Marein, a novel natural product for restoring chemo-sensitivity to cancer cells through competitive inhibition of ABCG2 function.

The pivotal roles of ATP-binding cassette (ABC) transporters in drug resistance have been widely appreciated. Here we report that marein, a natural product from Coreopsis tinctoria Nutt, is a potent chemo-sensitizer in drug resistant cancer cells overexpressing ABCG2 transporter. We demonstrate that marein can competitively inhibit efflux activity of ABCG2 protein and increase the intracellular accumulation of the chemotherapeutic drugs that belong to substrate of this transporter. We further show that marein can bind to the conserved amino acid residue F439 of ABCG2, a critical site for drug-substrate interaction. Moreover, marein can significantly sensitize the ABCG2-expressing tumor cells to chemotherapeutic drugs such as topotecan, mitoxantrone, and olaparib. This study reveals a novel role and mechanism of marein in modulating drug resistance, and may have important implications in treatment of cancers that are resistant to chemotherapeutic drugs that belong to the substrates of ABCG2 transporters.

Dexamethasone Pretreatment Potentiates a Folic Acid-Functionalized Delivery System for Enhanced Lung Cancer Therapy.

Folic acid (FA) has been widely engineered to promote the targeted delivery of FA-modified nanoparticles (NPs) by recognizing the folate receptor α (FRα). However, the efficacy of FA-targeted therapy significantly varied with the abundance of FRα and natural immunoglobulin levels in different tumors. Therefore, a sequential therapy of dexamethasone (Dex)-induced FRα amplification and immunosuppression combined with FA-functionalized doxorubicin (DOX) micelles to synergistically suppress tumor proliferation was proposed in this study. In brief, a pH/reduction-responsive FA-functionalized micelle (FCSD) was obtained by grafting FA, derivatization-modified cholesterol, and 2,3-dimethylmaleic anhydride onto a chitosan oligosaccharide. The obtained FCSD/DOX NPs can effectively deliver DOX in tumors, and their targeting efficiency can be further improved with Dex pretreatment to decrease the immunoglobulin M (IgM) content in serum and amplify FRα levels on the surface of M109 cells. After internalization, charge reversal and disulfide bond breakage of FCSD vectors under the stimulation of tumor extracellular pH (pHe) and intracellular glutathione (GSH) would contribute to the disintegration of vectors and the rapid release of DOX. The sequential therapy that combined Dex pretreatment and targeted chemotherapy by FCSD/DOX NPs demonstrated superior tumor suppression compared with monotherapy, which is expected to provide a potential strategy for FRα-positive lung cancer patients.

Cathepsin L promotes chemresistance to neuroblastoma by modulating serglycin.

Cathepsin L (CTSL), a lysosomal acid cysteine protease, is found to play a critical role in chemosencitivity and tumor progression. However, the potential roles and molecular mechanisms of CTSL in chemoresistance in neuroblastoma (NB) are still unclear. In this study, the correlation between clinical characteristics, survival and CTSL expression were assessed in Versteeg dataset. The chemoresistant to cisplatin or doxorubicin was detected using CCK-8 assay. Western blot was employed to detect the expression of CTSL, multi-drug resistance proteins, autophagy-related proteins and apoptosis-related proteins in NB cells while knocking down CTSL. Lysosome staining was analyzed to access the expression levels of lysosomes in NB cells. The expression of apoptosis markers was analyzed with immunofluorescence. Various datasets were analyzed to find the potential protein related to CTSL. In addition, a subcutaneous tumor xenografts model in M-NSG mice was used to assess tumor response to CTSL inhibition in vivo. Based on the validation dataset (Versteeg), we confirmed that CTSL served as a prognostic marker for poor clinical outcome in NB patients. We further found that the expression level of CTSL was higher in SK-N-BE (2) cells than in IMR-32 cells. Knocking down CTSL reversed the chemoresistance in SK-N-BE (2) cells. Furthermore, combination of CTSL inhibition and chemotherapy potently blocked tumor growth in vivo. Mechanistically, CTSL promoted chemoresistance in NB cells by up-regulating multi-drug resistance protein ABCB1 and ABCG2, inhibiting the autophagy level and cell apoptpsis. Furthermore, we observed six datasets and found that Serglycin (SRGN) expression was positively associated with CTSL expresssion. CTSL could mediate chemoresistance by up-regulating SRGN expression in NB cells and SRGN expression was positively correlated with poor prognosis of NB patients. Taken together, our findings indicate that the CTSL promotes chemoresistance to cisplatin and doxorubicin by up-regulating the expression of multi-drug resistance proteins and inhibiting the autophagy level and cell apoptosis in NB cells. Thus, CTSL may be a therapeutic target for overcoming chemoresistant to cisplatin and doxorubicin in NB patients.

The Correlation Between Busulfan Exposure and Clinical Outcomes in Chinese Pediatric Patients: A Population Pharmacokinetic Study.

Aims: The aims of the study were to 1) establish a population pharmacokinetic (Pop-PK) model for busulfan in Chinese pediatric patients undergoing hematopoietic stem cell transplantation (HSCT) and then estimate busulfan exposure and 2) explore the association between busulfan exposure and clinical outcomes. Methods: A total of 128 patients with 467 busulfan concentrations were obtained for Pop-PK modeling using nonlinear mixed effect model (NONMEM) software. Sixty-three patients who received the 16-dose busulfan conditioning regimen were enrolled to explore the correlations between clinical outcomes and the busulfan area under the concentration-time curve (AUC) using the Cox proportional hazards regression model, Kaplan-Meier method and logistic regression. Results: The typical values for clearance (CL) and distribution volume (V) of busulfan were 7.71 L h-1 and 42.4 L, respectively. The allometric normal fat mass (NFM) and maturation function (Fmat) can be used to describe the variability in CL, and the fat-free mass (FFM) can be used to describe the variability in V. Patients with AUCs of 950-1,600 µM × min had 83.7% (95% CI: 73.3-95.5) event-free survival (EFS) compared with 55.0% (95% CI: 37.0-81.8) for patients with low or high exposure (p = 0.024). The logistic regression analysis results showed no association between transplant-related toxicities and the busulfan AUC (p > 0.05). Conclusions: The variability in busulfan CL was related to the NFM and Fmat, while busulfan V was related to the FFM. Preliminary analysis results suggested that a busulfan AUC of 950-1,600 µM × min was associated with better EFS in children receiving the 16-dose busulfan regimen.

Osthole Increases the Sensitivity of Liver Cancer to Sorafenib by Inhibiting Cholesterol Metabolism.

Osthole is a natural product that has an inhibitory effect on liver cancer, but its effect on the sensitivity of liver cancer to sorafenib is poorly understood. Here, we investigated the effect of osthole and possible sensitization mechanisms. Our results showed that the combination of 2.5 µM sorafenib and 10 µM osthole had significantly synergistic inhibitory effects on proliferation, colony formation, and migration of HCCLM3, sorafenib-resistant HCCLM3 (HCCLM3-SR), and SK-Hep-1 cells. After treatment of HCCLM3 cells-inoculated subcutaneous xenotransplanted tumor mice with 100 mg/kg osthole, 70 mg/kg sorafenib or their combination for 24 day, the tumor volume, tumor weight, and tumor weight coefficient were significantly lower in the osthole + sorafenib group than in the sorafenib group. Compared with the control group, the total cholesterol and low density lipoprotein-cholesterol contents in serum and tumor tissue were significantly decreased in the osthole or osthole + sorafenib groups, the sterol regulatory element binding protein (SREBP)-2c, 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR), and low-density lipoprotein receptor (LDLR) protein expressions in tumor tissue were significantly downregulated as well. In conclusion, osthole can increase the sensitivity of liver cancer to sorafenib, and the mechanism is related to the downregulations of SREBP-2c, HMGCR, and LDLR protein expressions and subsequent inhibition of cholesterol metabolism.

Role of G Protein-Coupled Receptors in Microglial Activation: Implication in Parkinson's Disease.

Parkinson's disease (PD) is one of the prevalent neurodegenerative diseases associated with preferential loss of dopaminergic (DA) neurons in the substantia nigra compacta (SNc) and accumulation of α-synuclein in DA neurons. Even though the precise pathogenesis of PD is not clear, a large number of studies have shown that microglia-mediated neuroinflammation plays a vital role in the process of PD development. G protein-coupled receptors (GPCRs) are widely expressed in microglia and several of them act as regulators of microglial activation upon corresponding ligands stimulations. Upon α-synuclein insults, microglia would become excessively activated through some innate immune receptors. Presently, as lack of ideal drugs for treating PD, certain GPCR which is highly expressed in microglia of PD brain and mediates neuroinflammation effectively could be a prospective source for PD therapeutic intervention. Here, six kinds of GPCRs and two types of innate immune receptors were introduced, containing adenosine receptors, purinergic receptors, metabotropic glutamate receptors, adrenergic receptors, cannabinoid receptors, and melatonin receptors and their roles in neuroinflammation; we highlighted the relationship between these six GPCRs and microglial activation in PD. Based on the existing findings, we tried to expound the implication of microglial GPCRs-regulated neuroinflammation to the pathophysiology of PD and their potential to become a new expectation for clinical therapeutics.

Apigenin increases radiosensitivity of glioma stem cells by attenuating HIF-1α-mediated glycolysis.

Apigenin, a natural flavonoid compound present in a variety of edible plants and health foods, has an anti-tumor effect and inhibits hypoxia inducible factor-lα (HIF-1α) expression in hypertrophic cardiac tissues. However, whether or not apigenin has a radiosensitization effect on glioma stem cells (GSCs) is unknown. Our present study aimed to investigate the effect of apigenin and its possible mechanisms. The human GSCs SU3 and its radioresistance line SU3-5R were treated with apigenin, radiation, or their combination, and the cell proliferation, migration, colony formation, and intracellular lactic acid and glycolytic related protein expressions were determined. Additionally, a cell model with hypoxia-induced HIF-1α expression was used and treated with apigenin. The current results displayed that the combination of apigenin and radiation could synergically reduce the viability, colony formation, and migration of the both GSCs. Moreover, this combination could also decrease the radiation-induced increments of glycolytic production lactic acid in the both GSCs and related protein expressions, including HIF-1α, glucose transporter (GLUT)-1/3, nuclear factor kappa B (NF-κB) p65, and pyruvate kinase isozyme type M2 (PKM2). Further study confirmed that after treatment of hypoxia-cultured SU3 or SU3-5R cells with apigenin, the expression levels of HIF-1α, GLUT-1/3, NF-κB p65, and PKM2 proteins were reduced. These results demonstrated that apigenin could increase the radiosensitivity of GSCs and its radiosensitization mechanisms were attributable to the attenuation of glycolysis, which might result from the inhibition of HIF-1α expression and subsequent reductions of GLUT-1/3, NF-κB, and PKM2 expressions.

PPARα/γ antagonists reverse the ameliorative effects of osthole on hepatic lipid metabolism and inflammatory response in steatohepatitic rats.

Our previous studies have indicated that osthole may ameliorate the hepatic lipid metabolism and inflammatory response in nonalcoholic steatohepatitic rats, but the underlying mechanisms remain unclear. This study aimed to determine whether the effects of osthole were mediated by the activation of hepatic peroxisome proliferator-activated receptor α/γ (PPARα/γ). A rat model with steatohepatitis was induced by orally feeding high-fat and high-sucrose emulsion for 6 weeks. These experimental rats were then treated with osthole (20 mg/kg), PPARα antagonist MK886 (1 mg/kg) plus osthole (20 mg/kg), PPARγ antagonist GW9662 (1 mg/kg) plus osthole (20 mg/kg) and MK886 (1 mg/kg) plus GW9662 (1 mg/kg) plus osthole (20 mg/kg) for 4 weeks. The results showed that after osthole treatment, the hepatic triglycerides, free fatty acids, tumor necrosis factor-α, monocyte chemotactic protein-1, interleukin-6 (IL-6), IL-8 and liver index decreased by 52.3, 31.0, 32.4, 28.9, 36.3, 29.3 and 29.9%, respectively, and the score of steatohepatitis also decreased by 70.0%, indicating that osthole improved the hepatic steatosis and inflammation. However, these effects of osthole were reduced or abrogated after simultaneous addition of the specific PPARα antagonist MK886 or/and the PPARγ antagonist GW9662, especially in the co-PPARα/γ antagonists-treated group. Importantly, the osthole-induced hepatic expressions of PPARα/γ proteins were decreased, and the osthole-regulated hepatic expressions of lipogenic and inflammatory gene proteins were also reversed by PPARα/γ antagonist treatment. These findings demonstrated that the ameliorative effect of osthole on nonalcoholic steatohepatitis was mediated by PPARα/γ activation, and osthole might be a natural dual PPARα/γ activator.

Chrysanthemum morifolium extract improves hypertension-induced cardiac hypertrophy in rats by reduction of blood pressure and inhibition of myocardial hypoxia inducible factor-1alpha expression.

CONTEXT: Chrysanthemum morifolium Ramat. (Asteraceae) extract (CME) possesses a vasodilator effect in vitro. However, the use of polyphenol-rich CME in the treatment of hypertension-induced cardiac hypertrophy has not been reported. OBJECTIVE: We investigated the effect of polyphenol-rich CME on hypertension-induced cardiac hypertrophy in rats and its possible mechanism of action. MATERIALS AND METHODS: The Sprague-Dawley rat model with cardiac hypertrophy was induced by renovascular hypertension. The blood pressure, cardiac weight index, free fatty acids (FFA) in serum and myocardium, and protein expressions of myocardial hypoxia inducible factor-1α (HIF-1α), peroxisome proliferator-activated receptor α (PPARα), carnitine palmitoyltransferase-1a (CPT-1a), pyruvate dehydrogenase kinase-4 (PDK-4) and glucose transporter-4 (GLUT-4) were measured after treating hypertensive rats with polyphenol-rich CME of anthodia 75-150 mg/kg once daily for 4 weeks. A myocardial histological examination was also conducted. RESULTS: After CME treatment, the blood pressure, cardiac weight and cardiac weight index decreased by 5.7-9.6%, 9.2-18.4% and 10.9-20.1%, respectively, and the cardiomyocyte cross-sectional area also decreased by 8.3-30.4%. The CME treatment simultaneously decreased the FFA in serum and myocardium and protein expressions of myocardial HIF-1α and GLUT-4, and increased the protein expressions of myocardial PPARα, CPT-1a and PDK-4, especially in the CME 150 mg/kg group (p < 0.05 or p < 0.01). DISCUSSION AND CONCLUSION: Polyphenol-rich CME may alleviate hypertensive cardiac hypertrophy in rats. Its mechanisms may be related to the reduction of blood pressure and amelioration of the myocardial energy metabolism. The latter may be attributed to the inhibition of HIF-1α expression and subsequent modulation of PPARα-mediated CPT-1a, PDK-4 and GLUT-4 expressions.

Apigenin ameliorates hypertension-induced cardiac hypertrophy and down-regulates cardiac hypoxia inducible factor-lα in rats.

Apigenin is a natural flavonoid compound that can inhibit hypoxia-inducible factor (HIF)-1α expression in cultured tumor cells under hypoxic conditions. Hypertension-induced cardiac hypertrophy is always accompanied by abnormal myocardial glucolipid metabolism due to an increase of HIF-1α. However, whether or not apigenin may ameliorate the cardiac hypertrophy and abnormal myocardial glucolipid metabolism remains unknown. This study aimed to examine the effects of apigenin. Rats with cardiac hypertrophy induced by renovascular hypertension were treated with apigenin 50-100 mg kg(-1) (the doses can be achieved by pharmacological or dietary supplementation for an adult person) by gavage for 4 weeks. The results showed that after treatment with apigenin, the blood pressure, heart weight, heart weight index, cardiomyocyte cross-sectional area, serum angiotensin II, and serum and myocardial free fatty acids were reduced. It is important to note that apigenin decreased the expression level of myocardial HIF-1α protein. Moreover, apigenin simultaneously increased the expression levels of myocardial peroxisome proliferator-activated receptor (PPAR) α, carnitine palmitoyltransferase (CPT)-1, and pyruvate dehydrogenase kinase (PDK)-4 proteins and decreased the expression levels of myocardial PPARγ, glycerol-3-phosphate acyltransferase genes (GPAT), and glucose transporter (GLUT)-4 proteins. These findings demonstrated that apigenin could improve hypertensive cardiac hypertrophy and abnormal myocardial glucolipid metabolism in rats, and its mechanisms might be associated with the down-regulation of myocardial HIF-1α expression and, subsequently increasing the expressions of myocardial PPARα and its target genes CPT-1 and PDK-4, and decreasing the expressions of myocardial PPARγ and its target genes GPAT and GLUT-4.

Involvement of adenosine monophosphate-activated protein kinase in the influence of timed high-fat evening diet on the hepatic clock and lipogenic gene expression in mice.

A high-fat diet may result in changes in hepatic clock gene expression, but potential mechanisms are not yet elucidated. Adenosine monophosphate-activated protein kinase (AMPK) is a serine/threonine protein kinase that is recognized as a key regulator of energy metabolism and certain clock genes. Therefore, we hypothesized that AMPK may be involved in the alteration of hepatic clock gene expression under a high-fat environment. This study aimed to examine the effects of timed high-fat evening diet on the activity of hepatic AMPK, clock genes, and lipogenic genes. Mice with hyperlipidemic fatty livers were induced by orally administering high-fat milk via gavage every evening (19:00-20:00) for 6 weeks. Results showed that timed high-fat diet in the evening not only decreased the hepatic AMPK protein expression and activity but also disturbed its circadian rhythm. Accordingly, the hepatic clock genes, including clock, brain-muscle-Arnt-like 1, cryptochrome 2, and period 2, exhibited prominent changes in their expression rhythms and/or amplitudes. The diurnal rhythms of the messenger RNA expression of peroxisome proliferator-activated receptorα, acetyl-CoA carboxylase 1α, and carnitine palmitoyltransferase 1 were also disrupted; the amplitude of peroxisome proliferator-activated receptorγcoactivator 1α was significantly decreased at 3 time points, and fatty liver was observed. These findings demonstrate that timed high-fat diet at night can change hepatic AMPK protein levels, activity, and circadian rhythm, which may subsequently alter the circadian expression of several hepatic clock genes and finally result in the disorder of hepatic lipogenic gene expression and the formation of fatty liver.