Elsevier

Biochemical Pharmacology

Volume 72, Issue 6, 14 September 2006, Pages 729-738
Biochemical Pharmacology

Expression of transporters potentially involved in the targeting of cytostatic bile acid derivatives to colon cancer and polyps

https://doi.org/10.1016/j.bcp.2006.06.007Get rights and content

Abstract

Drug targeting might help to overcome resistance to chemotherapy. Here we investigated whether colon cancer and polyps do express functional carriers involved in the uptake of cytostatic bile acid derivatives, in this case Bamet-UD2 [cis-diammine-bisursodeoxycholate-platinum(II)], which has been reported to be taken up by colon cancer cells “in vitro”, efficiently induce apoptosis and overcome resistance to cisplatin. Although at lower levels than in ileum, a detectable expression of ASBT, OATP8/1B3, OCT1 and OSTα in colon tissue was found, which was not impaired in colon cancer or polyps. The expression of OATP-A/1A2 and OSTβ was also found in colon, but this was markedly decreased in neoplastic colon tissue. In contrast, the expression of OATP-C/1B1 was low in colon but significantly enhanced in neoplastic colon tissue. MDR1 and MRP2 were poorly expressed in colon as compared with ileum, whereas MRP3 expression was higher in colon than in ileum. The abundance of mRNA for these ABC proteins was not changed in colon cancer or polyps. When RNA from different tissues was injected to Xenopus laevis oocytes their ability to take up taurocholate and Bamet-UD2 was enhanced (healthy ileum > healthy colon  neoplastic colon tissue). In all cases, uptake was lower for taurocholate than for Bamet-UD2, probably due to that ASBT mediates sodium-dependent uptake of both substrates, whereas additional transporters expressed in these tissues can participate in Bamet-UD2 uptake. In conclusion, our results suggest that the use of cytostatic bile acid derivatives might be a good pharmacological strategy for the treatment of colon tumors.

Introduction

A large proportion of all cancers is constituted by those affecting tissues included in the enterohepatic circuit, i.e., the liver, the gallbladder, the biliary tree and the intestine [1], [2], [3]. Moreover, these tumors usually have a poor prognosis, aggravated by the lack of response to the available chemotherapy. Among the attempts made to overcome resistance by gastrointestinal tumors to chemotherapy are investigations aimed to evaluate the possibility of using cytostatic bile acid derivatives to enhance intracellular drug concentrations in liver tumors [4]. In this line of research, several groups have synthesized different compounds and have explored whether they have the required characteristics, i.e., to inhibit tumor cell growth, to be less toxic than the parent drug, and to be efficiently taken up by transporters involved in the uptake of cholephilic organic anions by the liver [4]. One of the most interesting compounds fulfilling these requirements is cis-diammine-bisursodeoxycholate-platinum(II) (Bamet-UD2), which was obtained by chemical coupling of cisplatin to ursodeoxycholic acid [5], and which has been recently shown to be taken up by colon cancer cells “in vitro”, efficiently induce apoptosis and overcome resistance to cisplatin [6]. Thus, this compound has been used in the present study as a model drug to evaluate the possible interest of using bile acid derivatives in drug targeting to colon tumors.

The major transporter involved in active bile acid uptake by the intestinal mucosa is the apical sodium-dependent bile salt transporter or ASBT (gene symbol SLC10A2) [7]. The ability of this carrier to transport bile acid derivatives obtained by coupling the active agent to the hydroxyl group at C3 of the bile acid moiety has been suggested previously [8]. Here, we investigated whether bile acid derivatives, in which the side chain was used to link the cytostatic moiety, such as happens in Bamet-UD2, were also transported by this carrier.

Several members of a large family of Na+-independent carriers, the organic anion transporting polypeptides (OATPs), are able to transport bile acids. Since the nomenclature of these proteins has recently been changed [9], during a transitory period we have decided to use both the previous and new nomenclatures. In the present study, we investigated the presence in healthy and neoplastic intestinal tissue of three isoforms, namely OATP-A/1A2, OATP-C/1B1 and OATP8/1B3. These transporters, expressed in different tissues, are able to carry out the uptake of a broad spectrum of structurally unrelated compounds, such as bile acids and anionic conjugated and neutral steroids, such as ouabain [10]. However, some of them, such as the human OATP-A/1A2, also seem to be able to transport type II organic cations [11]. Such transport is also accomplished mainly by carrier systems known as organic cation transporters (OCTs), whose isoform 1 was included in the present study. These carriers have been found to be involved in the hepatic clearance of many organic cations, including several chemotherapeutic drugs [12].

An important aspect regarding the handling of cytostatic drugs by tumor cells is the existence of mechanisms of extrusion. Whether OATPs and OCTs might also mediate drug efflux from intestinal cells is not known. However, when expressed, the heterodimeric protein OSTα/OSTβ has been suggested to likely play a role as an efflux system in cells handling bile acids [13]. In addition, the extrusion of compounds may partly be due to ATP-binding cassette (ABC) proteins belonging, for instance, to the families of multidrug resistance proteins (MDR), such as MDR1, and multidrug resistance-associated proteins, such as MRP2 and MRP3. These proteins are already expressed in normal gastrointestinal epithelial cells [14] and are often over-expressed in several tumors derived from these cells [15]. When this occurs, these ABC proteins may play an important role contributing to the failure of chemotherapy by efficiently pumping the cytostatic agents, including cisplatin, out of these cells [16]. Among the aims of the present study, we also investigated the expression of these efflux transporters that may mediate the extrusion of bile acid derivatives from healthy and tumor intestinal cells.

Section snippets

Chemicals and samples

Cisplatin, sodium taurocholate (TC) and ursodeoxycholic acid were obtained from Sigma–Aldrich (Madrid, Spain). [3H]-TC (1.85 TBq/mmol) was purchased from American Radiolabeled Chemicals (Itisa Biomedica, Madrid). Bamet-UD2 [cis-diammine-bisursodeoxycholate-platinum(II)] was synthesized by binding two ursodeoxycholic acid molecules to cisplatin [17]. All other reagents were of analytical grade and were readily available from commercial sources.

Human tissue samples were the remains of biopsies

Results

To obtain indirect evidence of the ability of neoplastic colon tissue to take up cytostatic drugs synthesized by using bile acids as targeting elements, the expression of ASBT, the major carrier involved in active sodium-dependent intestinal absorption of bile acids, was investigated (Fig. 1). The mRNA of this transporter was detected in healthy colon at levels of less than 10% of those found in normal ileum. No significant decrease in ASBT expression in colon cancer and polyps was detected (

Discussion

The expression in certain tissues of transporters able to carry out an efficient and relatively specific uptake of structurally related compounds has been the basis of a promising strategy for tissue-selective drug delivery. This consists of coupling pharmacological active agents, such as cytostatic drugs, to shuttle molecules that can be recognized as substrates by these carrier proteins [19]. In this respect, the restricted expression of bile acid carriers able to mediate the uptake of bile

Conflict of interest statement

In the period of research leading up to this publication we have not received any financial support that may affect in any way the conclusions of our article. Moreover, the authors have no direct or indirect commercial interest in any company that might be financially affected by the conclusions of the present article.

Acknowledgements

The authors thank L. Muñoz, J.F. Martin and J. Villoria for the care of the animals. Secretarial help by M.I. Hernandez and revision of the English version of the article by N. Skinner are also gratefully acknowledged.

Financial support: This study was supported in part by the Instituto de Salud Carlos III, FIS (Grants CP03/00093 and PI051547), Spain; Junta de Castilla y León (Grant SA059A05), Spain; Ministerio de Ciencia y Tecnología, Plan Nacional de Investigación Científica, Desarrollo e

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