Table 4 Global molecular profiling studies investigating VitC in the cancer context
From: High-dose intravenous vitamin C, a promising multi-targeting agent in the treatment of cancer
Cancer type(s) | Model system | Methodology | Treatment(s) | Type of combination therapy | VitC dosea | Aim | Omics results | Ref. |
|---|---|---|---|---|---|---|---|---|
Proteomics | ||||||||
Colorectal | DiFi (RS and XM Difi) cell lines | SILAC-based MS (LC–ESI–MS-MS) | 4 h and 24 h treatments with 1 mM VitCC and/or 50 μg/mL cetuximab | Targeted | high | Hypothesis that VitC in combination with cetuximab could restrain the emergence of secondary resistance to EGFR blockade in CRC RAS/BRAF wild-type models | - Identification of 4147 proteins Switch from glycolysis to oxidative phosphorylation in cetuximab and combo-treated cells at 4 h - downregulation of LDHA/LDHB - upregulation of PDHA1/PDHB and respiratory enzymes Perturbation of iron metabolism in VitC and combo-treated cells at 24 h - downregulation of TFRC - upregulation of FT | [159] |
Breast | MDA-MB-231 cell line | Biotin switch approach (enrichment of proteins containing oxidized thiols) followed by LC-MS/MS | 30 min treatment with 10 mM ascorbic acid | – | high | Identify early alterations of the redoxome in cellular response to AA that might be linked to AA-induced cell death | - Identification of 2910 cysteine-containing proteins Oxidized targets upon AA treatment: - antioxidant enzymes (eg. PRDX1) - glycolysis and gluconeogenesis pathway (eg. PGK1) - tricarboxylic acid cycle (eg. ACOT7) - DNA, RNA and protein metabolism Cell cycle arrest and translation inhibition associated with AA-induced cytotoxicity. PRDX1 expression levels correlated with AA differential cytotoxicity | [160] |
Breast | MCF7 cell line | LC-MS/MS | 24 h treatment with 2 mM VitC | – | high | Effect of VitC in itself at different concentration levels on MCF-7 breast cancer cell line | - Identification of 1694 proteins with differential regulation Processes impacted by VitC treatment included - unfolded protein response and inhibition of the cell translation (eIF2α, PKR/PKR pThr-446) - apoptotic process | [161] |
Neuroblastoma | SH-SY5Y cell line | SUMO-1 IP followed by ESI-FT ICR MS | 30 min treatment with 100 μM ascorbate (or 100 μM hydrogen peroxide) | – | low | Identify redox sensitive proteins of the conjugation machinery for SUMOylation. Oxidative stress (hydrogen peroxide), antioxidant (ascorbate) or control conditions were tested | - Identification of 169 proteins - Great overlap between all treatments - Proteins identified only in the ascorbate sample included DTD2 and MGAT5B - Proteins without predicted SUMOylation site indentified in both ascorabte and hydrogen peroxide treatments included TUBB4A, TUBB1, HNRNPH3, POLG2 and BUB3 | [162] |
Gastric | AGS cell line | MALDI-TOF MS | 24 h treatment with 300 μg/mL (~ 1.7 mM) VitC | – | high | Investigate the molecular mechanism of the inhibitory effect of VitC on AGS cell growth, and protein profiles in AGS cells after exposure to VitC treatment | - 20 differential proteins identified - downregulation eg. of TPM3 and TPM4 - upregulation of PRDX4 and TXND5 - Identified proteins are mainly involved in cell mobility, antioxidant and detoxification, signal transduction and protein metabolism | [163] |
Leukemia | NB4 cell line | MALDI–TOF | 30 min treatment with 0.5 mM LAA (ascorbic acid) | – | medium | Identification of early protein targets of LAA in leukemia cells | - 9 differential proteins identified - changes in pI as a result of phosphorylation of a TPM isoform) - downregulation eg of of SUPT6H and HSPA8 - upregulation eg. of MATN4 and NONO | [164] |
Sarcoma | BALB/C mice implanted with S-180 cancer cells | MALDI TOF-MS/MS | Treatment with 1.5 mg/g body weight ascorbate every three days | – | high | Identify proteins involved in the ascorbic acid-mediated inhibition of tumor progression | - 11 differential proteins identified - upregulastion of RKIP and ANXA5 | [165] |
Colorectal | BALB/C mice implanted with CT-26 cancer cells | MALDI TOF-MS/MS | Treatment with 1.5 mg/g body weight ascorbate every three days | – | high | Proteome changes of tumor tissue were investigated after intraperitoneal administration of a high concentration of ascorbic acid | - 18 differential proteins identified - upregulation eg. of EIF3I, NPM1 and VIM - regulation of cytoskeleton remodeling | [166] |
Breast | MCF7 cell line | LC-MS/MS | 18 h treatment with 1 μM DOX (doxorubicin) or DOX + 200 μM of VitC | Chemo | medium | Describe the changes in protein expression and proliferation of the MCF-7 cells induced by the VitC applied with doxorubicin | - Identification of 229 proteins - Downregulation of cytoskeletal (FLNA), ribosomal (eg. RPL27A), transcriptional (eg. HNRNPH1), immune system and antioxidant (HSP90AA1, SOD1) proteins in DOX + VitC-treated cells - Upregulation of GAPDH, GPI and ACTA1 | [167] |
Leukemia | HL-60 cell line | LC-MS/MS | 48 h treatment with 10 μM As2O3 (arsenic trioxide) or As2O3 + 100 μM L-AA (ascorbic acid) + 50 μM α-TOC (α-tocopherol) | Chemo + Dietary suppl. | low | Evaluate the synergistic mechanism of action of vitamins, such as L-ascorbic acid (L-AA) and a-tocopherol (a-TOC) in As2O3 chemotherapy | - Number of identified proteins n.s. - Downregulation of cell cycle and translation in cells treated with As2O3, L-AA, and a-TOC compared to As2O3-only - Identification of numerous proteins associated with apoptosis and cell stress in combination treatment | [96] |
Breast, Lung | A549 and MDA-MB-231 cell lines | SILAC-based MS (LC-MS/MS) | untreated (A549 cell line resistant to 1 mM AUF (auranofin) + 2.5 mM VitC, MDA-MB-231 cell line sensitive) | Anti-inflammatory | untreated | Decipher the underlying mechanisms for differential response of lung and breast cancer cell models to redox-modulating molecule auranofin (AUF) and to combinations of AUF and VitC | - Identification of f 4131 proteins common to both cell lines - proteins involved in GSH synthesis and reduction, the pentose phosphate pathway and those belonging to other metabolic pathways (eg PGDH and PTGR1) more abundant in A549 (resistant) cells | [97] |
Transcriptomics | ||||||||
Melanoma | A2058 cell line | RNA-seq | 48 h treatment with 0.1 mM VitC | – | low | Examined the possible mechanisms that could reveal how VitC suppresses cell migration and anchorage-independent growth of A2058 cells | - 66 genes differentially expressed - alterations predominantly in genes involved in extracellular matrix remodeling. - ARGHAP30, TRIM63 and PTPN7 among 10 most differential genes | [168] |
Melanoma | A2058 cell line | RNA-seq | 7 days treatment with 100 μM ascorbate | – | low | To elucidate potential mechanism of ascorbate in inducing apoptosis in A2058 cells. Re-analyse data of Gustafson et al., 2015 using updated algorithms | - 344 genes including 20 non-coding RNAs (ncRNA) differentially expressed - expression of CLU gene one of the most downregulated genes | [36] |
Breast | MDA-MB-231 cell line | RNA-seq | 3 days treatment with 100 μM VitC | – | low | Analysis of transcriptomic changes associated with increased 5hmC generation following exposure to VitC | - 778 differentially expressed genes - TNFSF10, TFRC and PGK1 among 10 most differential genes | [169] |
Renal Cell | 786-O cell line | RNA-seq | Treatment for 10 passages with 100 μM AsANa (sodium L-ascorbate; VitC) or 100 μM APM (oxidation-resistant VitC derivative) | – | low | Examine ccRCC phenotype changes at the global transcriptome level after treatment of VitC for 10 passages | - 81 differentially expressed genes - most notable genes positively enriched in VitC-treated cells belong to multiple metabolic pathways, such as peroxisome and pentose phosphate pathways - most notable gene sets negatively enriched in VitC-treated cells include DNA replication and mismatch repair genes | [170] |
Bladder | T24 cell line | RNA-seq | 0.25 mM VitC, time n.s. | – | medium | Explore the role of 5hmC in bladder cancer and the therapeutic efficacy of VitC in increasing the 5hmC pattern | - 1172 differentially expressed genes were identified - differential genes mainly associated with focal adhesion, DNA replication, cell cycle, and several cancer-related pathways. | [171] |
Hepatocellular | Huh-7 cell line xenograft tumour mouse model | Microarray | 3 days treatment of mice with IP injection of 4.0 g/kg or 2.0 g/kg ascorbate | – | high | Assess effects of high-dose ascorbate on hepatoma | - 192 genes/ncRNAs uniquely differentially expressed in HCC tumour tissue obtained from mice treated specifically with high-dose ascorbate (4.0 g/kg/3 days) - deregulated genes were involved in insulin receptor signalling, metabolism and mitochondrial respiration | [172] |
Lymphoma | JLPS and JLPR cell lines (sensitive/resistant to ascorbate) | Microarray | untreated (JLPR cell line resistant to VitC (incubation of JLPS cells with increasing ascorbate concentrations from 100 μM to 1 mM over 6 month), JLPRS cell line sensitive) | – | untreated | Identify possible mechanisms of ascorbate resistance | - Acquired ascorbate resistance associated with downregulation of eg. HMGB1 and MYC and upregulation of eg. ATF5 | [173] |
Leukemia | HL60 and MOLM13 cell lines | RNA-seq | 12 or 72 h treatment with 250 μM L-AA (ascorbic acid) | – | medium | Analyse expression of genes upregulated by Tet2 restoration in cKit+ cells in HL60 and MOLM13 cells treated with L-AA | - 14/50 genes upregulated by Tet2 restoration in mouse cKit+ cells also induced in both human leukemia lines after 12 h of VitC treatment, including genes involved in apoptotic and death receptor signaling (eg. BAX) and NOTCH signaling - Of the top genes downregulated by Tet2-restoration, 34/50 were downregulated in both leukemia lines after 12 h of VitC - Hence, VitC treatment can enhance TET2 function in human leukemia cells in a manner similar to the effects of Tet2 restoration in mouse HSPCs | [174] |
Breast | MCF-7 cell line | Microarray | 3 days treatment with 100 nM RA (retinoic acid) and/or 1 mM AA (ascorbic acid) | Chemo | high | Elucidate the mechanism by which RA + AA inhibits breast carcinoma proliferation | - 29 genes were up-regulated and 38 genes were down-regulated after RA + AA treatment - up-regulation of antioxidant enzymes (eg. GPX2) and proteins involved in apoptosis (eg. CDK11B), cell cycle regulation (eg. EDN1) and DNA repair (eg. RAD51C) - RA or AA on their own failed to upregulate antioxidant genes | [175] |
Metabolomics | ||||||||
Breast, Colorectal | MCF-7, MDA-MB231 and HT29 cell lines | LC-MS | 4 h treatment with 3 mM ascorbate | – | high | Gain insight into the cellular effects of high doses of ascorbate | - Metabolic shift, reversal of Warburg effect, disruption of redox homeostasis - Cell death dependent on ascorbate-induced oxidative stress and accumulation of ROS, DNA damage, and depletion of essential intracellular co-factors including NAD+/NADH - disruption of glycolysis, rapid drop in ATP levels - inhibition the TCA cycle and increased oxygen consumption | [176] |
Breast, Colorectal | MCF7 and HT29 cell lines | CE-TOF MS | 1 h treatment with VitC (0.2 mM, 1 mM or 10 mM) | – | high | Understand anticancer mechanisms of VitC | - Levels of upstream metabolites in the glycolysis pathway and TCA cycle were increased in both cell lines following treatment with VitC - ATP levels decreased concentration-dependently - VitC inhibited energy metabolism through NAD depletion, thereby inducing cancer cell death | [177] |
Colorectal | HCT116 and VACO432 cell lines | LC-MS/MS | 2 mM VitC for 30 min to 2 h | – | high | Clarify the mechanism by which VitC kills cancer cells while sparing normal cells. Profile metabolic changes following VitC treatment | - Glycolytic intermediates upstream of GAPDH accumulated while those downstream were depleted suggesting that GAPDH was inhibited - Oxidative PPP metabolites increased, indicating that the blockage may shift glycolytic flux into the oxidative PPP - Cysteine, the major limiting precursor for GSH biosynthesis, was also dramatically depleted following VitC treatment - As expected, VitC treatment induced a substantial increase in endogenous ROS in KRAS and BRAF mutant cells | [32] |
Hepatocellular | SMMC-7721 cell line | NMR spectroscopy | 48 h treatment with 50 μmol/L OXA (oxaliplatin) and/or 1 mmol/L VitC | Chemo | high | Assess the global metabolic changes in HCC cells following VitC treatment | - VitC treatment led to inhibition of energy metabolism via NAD+ depletion and amino acid deprivation - OXA caused significant perturbation in phospholipid biosynthesis and phosphatidylcholine biosynthesis pathways - Glutathione metabolism and pathways related to succinate and choline may play central roles in conferring the combined effect between OXA and VitC | [178] |