MRP3: a molecular target for human glioblastoma multiforme immunotherapy.

Abstract

BackgroundGlioblastoma multiforme (GBM) is refractory to conventional therapies. To overcome the problem of heterogeneity, more brain tumor markers are required for prognosis and targeted therapy. We have identified and validated a promising molecular therapeutic target that is expressed by GBM: human multidrug-resistance protein 3 (MRP3).MethodsWe investigated MRP3 by genetic and immunohistochemical (IHC) analysis of human gliomas to determine the incidence, distribution, and localization of MRP3 antigens in GBM and their potential correlation with survival. To determine MRP3 mRNA transcript and protein expression levels, we performed quantitative RT-PCR, raising MRP3-specific antibodies, and IHC analysis with biopsies of newly diagnosed GBM patients. We used univariate and multivariate analyses to assess the correlation of RNA expression and IHC of MRP3 with patient survival, with and without adjustment for age, extent of resection, and KPS.ResultsReal-time PCR results from 67 GBM biopsies indicated that 59/67 (88%) samples highly expressed MRP3 mRNA transcripts, in contrast with minimal expression in normal brain samples. Rabbit polyvalent and murine monoclonal antibodies generated against an extracellular span of MRP3 protein demonstrated reactivity with defined MRP3-expressing cell lines and GBM patient biopsies by Western blotting and FACS analyses, the latter establishing cell surface MRP3 protein expression. IHC evaluation of 46 GBM biopsy samples with anti-MRP3 IgG revealed MRP3 in a primarily membranous and cytoplasmic pattern in 42 (91%) of the 46 samples. Relative RNA expression was a strong predictor of survival for newly diagnosed GBM patients. Hazard of death for GBM patients with high levels of MRP3 RNA expression was 2.71 (95% CI: 1.54-4.80) times that of patients with low/moderate levels (p = 0.002).ConclusionsHuman GBMs overexpress MRP3 at both mRNA and protein levels, and elevated MRP3 mRNA levels in GBM biopsy samples correlated with a higher risk of death. These data suggest that the tumor-associated antigen MRP3 has potential use for prognosis and as a target for malignant glioma immunotherapy.

Prognostic significance of MRP5 immunohistochemical expression in glioblastoma

“…Nine MRP members have been identiWed so far. In gliomas, MRP1, 3, 4 and 5 have been reported to be expressed more than the other members [9,10]. Benyahia et al studied the MRP1 expression in surgical tumor samples from 17 patients with gliomas.…”

“…Calatozzolo et al [13] found immunohistochemical expression of glutathione-S-transferase pi, MRP1, MRP5, Pgp and MRP3 in glioma cells. Recently, Kuan et al [10] using real-time PCR and immunohistochemistry demonstrated that that the MRP3 gene is expressed both at the mRNA level and at the protein level in high-grade gliomas, but minimally in normal brain tissue. Patients with GBMs that expressed elevated MRP3 mRNA levels in tumor biopsy samples had a higher risk of death.…”

“…MRP1, MRP3 and MRP5 have been reported to be expressed more than the other members in gliomas [12][13][14]. Although MRP1 and MRP3 have been studied in glioma patients, little is known regarding the signiWcance of MRP5 expression [10,15]. In the present study, we investigated the prognostic signiWcance of MRP5 expression in patients with GBM.…”

“…In patients with GBM, epigenetic silencing via promoter hypermethylation of the DNA repair gene O6-methylguanine-DNA methyltransferase (MGMT) has been linked to better response to alkylating agents and longer survival [3]. Apart from MGMT, various other genes have been implicated in chemoresistance, among them MDR1 and MRPs [4][5][6][7][8][9][10]. The MDR1 gene encodes the P-glycoprotein (P-gp), which is an ATP-dependent drug eZux pump.…”

Recent Approaches and Novel Therapeutic Targets in Human Glioma

“…From the initial identification of chromosomal aberrations by karyotyping and comparative genomic hybridization, we have rapidly moved to expression array studies and to integrative genomic approaches which have allowed the stratification of several human brain tumors into molecular subgroups (Claudia et al, 2013;Kuan et al, 2010;Gerstner et al, 2009;Hormigo et al, 2011;Mischel et al, 2003). These data have not only increased our understanding of the molecular pathogenesis of human brain tumors, but have also identified prognostic markers and opened new avenues for targeted therapies.…”