TRPM2 Channels: A Potential Therapeutic Target in Melanoma?

The transient receptor potential, the melastatin (TRPM) subfamily, which consists of eight known members, appears to have significant importance in melanoma progression, treatment, and prognosis. As several members were originally cloned from cancerous tissue, initial studies aimed towards identifying TRPM involvement in cancer progression and tumorigenesis. For relevance in skin cancer, previous research has shown roles for several TRPM members in skin cancer progression, growth, and patient prognosis.

Methods for Investigating Transient Receptor Potential Melastatin-2 (TRPM2): A Cation Channel Activated by ADP-Ribose and Involved in Cell Death.

Transient receptor potential melastatin-2 (TRPM2) is an emerging chemotherapeutic target due to its involvement in poly(ADP-ribose) metabolism and the ability to induce anticancer effects after antagonism of its functions. Normally functioning as a nonspecific cation channel that is activated by free ADP-ribose, TRPM2 is involved with many cellular processes, including the induction of cell death after oxidative stress. What is becoming clear is that antagonism of TRPM2 selectively induces anticancer effects in several types of cancer.

Antagonism of the transient receptor potential melastatin‑2 channel leads to targeted antitumor effects in primary human malignant melanoma cells.

Melanoma continues to be the most aggressive and devastating form of skin cancer for which the development of novel therapies is required. The present study aimed to determine the effects of antagonism of the transient receptor potential melastatin‑2 (TRPM2) ion channel in primary human malignant melanoma cells. TRPM2 antagonism via use of the antifungal agent, clotrimazole, led to decreases in cell proliferation, as well as dose‑dependent increases in cell death in all melanoma cell lines investigated.

Enhanced cytotoxicity in triple-negative and estrogen receptor‑positive breast adenocarcinoma cells due to inhibition of the transient receptor potential melastatin-2 channel.

We previously demonstrated a unique protective role for the transient receptor potential, melastatin-2 (TRPM2) cation channel in breast cancer cells. In the present study, we investigated the chemotherapeutic effects elicited by inhibiting this protective role in metastatic breast adenocarcinoma cells. TRPM2 inhibition led to dose-dependent increases in MDA-MB-231 breast adenocarcinoma cell death after treatment with doxorubicin or the DNA-methylating agent, N-methyl-N'-nitro-N-nitrosoguanidine.

Inhibition of the transient receptor potential melastatin-2 channel causes increased DNA damage and decreased proliferation in breast adenocarcinoma cells.

Transient receptor potential, melastatin-2 (TRPM2) is a plasma membrane cation channel with important roles in sensory functions and promoting cell death. However, we demonstrated here that TRPM2 was present in the nuclei of MCF-7 and MDA-MB-231 human breast adenocarcinoma cells, and its pharmacologic inhibition or RNAi silencing caused decreased cell proliferation. Neither an effect on proliferation nor a localization of TRPM2 in the nucleus was observed in noncancerous HMEC and MCF-10A human mammary epithelial cells.

Quantitative proteomic analysis in HCV-induced HCC reveals sets of proteins with potential significance for racial disparity.

Background: The incidence and mortality of hepatitis C virus (HCV)-induced hepatocellular carcinoma (HCC) is higher in African Americans (AA) than other racial/ethnic groups in the U.S., but the reasons for this disparity are unknown.

Roles of poly(ADP-ribose) glycohydrolase in DNA damage and apoptosis.

Poly(ADP-ribose) glycohydrolase (PARG) is the primary enzyme that catalyzes the hydrolysis of poly(ADP-ribose) (PAR), an essential biopolymer that is synthesized by poly(ADP-ribose) polymerases (PARPs) in the cell. By regulating the hydrolytic arm of poly(ADP-ribosyl)ation, PARG participates in a number of biological processes, including the repair of DNA damage, chromatin dynamics, transcriptional regulation, and cell death. Collectively, the research investigating the roles of PARG in the cell has identified the importance of PARG and its value as a therapeutic target.

Inhibition of poly(ADP-ribose) polymerase-1 or poly(ADP‑ribose) glycohydrolase individually, but not in combination, leads to improved chemotherapeutic efficacy in HeLa cells.

The genome-protecting role of poly(ADP-ribose) (PAR) has identified PAR polymerase-1 (PARP-1) and PAR glycohydrolase (PARG), two enzymes responsible for the synthesis and hydrolysis of PAR, as chemotherapeutic targets. Each has been previously individually evaluated in chemotherapy, but the effects of combination PARP-1 and PARG inhibition in cancer cells are not known. Here we determined the effects of the inhibition of PARP-1 and the absence or RNAi knockdown of PARG on PAR synthesis, cell death after chemotherapy and long-term viability.

Silencing of Apoptosis-Inducing factor and poly(ADP-ribose) glycohydrolase reveals novel roles in breast cancer cell death after chemotherapy.

Background: Cell death induced by poly(ADP-ribose) (PAR) and mediated by apoptosis-inducing factor (AIF) is well-characterized in models of ischemic tissue injury, but their roles in cancer cell death after chemotherapy are less understood.

Methods: Here we investigated the roles of PAR and AIF by RNA interference (RNAi) in MDA-MB-231 and MCF-7 breast adenocarcinoma cells after chemotherapy. Differences in effects were statistically tested by analysis-of-variance and unpaired student's t-test.

Poly(ADP-ribosyl)ation pathways in mammals: the advantage of murine PARG null mutation.

Studies suggest that inhibiting the hydrolysis of poly(ADP-ribose) (PAR) by targeting the enzyme PAR glycohydrolase (PARG) is a potential chemotherapeutic strategy to induce cell death. However, the lack of structural data for PARG has hindered the discovery of specific PARG inhibitors and thus hampered the search for cellular effects dependent on the hydrolysis of PAR. We previously generated a murine PARG null cell model to identify the intracellular processes mediated by PARG.

Synergistic cytotoxicity of N-methyl-N'-nitro-N-nitrosoguanidine and absence of poly(ADP-ribose) glycohydrolase involves chromatin decondensation.

DNA-alkylating agents in combination with poly (ADP-ribose) (PAR) synthesis inhibitors are a promising treatment for cancer. In search of other efficacious alternatives, we hypothesized that the absence of poly(ADP-ribose) glycohydrolase (PARG), which leads to the inhibition of PAR hydrolysis, would lead to increased DNA alkylation after treatment with low doses of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). At a sublethal dose, MNNG shows synergistic cytotoxicity in PARG-null embryonic trophoblast stem (TS) cells.

Activation of cell death mediated by apoptosis-inducing factor due to the absence of poly(ADP-ribose) glycohydrolase.

We previously demonstrated that the absence of poly(ADP-ribose) glycohydrolase (PARG) led to increased cell death following DNA-damaging treatments. Here, we investigated cell death pathways following UV treatment. Decreased amounts of PARG-null embryonic trophoblast stem (TS) cells were observed following doses of 10-100 J/m2 as compared to wild-type cells.

Enhanced DNA accessibility and increased DNA damage induced by the absence of poly(ADP-ribose) hydrolysis.

Poly(ADP-ribose) (PAR) is a therapeutic target primarily identified through inhibiting its synthesis by PAR polymerase-1 (PARP-1). However, inhibiting its hydrolysis by PAR glycohydrolase (PARG) has therapeutic potential in cancer. Unknown is the effect of elevated PAR levels on cellular processes and if this effect can enhance the therapeutic value of PARG.

Poly(ADP-ribose) (PAR) polymer is a death signal.

Excessive activation of the nuclear enzyme, poly(ADP-ribose) polymerase-1 (PARP-1) plays a prominent role in various of models of cellular injury. Here, we identify poly(ADP-ribose) (PAR) polymer, a product of PARP-1 activity, as a previously uncharacterized cell death signal. PAR polymer is directly toxic to neurons, and degradation of PAR polymer by poly(ADP-ribose) glycohydrolase (PARG) or phosphodiesterase 1 prevents PAR polymer-induced cell death.

Differential effect of PARP-2 deletion on brain injury after focal and global cerebral ischemia.

Poly(ADP-ribose) polymerase-2 (PARP-2) is a member of the PARP enzyme family, and, similarly to PARP-1, catalyzes the formation of ADP-ribose polymers in response to DNA damage. While PARP-1 overactivation contributes to ischemic cell death, no information is available regarding the role of PARP-2. In this study, we evaluated the impact of PARP-2 deletion on histopathological outcome from two different experimental models of cerebral ischemia.

Mediation of cell death by poly(ADP-ribose) polymerase-1.

Poly(ADP-ribosyl)ation plays an important role in modulating the cellular response to stress. The extent of poly(ADP-ribosyl)ation, chiefly via the activation of the poly(ADP-ribose) polymerase-1 (PARP-1), correlates with the severity of genotoxic stress and this determines the cellular response. Under mild and moderate stress, it plays important roles in DNA processing and it participates in the proinflammatory/cellular defense via transcriptional regulation.

The road to survival goes through PARG.

Unlike poly(ADP-ribose) polymerase-1 (PARP-1), poly(ADP-ribose) glycohydrolase (PARG) has long been a difficult protein to study. However, the complete absence of PARG activity was recently characterized in mice via disruption of the murine PARG gene. As expected, PARG is critical for the maintenance of steady-state poly(ADP-ribose) levels.

Identification of three critical acidic residues of poly(ADP-ribose) glycohydrolase involved in catalysis: determining the PARG catalytic domain.

PARG [poly(ADP-ribose) glycohydrolase] catalyses the hydrolysis of alpha(1''-->2') or alpha(1'''-->2'') O-glycosidic linkages of ADP-ribose polymers to produce free ADP-ribose. We investigated possible mechanistic similarities between PARG and glycosidases, which also cleave O-glycosidic linkages. Glycosidases typically utilize two acidic residues for catalysis, thus we targeted acidic residues within a conserved region of bovine PARG that has been shown to contain an inhibitor-binding site.

Failure to degrade poly(ADP-ribose) causes increased sensitivity to cytotoxicity and early embryonic lethality.

The metabolism of poly(ADP-ribose) (PAR) is critical for genomic stability in multicellular eukaryotes. Here, we show that the failure to degrade PAR by means of disruption of the murine poly(ADP-ribose) glycohydrolase (PARG) gene unexpectedly causes early embryonic lethality and enhanced sensitivity to genotoxic stress. This lethality results from the failure to hydrolyze PAR, because PARG null embryonic day (E) 3.

Apoptosis-inducing factor substitutes for caspase executioners in NMDA-triggered excitotoxic neuronal death.

The profound neuroprotection observed in poly(ADP-ribose) polymerase-1 (PARP-1) null mice to ischemic and excitotoxic injury positions PARP-1 as a major mediator of neuronal cell death. We report here that apoptosis-inducing factor (AIF) mediates PARP-1-dependent glutamate excitotoxicity in a caspase-independent manner after translocation from the mitochondria to the nucleus. In primary murine cortical cultures, neurotoxic NMDA exposure triggers AIF translocation, mitochondrial membrane depolarization, and phosphatidyl serine exposure on the cell surface, which precedes cytochrome c release and caspase activation.

SAR analysis of adenosine diphosphate (hydroxymethyl)pyrrolidinediol inhibition of poly(ADP-ribose) glycohydrolase.

Polyadenosine diphosphoribose glycohydrolase (PARG) catalyzes the intracellular hydrolysis of adenosine diphosphoribose polymers. Because structure-activity data are lacking for PARG, the specific inhibitor adenosine diphosphate (hydroxymethyl)pyrrolidinediol (ADP-HPD) was utilized to determine the effects of structure on inhibitor potency using PARG isolated from bovine thymus (bPARG) and recombinant bovine PARG catalytic fragment (rPARG-CF). Both enzymes were strongly inhibited by submicromolar levels of ADP-HPD, but ADP and the phosphorylated pyrrolidine displayed no activity.

Identification of an inhibitor binding site of poly(ADP-ribose) glycohydrolase.

Polymers of ADP-ribose involved in the maintenance of genomic integrity are converted to free ADP-ribose by the action of poly(ADP-ribose) glycohydrolase (PARG). As an approach to mapping functions of PARG onto the amino acid sequence of the protein, we report here experiments that identify an amino acid residue involved in the binding of potent PARG inhibitors. A photoreactive inhibitor, [alpha-(32)P]-8-azidoadenosine diphosphate (hydroxymethyl)pyrrolidinediol (8-N(3)-ADP-HPD), was used to photolabel a recombinant bovine PARG catalytic fragment (rPARG-CF).