www.elsevier.com/locate/jneuroim

Journal of Neuroimmunology 147 (2004) 95–98

Cocaine modulates cytokine and enhances tumor growth

through sigma receptors

Brian Gardnera,b,*, Li X. Zhua, Michael D. Rotha,c, Donald P. Tashkina,c,Steven M. Dubinetta,b,c, Sherven Sharmaa,b,*

aDivision of Pulmonary and Critical Care Medicine, David Geffen School of Medicine at the University of California-Los Angeles,

Los Angeles, CA 90095, USAbDivision of Pulmonary and Critical Care Medicine, Veteran Administration Greater Los Angeles Healthcare System, Los Angeles, CA 90073, USA

cUCLA Lung Cancer Research Program of the Jonsson Comprehensive Cancer Center, Los Angeles, CA 90095, USA

Abstract

Sigma receptors are intracellular receptors that interact with a variety of psychotropic ligands, including cocaine. Administration of

cocaine to mice promoted the in vivo growth of a syngeneic lung cancer cell line and identical effects were observed with PRE 084, a

selective sigma1 receptor agonist. Increased tumor growth was accompanied by an increase in IL-10 and a decrease in IFN-g production in

splenocytes and at the tumor site. The tumor-promoting effects produced by both cocaine and PRE 084 were abrogated by administration of

specific antibodies to IL-10, or by administration of a sigma1 receptor antagonist. We conclude that sigma1 receptor ligands, including

cocaine, augment tumor growth via a cytokine-dependent, receptor-mediated mechanism that involves regulation of T helper 1/T helper 2

cytokine balance.

D 2003 Elsevier B.V. All rights reserved.

Keywords: Cocaine; Sigma receptor; Tumor; Cytokine; Interleukin-10; Immunity

1. Introduction

Sigma receptors are a unique family of intracellular

receptors found in neuronal tissues, cells of the immune

system, (Liu et al., 1995; Ganapathy et al., 1999) and also in

human and murine tumors (John et al., 1995; Vilner et al.,

1995). These receptors interact with a variety of compounds

including cocaine, dextromethorphan and benzomorphans

(Walker et al., 1990). At least two subtypes of sigma

receptors, classified as sigma1 and sigma2, are distinguish-

able by their physiological function and pharmacological

response.

Sigma ligands have potent regulatory effects on cyto-

kines including the induction of IL-10 (Bourrie et al.,

0165-5728/$ - see front matter D 2003 Elsevier B.V. All rights reserved.

doi:10.1016/j.jneuroim.2003.10.020

* Corresponding authors. UCLA Lung Cancer Research Program,

David Geffen School of Medicine at UCLA, 37-131 Center for Health

Sciences, 10833 Le Conte Avenue, Los Angeles, CA 90095-1690, USA.

Tel.: +1-310-478-3711x41863; fax: +1-310-268-4164.

E-mail addresses: bgardner@mednet.ucla.edu (B. Gardner),

sharmasp@ucla.edu (S. Sharma).

1995) and the suppression of IFN-g and GM-CSF (Carayon

et al., 1995). In murine studies, treatment with sigma

ligands prevented both graft-versus-host disease and

delayed-type hypersensitivity granuloma formation (Car-

ayon et al., 1995). These studies indicate that sigma

receptor signaling can regulate immune responses. Co-

caine has been reported to be a sigma1 receptor ligand

and to modulate immune function in vivo and in vitro

(Pellegrino and Bayer, 1998; Xu et al., 1999). Cocaine

down-regulates the production of IFN-g by human pe-

ripheral blood leukocytes and increases the production of

TGF-h by endothelial cells and macrophages (Mattana et

al., 1994; Mao et al., 1996; Mao et al., 1997). Alveolar

macrophages isolated from crack cocaine smokers exhibit

depressed antimicrobial activity compared to nonsmokers

(Baldwin et al., 1997). Their capacity to kill Staphylo-

coccus aureus was restored by the addition of exogenous

GM-CSF or IFN-g, suggesting that exposure to crack

cocaine inhibited the production of these cytokines in

vivo (Shay et al., 2003). Cocaine also enhances HIV-1

replication in stimulated peripheral blood mononuclear

cells in a TGF-h dependent manner (Peterson et al.,

Table 1

Enhanced in vivo tumor growth produced by administration of cocaine or

PRE 084 was abrogated by BD1047, a selective sigma1 receptor antagonist

Groupa Pretreatment

with BD1047bTumor size at

day 21 (cm3)cP

valued

Control � 817F 60

Cocaine-treated � 1130F 58 < 0.01

+ 803F 70 < 0.01

PRE 084-treated � 1143F 16 < 0.01

+ 740F 124 < 0.01

a Mice were pretreated 5 days per week for 2 weeks with saline

(control), cocaine (5 mg/kg), or PRE 084 (20 mg/kg), implanted with 105

LIC2 tumor cells by subcutaneous injection, and then continued on drug

B. Gardner et al. / Journal of Neuroimmunology 147 (2004) 95–9896

1991). Thus, sigma ligands such as cocaine might pro-

mote a Th2 type immune response capable of regulating

immune function in vivo. To investigate this hypothesis,

we evaluated the capacity for both cocaine and another

well-characterized sigma ligand to regulate antitumor

immunity, cytokine production, and tumor growth in vivo

in a murine tumor model. These studies reveal that

cocaine, acting via a sigma1 receptor pathway, modulated

the balance of T helper type 1 (Th1) and type 2 (Th2)

cytokines, suppressed antitumor immune responses and,

as a consequence of these effects, enhanced the rate of

tumor growth.

treatment.

b Mice were either pretreated with BD1047 (20 mg/kg) or saline prior to

each injection of drug.c Tumor size was measured at 21 days after tumor implantation and

volume recorded from measurement with calipers in three dimensions.d P value comparing treatment group to control animals receiving saline

alone or treatment groups compared to treatment plus BD 1047. N = 6 per

group.

2. Results and discussion

2.1. Sigma1 receptor agonists promote tumorigenicity in

immunocompetent mice

To determine how sigma1 receptor agonists modulate

tumor growth, we evaluated their effects on the growth of

weakly immunogenic Line 1 alveolar cell carcinoma

(L1C2, H-2d) cells in vivo in BALB/c mice. Before

implantation of tumors, mice were pretreated for 2 weeks

with intraperitoneal injections of cocaine (5 mg/kg,

obtained from NIDA), a selective sigma1 receptor agonist,

PRE 084 (20 mg/kg; TOCRIS Cookson, Ellisville, MO),

or diluent control (saline) five times per week. L1C2

tumor cells (105) were then inoculated subcutaneously

and tumor growth over time was compared in mice

receiving continued injections with cocaine or PRE 084

as compared that observed in diluent-treated controls.

There was a significant enhancement of L1C2 tumor

growth in mice treated with PRE 084 or cocaine,

compared to control mice. The cocaine-mediated increase

in tumor growth was sigma receptor-dependent as dem-

onstrated by treatment with BD1047 (20 mg/kg; TOCRIS

Cookson), a sigma receptor antagonist that abrogated the

increase in tumor growth (Table 1). Treatment with

BD1047 alone did not alter tumor growth.

2.2. Cocaine and sigma receptor agonists regulate Th1/Th2

cytokine profiles

Prior studies demonstrated that sigma1 receptor ligands

modulate cell-mediated immune responses (Wang et al.,

1994; Bourrie et al., 1995; Carayon et al., 1995; Derocq

et al., 1995; Liu et al., 1995), leading us to speculate that

treatment with sigma1 receptor ligands might act by limiting

the production of Th1 cytokines and increasing the produc-

tion of Th2 cytokines. This shift in cytokine profile can limit

effective immune responses to a tumor challenge and

thereby allow increased tumor growth. We recently reported

(Zhu et al., 2003) that cocaine treatment resulted in approx-

imately a twofold decrease in IFN-g and twofold increase in

IL-10 production at the tumor site compared to diluent

treatment. Splenocytes isolated from cocaine-treated, non-

tumor-bearing mice showed a similar shift in cytokine

pattern (Zhu et al., 2003). Thus, sigma receptor ligands

such as cocaine can result in a shift towards a Th2 dominant

immune response (Fig. 1). In addition to changes in IL-10

and IFN-g production, we found that cocaine administration

augments the production of the immune suppressive cyto-

kine TGF-h. Splenocyte TGF-h production increased two-

fold in cocaine-treated compared to diluent-treated tumor-

bearing mice (2.2 vs. 1.0 ng/ml/106 cells).

2.3. Administration of anti-IL-10 mAb prevents the cocaine

mediated increase in tumor growth in vivo

Based on the known detrimental effects of IL-10 in this

tumor model (Huang et al., 1998), we speculated that the

induction of IL-10 by sigma1 receptor ligands was respon-

sible for the observed enhancement of tumor growth in vivo.

IL-10 inhibits a broad array of immune parameters that

include antigen presentation, antigen-specific T cell prolif-

eration (de Waal-Malefyt et al., 1991; Taga and Tosato,

1992), and type 1 cytokine production (Fiorentino et al.,

1991; Mosmann and Moore, 1991). IL-10 is also a potent

inhibitor of host immunity (Rohrer and Coggin, 1995;

Sharma et al., 1999) and may act at several points to

interfere with either the generation or maintenance of

antitumor immune responses. Neutralizing anti-IL-10 mAb

was therefore administered to mice receiving PRE 084 or

cocaine in order to block the biological effects resulting

from the heightened production of IL-10. Administration of

anti-IL-10 mAb, but not control antibody, prevented these

sigma1 receptor agonists, including both cocaine and PRE

084, from increasing the rate of tumor growth.

The production of immunoregulatory cytokines is a

critical element in the generation of an effective immune

response. The tumor-bearing state is associated with an up-

Fig. 1. Cocaine or the sigma receptor agonist PRE 084 promote a Th1 cytokine profile. Mice were treated with PRE 084, cocaine or diluent control using the

dosing and injection schedule as described in the text. L1C2 tumor cells (105) were injected subcutaneously and non-necrotic tumors recovered from L1C2

tumor-bearing mice after 4 weeks of continued drug treatment. Tumors were homogenized and the concentrations of IL-10 and IFN-g in the resulting

supernatant determined by cytokine-specific ELISA (N= 6 per group). Mean values of the ratio of IFN-g to IL-10 (F S.E.M.) are shown.

B. Gardner et al. / Journal of Neuroimmunology 147 (2004) 95–98 97

regulation of immune suppressive cytokines including IL-10

and TGF-h (Alleva et al., 1994; Maeda and Shiraishi, 1996).

As a result, host immunity against tumors is down-regulated

(Huang et al., 1998). Tumors may either directly release

suppressive cytokines or orchestrate host immune cells to

produce them (Huang et al., 1998; Seo et al., 1999).

Consistent with these conclusions, we previously found that

production of IL-10 by cutaneous carcinomas provides a

mechanism for evasion of T cell immunity (Kim et al.,

1995). We also found that transgenic mice over-expressing

IL-10 under the control of the IL-2 promoter were unable to

limit the growth of immunogenic tumors (Hagenbaugh et

al., 1997). Administration of blocking IL-10 mAbs restored

in vivo antitumor responses in these transgenic mice. Our

finding that sigma1 receptor agonists, such as cocaine,

increased the release of IL-10 in vivo at the tumor site,

and increased lymphocyte IL-10 production in vitro, sug-

gested a mechanism by which these agents exaggerate

tumor-induced immune suppression and promoted tumor

growth. The capacity for anti-IL-10 mAb to reverse the

detrimental effects produced by sigma1 receptor ligands

lends strong support to this hypothesis.

These findings may have important clinical implications

for both medically prescribed drugs as well as those used in

abuse. For example, cocaine smokers have increased histo-

pathologic abnormalities in their lungs including a higher

incidence of hyperplasia and squamous cell metaplasia

compared to nonsmokers (Fligiel et al., 1997). In addition,

cocaine smokers have elevated expression of molecular

markers that are associated with increased cancer risk such

as Ki-67 and epidermal growth factor receptor (Barsky et al.,

1998). Thus cocaine smoking, like tobacco, has been impli-

cated in field cancerization of bronchial epithelial cells

(Barsky et al., 1998). These findings raise a concern that

the combination of these genetic alterations and sigma

ligand-induced immune suppression may have additive or

synergistic effects in promoting tumorigenesis. In addition,

there are epidemiological studies that associate cocaine

abuse with an increase in opportunistic infections and

progression of HIV (Chaisson et al., 1989; Caiaffa et al.,

1994). In an experimental model of AIDS, we have found

that cocaine administration significantly enhanced HIV rep-

lication (Roth et al., 2002). Further studies are warranted to

determine if immune suppression, opportunistic infections,

and cancer potentiation result from abuse of crack cocaine

and whether these in vivo effects related to activation of

sigma receptors.

Acknowledgements

This work was supported by National Institutes of Health

Grants R01 DA08254, RO1 CA71818, and P50 90388, and

the Merit Review Research Funds from the Department of

Veterans Affairs.

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