Journal Article

The glutamate transport inhibitor DL-Threo-β-Benzyloxyaspartic acid (DL-TBOA) differentially affects SN38- and oxaliplatin-induced death of drug-resistant colorectal cancer cells

Romer, Maria Unni, Jensen, Anders A., Stenvang, Jan, Jensen, Niels Frank, Pedraz-Cuesta, Elena, Brünner, Nils, Christensen, Sandra, Pedersen, Stine Falsig, Bunch, Lennart

2015

Overview

Background Colorectal cancer (CRC) is a leading cause of cancer death globally and new biomarkers and treatments are severely needed. Methods Here, we employed HCT116 and LoVo human CRC cells made resistant to either SN38 or oxaliplatin, to investigate whether altered expression of the high affinity glutamate transporters Solute Carrier (SLC)-1A1 and -1A3 (EAAT3, EAAT1) is associated with the resistant phenotypes. Analyses included real-time quantitative PCR, immunoblotting and immunofluorescence analyses, radioactive tracer flux measurements, and biochemical analyses of cell viability and glutathione content. Results were evaluated using one- and two-way ANOVA and Students two-tailed t- test, as relevant. Results In SN38-resistant HCT116 and LoVo cells, SLC1A1 expression was down-regulated ~60 % and up-regulated ~4-fold, respectively, at both mRNA and protein level, whereas SLC1A3 protein was undetectable. The changes in SLC1A1 expression were accompanied by parallel changes in DL-Threo-β-Benzyloxyaspartic acid (TBOA)-sensitive, UCPH101-insensitive [ 3 H]-D-Aspartate uptake, consistent with increased activity of SLC1A1 (or other family members), yet not of SLC1A3. DL-TBOA co-treatment concentration-dependently augmented loss of cell viability induced by SN38, while strongly counteracting that induced by oxaliplatin, in both HCT116 and LoVo cells. This reflected neither altered expression of the oxaliplatin transporter Cu 2+ -transporter-1 (CTR1), nor changes in cellular reduced glutathione (GSH), although HCT116 cell resistance per se correlated with increased cellular GSH. DL-TBOA did not significantly alter cellular levels of p21, cleaved PARP-1, or phospho-Retinoblastoma protein, yet altered SLC1A1 subcellular localization, and reduced chemotherapy-induced p53 induction. Conclusions SLC1A1 expression and glutamate transporter activity are altered in SN38-resistant CRC cells. Importantly, the non-selective glutamate transporter inhibitor DL-TBOA reduces chemotherapy-induced p53 induction and augments CRC cell death induced by SN38, while attenuating that induced by oxaliplatin. These findings may point to novel treatment options in treatment-resistant CRC.

• Publisher: BioMed Central
• Subject: Amino Acid Transport System X-AG - antagonists & inhibitors / Amino Acid Transport System X-AG - genetics / Amino Acid Transport System X-AG - metabolism / Antineoplastic Agents - pharmacology / Aspartic Acid - pharmacology / Biomedical and Life Sciences / Biomedicine / Camptothecin - analogs & derivatives / Camptothecin - pharmacology / Cancer Research / Cell Death - drug effects / Cell Line, Tumor / Cell Survival - drug effects / Colorectal Neoplasms - genetics / Colorectal Neoplasms - metabolism / Copper - metabolism / Drug Resistance, Neoplasm / Excitatory Amino Acid Transporter 1 - genetics / Excitatory Amino Acid Transporter 1 - metabolism / Excitatory Amino Acid Transporter 3 - genetics / Excitatory Amino Acid Transporter 3 - metabolism / Experimental therapeutics and drug development / Gene Expression / Gene Knockdown Techniques / Glutathione - metabolism / HCT116 Cells / Health Promotion and Disease Prevention / Humans / Medicine/Public Health / Oncology / Organoplatinum Compounds - pharmacology / Protein Transport / Research Article / Surgical Oncology / Tumor Suppressor Protein p53 - metabolism