A phase II randomized, double-blind, placebo-controlled study of Nuvastatic (C50SEW505OESA), a standardized rosmarinic acid-rich polymolecular botanical extract formulation to reduce cancer-related fatigue in patients with solid tumors.

AIM: We evaluated the efficacy and safety of Nuvastatic™ (C5OSEW5050ESA) in improving cancer-related fatigue (CRF) among cancer patients. METHODS: This multicenter randomized double-blind placebo-controlled phase 2 trial included 110 solid malignant tumor patients (stage II-IV) undergoing chemotherapy. They were randomly selected and provided oral Nuvastatic™ 1000 mg (N = 56) or placebo (N = 54) thrice daily for 9 weeks. The primary outcomes were fatigue (Brief Fatigue Inventory (BFI)) and Visual Analog Scale for Fatigue (VAS-F)) scores measured before and after intervention at baseline and weeks 3, 6, and 9. The secondary outcomes were mean group difference in the vitality subscale of the Medical Outcome Scale Short Form-36 (SF-36) and urinary F2-isoprostane concentration (an oxidative stress biomarker), Eastern Cooperative Oncology Group scores, adverse events, and biochemical and hematologic parameters. Analysis was performed by intention-to-treat (ITT). Primary and secondary outcomes were assessed by two-way repeated-measures analysis of variance (mixed ANOVA). RESULTS: The Nuvastatic™ group exhibited an overall decreased fatigue score compared with the placebo group. Compared with the placebo group, the Nuvastatic™ group significantly reduced BFI-fatigue (BFI fatigue score, F (1.4, 147) = 16.554, p < 0.001, partial η2 = 0.333). The Nuvastatic™ group significantly reduced VAS-F fatigue (F (2, 210) = 9.534, p < 0.001, partial η2 = 0.083), improved quality of life (QoL) (F (1.2, 127.48) = 34.07, p < 0.001, partial η2 = 0.243), and lowered urinary F2-IsoP concentrations (mean difference (95% CI) = 55.57 (24.84, 86.30)), t (55) = 3.624, p < 0.001, Cohen's d (95% CI) = 0.48 (0.20, 0.75)). Reported adverse events were vomiting (0.9%), fever (5.4%), and headache (2.7%). CONCLUSION: Nuvastatic™ is potentially an effective adjuvant for CRF management in solid tumor patients and worthy of further investigation in larger trials. TRIAL REGISTRATION: ClinicalTrial.gov ID: NCT04546607. Study registration date (first submitted): 11-05-2020.

A study on the antioxidant activity of rosmarinic acid against carbon tetrachloride-induced liver toxicity in adult male albino rats / Estudio sobre la actividad antioxidante del ácido rosmarínico contra la toxicidad hepática inducida por tetracloruro de carbono en ratas albinas macho adultas

SUMMARY: Carbon tetrachloride (CCl4) is a manufactured chemical and does not occur naturally in the environment. CCl4 is a clear liquid that evaporates very easily. It has a sweet odor. CCl4 is toxic to the mammalian liver and is hepatocarcinogenic in both rats and mice. Rosemary (Rosmarinus Officinalis) is commonly used as a spice and flavoring agent in food processing. It is known for its antioxidant properties. The present study aims to investigate the antioxidant activity of rosmarinic acid (RA) on CCl4-induced liver toxicity in adult male albino rats. Forty adult male albino rats were divided into 4 groups with 10 rats in each group. Group I (control group). Group II animals received RA at a dose of 50 mg/kg/day by oral gavage for 4 weeks. Group III animals received CCl4 intraperitoneally at a dose of 3ml/kg twice weekly for 4 weeks. Group IV animals received CCl4 Plus RA. At the end of the experiment, liver specimens are processed for histological, immunohistochemical, EM and biochemical studies. Administration of RA deceased the elevated serum liver enzymes (AST, ALT, and ALP), elevated MDA level and immunoexpression of the proapoptotic protein (Bax) induced by CCl4. It increased reduced glutathione (GSH), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), and immunoexpression of the antiapoptotic protein (Bcl2). It also improved the histological and ultrastructural changes induced by CCl4. It appears that Rosmarinic acid has protective effects against CCl4-induced hepatotoxicity as indicated by biochemical, histological, immunohistochemical and ultrastructural results.

RESUMEN: El tetracloruro de carbono (CCl4) es un producto químico fabricado y no se encuentra de forma natural en el medio ambiente. CCl4 es un líquido transparente que se evapora fácilmente; tiene un olor dulce. CCl4 es tóxico para el hígado de los mamíferos y es hepatocarcinogénico tanto en ratas como en ratones. El romero (Rosmarinus officinalis) se usa comúnmente como condimento y agente aromatizante en el procesamiento de alimentos. Es conocido por sus propiedades antioxidantes. El presente estudio tuvo como objetivo investigar la actividad antioxidante del ácido rosmarínico (RA) sobre la toxicidad hepática inducida por CCl4 en ratas albinas macho adultas. Se dividieron cuarenta ratas albinas macho adultas en 4 grupos con 10 ratas en cada grupo. Grupo I (grupo control). Los animales del grupo II recibieron AR a una dosis de 50 mg / kg / día por sonda oral durante 4 semanas. Los animales del grupo III recibieron CCl4 por vía intraperitoneal a una dosis de 3 ml / kg dos veces por semana durante 4 semanas. Los animales del grupo IV recibieron CCl4 Plus RA. Al final del experimento, las muestras de hígado se procesaron para estudios histológicos, inmunohistoquímicos, EM y bioquímicos. La administración de AR eliminó las enzimas hepáticas séricas elevadas (AST, ALT y ALP), el nivel elevado de MDA y la inmunoexpresión de la proteína proapoptótica (Bax) inducida por CCl4. Aumentó el glutatión reducido (GSH), glutatión peroxidasa (GSH-Px), la superóxido dismutasa (SOD) y la inmunoexpresión de la proteína antiapoptótica (Bcl2). También mejoró los cambios histológicos y ultraestructurales inducidos por CCl4. El ácido rosmarínico puede tener efectos protectores contra la hepatotoxicidad inducida por CCl4, tal como lo indican los resultados bioquímicos, histológicos, inmunohistoquímicos y ultraestructurales.

NLRP3 inflammasome in rosmarinic acid-afforded attenuation of acute kidney injury in mice.

Cisplatin (CP) is a well-known anticancer drug used to effectively treat various kinds of solid tumors. CP causes acute kidney injury (AKI) and unfortunately, there is no therapeutic approach in hand to prevent AKI. Several signaling pathways are responsible for inducing AKI which leads to inflammation in proximal convoluted tubule cells in the kidney. Furthermore, the nucleotide-binding oligomerization domain (NOD)-like receptor containing pyrin domain 3 (NLRP3) inflammasome is involved in the CP-induced AKI. In this study, we investigated therapeutic effects of rosmarinic acid (RA) against inflammation-induced AKI. RA was orally administered at the dose of 100 mg/kg for two consecutive days after 24 h of a single injection of CP at the dose of 20 mg/kg administered intraperitoneally in Swiss albino male mice. Treatment of RA inhibited the activation of NLRP3 signaling pathway by blocking the activated caspase-1 and downstream signal molecules such as IL-1ß and IL18. CP activated HMGB1-TLR4/MyD88 axis was also found to be downregulated with the RA treatment. Activation of nuclear factor-κB and elevated protein expression of cyclooxygenase-2 (COX-2) were also found to be downregulated in RA-treated animals. Alteration of early tubular injury biomarker, kidney injury molecule-1 (KIM-1), was found to be subsided in RA-treated mice. RA has been earlier reported for antioxidant and anti-inflammatory properties. Our findings show that blocking a critical step of inflammasome signaling pathway by RA treatment can be a novel and beneficial approach to prevent the CP-induced AKI.

Suppression of alveolar bone resorption by salubrinal in a mouse model of periodontal disease.

AIMS: The relationship between stress to endoplasmic reticulum (ER) and periodontitis has been known, and ER stress induced by Porphyromonas gingivalis results in the loss of alveolar bone. Salubrinal is a small synthetic compound and attenuates ER stress through inhibition of de-phosphorylation of eukaryotic translation initiation factor 2 alpha (eIF2α). In this study, we examined whether salubrinal attenuates periodontitis in a mouse model of experimental periodontal disease. MATERIALS AND METHODS: We evaluated loss of alveolar bone and attachment levels in periodontium using micro-computed tomography (µCT) and hematoxylin-eosin (HE) staining, respectively. Furthermore, we measured osteoclast numbers using tartrate-resistant acid phosphatase (TRAP) staining and osteoblast numbers using HE staining for bone resorption and for bone formation, respectively. To examine the inhibitory effects of salubrinal against pro-inflammatory cytokines, we measured TNF-α and IL1-ß score in periodontium using immunohistostaining. KEY FINDINGS: The results revealed that salubrinal suppressed loss of alveolar bone and attachment levels in periodontium induced by periodontitis. It decreased osteoclast numbers and increased osteoblasts. It also suppressed the expression levels of TNF-α in periodontium. SIGNIFICANCE: These results show that salubrinal alleviates periodontitis through suppression of alveolar bone resorption and the pro-inflammatory cytokine, and promotion of the bone formation. Since salubrinal has been shown to have these beneficial effects for periodontal disease, it may provide a novel therapeutic possibility for the disease.

Rosmarinic acid ameliorates acetaminophen-induced acute liver injury in mice via RACK1/TNF-α mediated antioxidant effect.

CONTEXT: Rosmarinic acid (RA) dose-dependently ameliorates acetaminophen (APAP) induced hepatotoxicity in rats. However, whether RA hepatoprotective effect is by regulating RACK1 and its downstream signals is still unclear. OBJECTIVE: This study explores the RA protective effect on APAP-induced ALI and its mechanism. MATERIALS AND METHODS: Sixty Kunming mice 6-8 weeks old were randomly separated into six groups (n = 10) and pre-treated with normal saline, ammonium glycyrrhetate (AG) or RA (10, 20 or 40 mg/kg i.p./day) for two consecutive weeks. Then, APAP (300 mg/kg, i.g.) was administrated to induce ALI, except for the control. Serum alanine/aspartate aminotransferases (ALT and AST), malondialdehyde (MDA), superoxide dismutase (SOD) and histopathology were used to authenticate RA effect. The liver RACK1 and TNF-α were measured by western blot. RESULTS: Compared with the APAP group, different dosages RA significantly decreased ALT (52.09 ± 7.98, 55.13 ± 10.19, 65.08 ± 27.61 U/L, p < 0.05), AST (114.78 ± 19.87, 115.29 ± 31.91, 101.78 ± 21.85 U/L, p < 0.05), MDA (2.37 ± 0.87, 2.13 ± 0.87, 1.86 ± 0.39 nmol/mg, p < 0.01) and increased SOD (306.178 ± 90.80, 459.21 ± 58.54, 444.01 ± 78.09 U/mg, p < 0.05). With increasing doses of RA, RACK1 and TNF-α expression decreased. Moreover, the RACK1 and TNF-α levels were positively correlated with MDA (r = 0.8453 and r = 0.9391, p < 0.01). DISCUSSION AND CONCLUSIONS: Our findings support RA as a hepatoprotective agent to improve APAP-induced ALI and the antioxidant effect mediated through RACK1/TNF-α pathway.

Synthesis, Antibacterial and Pharmacokinetic Evaluation of Novel Derivatives of Harmine N9-Cinnamic Acid.

A series of 16 new derivatives of harmine N9-Cinnamic acid were synthesized and fully characterized using NMR and MS. The in vitro antibacterial evaluation revealed that most of the synthesized harmine derivatives displayed better antibacterial activities against Gram-positive strains (S. aureus, S. albus and MRSA) than Gram-negative strains (E. coli and PA). In particular, compound 3c showed the strongest bactericidal activity with a minimum inhibitory concentration of 13.67 µg/mL. MTT assay showed that compound 3c displayed weaker cytotoxicity than harmine with IC50 of 340.30, 94.86 and 161.67 µmol/L against WI-38, MCF-7 and HepG2 cell lines, respectively. The pharmacokinetic study revealed that the distribution and elimination of 3c in vivo were rapid in rats with an oral bioavailability of 6.9%.

Rosmarinic acid attenuates acrylamide induced apoptosis of BRL-3A cells by inhibiting oxidative stress and endoplasmic reticulum stress.

Acrylamide (AA) is a common endogenous contaminant in food, with a complex toxicity mechanism. The study on liver damage to experimental animals caused by AA has aroused a great attention. Rosmarinic acid (RosA) as a natural antioxidant shows excellent protective effects against AA-induced hepatotoxicity, but the potential mechanism is still unclear. In the current study, the protective effect of RosA on BRL-3A cell damage induced by AA was explored. RosA increased the activity of SOD and GSH, reduced the content of ROS and MDA, and significantly reduced the oxidative stress (OS) damage of BRL-3A cells induced by AA. RosA pretreatment inhibited the MAPK signaling pathway activated by AA, and down-regulated the phosphorylation of JNK, ERK and p38. RosA pretreatment also reduced the production of calcium ions caused by AA. In addition, the key proteins p-IRE1α, XBP-1s, TRAF2 of the IRE1 pathway, and the expression of endoplasmic reticulum stress (ERS) characteristic proteins GRP78, p-ASK1, Caspase-12 and CHOP were also down-regulated by RosA. NAC blocked the activation of the MAPK signaling pathway and inhibited the ERS pathway. RosA reduced the rate of apoptosis and down-regulated the expression of Bax/Bcl-2 and Caspase-3, thereby inhibiting AA-induced apoptosis. In conclusion, RosA reduced the OS and ERS induced by AA in BRL-3A cells, thereby inhibiting cell apoptosis, and it could be used as a potential protective agent against AA toxicity.

Rosmarinic acid prevents gastric ulcers via sulfhydryl groups reinforcement, antioxidant and immunomodulatory effects.

Rosmarinic acid (RA) is a secondary metabolite present in several plant species that has already demonstrated antioxidant, antiallergic, anticancer, antimicrobial, neuroprotective, and hepatoprotective effects experimentally. Due to the promising pharmacological properties found previously, this study aimed to assess the oral acute toxicity and the gastroprotective effect of RA using animal models. Acute toxicity was assessed according to OECD guide 423. Ethanol, stress, NSAIDs, and pylorus ligature-induced gastric ulcer models were used to investigate antiulcer properties. The related mechanisms of action were also evaluated from ethanol-induced gastric lesions protocol. RA (300 and 2000 mg/kg) showed no changes in behavioral, water and food intake, body and organs weight parameters with LD50 set around 2500 mg/kg. RA presented gastroprotective activity in all assessed doses (25, 50, 100, and 200 mg/kg) using different animal models. Besides, it was observed that this effect is not related to the modulation of gastric juice parameters (pH, volume, and [H+]), the participation of nitric oxide, mucus, and prostaglandins. However, increased sulfhydryl groups, GSH and IL-10 levels as well as reduced of proinflammatory cytokine (TNF-α and IL-1ß) levels were found for RA-treated groups. RA presents low acute toxicity and gastroprotective activity, preventing ulcer formation via cytoprotective, antioxidant, and anti-inflammatory mechanisms. Graphical abstract.

Bioactivation of α,ß-Unsaturated Carboxylic Acids Through Acyl Glucuronidation.

After oral administration to monkeys of [14C]GDC-0810, an α,ß-unsaturated carboxylic acid, unchanged parent and its acyl glucuronide metabolite, M6, were the major circulating drug-related components. In addition, greater than 50% of circulating radioactivity in plasma was found to be nonextractable 12 hours post-dose, suggesting possible covalent binding to plasma proteins. In the same study, one of the minor metabolites was a cysteine conjugate of M6 (M11) that was detected in plasma and excreta (urine and bile). The potential mechanism for the covalent binding to proteins was further investigated using in vitro methods. In incubations with glutathione (GSH) or cysteine (5 mM), GSH and cysteine conjugates of M6 were identified, respectively. The cysteine reaction was efficient with a half-life of 58.6 minutes (k react = 0.04 1/M per second). Loss of 176 Da (glucuronic acid) followed by 129 Da (glutamate) in mass fragmentation analysis of the GSH adduct of M6 (M13) suggested the glucuronic acid moiety was not modified. The conjugation of N-glucuronide M4 with cysteine in buffer was >1000-fold slower than with M6. Incubations of GDC-0810, M4, or M6 with monkey or human liver microsomes in the presence of NADPH and GSH did not produce any oxidative GSH adducts, and the respective substrates were qualitatively recovered. In silico analysis quantified the inherent reactivity differences between the glucuronide and its acid precursor. Collectively, these results show that acyl glucuronidation of α,ß-unsaturated carboxylic acids can activate the compound toward reactivity with GSH, cysteine, or other biologically occurring thiols and should be considered during the course of drug discovery. SIGNIFICANCE STATEMENT: Acyl glucuronidation of the α,ß-unsaturated carboxylic acid in GDC-0810 activates the conjugated alkene toward nucleophilic addition by glutathione or other reactive thiols. This is the first example that a bioactivation mechanism could lead to protein covalent binding to α,ß-unsaturated carboxylic acid compounds.

Evaluation of drug-drug interaction of lusutrombopag, a thrombopoietin receptor agonist, via metabolic enzymes and transporters.

PURPOSE: Drug-drug interaction (DDI) potentials of lusutrombopag, a thrombopoietin receptor agonist, on the activity of cytochrome P450 (CYP) 3A and of cyclosporine, which inhibits P-glycoprotein and breast cancer resistance protein, on lusutrombopag pharmacokinetics were assessed via clinical studies and physiologically based pharmacokinetic (PBPK) modeling. METHODS: The effect of lusutrombopag on midazolam (a CYP3A probe substrate) pharmacokinetics was assessed in 15 healthy subjects receiving a single midazolam 5-mg dose with or without coadministration of lusutrombopag 0.75 mg for 6 days (first dose: 1.5-mg dose). The effect of cyclosporine on lusutrombopag pharmacokinetics was assessed in 16 healthy subjects receiving a single lusutrombopag 3-mg dose with or without a single cyclosporine 400- to 600-mg dose. PBPK modeling was employed to extrapolate the effect of lusutrombopag at the clinical dose (3 mg once daily) on midazolam pharmacokinetics. RESULTS: In the clinical study, mean ratios (90% confidence intervals [CIs]) of with/without lusutrombopag for maximum plasma concentration (Cmax) and area under the plasma concentration-time curve (AUC) of midazolam were 1.01 (0.908-1.13) and 1.04 (0.967-1.11), respectively, indicating no effect of lusutrombopag on midazolam pharmacokinetics. PBPK modeling suggested no effect of lusutrombopag at the clinical dose on midazolam pharmacokinetics. Mean ratios (90% CIs) of with/without cyclosporine for lusutrombopag Cmax and AUC were 1.18 (1.11-1.24) and 1.19 (1.13-1.25), respectively, indicating a slight increase in lusutrombopag exposure. CONCLUSIONS: In consideration with in vitro data, the in vivo and in silico results suggested no clinically significant DDI potential of lusutrombopag with other medical products via metabolic enzymes and transporters.

S(+)-(2E)-N-(2-Hydroxypropyl)-3-Phenylprop-2-Enamide (KM-568): A Novel Cinnamamide Derivative with Anticonvulsant Activity in Animal Models of Seizures and Epilepsy.

Epilepsy is one of the most frequent neurological disorders affecting about 1% of the world's human population. Despite availability of multiple treatment options including antiseizure drugs, it is estimated that about 30% of seizures still remain resistant to pharmacotherapy. Searching for new antiseizure and antiepileptic agents constitutes an important issue within modern medicinal chemistry. Cinnamamide derivatives were identified in preclinical as well as clinical studies as important drug candidates for the treatment of epilepsy. The cinnamamide derivative presented here: S(+)-(2E)-N-(2-hydroxypropyl)-3-phenylprop-2-enamide (S(+)-N-(2-hydroxypropyl)cinnamamide, compound KM-568) showed anticonvulsant activity in several models of epilepsy and seizures in mice and rats. It was active in a genetic animal model of epilepsy (Frings audiogenic seizure-susceptible mouse model, ED50 = 13.21 mg/kg, i.p.), acute seizures induced electrically (maximal electroshock test ED50 = 44.46 mg/kg mice i.p., ED50 = 86.6 mg/kg mice p.o., ED50 = 27.58 mg/kg rats i.p., ED50 = 30.81 mg/kg rats p.o., 6-Hz psychomotor seizure model 32 mA ED50 = 71.55 mg/kg mice i.p., 44 mA ED50 = 114.4 mg/kg mice i.p.), chronic seizures induced electrically (corneal kindled mouse model ED50 = 79.17 mg/kg i.p., hippocampal kindled rat model ED50 = 24.21 mg/kg i.p., lamotrigine-resistant amygdala kindled seizure model in rats ED50 = 58.59 mg/kg i.p.), acute seizures induced chemically (subcutaneous metrazol seizure threshold test ED50 = 104.29 mg/kg mice i.p., ED50 = 107.27 mg/kg mice p.o., ED50 = 41.72 mg/kg rats i.p., seizures induced by picrotoxin in mice ED50 = 94.11 mg/kg i.p.) and the pilocarpine-induced status epilepticus model in rats (ED50 = 279.45 mg/kg i.p., ED97 = 498.2 mg/kg i.p.). The chemical structure of the compound including configuration of the chiral center was confirmed by NMR spectroscopy, LC/MS spectroscopy, elemental analysis, and crystallography. Compound KM-568 was identified as a moderately stable derivative in an in vitro mouse liver microsome system. According to the Ames microplate format mutagenicity assay performed, KM-568 was not a base substitution or frameshift mutagen. Cytotoxicity evaluation in two cell lines (HepG2 and H9c2) proved the safety of the compound in concentrations up to 100 µM. Based on the results of anticonvulsant activity and safety profile, S(+)-(2E)-N-(2-hydroxypropyl)-3-phenylprop-2-enamide could be proposed as a new lead compound for further preclinical studies on novel treatment options for epilepsy.

Improvement of antioxidant activity of oregano (Origanum vulgare L.) with an oral pharmaceutical form.

Aging-related diseases can be triggered by multiple factors such as oxidative stress. Oxidative stress is an imbalance between free radicals and antioxidants, so today, compounds capable of reducing or neutralizing free radicals are being studied for a therapeutic use. Origanum vulgare L. is a traditional medicinal plant used for a wide number of health problems due to its antimicrobial, carminative and antioxidant activities. However, when administered orally, gastrointestinal digestion can modify some of therapeutical properties. To avoid this, two different solid oral formulations have been designed for an O. vulgare extract evaluating their antioxidant behaviours in vitro and in vivo after a simulation of gastrointestinal digestion. The results showed that the divided powder has a lower antioxidant activity both in vitro and in vivo than the encapsulated extract. The quantitative difference of polyphenols found on HPLC-DAD (especially luteolin, apigenin and caffeic acid) may explain the differences in pharmacological activity. Thus, we propose that the best form to administrate O. vulgare extracts to maintain the antioxidant properties is the encapsulated form, that is, two capsules of 250 mg of a hydroalcoholic extract of O. vulgare with a minimum of 33 % of rosmarinic acid as a daily dose.

A Polyphenolic Complex Attenuates Inflammatory Response and Blood- Brain Barrier Disruption.

BACKGROUND: Cerebral ischemia causes a strong inflammatory response. Neumentix is a dietary supplement containing 14.9% rosmarinic acid and 29.9% total phenolic content, which has been proved to be beneficial against inflammatory response. Therefore, Neumentix's effect on anti-inflammatory and blood brain barrier (BBB) disruption in transient middle cerebral artery occlusion (tMCAO) model mice is investigated in this study. METHODS: After the pretreatment of vehicle or Neumentix 134 mg/kg/d, intraperitoneal injection (i.p.) (containing rosmarinic acid 20 mg/kg/d) for 14 days, mice were subjected to tMCAO for 60 min and kept receiving vehicle or Neumentix daily 5 days afterward. RESULTS: Neumentix treatment ameliorated neurobehavioral impairment in the corner test (5d after tMCAO, **P<0.01), reduced infarct volume (#P<0.05), suppressed expression of ionized calciumbinding adapter molecule-1 (Iba-1), tumor necrosis factor alpha (TNF-α) and monocyte chemoattractant protein-1 (MCP-1) (###P<0.001), and improved the integrity of BBB (§P<0.05) at 5 days after tMCAO. CONCLUSION: The present study provided an evidence of Neumentix's anti-inflammatory and neuroprotection effect against BBB disruption on experimental tMCAO model mice, suggesting that Neumentix could be a potential therapeutic agent for stroke.

A Randomized, Open-label, Presurgical, Window-of-Opportunity Study Comparing the Pharmacodynamic Effects of the Novel Oral SERD AZD9496 with Fulvestrant in Patients with Newly Diagnosed ER+ HER2- Primary Breast Cancer.

PURPOSE: Fulvestrant, the first-in-class selective estrogen receptor (ER) degrader (SERD), is clinically effective in patients with ER+ breast cancer, but it has administration and pharmacokinetic limitations. Pharmacodynamic data suggest complete ER degradation is not achieved at fulvestrant's clinically feasible dose. This presurgical study (NCT03236974) compared the pharmacodynamic effects of fulvestrant with AZD9496, a novel, orally bioavailable, nonsteroidal, potent SERD, in treatment-naïve patients with ER+ HER2- primary breast cancer awaiting curative intent surgery. PATIENTS AND METHODS: Patients were randomized 1:1 to receive AZD9496 250 mg twice daily from day 1 for 5-14 days, or fulvestrant 500 mg on day 1. On-treatment imaging-guided core tumor biopsies were taken between day 5 and 14 and compared with pretreatment diagnostic biopsies. The primary objective was to compare the effects of AZD9496 and fulvestrant on ER expression. Secondary objectives included changes in progesterone receptor (PR) and Ki-67 pharmacokinetic/pharmacodynamic relationships and safety. RESULTS: Forty-six women received treatment (AZD9496 n = 22; fulvestrant n = 24); 35 paired biopsies were evaluable (AZD9496 n = 15; fulvestrant n = 20). The least square mean estimate for ER H-score reduction was 24% after AZD9496 versus 36% after fulvestrant treatment (P = 0.86). AZD9496 also reduced PR H-scores (-33.3%) and Ki-67 levels (-39.9%) from baseline, but was also not superior to fulvestrant (PR: -68.7%, P = 0.97; Ki-67: -75.4%, P = 0.98). No new safety findings were identified. CONCLUSIONS: This was the first presurgical study to demonstrate that an oral SERD affects its key biological targets. However, AZD9496 was not superior to fulvestrant at the dose tested.

Dual functional liposomes carrying antioxidants against tau hyperphosphorylation and apoptosis of neurons.

Quercetin (QU) and rosmarinic acid (RA) were loaded in phosphatidic acid-liposomes (QU/RA-PA-liposomes) with surface apolipoprotein E (ApoE) using a process of thin-film hydration, followed by covalent crosslinking to activate biological pathways for penetrating the blood-brain barrier (BBB) and redeeming the neuronal apoptosis from attack of ß-amyloid 1-42 (Aß1-42) and neurofibrillary tangles. The conjugation of liposomes with PA improved the activity of QU and RA against neurotoxicity of Aß1-42. The fluorescent images of brain capillaries revealed that surface modification with ApoE improved the permeation ability of QU/RA-PA-ApoE-liposomes across the BBB. In addition, the highest therapeutic efficacy was obtained in the case of QU/RA-PA-ApoE-liposomes, compared to other QU/RA formulations studied using in vivo Aß1-42-insulted rats mimicking Alzheimer's disease (AD). The cellular and molecular evidence from AD rats included the decrease in Aß1-42 plaque formation and interleukin-6 secretion, increase in the neuronal count in Nissl staining, and reduction in the expression of phosphorylated extracellular signal-regulated kinase 1/2, c-Jun N-terminal kinase, p38 kinase and tau protein at serine 202 as well as caspase-3. The use of PA-ApoE-liposomes as a dual targeting formulation enhances the QU and RA ability to infiltrate the BBB, docks Aß1-42 plaques and can be a potent approach to rescue degenerated neurons from AD.

Rosmarinic acid, the active component of Salvia miltiorrhizae, improves gliquidone transport by regulating the expression and function of P-gp and BCRP in Caco-2 cells.

Salvia miltiorrhiza (Danshen) is typically used in the treatment of diabetic complications and is often co-prescribed with gliquidone in China. However, whether danshen affects the absorption of gliquidone has not been elucidated. In this study, the effects of an aqueous extract of danshen (danshen injection, DSI) and its primary compounds (danshensu, protocatechuic aldehyde, rosmarinic acid and salvianolic acid B) on gliquidone transport across Caco-2 monolayer cells was investigated. DSI enhanced the transport of gliquidone in Caco-2 cell monolayers from the apical (AP) to basolateral (BL) sides and from the BL to AP sides. Rosmarinic acid (RA) also significantly increased the Papp (AP-BL) value for gliquidone transport. Verapamil (a P-gp inhibitor) and Ko143 (a BCRP inhibitor) inhibited the BL-AP transport of gliquidone and promoted the AP-BL transport of gliquidone, whereas MK571 (an MRP1 inhibitor), probenecid (an MRP2 inhibitor), and benzbromarone (an MRP3 inhibitor) had no effect on gliquidone transport. RA also enhanced the intracellular accumulation of Rho123 and Hoechst 33342. The expression of P-gp and BCRP was significantly downregulated, and P-gp ATPase activity was promoted by RA in a dose-dependent manner. These results indicate that an aqueous extract of danshen can increase the transport of gliquidone in Caco-2 cell monolayers and that RA may be the primary compound associated with this activity, which is in agreement with RA simultaneously suppressing the function and expression of P-gp and BCRP.

Role of severe thrombocytopenia in preventing platelet count recovery in thrombocytopenic patients with chronic liver disease.

BACKGROUND AND AIM: Certain thrombocytopenic patients with chronic liver disease have inadequate platelet count recovery after platelet transfusion or lusutrombopag administration. We aimed to identify the reasons for this phenomenon. METHODS: We investigated 58 and 86 thrombocytopenic patients with chronic liver disease who received lusutrombopag (3 mg orally for up to 7 days) or underwent blood transfusions, respectively. Thirty patients underwent simultaneous hepatic surgery and splenectomy. Factors preventing platelet count recovery above 50 × 103 /µL were identified. RESULTS: The median patient age was 64 years. Eleven, 78, and 55 patients had hepatitis B, hepatitis C, or another etiology, respectively; 59, 69, and 16 had Child-Pugh classes A, B, and C, respectively. The median spleen volume was 432 mL, and a median of 10 blood units were transfused per patient. The median platelet count rose significantly (from 41.5 × 103 /µL to 81.0 × 103 /µL) after lusutrombopag administration but not after blood transfusion before invasive procedures. However, maximum platelet counts in patients who underwent splenectomy before platelet transfusion were markedly improved over those who did not. Increasing platelet counts above 50 × 103 /µL required baseline platelets > 30 × 103 /µL and lusutrombopag administration for all patients. Platelet count recovery was dependent on a spleen volume of < 300 mL and baseline platelets of > 40 × 103 /µL in patients who underwent platelet transfusions, while a baseline platelet count of > 30 × 103 /µL was required for patients administered with lusutrombopag. CONCLUSION: Neither blood transfusion nor lusutrombopag improves thrombocytopenia in patients with severe conditions; however, the degree of platelet count elevation following lusutrombopag administration is higher than that following blood transfusion.

The cytotoxic concentration of rosmarinic acid increases MG132-induced cytotoxicity, proteasome inhibition, autophagy, cellular stresses, and apoptosis in HepG2 cells.

Rosmarinic acid (RA) is a natural polyphenolic compound derived from many common herbal plants. Although it is known that RA has many important biological activities, its effect on proteasome inhibitor-induced changes in cancer treatment or its effects on any experimental proteasome inhibition model is unknown. The aim of the study was to investigate the effect of RA on MG132-induced cytotoxicity, proteasome inhibition, autophagy, cellular stresses, and apoptosis in HepG2 cells. HepG2 cells were treated with 10, 100, and 1000 µM RA in the presence of MG132 for 24 h; 10 and 100 µM RA did not affect but 1000 µM RA decreased cell viability in HepG2 cells. MG132 caused a significant decrease in cell viability and phosphorylation of mammalian target of rapamycin and a significant increase in levels of polyubiquitinated protein, microtubule-associated proteins 1A/1B light chain 3B-II (LC3B-II), heat shock protein 70 (HSP70), binding immunoglobulin protein (BiP), activating transcription factor 4 (ATF4), protein carbonyl, and cleaved poly(adenosine diphosphate-ribose) polymerase 1 (PARP1); 10 and 100 µM RA did not significantly change these effects of MG132 in HepG2 cells; 1000 µM RA caused a significant decrease in cell viability and a significant increase in polyubiquitinated protein, LC3B-II, HSP70, BiP, ATF4, protein carbonyl, and cleaved PARP1 levels in MG132-treated cells. Our study showed that only 1000 µM RA increased MG132-induced cytotoxicity, proteasome inhibition, autophagy, cellular stresses, and apoptosis in HepG2 cells. According to our results, cytotoxic concentration of RA can potentiate the effects of MG132 in hepatocellular carcinoma treatment.

GDC-0810 Pharmacokinetics and Transporter-Mediated Drug Interaction Evaluation with an Endogenous Biomarker in the First-in-Human, Dose Escalation Study.

GDC-0810 (Cheeti et al., 2018) is an orally bioavailable, selective estrogen receptor (ER) degrader developed to treat ER-positive breast cancer. A first-in-human (FIH) dose escalation phase I study (n = 41) was conducted to characterize the pharmacokinetics (PK) of GDC-0810 and its two major metabolites. GDC-0810 demonstrated linear PK from 100 to 600 mg given once daily. The mean terminal half-life following a single 600 mg dose was approximately 8 hours. Since GDC-0810 is a potent in vitro inhibitor of organic anion transporting polypeptide (OATP) 1B1/3, the kinetic profile of coproporphyrin I (CPI), a promising endogenous biomarker for OATP1B1/3, was analyzed retrospectively in a subset of the plasma samples collected in the same FIH study. CPI exhibited a GDC-0810 dose-dependent increase, suggesting in vivo inhibition of OATP1B transporters. To quantitatively predict the magnitude of OATP1B-mediated drug-drug interactions (DDIs) with pravastatin (a known OATP1B substrate), the in vivo unbound inhibition constant was first estimated using a one-compartment model, and then incorporated to a physiologically based pharmacokinetic model. The model showed some underestimation of the magnitude of the DDI when compared with a clinical DDI study result, while prediction had a relatively large uncertainty due to the small effect size, limited sample size, and variability in CPI kinetics. In conclusion, this study characterized the pharmacokinetic profiles of GDC-0810 in breast cancer patients and demonstrated the utility of CPI in detecting OATP1B-mediated DDIs of a new molecular entity as early as FIH study. SIGNIFICANCE STATEMENT: Endogenous biomarkers of transporters have recently been shown to be promising tools in evaluating the risk of clinical transporter-mediated DDIs. This is the first study to report a pharmacokinetic interaction between an investigational molecule and a transporter biomarker in a first-in-human study. The observed interaction and model-based analysis and the prediction provide important insights on the novel approach to quantitatively predict transporter-mediated DDIs as early as FIH studies in the clinical development.

A randomized controlled trial of lusutrombopag in Japanese patients with chronic liver disease undergoing radiofrequency ablation.

BACKGROUND: Thrombocytopenia represents an obstacle for invasive procedures in chronic liver disease (CLD) patients. We aimed to estimate the appropriate dose and evaluate the efficacy and safety of lusutrombopag for the treatment of thrombocytopenia before percutaneous liver radiofrequency ablation (RFA) for primary hepatic cancer in patients with CLD. METHODS: In this multicenter, randomized, double-blind, placebo-controlled study conducted in Japan, 61 CLD patients with platelet count < 50 × 103/µL at screening were randomized to placebo or lusutrombopag 2, 3, or 4 mg once daily for 7 days, followed by a 28-day post-treatment assessment period. The primary efficacy endpoint was the proportion of patients who did not require platelet transfusion before RFA. The pre-specified key secondary efficacy endpoint was the proportion of responders. Adverse events (AEs) and thrombosis-related AEs were evaluated. RESULTS: The proportion of patients who did not require platelet transfusion before RFA and that of responders were significantly higher (p < 0.01) in the 2-mg (80.0, 66.7%), 3-mg (81.3, 68.8%), and 4-mg groups (93.3, 80.0%) compared with the placebo group (20.0, 6.7%) and showed a dose-dependent effect. The incidence of AEs was 97.8 and 100% in the lusutrombopag (all groups) and placebo groups, respectively; no dose-related increase was observed. Four patients experienced thrombosis-related events (one each in the placebo and 2-mg groups, and two in the 4-mg group). A total of 16 (18%) adverse drug reactions occurred in the safety analysis set. CONCLUSIONS: Lusutrombopag 3 mg once daily for 7 days was effective without raising concerns about excessive increases in platelet count. CLINICAL TRIAL REGISTRATION: The study is registered at JapicCTI-121944.