Cannabinoids increase lung cancer cell lysis by lymphokine-activated killer cells via upregulation of ICAM-1

This article originally appeared at https://www.sciencedirect.com/science/article/abs/pii/S0006295214004201

Authors

Institute of Toxicology and Pharmacology, University of Rostock, Rostock, Germany^bSection of Molecular Oncology and Immunotherapy, Department of General Surgery, University of Rostock, Rostock, Germany^cDepartment of Radiotherapy and Radiation Oncology, University of Rostock, Rostock, Germany

https://doi.org/10.1016/j.bcp.2014.07.014

Abstract

Cannabinoids have been shown to promote the expression of the intercellular adhesion molecule 1 (ICAM-1) on lung cancer cells as part of their anti-invasive and antimetastatic action. Using lung cancer cell lines (A549, H460) and metastatic cells derived from a lung cancer patient, the present study addressed the impact of cannabinoid-induced ICAM-1 on cancer cell adhesion to lymphokine-activated killer (LAK) cells and LAK cell-mediated cytotoxicity. Cannabidiol (CBD), a non-psychoactive cannabinoid, enhanced the susceptibility of cancer cells to adhere to and subsequently be lysed by LAK cells, with both effects being reversed by a neutralizing ICAM-1 antibody. Increased cancer cell lysis by CBD was likewise abrogated when CBD-induced ICAM-1 expression was blocked by specific siRNA or by antagonists to cannabinoid receptors (CB₁, CB₂) and to transient receptor potential vanilloid 1. In addition, enhanced killing of CBD-treated cancer cells was reversed by preincubation of LAK cells with an antibody to lymphocyte function associated antigen-1 (LFA-1) suggesting intercellular ICAM-1/LFA-1 crosslink as crucial event within this process. ICAM-1-dependent pro-killing effects were further confirmed for the phytocannabinoid Δ⁹-tetrahydrocannabinol (THC) and R(+)-methanandamide (MA), a hydrolysis-stable endocannabinoid analogue. Finally, each cannabinoid elicited no significant increase of LAK cell-mediated lysis of non-tumor bronchial epithelial cells, BEAS-2B, associated with a far less pronounced (CBD, THC) or absent (MA) ICAM-1 induction as compared to cancer cells. Altogether, our data demonstrate cannabinoid-induced upregulation of ICAM-1 on lung cancer cells to be responsible for increased cancer cell lysis by LAK cells. These findings provide proof for a novel antitumorigenic mechanism of cannabinoids.

Graphical abstract

Introduction

Cannabinoids have been demonstrated to exert anticarcinogenic effects via multiple mechanisms (for review see [1], [2]). However, despite increasing knowledge on cannabinoids’ effects on cancer cell apoptosis, invasion, metastasis and cancer-associated angiogenesis, its role in immunological antitumor responses is barely investigated. Most of the work published in this field has focussed on the modulation of immune cell migration by cannabinoids. Accordingly, the endocannabinoid 2-arachidonylglycerol has been demonstrated to induce the migration of natural killer cells [3], splenocytes, B lymphoid cells as well as myeloid leukaemia cells [4]. By contrast, Δ⁹-tetrahydrocannabinol (THC), the major psychoactive ingredient of marijuana, was associated with inhibition of antitumor immunity via a CB₂ receptor-mediated, cytokine-dependent pathway [5]. Other work addressed the impact of cannabinoids on the induction of cell death or apoptosis of diverse immune cell populations (for review see [6]). However, the susceptibility of cancer cells to the cytolytic action of lymphokine-activated killer (LAK) cells in response to cannabinoid treatment has not been studied as yet. In line with this notion, comprehensive in vitro studies addressing the mechanisms by which anticancer drugs elicit tumor-immune interactions have not been published so far.

Recently, our group has shown that the cannabinoids cannabidiol (CBD), THC as well as R(+)-methanandamide (MA), a hydrolysis-stable anandamide analogue, induce the expression of the intercellular adhesion molecule 1 (ICAM-1) in several lung cancer cells as well as in metastatic cells of a lung cancer patient thereby conferring increased levels of the tissue inhibitor of matrix metalloproteinases-1 (TIMP-1) and a subsequent decreased cancer cell invasiveness [7]. In athymic nude mice the non-psychoactive CBD elicited an increase of ICAM-1 and TIMP-1 protein in A549 xenografts and an antimetastatic effect that was fully reversed by a neutralizing antibody against ICAM-1 [7]. Apart from this and other evidence [8] pointing to a function for ICAM-1 in cellular signal transduction pathways, the glycoprotein containing 5 extracellular immunoglobulin-like domains, a transmembrane and a C-terminal intracellular domain [9] is known to play a crucial role as an adhesion molecule in trafficking of inflammatory cells and in cell-to-cell interactions during antigen presentation [8].

In context with the endogenous tumor immune surveillance an increasing number of data suggests a functional role of ICAM-1 on the cancer cell surface. Accordingly, several studies revealed increased tumor susceptibility to lymphocyte adhesion and cell-mediated cytotoxicity following transfection or upregulation of ICAM-1 [10], [11], [12], [13]. Vice versa, downregulation of ICAM-1 by transforming growth factor β₁ has been demonstrated to decrease both lymphocyte adhesion to cancer cells as well as cytotoxicity on cancer cells [14]. In line with this notion, ICAM-1 expression has been reported to be negatively correlated to metastasis of several cancer types in clinical studies [15], [16], [17].

In view of this data the present study addressed the impact of cannabinoids on tumor immune surveillance with special reference to the role of ICAM-1 in this response. Here we demonstrate that cannabinoid-induced upregulation of ICAM-1 on lung cancer cells is responsible for increased cancer cell susceptibility to LAK cell-mediated cytolysis. These findings provide a novel mechanism within the diverse antitumorigenic effects of cannabinoids.

Section snippets

Materials

AM-251, AM-630 and leupeptin were bought from Biomol (Hamburg, Germany). Aprotinin, calcein-AM, capsazepine, p-coumaric acid, luminol, orthovanadate and phenylmethylsulfonyl fluoride (PMSF) were purchased from Sigma–Aldrich (Taufkirchen, Germany). (−)-CBD was supplied by Biotrend AG (Cologne, Germany). THC and MA were bought from Lipomed (Weil am Rhein, Germany) and Tocris Bioscience (Wiesbaden-Nordenstadt, Germany), respectively. Dimethyl sulfoxide (DMSO), ethylenediaminetetraacetic acid

Role of ICAM-1 in CBD-induced adhesion of lung cancer cells to LAK cells

In line with the data published recently by our group [7] CBD was found to induce the expression of ICAM-1 protein in both lung cell lines (A549, H460) as well as in metastatic cells derived from a lung cancer patient (Fig. 1A–C, left).

To investigate a possible proadhesive action of CBD and a potential involvement of ICAM-1 in this response, calcein AM-labeled LAK cells were added to cancer cells. Adherent LAK cells were quantified as calcein fluorescence released by remaining and thus adherent

Discussion

Recently, ICAM-1 was identified as an essential link within the signal transduction pathway conferring the TIMP-1-dependent anti-invasive action of the cannabinoids CBD, THC and MA on human lung tumor cells [7]. Using the same cell lines (A549, H460) as well as metastatic cells from a lung cancer patient, the present study analyzed the contribution of cannabinoid-induced ICAM-1 on cancer cell susceptibility to cytolytic LAK cells. As a major result the data from the present study suggest

Acknowledgments

The authors are grateful to Prof. Dr. V. Kiefel and his associates of the Institute of Transfusion Medicine, Medical Faculty, University of Rostock, for providing the buffy coats.

References (50)

A.K. Hubbard et al.Intercellular adhesion molecule-1 (ICAM-1) expression and cell signaling cascadesFree Radic Biol Med(2000)
A.M. Hopkins et al.ICAM-1 targeted docking for exogenous as well as endogenous ligandsAdv Drug Deliv Rev(2004)
S. Mukherjee et al.Species comparison and pharmacological characterization of rat and human CB2 cannabinoid receptorsEur J Pharmacol(2004)
R. Ramer et al.Cannabidiol inhibits cancer cell invasion via upregulation of tissue inhibitor of matrix metalloproteinases-1Biochem Pharmacol(2010)
W.M. Sartor et al.Enhanced expression of ICAM-1 in a murine fibrosarcoma reduces tumor growth rateJ Surg Res(1995)
A.T. Lefor et al.Enhanced cytolytic activity of tumor infiltrating lymphocytes (TILs) derived from an ICAM-1 transfected tumor in a murine modelJ Surg Res(1998)
S.B. Hosch et al.Expression and prognostic significance of HLA class I. ICAM-1, and tumor-infiltrating lymphocytes in esophageal cancerJ Gastrointest Surg(1997)
P. Sacerdote et al.The nonpsychoactive component of marijuana cannabidiol modulates chemotaxis and IL-10 and IL-12 production of murine macrophages both in vivo and in vitroJ Neuroimmunol(2005)
R.J. McKallip et al.Targeting CB2 cannabinoid receptors as a novel therapy to treat malignant lymphoblastic diseaseBlood(2002)
N. Freimuth et al.Antitumorigenic effects of cannabinoids beyond apoptosisJ Pharmacol Exp Ther(2010)

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