Abstract
BackgroundColorectal cancer (CRC) is a leading cause of cancer death globally and new biomarkers and treatments are severely needed.MethodsHere, 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.ResultsIn 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 [3H]-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 Cu2+-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.ConclusionsSLC1A1 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1405-8) contains supplementary material, which is available to authorized users.
Highlights
Colorectal cancer (CRC) is a leading cause of cancer death globally and new biomarkers and treatments are severely needed
The glutamate transporter inhibitor DL-Threo-β-Benzyloxyaspartic acid (DL-Threo-β-Benzyloxyaspartic acid (TBOA)) reduces chemotherapy-induced p53 induction and augments CRC cell death induced by SN38, while strongly attenuating that induced by oxaliplatin
We asked whether changes in SLC1A1 and SLC1A3 expression were involved in resistance development in HCT116 and LoVo cells
Summary
Colorectal cancer (CRC) is a leading cause of cancer death globally and new biomarkers and treatments are severely needed. Oxaliplatin is a diaminocyclohexane platinum derivative which induces formation of DNA adducts, and irinotecan is the precursor of the topoisomerase-I inhibitor 7-ethyl-10-hydroxycamptothecin (SN38) Both compounds induce DNA damage, upregulation of p53 and p21WAF1/Cip, cell cycle arrest, and cell death [9,10,11]. The majority of patients with metastatic CRC, whether on FOLFOX or FOLFIRI, will experience treatment resistance and disease progression upon treatment, leaving only limited additional treatment options. Possible remedies to this include the development of drugs that do not exhibit cross-resistance with those currently used, and of predictive biomarkers ensuring that patients receive the treatment with the highest likelihood of effect [5]. Progress has been made in recent years, strong biomarkers predicting response to oxaliplatin or irinotecan are lacking and urgently needed [3, 4, 12]
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