The Rescue on Reperfusion Damage in Cerebral Infarction by Nelonemdaz (RODIN) Trial: Protocol for a Double-Blinded Clinical Trial of Nelonemdaz in Patients with Hyperacute Ischemic Stroke and Endovascular Thrombectomy.

BACKGROUND AND PURPOSE: Nelonemdaz (Neu2000) has both selective antagonism against 2B subunit of N-methyl-D-aspartate receptor and antioxidant activity. This drug provides sufficient evidence of neuroprotection in acute cerebral ischemia/reperfusion models. This phase III trial aims to determine this effect in patients. DESIGN: The Rescue on Reperfusion Damage in Cerebral Infarction by Nelonemdaz is a multicenter, double-blinded clinical trial. A total of 496 patients will be randomly assigned into the nelonemdaz (a total of 5,250 mg divided by 10 times for 5 days) and placebo groups. Patients will be included if they have an acute ischemic stroke (National Institutes of Health Stroke Scale score ≥8) caused by intracranial large vessel occlusion in the anterior circulation (Alberta Stroke Program Early CT Score ≥4), and if they are expected to undergo endovascular thrombectomy within 12 hours after stroke onset. ENDPOINTS: The primary endpoint is a favorable shift in the modified Rankin Scale (mRS) score at 90 days after the first dose of drug. The data will be analyzed by the Cochran-Mantel-Haenszel shift test. The secondary endpoints include functional independence (mRS 0-2) at 35 and 90 days, the favorable shift of mRS at 35 days, the proportion of mRS 0 at 35 and 90 days, and the occurrence rates of symptomatic intracranial hemorrhage within 7 days. CONCLUSION: This trial will clarify the efficacy and safety of nelonemdaz in patients with acute ischemic stroke and endovascular thrombectomy. This study has been registered at ClinicalTrials. gov (NCT05041010).

Rationale and methods of the Antioxidant and NMDA receptor blocker Weans Anoxic brain damage of KorEa OHCA patients (AWAKE) trial.

BACKGROUND: Ischemic brain injury is a major hurdle that limits the survival of resuscitated out-of-hospital cardiac arrest (OHCA). METHODS: The aim of this study is to assess the feasibility and potential for reduction of ischemic brain injury in adult OHCA patients treated with high- or low-dose Neu2000K, a selective blocker of N-methyl-D-aspartate (NMDA) type 2B receptor and also a free radical scavenger, or given placebo. This study is a phase II, multicenter, randomized, double-blinded, prospective, intention-to-treat, placebo-controlled, three-armed, safety and efficacy clinical trial. This trial is a sponsor-initiated trial supported by GNT Pharma. Successfully resuscitated OHCA patients aged 19 to 80 years would be included. The primary outcome is blood neuron-specific enolase (NSE) level on the 3rd day. The secondary outcomes are safety, efficacy defined by study drug administration within 4 h in > 90% of participants, daily NSE up to 5th day, blood S100beta, brain MRI apparent diffusion coefficient imaging, cerebral performance category (CPC), and Modified Rankin Scale (mRS) at 5th, 14th, and 90th days. Assuming NSE of 42 ± 80 and 80 ± 80 µg/L in the treatment (high- and low-dose Neu2000K) and control arms with 80% power, a type 1 error rate of 5%, and a 28% of withdrawal prior to the endpoint, the required sample size is 150 patients. DISCUSSION: The AWAKE trial explores a new multi-target neuroprotectant for the treatment of resuscitated OHCA patients. TRIAL REGISTRATION: ClinicalTrials.gov NCT03651557 . Registered on August 29, 2018.

Chronic blockade of the angiotensin II receptor has a differential effect on adipose and vascular PAI-1 in OLETF rats.

Angiotensinogen (AGT) and plasminogen activator inhibitor-1 (PAI-1) are expressed in both vascular and adipose tissues. Angiotensin II (AG II) has an adipogenic effect and increases PAI-1 expression. To evaluate the chronic effects of AG II type 1 receptor (AT(1)R) antagonism on adipose mass and PAI-1 expression in vascular and adipose tissues, losartan (30mg/kg/day) was administered to Otsuka Long-Evans Tokushima Fatty (OLETF) rats, a model of type 2 diabetes, for 20 weeks. Adipose mass and regional fat distribution in the abdomen did not change after chronic AT(1)R antagonism in OLETF rats. AGT and PAI-1 mRNA expressions in adipose tissue of OLETF rats were significantly increased compared with Long-Evans Tokushima Otsuka (LETO) rats, the normal control. Chronic losartan therapy further increased the level of adipose AGT in OLETF rats, but did not affect the level of adipose PAI-1 mRNA. In contrast, aortic PAI-1 expression in OLETF rats was attenuated by chronic losartan therapy. Our results have two implications. First, adipose tissue may be an important source of AG II in metabolic syndrome even after chronic losartan therapy. Second, chronic AT(1)R antagonism with losartan causes differential effects on vascular and adipose PAI-1 expression.

The effects of thiazolidinedione treatment on the regulations of aquaglyceroporins and glycerol kinase in OLETF rats.

Aquaporins (AQPs) that transport glycerol in addition to water are classified as aquaglyceroporins (AQP3, 7, 9). AQP7 in the adipose tissue and AQP9 in the liver may coordinately contribute to the increase in hepatic gluconeogenesis in states of insulin resistance. Thiazolidinedione (TZD) has been shown to increase adipose AQP7 and induce glycerol kinase (GlyK) which is nearly absent in adipocytes. In the present study, we analyzed both GlyK and AQP gene expression in adipose and hepatic tissues, and AQP3 in kidneys from Long-Evans Tokushima Otsuka (LETO), Otsuka Long-Evans Tokushima Fatty (OLETF), and rosiglitazone (RSG)-treated OLETF (RSG-OLETF) rats. We also evaluated AQP9 protein expression in cultured human hepatoma cells treated with oleic acid, Wy14643, or RSG. A 2-week RSG treatment increased AQP7 mRNA levels in the mesenteric fat, but not in the epididymal fat of OLETF rats. Rosiglitazone treatment markedly increased GlyK expression in both fat depots, with a greater increase in the mesenteric fat. The magnitudes of GlyK induction by RSG were greater than that of AQP7 in both adipose tissues (P < .05, each). AQP9 and GlyK levels in the liver were not affected by RSG treatment in OLETF rats. Oleic acid and Wy14643 upregulated AQP9 protein expression in cultured human hepatoma cells in a dose-dependent manner. AQP3 mRNA levels tended to increase in the outer medulla of the RSG-OLETF rats. These results indicate that in the adipose tissue TZD has an important role in the glycerol metabolic pathway through the regulation of AQP and GlyK, especially by GlyK induction. Free fatty acids may directly enhance glycerol availability in the liver via the upregulation of AQP9 levels. Renal AQP3 may be related to the fluid retention caused by TZD.

Molecular mechanisms of heptaplatin effective against cisplatin-resistant cancer cell lines: less involvement of metallothionein.

BACKGROUND: Heptaplatin is a new platinum derivative with anticancer activity against various cancer cell lines, including cisplatin-resistant cancer cell lines (Cancer Chemother Pharmacol 1995; 35: 441). METHODS: Molecular mechanisms of heptaplatin effective against cisplatin-resistant cancer cell lines has been investigated in connection with metallothionein (MT). Cytotoxicity was determined by an MTT assay. MT mRNA, was determined by RT-PCR assay. Transfection study was carried out to examine the function of MT. RESULTS: Of various gastric cancer cell lines, SNU-638 and SNU-601 showed the highest and lowest levels of MT mRNA, respectively, showing 80-fold difference. The IC50 values of SNU-638 to cisplatin, carboplatin and heptaplatin were 11.2-fold, 5.1-fold and 2.0-fold greater than those of SNU-601, respectively. Heptaplatin was more effective against cisplatin-resistant and MT-transfected gastric cancer sublines than cisplatin or carboplatin was. In addition, heptaplatin attenuated cadmium, but not zinc, induction of MT. CONCLUSION: These results indicate that molecular mechanisms of heptaplatin effective against cisplatin-resistant gastric cancer sublines is at least in part due to the less involvement of MT in heptaplatin resistance as well as its attenuation of MT induction.

Prevention effects of ND-07, a novel drug candidate with a potent antioxidative action and anti-inflammatory action, in animal models of severe acute pancreatitis.

Oxidative stress and inflammation both play major roles in the development of the acute pancreatitis. Currently, a pancreatic enzyme inhibitor with limited efficacy is only clinically available in a few countries, and antioxidants or non-steroidal anti-inflammatory drugs (NSAIDs) provide only partial tissue protection in acute pancreatitis animal models. Here, we introduce a new drug candidate for treating acute pancreatitis named ND-07 [chemical name: 2-acetoxy-5-(2-4-(trifluoromethyl)-phenethylamino)-benzoic acid] that exhibits both potent antioxidative and anti-inflammatory activities. In an electron spin resonance (ESR) study, ND-07 almost blocked hydroxyl radical generation as low as 0.05 µM and significantly suppressed DNA oxidation and cell death in a lipopolysaccharide (LPS)-stimulated pancreatic cell line. In a cerulein plus LPS-induced acute pancreatitis model, ND-07 pretreatment showed significant tissue protective effects, with reductions of serum amylase and lipase levels and pancreatic wet weights. ND-07 not only diminished the plasma levels of malondialdehyde (MDA) and nitric oxide but also significantly decreased prostaglandin E2 (PGE2) and expression of tumor necrotizing factor-alpha (TNF-α) in the pancreatic tissue. In a severe acute necrotizing pancreatitis model induced by a choline deficient, ethionine-supplemented (CDE) diet, ND-07 dramatically protected the mortality even without any death, providing attenuation of pancreas, lung, and liver damages as well as the reductions in serum levels of lactate dehydrogenase (LDH), amylase and lipase, MDA levels in the plasma and pancreatic tissues, plasma levels of TNF-α, and interleukin-1 (IL-1ß). These findings suggest that current dual synergistic action mechanisms of ND-07 might provide a superior protection for acute pancreatitis than conventional drug treatments.

Concentration-dependent collateral sensitivity of cisplatin-resistant gastric cancer cell sublines.

The cisplatin-resistant gastric cancer cell sublines, SNU-601/Cis2 and /Cis10, were 49 and >530 times more resistant to cisplatin, respectively, compared with the drug-sensitive cells, SNU-601/WT. The SNU-601/Cis2 showed cross-resistance to carboplatin, heptaplatin, doxorubicin, mitomycin C, and 5-fluorouracil compared with the SNU-601/WT whereas the SNU-601/Cis10 displayed collateral sensitivity to these drugs with the exception of cisplatin compared with the SNU-601/Cis2, suggesting that the cross-resistance and collateral sensitivity of cisplatin-resistant gastric cancer cells are dependent upon cisplatin concentrations. Altered expression of the antioxidant and transporter genes (metallothionein, catalase, superoxide dismutases, P-glycoprotein, and the breast cancer resistance protein) was involved in these phenotypes of the cisplatin-resistant gastric cancer cell lines.

Reversal of P-glycoprotein-mediated multidrug resistance by 5,6,7,3',4'-pentamethoxyflavone (Sinensetin).

Multidrug resistance (MDR) cells can be sensitized to anticancer drugs when treated concomitantly with chemosensitizers. In this study, chemosensitizing effects of 5,6,7,3',4'-pentamethoxyflavone (sinensetin) and its analogs were investigated with respect to in vitro efficacy and structure-activity relationship. Sinensetin reversed the resistance of P-glycoprotein (Pgp)-overexpressing AML-2/D100 to vincristine in a concentration-dependent manner. Chemosensitizing effect of sinensetin was 10- and 18-fold higher than those of 5,7,3',4'-tetramethoxyflavone and 3,7-dihydroxy-3',4'-dimethoxyflavone, respectively. Sinensetin cytotoxicity in AML-2/D100 was not changed by the complete inhibition of Pgp, suggesting that it is not a substrate for Pgp. Flow cytometry showed that sinensetin increased drug accumulation in the AML-2/D100 in a concentration-dependent manner. Unlike verapamil and cyclosporin A, the maximum non-cytotoxic concentrations of sinensetin were found to decrease the Pgp levels. Azidopine-binding assay showed that cyclosporin A or verapamil inhibited azidopine binding on Pgp partially but sinensetin did not. Taken together, these results suggest that sinensetin has a chemosensitizing effect in reversing Pgp-mediated MDR by increasing the intracellular accumulation of drugs without competition in a binding site of azidopine. Thus, sinensetin is anticipated as a novel and highly potent second-generation flavonoid chemosensitizer, since sinensetin has significant advantages of having a high therapeutic index, of being a non-transportable inhibitor, and of effecting no induction of Pgp.