Glyoxalase 1 is up-regulated in hepatocellular carcinoma and is essential for HCC cell proliferation

Glyoxalase 1 (Glo1), belonging to the glyoxalase system, participates in the detoxification of methylglyoxal (MG), a byproduct of glycolysis. Glo1 is associated with the progression of many human malignancies. However, the role of Glo1 in hepatocellular carcinoma (HCC) is unclear. We have discovered that the expression of Glo1 is up-regulated in HCC tissues compared with adjacent non-tumorous tissues, and knockdown of Glo1 expression by RNA interference significantly inhibited the proliferation of human HCC cell lines. Glo1 knockdown resulted in the accumulation of its cytotoxic substrate, MG. Overall, thus Glo1 might be essential for HCC progression and can be designated as a potential therapeutic target for HCC in the future.     

Glyoxalase 1 sustains the metastatic phenotype of prostate cancer cells via EMT control

Metastasis is the primary cause of death in prostate cancer (PCa) patients. Effective therapeutic intervention in metastatic PCa is undermined by our poor understanding of its molecular aetiology. Defining the mechanisms underlying PCa metastasis may lead to insights into how to decrease morbidity and mortality in this disease. Glyoxalase 1 (Glo1) is the detoxification enzyme of methylglyoxal (MG), a potent precursor of advanced glycation end products (AGEs). Hydroimidazolone (MG‐H1) and argpyrimidine (AP) are AGEs originating from MG‐mediated post‐translational modification of proteins at arginine residues. AP is involved in the control of epithelial to mesenchymal transition (EMT), a crucial determinant of cancer metastasis and invasion, whose regulation mechanisms in malignant cells are still emerging. Here, we uncover a novel mechanism linking Glo1 to the maintenance of the metastatic phenotype of PCa cells by controlling EMT by engaging the tumour suppressor miR‐101, MG‐H1‐AP and TGF‐β1/Smad signalling. Moreover, circulating levels of Glo1, miR‐101, MG‐H1‐AP and TGF‐β1 in patients with metastatic compared with non‐metastatic PCa support our in vitro results, demonstrating their clinical relevance. We suggest that Glo1, together with miR‐101, might be potential therapeutic targets for metastatic PCa, possibly by metformin administration.     

Unease on the role of glyoxalase 1 in high-anxiety-related behaviour

Overexpression and silencing of glyoxalase 1 (Glo1) in brains of mice and behavioural analyses have suggested a link between Glo1 and anxiety, making Glo1 a possible novel target for anxiolytic-drug development. However, this finding is discordant with others, and further research at metabolic level, particularly glycation of neuronal proteins by dicarbonyl substrates of Glo1, is required. Therefore, it remains to be established whether Glo1 is a risk marker or a risk factor of increased anxiety, how applicable the association between Glo1 and anxiety is and whether it can be translated into clinical diagnostic and therapeutic applications.     

The glucose metabolite methylglyoxal inhibits expression of the glucose transporter genes by inactivating the cell surface glucose sensors Rgt2 and Snf3 in yeast

Methylglyoxal (MG) is a cytotoxic by-product of glycolysis. MG has inhibitory effect on the growth of cells ranging from microorganisms to higher eukaryotes, but its molecular targets are largely unknown. The yeast cell-surface glucose sensors Rgt2 and Snf3 function as glucose receptors that sense extracellular glucose and generate a signal for induction of expression of genes encoding glucose transporters (HXTs). Here we provide evidence that these glucose sensors are primary targets of MG in yeast. MG inhibits the growth of glucose-fermenting yeast cells by inducing endocytosis and degradation of the glucose sensors. However, the glucose sensors with mutations at their putative ubiquitin-acceptor lysine residues are resistant to MG-induced degradation. These results suggest that the glucose sensors are inactivated through ubiquitin-mediated endocytosis and degraded in the presence of MG. In addition, the inhibitory effect of MG on the glucose sensors is greatly enhanced in cells lacking Glo1, a key component of the MG detoxification system. Thus the stability of these glucose sensors seems to be critically regulated by intracellular MG levels. Taken together, these findings suggest that MG attenuates glycolysis by promoting degradation of the cell-surface glucose sensors and thus identify MG as a potential glycolytic inhibitor.     

Determination of the Effects of Bee Venom on Triple Negative Breast Cancer Cells in Vitro

Honeybees provide multiple products such as bee venom (BV) which are used for various nutritional and medicinal purposes. BV has received great attention due to its wide range of bioactive components with potential anti-cancer effects on different cancers. Triple negative breast cancer (TNBC) is defined as an aggressive type of breast cancer and new therapeutic targets are required for its treatment. In the current literature information is varied about the composition and quantity of BV bioactive compounds as well as the origin of BV and its significance. In this context, the cytotoxic and apoptotic effects of BV with a higher rate of mellitin from Apis mellifera anatoliaca (Muğla ecotype) on MDA-MB-231 cells was evaluated, in vitro. The cytotoxic, apoptotic and morphological effects of BV were determined by WST-1, Annexin V, cell cycle analysis and Acridine Orange staining. The results showed that BV caused apoptotic cell death in TNBC cells at a lower dose (0.47 μg/mL, p<0.01). This study suggests that BV could be developed as a potential therapeutic agent for cancer treatment. However, the mechanism of BV-induced apoptosis death should be clarified at the molecular level.     

SIRT1 participates in the response to methylglyoxal-dependent glycative stress in mouse oocytes and ovary

Methylglyoxal (MG), a highly reactive dicarbonyl derived from metabolic processes, is the most powerful precursor of advanced glycation end products (AGEs). Glycative stress has been recently associated with ovarian dysfunctions in aging and PCOS syndrome. We have investigated the role of the NAD⁺-dependent Class III deacetylase SIRT1 in the adaptive response to MG in mouse oocytes and ovary. In mouse oocytes, MG induced up-expression of glyoxalase 1 (Glo1) and glyoxalase 2 (Glo2) genes, components of the main MG detoxification system, whereas inhibition of SIRT1 by Ex527 or sirtinol reduced this response. In addition, the inhibition of SIRT1 worsened the effects of MG on oocyte maturation rates, while SIRT1 activation by resveratrol counteracted MG insult. Ovaries from female mice receiving 100 mg/kg MG by gastric administration for 28 days (MG mice) exhibited increased levels of SIRT1 along with over-expression of catalase, superoxide dismutase 2, SIRT3, PGC1α and mtTFA. Similar levels of MG-derived AGEs were observed in the ovaries from MG and control groups, along with enhanced protein expression of glyoxalase 1 in MG mice. Oocytes ovulated by MG mice exhibited atypical meiotic spindles, a condition predisposing to embryo aneuploidy. Our results from mouse oocytes revealed for the first time that SIRT1 could modulate MG scavenging by promoting expression of glyoxalases. The finding that up-regulation of glyoxalase 1 is associated with that of components of a SIRT1 functional network in the ovaries of MG mice provides strong evidence that SIRT1 participates in the response to methylglyoxal-dependent glycative stress in the female gonad.     

Antibody-drug therapeutic conjugates: Potential of antibody-siRNAs in cancer therapy

Codelivery is a promising strategy of targeted delivery of cytotoxic drugs for eradicating tumor cells. This rapidly growing method of drug delivery uses a conjugate containing drug linked to a smart carrier. Both two parts usually have therapeutic properties on the tumor cells. Monoclonal antibodies and their derivatives, such as Fab, scFv, and bsAb due to targeting high potent have now been attractive candidates as drug targeting carrier systems. The success of some therapeutic agents like small interfering RNA (siRNA), a small noncoding RNAs, with having problems such as enzymatic degradation and rapid renal filtration need to an appropriate carrier. Therefore, the aim of this study is to review the recent enhancements in development of antibody drug conjugates (ADCs), especially antibody–siRNA conjugates (SRCs), its characterizations and mechanisms in innovative cancer therapy approaches.     

Curcumin inhibits glyoxalase 1: a possible link to its anti-inflammatory and anti-tumor activity

Background

Glyoxalases (Glo1 and Glo2) are involved in the glycolytic pathway by detoxifying the reactive methylglyoxal (MGO) into D-lactate in a two-step reaction using glutathione (GSH) as cofactor. Inhibitors of glyoxalases are considered as anti-inflammatory and anti-carcinogenic agents. The recent finding that various polyphenols modulate Glo1 activity has prompted us to assess curcumin''s potency as an Glo1 inhibitor.

Methodology/Principal Findings

Cultures of whole blood cells and tumor cell lines (PC-3, JIM-1, MDA-MD 231 and 1321N1) were set up to investigate the effect of selected polyphenols, including curcumin, on the LPS-induced cytokine production (cytometric bead-based array), cell proliferation (WST-1 assay), cytosolic Glo1 and Glo2 enzymatic activity, apoptosis/necrosis (annexin V-FITC/propidium iodide staining; flow cytometric analysis) as well as GSH and ATP content. Results of enzyme kinetics revealed that curcumin, compared to the polyphenols quercetin, myricetin, kaempferol, luteolin and rutin, elicited a stronger competitive inhibitory effect on Glo1 (K_i = 5.1±1.4 µM). Applying a whole blood assay, IC₅₀ values of pro-inflammatory cytokine release (TNF-α, IL-6, IL-8, IL-1β) were found to be positively correlated with the K_i-values of the aforementioned polyphenols. Moreover, whereas curcumin was found to hamper the growth of breast cancer (JIMT-1, MDA-MB-231), prostate cancer PC-3 and brain astrocytoma 1321N1 cells, no effect on growth or vitality of human primary hepatocytes was elucidated. Curcumin decreased D-lactate release by tumor cells, another clue for inhibition of intracellular Glo1.

Conclusions/Significance

The results described herein provide new insights into curcumin''s biological activities as they indicate that inhibition of Glo1 by curcumin may result in non-tolerable levels of MGO and GSH, which, in turn, modulate various metabolic cellular pathways including depletion of cellular ATP and GSH content. This may account for curcumin''s potency as an anti-inflammatory and anti-tumor agent. The findings support the use of curcumin as a potential therapeutic agent.     

Monovalency Unleashes the Full Therapeutic Potential of the DN-30 Anti-Met Antibody

Met, the high affinity receptor for hepatocyte growth factor, is one of the most frequently activated tyrosine kinases in human cancer and a validated target for cancer therapy. We previously developed a mouse monoclonal antibody directed against the extracellular portion of Met (DN-30) that induces Met proteolytic cleavage (receptor “shedding”) followed by proteasome-mediated receptor degradation. This translates into inhibition of hepatocyte growth factor/Met-mediated biological activities. However, DN-30 binding to Met also results in partial activation of the Met kinase due to antibody-mediated receptor homodimerization. To safely harness the therapeutic potential of DN-30, its shedding activity must be disassociated from its agonistic activity. Here we show that the DN-30 Fab fragment maintains high affinity Met binding, elicits efficient receptor shedding and down-regulation, and does not promote kinase activation. In Met-addicted tumor cell lines, DN-30 Fab displays potent cytostatic and cytotoxic activity in a dose-dependent fashion. DN-30 Fab also inhibits anchorage-independent growth of several tumor cell lines. In mouse tumorigenesis assays using Met-addicted carcinoma cells, intratumor administration of DN-30 Fab or systemic delivery of a chemically stabilized form of the same molecule results in reduction of Met phosphorylation and inhibition of tumor growth. These data provide proof of concept that monovalency unleashes the full therapeutic potential of the DN-30 antibody and point at DN-30 Fab as a promising tool for Met-targeted therapy.     

细胞自噬与肿瘤耐药

细胞自噬是一种细胞自我降解的过程,在适应代谢应激、保持基因组完整性及维持内环境稳定方面发挥重要作用. 在肿瘤治疗中,凋亡耐受是产生肿瘤耐药的重要机制. 细胞自噬可防止抗肿瘤药诱导的凋亡,促进肿瘤耐药. 然而,自噬性细胞死亡可能是凋亡耐受肿瘤细胞的一种死亡方式. 因此,细胞自噬对肿瘤细胞的耐药性有双重影响. 本文综述了细胞自噬的分子机制、细胞自噬与凋亡的关系、细胞自噬与肿瘤耐药以及治疗的主要研究进展.     

Inhibitory effects of S-nitrosoglutathione on cell proliferation and DNA synthesis: possible role of glyoxalase I inactivation

We investigated the inhibitory effects of S-nitrosoglutathione (GSNO) on cell proliferation, DNA synthesis and several enzymatic activities using spontaneously immortalized human endothelial cells (ECV304). Proliferation of ECV304 was inhibited by GSNO in a dose-dependent manner (125-1000 microM). DNA synthesis was decreased 2 h after addition of GSNO to cells and was markedly repressed from 20 h after the addition. The activity of ribonucleotide reductase, a rate-limiting enzyme for DNA synthesis, was unchanged in GSNO-treated cells. GSNO inhibited less than 40% of mitochondrial respiration activity, and the membrane potential and cellular levels of ATP were not significantly decreased by GSNO. GSNO had no inhibitory effect on activities of glutathione peroxidase, glutathione S-transferase and glutathione reductase. However, glyoxalase I (Glo I) activity was decreased to 20% of the control level within 60 min, and was consistently repressed during exposure to GSNO for 20 h. A membrane-permeable Glo I inhibitor, S-bromobenzylglutathione diethylester, inhibited proliferation of ECV304 cells, while methylglyoxal (MG), a toxic metabolite generated during glycolysis and a substrate for Glo I, failed to inhibit the cell growth even at 100 microM. Glo I in several mammalian cell lines was inactivated by GSNO with a pI shift. Although we failed to detect accumulation of MG under conditions of Glo I inactivation, these results suggest that the inhibitory effects of GSNO on cell proliferation and DNA synthesis might be at least partly due to inactivation of Glo I.     

Modulator of Apoptosis 1 (MOAP-1) Is a Tumor Suppressor Protein Linked to the RASSF1A Protein

Modulator of apoptosis 1 (MOAP-1) is a BH3-like protein that plays key roles in cell death or apoptosis. It is an integral partner to the tumor suppressor protein, Ras association domain family 1A (RASSF1A), and functions to activate the Bcl-2 family pro-apoptotic protein Bax. Although RASSF1A is now considered a bona fide tumor suppressor protein, the role of MOAP-1 as a tumor suppressor protein has yet to be determined. In this study, we present several lines of evidence from cancer databases, immunoblotting of cancer cells, proliferation, and xenograft assays as well as DNA microarray analysis to demonstrate the role of MOAP-1 as a tumor suppressor protein. Frequent loss of MOAP-1 expression, in at least some cancers, appears to be attributed to mRNA down-regulation and the rapid proteasomal degradation of MOAP-1 that could be reversed utilizing the proteasome inhibitor MG132. Overexpression of MOAP-1 in several cancer cell lines resulted in reduced tumorigenesis and up-regulation of genes involved in cancer regulatory pathways that include apoptosis (p53, Fas, and MST1), DNA damage control (poly(ADP)-ribose polymerase and ataxia telangiectasia mutated), those within the cell metabolism (IR-α, IR-β, and AMP-activated protein kinase), and a stabilizing effect on microtubules. The loss of RASSF1A (an upstream regulator of MOAP-1) is one of the earliest detectable epigenetically silenced tumor suppressor proteins in cancer, and we speculate that the additional loss of function of MOAP-1 may be a second hit to functionally compromise the RASSF1A/MOAP-1 death receptor-dependent pathway and drive tumorigenesis.