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Pharmacological Ascorbate Suppresses Growth of Gastric Cancer Cells with GLUT1 Overexpression and Enhances the Efficacy of Oxaliplatin Through Redox Modulation.

ABSTRACT: Rationale: The antitumor activity of high-dose ascorbate has been re-evaluated recently, but the mechanism underlying cell-specific sensitivity to ascorbate has not yet been clarified. Methods: The effects of high-dose ascorbate on gastric cancer were assessed using cancer cell lines with high and low expression of GLUT1 via flow cytometry and colony formation assays in vitro and patient-derived xenografts in vivo. Results: In this study, we demonstrated that gastric cancer cells with high GLUT1 expression were more sensitive to ascorbate treatment than cells with low GLUT1 expression. GLUT1 knockdown significantly reversed the therapeutic effects of pharmacological ascorbate, while enforced expression of GLUT1 enhanced the sensitivity to ascorbate treatment. The efficacy of pharmacological ascorbate administration in mice bearing cell line-based and patient-derived xenografts was influenced by GLUT1 protein levels. Mechanistically, ascorbate depleted intracellular glutathione, generated oxidative stress and induced DNA damage. The combination of pharmacological ascorbate with genotoxic agents, including oxaliplatin and irinotecan, synergistically inhibited gastric tumor growth in mouse models. Conclusions: The current study showed that GLUT1 expression was inversely correlated with sensitivity of gastric cancer cells to pharmacological ascorbate and suggested that GLUT1 expression in gastric cancer may serve as a marker for sensitivity to pharmacological ascorbate.

SUBMITTER: Lu YX

PROVIDER: S-EPMC5835938 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Abstract Pharmacological ascorbate (P-AscH-; high dose intravenous infusions of vitamin C generating milli-molar plasma concentrations) has re-emerged as an anti-cancer therapy. Phase 1 clinical trials combining P-AscH- with chemotherapy and ionizing radiation demonstrate safety and promising clinical outcomes in a variety of malignancies. In a first-in-human trial, subjects with newly diagnosed glioblastoma (GBM) and undetectable MGMT promoter methylation were treated with P-AscH-, ionizing radiation, and temozolomide. Results demonstrate median progression-free survival (PFS) of 10 months and median overall survival (OS) of 23 months, comparing favorably to historical GBM patients expressing MGMT. P-AscH-‘s anti-cancer mechanism is dependent upon the presence of redox active labile iron. In the presence of redox active iron, the formation of hydrogen peroxide, which causes oxidative stress and eventual cell death, selectively forms in cancer cells. Treatment with P-AscH- increased cancer cells’ labile iron pool, further enhancing sensitivity to P-AscH-. We investigated the capability of MR imaging (T2* relaxation time) to measure the redox active iron and predict response to P-AscH-. T2* relaxation time is influenced by in-field inhomogeneities, such as redox active paramagnetic iron. The active phase 2 trial evaluating P-AscH-, radiation, and temozolomide for GBM, obtains T2* imaging prior to (baseline) and immediately after ascorbate infusion (NCT02344355). A preliminary analysis of the baseline scan for the first 15 subjects suggests those with faster GBM T2* relaxation times (≤ 58 ms) have more redox active labile iron pools as well as an improved median PFS (11.4 months) compared to those with slower T2* relaxation times (> 58 ms; median PFS of 8.5 months). Pre-clinical studies evaluating the effectiveness of iron nano-particle supplementation in GBM animal models are on-going. (Supported by P01 CA217797, R01 CA169046, U01 CA140206, T32 CA078586, P30 CA086862, as well as the Gateway for Cancer Research grant G-17–1500.)

Project description:Gastric adenocarcinoma most often presents at an advanced stage and overall five-year survival of ∼30%. Pharmacological ascorbate (high-dose IV ascorbate) has been proposed as a promising nontoxic adjuvant to standard radio-chemotherapies in several cancer types. In the current study, pharmacological ascorbate (0.5-2 m M) caused a dose-dependent decrease (70-85% at 2 m M) in clonogenic survival of gastric adenocarcinoma cells (AGS and MNK-45), but was relatively nontoxic to a small intestinal epithelial nonimmortalized human cell isolate (FHs 74 Int). The addition of pharmacological ascorbate (1 m M) to standard radio-chemotherapies [i.e., 5-FU (5 μ M); cisplatin (0.5 μ M); irinotecan (2.5 μ M); carboplatin (5 μ M); paclitaxel (2-4 n M); and X rays (1.8 Gy)] also potentiated gastric cancer clonogenic cell killing [additional decreases were noted with: ascorbate plus 5-FU/radiation (1%); ascorbate plus cisplatin/irinotecan (9-19%); and ascorbate plus paclitaxel/carboplatin (6-7%)]. The gastric cancer cell toxicity and chemosensitization seen with pharmacological ascorbate was dependent on H2O2 and the presence of catalytic metal ions. In addition, pharmacological ascorbate dosing resulted in a concentration-dependent decrease (64% at 20 m M, P ≤ 0.0001) in cancer cell invasion and migration that was inhibited by catalase. Finally, pharmacological ascorbate significantly increased the overall survival of mice with gastric cancer xenografts when used in combination with paclitaxel, carboplatin and radiation ( P = 0.019). These results demonstrate that pharmacological ascorbate is selectively cytotoxic to gastric adenocarcinoma cells (relative to normal intestinal epithelial cells) by a mechanism involving H2O2 and redox active metal ions. Furthermore, pharmacological ascorbate significantly enhances gastric cancer xenograft responses to radio-chemotherapy as well as inhibiting tumor cell migration and invasiveness. Overall, these results support the hypothesis that pharmacological ascorbate can be used as an adjuvant with standard-of-care radio-chemotherapies for the treatment of gastric adenocarcinomas.

Project description:Pharmacological ascorbate (P-AscH-) combined with standard of care (SOC) radiation and temozolomide is being evaluated in a phase 2 clinical trial (NCT02344355) in the treatment of glioblastoma (GBM). Previously published data demonstrated that paramagnetic iron (Fe3+) catalyzes ascorbate's oxidation to form diamagnetic iron (Fe2+). Because paramagnetic Fe3+ may influence relaxation times observed in MR imaging, quantitative MR imaging of P-AscH--induced changes in redox-active Fe was assessed as a biomarker for therapy response. Gel phantoms containing either Fe3+ or Fe2+ were imaged with T2* and quantitative susceptibility mapping (QSM). Fifteen subjects receiving P-AscH- plus SOC underwent T2* and QSM imaging four weeks into treatment. Subjects were scanned: pre-P-AscH- infusion, post-P-AscH- infusion, and post-radiation (3-4 h between scans). Changes in T2* and QSM relaxation times in tumor and normal tissue were calculated and compared to changes in Fe3+ and Fe2+ gel phantoms. A GBM mouse model was used to study the relationship between the imaging findings and the labile iron pool. Phantoms containing Fe3+ demonstrated detectable changes in T2* and QSM relaxation times relative to Fe2+ phantoms. Compared to pre-P-AscH-, GBM T2* and QSM imaging were significantly changed post-P-AscH- infusion consistent with conversion of Fe3+ to Fe2+. No significant changes in T2* or QSM were observed in normal brain tissue. There was moderate concordance between T2* and QSM changes in both progression free survival and overall survival. The GBM mouse model showed similar results with P-AscH- inducing greater changes in tumor labile iron pools compared to the normal tissue. CONCLUSIONS: T2* and QSM MR-imaging responses are consistent with P-AscH- reducing Fe3+ to Fe2+, selectively in GBM tumor volumes and represent a potential biomarker of response. This study is the first application using MR imaging in humans to measure P-AscH--induced changes in redox-active iron.

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Pharmacological Ascorbate Suppresses Growth of Gastric Cancer Cells with GLUT1 Overexpression and Enhances the Efficacy of Oxaliplatin Through Redox Modulation.

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