Differential Expression of Multi Drug Resistance-Associated Protein (MRP 1) in phenotypically distinct neuroblastoma cell lines

Sonal Kashyap

Introduction

Neuroblastoma is the most common extracranial solid tumor of infancy which arises from embryonal neural crest cells. Neuroblastomas are categorized clinically by their location, age at diagnosis, spread/metastasis and degree of cellular maturation and heterogeneity. Long term studies undertaken in human cell lines have shown that there are three distinct neuroblastoma cell types—I-type stem cells, N-type neuroblastic/neuroendocrine precursors, S-type schwannian cell/melanoblastic precursors(1). The I-type stem cells are malignant neural crest cells and form tumors in the nude mice. The I-type cells show features common to both N and S type cells. The N-type cells adhere tightly to other cells forming structures like pseudoganglia and they form tumors only in athymic mice. The S type cells adhere tightly to the substrate and do not form tumors in nude mice. Studies have also shown that neuroblastomas contain multiple cell phenotypes and this feature is often used for the prognosis of the disease. I-cell types are present in tumors of all stages. Higher incidence of I type cells in most neuroblastoma is consistent with their high malignant potential in vitro (1, 6). The phenotypically distinct neuroblastoma cell lines have been reported to show varying levels and differential expression of MYCN gene. MYCN is the most widely used biomarker for neuroblastoma, higher the expression of MYCN, poorer is the disease prognosis. MYCN belongs to class of bHLH leucine zipper transcription factors which are involved in cell differentiation, apoptosis, cell growth and genomic instability (4). Some N and I type cell lines have higher levels of MYCN and some show overexpression of the MYCN gene. In the S-type cell line MYCN expression is reported to be down regulated (5, 6).
Some malignant neuroblastomas display chemoresistance. One of the factors causing chemoresistance in these tumors is the expression of Multidrug Resistance-associated Protein (MRP). MRP is overexpressed in most malignant and chemoresistant neuroblastomas. It is an ATP binding cassette protein which acts as an efflux pump. MRP is expressed at normally in many tissues and plays a significant role in efflux of unconjugated bilirubin in fetal hepatocytes and efflux of heavy metal ions from other tissues. Overexpression of MRP in different tumors results in pumping out of unrelated hydrophobic drugs from the cancer cells thereby resulting in chemotherapy failure. Overexpression of MRP have been reported in many cancers—Leukemia and Neuroblastoma. Alternative splicing of MRP gene can generate many variants.
It has been reported that MYCN may upregulate the expression of MRP gene, especially in the poor prognostic tumor types (2, 3, and 4). There is considerable literature demonstrating concomitant overexpression of MRP with MYCN gene in human neuroblastoma. It is possible that MYCN may act as transcriptional factor in regulating MRP gene expression but the exact mechanism still needs to be elucidated(2). Amplification of the MYCN gene is not always the primary factor involved in the neuroblastoma malignancy, presence of distinct cell phenotypes can also affect cancer prognosis. Differential cell lineages and states within a particular lineage have been reported to modulate the MYCN expression (5). The main objective of the present project was whether the MRP-1 expression correlates with a) overexpression of MYCN gene, or b) correlates with distinct neuroblastoma cell types, Resolution of this matter could help physicians select a more effective treatment of neuroblastoma.