerted through ERK and AKT pathway activation.

Introduction

Colon cancer is one of the most common malignancies and a leading cause of cancer-related deaths in the western world (Jemal et al., 2010). Although surgical resection currently remains the primary mode of colon cancer treatment, chemotherapy can be provided after surgery as adjuvant to reduce tumor recurrence and prolong postoperative patient survival. Among the most common chemotherapeutic drugs used as adjuvant treatment of colon cancer are 5-Fluorouracil (5-FU) and Camptothecin (CPT) derivatives (Nordman et al., 2006). 5-FU is an analog of uracil with a fluorine atom at the C-5 position in place of hydrogen. It exerts its anticancer effects through inhibition of the thymidylate synthase activity and incorporation of its active metabolites into RNA and DNA (Longley et al., 2003). CPT, a cytotoxic alkaloid extracted from the Chinese tree Camptotheca acuminata, exerts its potent antitumor activity through inhibition of DNA topoisomerase I. However, since the high CPT toxicity and instability, many CPT derivatives have been developed and commonly used in anticancer therapy (Pommier, 2006). In vitro treatment of cancer cells, derived from different tissue types, with both 5-FU and CPT induces growth inhibition, cell cycle arrest, and apoptosis (Li et al., 2009, Li et al., 2003).

Cigarette smoking has been implicated in several human tumors, including colon cancer. Clinical studies showed that cancer patients who smoke have worse survival profiles than those who do not when they are treated with chemotherapeutic drugs (Cataldo et al., 2010). In advanced colorectal cancer patients, Vincenzi et al. (2009) showed that cigarette smoking, during anticancer treatment with a centuximab-based regimen, induced a reduction in the response rate and led to a lower time to progression. In particular, nicotine, one of the major components of cigarette smoking, could decrease the efficacy of certain drugs by increasing their metabolism through the induction of hepatic enzymes (Zevin and Benowitz, 1999) or by inhibiting the apoptotic potential of chemotherapeutic agents (Xu et al., 2007, Dasgupta et al., 2006, Shen et al., 2010). In a previous study we have shown that nicotine, at a concentration similar to that present in the bloodstream of smokers (Hukkanen et al., 2005), induced an increase in cell proliferation and suppression of apoptosis in two colon cancer cell lines, Caco-2 and HCT-8, cultured in standard and serum deprivation conditions (Cucina et al., 2012). Both these processes appear to be activated by the binding of nicotine to α7-nicotinic acetylcholine receptor (α7-nAChR) and likely mediated by the overexpression of pro-survival factors, as survivin and P-Bcl2, through the activation of both PI3K/AKT and PKC/ERK signaling pathways (Cucina et al., 2012).

An open issue, according these results, is whether nicotine could interfere with colorectal cancer pharmacological treatment, by inhibiting cell death. To assess this hypothesis, we evaluated herein the nicotine effect on colon cancer cell lines treated with 5-FU and CPT, two of the most commonly used chemotherapeutic drugs utilized as adjuvant treatment of colon cancer. Nicotine effects on cell proliferation and apoptosis have been therefore investigated in order to discover the possible molecular factors involved.

Section snippets

Cell culture

The human colorectal cancer cell lines Caco-2 and HCT-8 were obtained from European Collection of Cell Cultures (ECACC). Caco-2 and HCT-8 cells were seeded into 25-cm2 flasks (Falcon; Becton Dickinson Labware; Franklin Lakes NJ, USA) in Dulbecco modified Eagle medium (DMEM) supplemented with 10% Fetal Calf Serum (FCS) and antibiotics (Penicillin 100 IU/ml, Streptomycin 100 μg/ml, and Gentamycin 200 μg/ml all from Euroclone Ltd., Cramlington, UK). The cells were cultured at 37 °C in an atmosphere of

Effects of nicotine on cell proliferation

Caco-2 and HCT-8 cells, treated for 24, 48 and 72 h with either 5-FU or CPT, showed a marked suppression of proliferation in a time-dependent manner with respect to control cells cultured in a complete medium (Figs. 1A ,2A). Addition of 1 μM nicotine to the cells cultured in presence of either 5-FU or CPT induced a statistically significant increase of cell proliferation (Figs. 1B, C, 2B, C). Addition of α-BTX to Caco-2 and HCT-8 cells co-treated with chemotherapeutic drugs and nicotine induced a

Discussion

Cigarette smoking has been associated with increased incidence of colon cancer (Tsoi et al., 2009, Liang et al., 2009), and nicotine, the principal active component of tobacco smoke, has been shown to play a role in increasing cell proliferation in human colon carcinoma cells (Wong et al., 2007, Ye et al., 2004). Our previous study demonstrated that nicotine, at a concentration comparable to that found in plasma of active smokers, enhanced cell proliferation and hinders apoptosis in Caco-2 and

Conflict of Interest

All authors have no personal or financial conflicts of interest and they have not entered into any agreement that could interfere with our access to the data on the research or on our ability to analyze the data independently, to prepare manuscripts and to publish them.

Acknowledgement

No funding was received for the present study.

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