Cancer Letters

Cancer Letters

Volume 356, Issue 2, Part B, 28 January 2015, Pages 517-524
Cancer Letters

Original Articles
Specific expression of OATPs in primary small cell lung cancer (SCLC) cells as novel biomarkers for diagnosis and therapy

https://doi.org/10.1016/j.canlet.2014.09.025Get rights and content

Abstract

The expression of organic anion transporting polypeptides (OATPs) was elucidated in cell lines from small cell lung cancer (SCLC) and lung carcinoids and in paraffin-embedded samples from primary and metastatic SCLCs. We found a strong relationship between OATP expression and the origin of the cells, as cells from primary or metastatic SCLC and carcinoid tumors differ with respect to OATP levels. OATP4A1 is most prominent in non-malignant lung tissue and in all SCLC and carcinoid cell lines and tissues, OATP5A1 is most prominent in metastatic cells, and OATP6A1 is most prominent in SCLC cell lines and tumors. Treatment with topotecan, etoposide and cisplatin caused significant changes in the expression patterns of OATP4A1, OATP5A1, OATP6A1, chromogranin and synaptophysin. This effect was also evident in GLC-14 cells from an untreated SCLC patient before chemotherapy compared to GLC-16/-19 chemoresistant tumor cells from this patient after therapy. mRNA expression of OATP4A1, 5A1 and 6A1 correlates with protein expression as confirmed by quantitative microscopic image analysis and Western blots. OATPs might be novel biomarkers for tumor progression and the development of metastasis in SCLC patients.

Introduction

Lung cancer is the leading cause of cancer-related death in western countries and small-cell lung cancer (SCLC) accounts for 15%–20% of all lung cancer types [1]. SCLC is characterized by rapid tumor doubling time, a high growth fraction and the development of widespread metastases, especially to the brain, at a rather early stage [2]. In contrast to the majority of non-small cell lung cancers (NSCLC), SCLCs express neuroendocrine markers (e.g., chromogranin A, synaptophysin) and are thought to originate from lung neuronal precursor cells, from which other neuroendocrine lung tumors (e.g., carcinoids) are also derived [3], [4]. Importantly, SCLC cells, but not carcinoids, are usually highly responsive to initial chemotherapy, usually with a platin derivative-containing regimen in combination with etoposide. However, a subsequent relapse of SCLC mostly occurs within 2 years. Recurrent tumors are than often resistant or has only a mild response to further treatment [5], [6]. Only 10% of patients with advanced tumors survive longer than 2 years, leading to a five-year survival rate of only 3%–7% [7], [8], [9]. Therefore, clues to a more effective treatment for primary SCLC and its metastases are of utmost interest. In lung cancer, the role of transporters in drug absorption, distribution and elimination processes as well as in drug–drug interactions is increasingly being recognized [10]. One of the most important cellular uptake mechanisms for anticancer drugs and endogenous compounds is via the organic anion-transporting polypeptides (OATP proteins/SLCO genes) [11], [12]. The term OATP is used for both genes and proteins throughout the manuscript [13], [14], [15], [16], [17], [18]. Eleven human OATPs, divided into six distinct subfamilies (OATP1-6) [19], were found in human tissues mainly at biological barriers with a specific pattern [16], [20]. Some OATPs, such as OATP4A1, are widely expressed in the human body and were also identified at the mRNA level in bronchial epithelial cell models and might contribute to the uptake of solutes into lung cells [10], [21], [22], [23]. The malignant transformation of cells is known to alter the OATP expression pattern in organs. Indeed, the gonad-specific OATP6A1 was identified as a cancer-testis antigen in lung tumors and lung tumor cell lines. The altered uptake of OATP substrates including anticancer drugs may lead to changes in the activity of drugs and may therefore play a role in the chemosensitivity of cancer cells. For example, OATP5A1 seems to play a role in the resistance of lung cancer cell lines to satraplatin [20], [24]. Otherwise, treatment with chemotherapeutic agents alters the expression pattern of OATPs in the cells, which again influences the response to specific cancer chemotherapies. Therefore, this study aimed to investigate the OATP expression of SCLC cell lines. We used cell lines from primary and metastatic SCLC tumors as well as pulmonary carcinoid cells and performed immunofluorescence and immunohistochemistry on paraffin-embedded SCLC samples. Furthermore, the mRNA expression of OATP4A1, OATP5A1 and OATP6A1 was assessed in cell lines that were exposed to the chemotherapeutic drugs cisplatin, etoposide and topotecan, which are all applied in SCLC therapy [10], [21], [23]. Chromogranin A and synaptophysin were applied as markers for neuroendocrine differentiation of the tumor cells, and cadherin-1 was applied as a marker for the epithelial origin of cells [25], [26].

Section snippets

Reagents

Cisplatin, etoposide, and topotecan were obtained from Eubio (Vienna, Austria).

Other reagents and solvents (if not otherwise stated), obtained from Sigma (Munich, Germany), were of analytical grade.

Cancer cell lines and treatment

NCI-H417 cells, the NSCLC cell line A549 [25] and the two pulmonary carcinoid cell lines NCI-H727 and NCI-H835 [26] were obtained from the American Type Culture Collection (Manassas, VA, USA), DMS-153 cells from ECACC (Porton Down, Salisbury, UK). Other SCLC cell lines were obtained from Dr. Nina

OATP mRNA expression studies in lung tumor cell lines

The relative mRNA expression of OATPs was assessed in cells from primary SCLC tumors (DMS-114, NCI-H417), SCLC metastatic tumors (DMS-153, NCI-526), carcinoid cell lines (NCI-H727, NCI-H835) and cells isolated from pleural SCLC effusions (NCI-H69, SCLC26A) (Fig. 1A and 1B). Normal lung tissue, human bronchial epithelia (HBE) and the NSCLC cell line A549 [43] were used for comparison. We showed that the OATP4A1 gene was the most abundant OATP in normal lung tissue and in cancer cells (Fig. 1B,

Discussion

In the present study we elucidated the mRNA expression pattern of all 11 OATPs in primary and metastatic SCLC and lung carcinoid tumor cell lines in comparison to normal non-malignant lung tissue and non-malignant human broncho-epithelial cells. Then we evaluated the effect of chemotherapeutic agents on the expression of selected OATPs. OATP4A1 was most abundant in normal lung tissue and expressed in lung cancer cell lines derived from SCLC, carcinoid and NSCLC tumors. Similar to other OATPs,

Conflict of interest

The authors declare that they have no conflict of interest.

Acknowledgements

This study was supported by the Medical-Scientific Funds of the Mayor of the City of Vienna (P1004), and grants from the University of Vienna (“BioProMoTION” Bioactivity Profiling and Metabolism) and the Ludwig Boltzmann Society.

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