Structurally diverse amides from Chloranthus henryi var. hupehensis and their anti-inflammatory activities by blocking Akt phosphorylation.

Eleven new amides, four racemic pairs of (±)-chlorahupetamides A, B, D, E (1, 2, 4, 5) and chlorahupetamides C, F, G (3, 6, 7), have been isolated from Chloranthus henryi var. hupehensis. Compounds 1-3 are the first naturally occurring dimers via an unprecedented [2 + 2] cycloaddition derived from two dissimilar cinnamic acid amides, while compounds 4 and 5 represent the first examples of lignanamides in Chloranthus; together with two new hydroxycinnamic acid amide monomers (6-7), these compounds were obtained. Their structures were characterized by nuclear magnetic resonance (NMR), electronic circular dichroism (ECD), and X-ray diffraction analysis. Meanwhile, an LPS-induced BV-2 cell inflammatory model was used to determine the potential anti-inflammatory activity of all the isolated compounds. Intriguingly, compound -1 treatment showed a much greater inhibition of TNF-α expression with an EC50 value of 1.80 µM, while compound + 1 had more advantages in reducing IL-1ß expression with an EC50 value of 19.93 µM. Moreover, compounds + 1 and -1 could significantly suppress inflammation and inhibit the Akt signaling pathway by decreasing the phosphorylated protein levels of Akt.

Discovery of sesquiterpenoids from the roots of Chloranthus henryi Hemsl. var. hupehensis (Pamp.) K. F. Wu and their anti-inflammatory activity by IKBα/NF-κB p65 signaling pathway suppression.

Phytochemical analysis of Chloranthus henryi var. hupehensis roots led to the identification of a new eudesmane sesquiterpenoid dimer, 18 new sesquiterpenoids, and three known sesquiterpenoids. Among the isolates, 1 was a rare sesquiterpenoid dimer that is assembled by a unique oxygen bridge (C11-O-C8') of two highly rearranged eudesmane-type sesquiterpenes with the undescribed C16 carbon framework. (+)-2 and (-)-2 were a pair of new skeleton dinorsesquiterpenoids with a remarkable 6/6/5 tricyclic ring framework including one γ-lactone ring and the bicyclo[3.3.1]nonane core. Their structures were elucidated using spectroscopic data, single-crystal X-ray diffraction analysis, and quantum chemical computations. In the LPS-induced BV-2 microglial cell model, 17 suppressed IL-1ß and TNF-α expression with EC50 values of 6.81 and 2.76 µM, respectively, indicating its excellent efficacy in inhibiting inflammatory factors production in a dose dependent manner and without cytotoxicity. In subsequent mechanism studies, compounds 3, 16, and 17 could reduce IL-1ß and TNF-α production by inhibiting IKBα/p65 pathway activation.

Bibenzyl and naphthalene derivatives from Dendrobium chrysanthum and their anti-hepatic-steatosis activities.

In this study, 16 new compounds, six bibenzyls (1-6) and 10 naphthalenes (7-13), including three pairs of naphthalene enantiomers and three known compounds (14-16), were isolated from Dendrobium chrysanthum. Structurally, compounds 1-5 are previously undescribed dimeric bibenzyls, uniquely linked by unusual carbon bonds. The structures of the compounds were determined using spectroscopy and X-ray crystallography. The screening results indicated that 1, 2, and 5 showed remarkable lipid-lowering activities in FFA-induced HepG2 cells, with EC50 values ranging from 3.13 to 6.57 µM. Moreover, 1, 2, and 5 significantly decreased both the mRNA and protein levels of the target SREBP-1c, and 5 also reduced PPARα mRNA and protein levels. Therefore, 1, 2, and 5 are potential drugs against hepatic steatosis by targeting PPARα or SREBP-1c.

[Traditional Chinese medicine health care from "long term taking" drugs in Shen Nong's Classic of Materia Medica].

"Taking drugs for a long term" is a qualitative expression of medication method based on the efficacy and safety of Chinese medicine, and the study on it is conducive to the full utilization of the efficacy and rational use of drugs. There are 148 drugs that can be taken for a long time recorded in Shen Nong's Classic of Materia Medica, accounting for 41% of the total drugs. This paper analyzed three-grade classification, natural qualities, four properties and five flavors, and efficacy features of the "long-term taking" drugs(LTTD), thus exploring the herbal source of traditional Chinese medicine health care and the rationality of effect accumulation by long-term taking. It was found that there were more than 110 top-grade LTTD in Shen Nong's Classic of Materia Medica, most of which were herbs, with sweet flavor, flat property, and no toxicity. The efficacies were mainly making body feel light and agile(Qingshen) and prolonging life. Eighty-three LTTD were included in the Chinese Pharmacopoeia(2020 edition). In the modern classification, tonic LTTD accounted for the most, followed by damp-draining diuretic LTTD and exterior-releasing LTTD. Twenty LTTD were included in the "List of Medicinal and Edible Products" and 21 were in the "List of Products Used for Health-care Food", involving in various modern health care effects, such as enhancing immunity, assisting in reducing blood lipids, and anti-oxidation. Shen Nong's Classic of Materia Medica is the classic source of traditional Chinese medicine health care, and its medication thought of taking drugs for a long term to accumulate effects has guiding significance for the regulation of sub-health and chronic diseases nowadays. The efficacy and safety of LTTD have been examined in practice for a long time, and some of the drugs are edible, which is unique in the whole cycle of health-care service, especially in line with the health-care needs in the aging society under the concept of Big Health. However, some records in the book are limited by the understanding of the times, which should be scientifically studied according to the Chinese Pharmacopoeia and the related regulations and technical requirements, under the attitude of eliminating falsifications and preserving the truth and keeping the right essence, so as to achieve further improvement, innovation, and development.

Chlorahupetenes A-D, Four Eudesmane-Type Sesquiterpenoid Dimer Enantiomers with Two Unusual Carbon Skeletons from Chloranthus henryi Var. hupehensis.

(+)- and (-)-Chlorahupetenes A (1a and 1b), B (2a and 2b), C (3a and 3b), and D (4a and 4b), four unique enantiomeric pairs of eudesmane-type sesquiterpenoid dimers with two new carbon skeletons, were isolated from the aerial parts of Chloranthus henryi var. hupehensis. Compounds 1 and 2 possess an unprecedented 6/6/5/6/6 pentacyclic carbon skeleton with a new dimerization pattern of two eudesmane-type sesquiterpenoids. Compounds 3 and 4, which are fused with two eudesmane-type sesquiterpenoids via an unprecedented five-membered O-heterocyclic ring, represent a new 6/6/5/5/6/6/5 heptacyclic ring system. The structures of the compounds were determined through spectroscopic data and X-ray crystallography. Compounds 1a-3b significantly inhibited NO production with IC50 values ranging from 9.62 to 12.91 µM. Moreover, compounds 1b and 3a suppressed the production of a proinflammatory mediator (TNF-α) and enzyme expression (iNOS) at the mRNA level.

Transient 2D Junction Temperature Distribution Measurement by Short Pulse Driving and Gated Integration with Ordinary CCD Camera.

The time resolution of the transient process is usually limited by the minimum exposure time of the high-speed camera. In this work, we proposed a method that can achieve nanosecond temporal resolution with an ordinary CCD camera by driving the LED under test with a periodic short-pulse signal and multiple-cycle superposition to obtain two-dimensional transient junction temperature distribution of the heating process. The temporal resolution is determined by the pulse width of the drive source. In the cooling process, the Boxcar gated integration principle is adopted to complete the two-dimensional transient junction temperature distribution with temporal resolution subject to the minimum exposure time of the CCD camera, i.e., 1 µs in this case. To demonstrate the validity of this method, we measured the two-dimensional transient junction temperature distribution of the blue LEDs according to the principle of thermoreflectance and compared it with the thermal imaging method.

A novel function of CYP21A2 in regulating cell migration and invasion via Wnt signaling.

CYP21A2, which is responsible for 21-hydroxylase activity, is prominent to the development of congenital adrenal hyperplasia (CAH). The aim of our current study is to investigate the role of CYP21A2 in the tumor processes. Here, we used HepG2 cell lines and generated CYP21A2 overexpressing vector and siRNA to investigate the effect of CYP21A2 on the tumor development processes, particularly cell migration and invasion; genes expression related to these processes were further examined. Results showed that CYP21A2 over-expressed or silenced had no effects on cell viability as well as the process of cell apoptosis. Further study suggested that CYP21A2 silenced significantly decreased the G0/G1 phase and increased the S phase of the cell cycle. However, no differences were observed when CYP21A2 was overexpressed. Moreover, we found that cell migration and invasion significantly improved with CYP21A2 overexpressed and impaired with silenced CYP21A2. Finally, we examined the expression of genes related to tumor processes and found that the Wnt signaling genes were changed. Taken together, our results demonstrated a novel function of CYP21A2 in the regulation of tumor processes, particularly cell migration and invasion, which this may be mediated by the Wnt signaling pathway.

Primate-specific miRNA-637 inhibited tumorigenesis in human pancreatic ductal adenocarcinoma cells by suppressing Akt1 expression.

As a primate-specific microRNA, miR-637 has been discovered for nearly 10 years. Our previous study demonstrated that miR-637 acted as a suppressor in hepatocellular carcinoma. However, its biomedical significance in pancreatic cancer remains obscure. In the present study, miR-637 was found to be significantly downregulated in pancreatic ductal adenocarcinoma (PDAC) cell lines and most of the PDAC specimens. Furthermore, the enforced overexpression of miR-637 dramatically inhibited cell proliferation and induced apoptosis of PDAC cells. Akt1, as a serine/threonine-protein kinase, has been identified as an oncogene in multiple cancers including pancreatic cancer. Our data confirmed that Akt1 was a novel target for miR-637, and its knockdown also induced cell growth inhibition and apoptosis in PDAC cells. In conclusion, our data indicated that miR-637 acted as a tumor-suppressor in PDAC, and the suppressive effect was mediated, at least partially, by suppressing Akt1 expression.

Comparative pharmacokinetics and metabolites study of seven major bioactive components of Shaoyao-Gancao decoction in normal and polycystic ovary syndrome rats by ultra high pressure liquid chromatography with tandem mass spectrometry.

A simple and sensitive liquid chromatography with tandem mass spectrometry method was developed for simultaneous quantification of paeoniflorin, albiflorin, oxypaeoniflorin, liquiritin, liquiritigenin, glycyrrhetinic acid, and glycyrrhizin in rat plasma after oral administration of Shaoyao-Gancao decoction, which is traditionally used in the treatment of polycystic ovary syndrome. The plasma samples were pretreated with methanol as precipitant. The method exhibited good linearity (correlation coefficient (R2 ) > 0.99) with lower quantification limits of 0.595-4.69 ng/mL for all analytes. Intra- and interbatch precision, accuracy, recovery, and stability of the method were all within accepted criteria. The results showed that the pharmacokinetic behaviors of the seven compounds were altered in the pathological status of polycystic ovary syndrome. Furthermore, a total of 36 metabolites were structurally identified based on their accurate masses and fragment ions. The major metabolic pathway involves phase I metabolic reactions (such as hydroxylation), phase II metabolic reactions (such as sulfation and glucuronidation conjugation) as well as the combined multiple-step metabolism. This study is the first report on the pharmacokinetic and metabolic information of Shaoyao-Gancao decoction in both normal and model rats, which would provide scientific evidences for the bioactive chemical basis of herbal medicines and also promote the clinical application of Shaoyao-Gancao decoction for treating polycystic ovary syndrome.

[Toxic effects of combination of Aconiti Lateralis Radix Praeparata with Trichosanthis Fructus on inflammatory response and myocardial fibrosis in pressure overload rats based on ß2-AR/PKA signal].

To study the effects of combination of Aconiti Lateralis Radix Praeparata( Fuzi) with Trichosanthis Fructus( Gualou) on cardiac function,electrocardiogram,inflammatory response and myocardial fibrosis in pressure overload( PO) rats,and further explore the mechanism based on ß2-AR/PKA signaling. PO rat model was established by constricting the abdominal aorta. Twelve weeks after the operation,these rats were randomly divided into model goup( PO),low dose Fuzi group( FL,5. 4 g·kg-1·d-1),Gualou group( GL,5. 4 g·kg-1·d-1),Fuzi and Gualou combination group( FG,5. 4 g·kg-1·d-1+5. 4 g·kg-1·d-1) and high dose Fuzi group( FH,10. 8 g·kg-1·d-1). At the same time,sham operation group was set. After intervention for 6 weeks,carotid blood pressure,cardiac function,electrocardiogram and heart mass index were measured. HE staining was used to observe the inflammatory response in the rat heart and kidney. Masson staining was used to determine the myocardial fibrosis. Western blot was used to detect the protein expression of ß2-AR and PKA. As compared with sham operation group,the blood pressure and heart mass index were obviously increased in PO model group,but there was no significant difference in various treatment groups in the above indexes. As compared with PO model group,FH treatment significantly increased the ejection fraction( EF) and GL treatment effectively enhanced the cardiac output( CO),but other treatment groups had no significant effect on these parameters. Moreover,FG treatment can synergistically attenuate QT and QTc internal prolongation,but it also aggravated inflammatory response in the heart and kidney tissues and promoted myocardial fibrosis as compared to FZ or GL alone treatment,with toxic effects equivalent to FH treatment group. Following FG and FH treatment,simultaneously,ß2-AR and PKA protein levels were significantly elevated,indicating that the increasing toxicity of FG could be associated with activation of ß2-AR/PKA signaling. These results suggested that combination of FZ and GL could synergistically enhance toxicity of FZ in special pathological states such as pressure overload,and caution should be taken in clinical application.

Elevated serum levels of TNF-α, IL-8, and ECP can be involved in the development and progression of bronchial asthma.

OBJECTIVE: This study aimed to explore the value of elevated serum levels of tumor necrosis factor-α (TNF-α), interleukin-8 (IL-8), and eosinophil cationic protein (ECP) in the diagnosis of bronchial asthma (BA). METHODS: A total of 170 patients with BA (case group, 85 patients in acute attack and 85 patients in clinical remission) and 150 healthy individuals (control group) were enrolled in this study. Enzyme-linked immunosorbent assay and receiver operating characteristic (ROC) curves were calculated for the contents and diagnostic values of serum TNF-α, IL-8, and ECP in BA. RESULTS: Compared with the control group, patients in acute attack and clinical remission had higher TNF-α, IL-8, and ECP levels (p < 0.05). The serum level of TNF-α was positively correlated with IL-8 and ECP (p < 0.05). ROC curves showed that the diagnostic threshold value of IL-8 was 13.53 ng/ml, its area under the curve (AUC) was 0.87, its specificity was 99.3%, and its sensitivity was 57.6%. The diagnostic threshold value of TNF-α was 1.29 ng/ml with AUC being 0.94, specificity was 89.3%, and sensitivity was 83.5%. ECP showed 7.22 ng/ml diagnostic threshold value (AUC = 0.88, specificity = 74.0%, sensitivity = 86.5%). The FEV1/pre(%) and FEV1/FVC were negatively correlated and the Z5/pre(%) and resonance frequency (Fres) were positively correlated with the serum levels of TNF-α, IL-8, and ECP in patients in acute attack and in clinical remission (all p < 0.05). CONCLUSION: Our findings reveal that elevated serum levels of TNF-α, IL-8, and ECP can be involved in the development and progression of BA.

Bile acid malabsorption is associated with diarrhea in acute phase of colitis.

The enterohepatic circulation of bile acids (BAs) critically depends on BA transporters and enzymes, which can be affected by inflammatory bowel disease. Diarrhea in colitis is believed to result in part from BA malabsorption. The work aimed to investigate whether diarrhea in colitis was associated with the expression of BA transporters, enzymes, and nuclear receptors. RT-qPCR and Western blot techniques were used to evaluate the gene and protein levels of Cyp7a1, Asbt, SHP, FXR, Ostß in a 2,4,6-trinitrobenzenesulfonic-acid-induced rat model of colitis. The total BAs (TBAs) levels were assayed using ELISA kits, and the individual BAs were measured by LC-MS/MS. Results showed that the fecal excretions of TBAs were significantly increased by 1.6-fold in acute stage of colitis. In ileum, Asbt was significantly decreased; however, there was a compensatory increase in Cyp7a1 level in liver. Moreover, FXR has a decreased tendency and the downstream target gene SHP was downregulated. Contrary to acute stage, molecular changes were completely reversible during the remission phase. Our results indicated that the expression of Asbt and Cyp7a1 were altered in acute colitis, which performed vital roles in maintaining BA homeostasis. Early medical manipulation of BA transporters and enzymes may help prevent diarrhea.

Synergistic Promotion on Tyrosinase Inhibition by Antioxidants.

When exposed to ultraviolet radiation, the human skin produces profuse reactive oxygen species (ROS), which in turn activate a variety of biological responses. Mounting ROS levels activate tyrosinase by mobilizing α-melanocyte-stimulating hormone in the epidermis and finally stimulates the melanocytes to produce melanin. Meanwhile, the Keap1-Nrf2/ARE pathway, which removes ROS, is activated at increased ROS levels, and antioxidant compounds facilitates the dissociation of Nrf2. In this study, we explored the possible suppressing effects of antioxidant compounds and tyrosine inhibitors on melanin formation and the promotory effects of these compounds on ROS scavenging. The antioxidant activity of glabridin (GLA), resveratrol (RES), oxyresveratrol (OXYR), and phenylethylresorcinol (PR) were investigated via the stable free radical 2,2-diphenyl-1-picrylhydrazyl method. The inhibitory effects of the four compounds and their mixtures on tyrosinase were evaluated. l-Tyrosine or 3-(3,4-dihydroxyphenyl)-l-alanine (l-DOPA) was used as a substrate. The results showed that all mixtures did not exhibit synergistic effects with the l-tyrosine as a substrate, suggesting that l-tyrosine is not suitable as a substrate. However, the mixtures of "GLA:RES," "GLA:OXYR," "OXYR:RES," and "PR:RES" demonstrated synergistic effects (CI < 0.9, p < 0.05), whereas "GLA:RES" and "PR:OXYR" indicated an additive effect (0.9 ditive1, p < 0.05). Furthermore, we used a molecular docking strategy to study the interactions of the four compounds with tyrosinase and l-DOPA. The molecular docking result is consistent with that of the experiment. Finally, we selected RES + OXYR and used PIG1 cells to verify whether OXYR synergistically promotes RES activity on tyrosinase. The two agents had a synergistic inhibitory effect on tyrosinase activity. These results provided a novel synergistic strategy for antioxidants and tyrosinase inhibitors, and this strategy is useful in skin injury treatment.

[Study on chemical constituents of Codonopsis pilosula].

A new neolignan, (-)-(7R,8S,7'E)-3',4-dihydroxy-3-methoxy-8,4'-oxyneoligna-7'-ene-7,9,9'-triol(1), and seven known compounds, 9-(tetrahydropyran-2-yl)-nona-trans,trans-2,8-diene-4,6-diyn-1-ol (2), 9-(tetrahydropyran-2-yl)-trans-non-8-ene-4,6-diyn-1-ol (3), lobetyol (4), lobetyolin (5),dehydrodieoniferyl alcohol (6), 5-hydroxymethylfurfural (7), and 4, 4'-dihydroxy-3, 3'-dimethoxy-trans-stilbene (8), were isolated from the H2O extract of Codonopsis pilosula. The structures of 1-8 were elucidated by spectroscopic methods including NMR, HR-ESI-MS, and CD. In addition, compounds 2 and 3 were isolated from the genus Codonopsis for the first time.

[Effect of Qishen Yiqi droplet on adenylate contents in myocardium of cor pulmonale rats].

Cor pulmonale rat models were induced by a single intraperitoneal injection of monocrotaline(MCT), and the sham group received a single intraperitioneal injection of normal saline. After the model rats received intragastric administration of Qishen Yiqi droplet(QS) for 6 weeks, the contents of adenylate(ATP, ADP and AMP) in right myocardial tissues were measured by HPLC, and then the metabolism changes in myocardium of cor pulmonale rats with QS were investigated. The results showed that ATP, ADP, and AMP were well separated, with a good linearity within a certain range of concentration; and the recovery rates were within the range of 90%-108%. As compared with model group, the level of ATP was significantly elevated in high-dose treatment group; ADP contents showed an increasing trend and AMP contents showed a decreasing trend, indicating that QS could significantly improve energy metabolism system in myocardium. By using the HPLC, a qualitative and quantitative analysis method was given for the determination of ATP, ADP and AMP contents in myocardium, providing a method for energy metabolism measurement in biological samples.

Looking for agonists of ß2 adrenergic receptor from Fuzi and Chuanwu by virtual screening and dual-luciferase reporter assay.

More and more studies demonstrated that ß2 adrenergic receptor (ß2-AR) plays a crucial role for the treatment of heart failure. Chuanwu and Fuzi have been used over thousands of years in China for the treatment of heart failure. Considering the effects of these herbs are very similar to ß2-AR agonists, we presume whether ß2-AR agonists can be found from Fuzi and Chuanwu. Fuzi and Chuanwu decoction were used to receive the luciferase reporter activity assay to verify the hypothesis, and the result is positive and encouraging. For it is very difficult to get all of the monomer compounds of Fuzi and Chuanwu, virtual screening was used to find potential ß2-AR agonists and a cell-based ß2-AR agonist functional evaluation model, combined with a luciferase reporter assay system, was used to confirm the final result. In this research, 45 compounds were identified as ß2-AR agonists, and four compounds were verified and the rest need further experiment.

Higher proliferation of peritumoral endothelial cells to IL-6/sIL-6R than tumoral endothelial cells in hepatocellular carcinoma.

BACKGROUND: This study aimed to explore the responses to the interleukin-6 (IL-6)/soluble interleukin-6 receptor (sIL-6R) complex in peritumoral endothelial cells (PECs) and tumor endothelial cells (TECs), as well as determine the signaling pathways in the angiogenesis of hepatocellular carcinoma (HCC). METHODS: The expression of IL-6, IL-6R, gp130, CD68, HIF-1α, and microvessel density (MVD) were assessed with an orthotopic xenograft model in nude mice. ECs were incubated under hypoxic conditions to detect IL-6 and gp130. The proliferation of PECs and TECs in the presence of IL-6 and sIL-6R, as well as the expression of gp130, JAK2/STAT3, PI3K/AKT in endothelial cells were measured. RESULTS: Peritumoral IL-6, IL-6R, gp130, CD68, and HIF-1α expression, as well as MVD, gradually increased during tumor growth. Hypoxia could directly induce IL-6 expression, but not gp130 in PECs. The co-culture of IL-6/sIL-6R induced much higher PEC proliferation and gp130 expression, as well as the elevated phosphorylation of JAK2 and STAT3, however not the phosphorylation of PI3K and AKT. CONCLUSIONS: PECs exhibited higher proliferation in response to IL-6/sIL-6R co-treatment compared with TECs in HCC via the up-regulation of gp130 /JAK2/STAT3. PEC and its associated peritumoral angiogenesis microenvironment may be a potential novel target for anti-angiogenic treatment.

ZBTB20 is involved in liver regeneration after partial hepatectomy in mouse.

BACKGROUND: A better understanding of the molecular mechanisms in liver regeneration holds promise for exploring the new potential therapy for liver failure. The present study was to investigate the role of zinc finger and BTB domain-containing protein 20 (ZBTB20), a potential factor associated with liver regeneration, in a model of 70% hepatectomy in mice. METHODS: Parameters for liver proliferation such as liver/body ratio and BrdU positivity were obtained via direct measurement and immunohistochemistry. The levels of zinc fingers and homeoboxes 2 (ZHX2), ZBTB20, alpha-fetoprotein (AFP) and glypican 3 (GPC3) transcripts in the regenerating liver tissue of a 70% hepatectomy rodent model were monitored by real-time PCR analysis at different time points. Knockdown of ZBTB20 was performed to characterize its regulatory function. RESULTS: A negatively regulating relationship between ZHX2, ZBTB20 and AFP, GPC3 was revealed from 24 to 72 hours after 70% hepatectomy. ZBTB20 appears to negatively regulate AFP and GPC3 transcription since the knockdown of ZBTB20 promoted the proliferation of hepatocytes and the expression of AFP and GPC3. CONCLUSION: In addition to AFP, GPC3 and ZHX2, ZBTB20 is a new regulator in liver regeneration and the decrease of ZBTB20 expression following 70% hepatectomy promotes AFP and GPC3 expression.

Antiangiogenic therapy promoted metastasis of hepatocellular carcinoma by suppressing host-derived interleukin-12b in mouse models.

Antiangiogenic therapy, specially sorafenib, has become the standard of care for patients with advanced hepatocellular carcinoma (HCC), however, the improvement in survival time is not satisfactory. Previous studies have found that, in some circumstances, antiangiogenic therapy promoted tumor metastasis and the mechanistic studies were mainly focus on cancer-cell-autonomous manners. In two experimental metastasis models with tail-vein injection with hepatoma cells and an orthotopic HCC mouse model, we found that pretreatment with two vascular endothelial growth factor receptor (VEGFR) inhibitors, sunitinib and sorafenib, facilitated tumor cell survival in blood stream and promoted lung metastasis from tumors that were subsequently incubated after drug discontinuation, indicating that host response joined into the pro-metastatic effects. An antibody microarray identified that interleukin (IL)-12b was decreased in the peripheral blood of the mice treated with the two VEGFR inhibitors. IL-12b suppression in macrophages and dendritic cells from host organs was found to play a crucial role in treatment-induced metastasis. Supplement with recombinant mouse IL-12b or restoration of IL-12b expression in the host by zoledronic acid, which was previously reported to enhance IL-12 expression in vitro and in vivo, alleviated the metastasis-promoting effects of sunitinib and sorafenib. These studies suggest that host response to VEGFR inhibitors facilitates HCC metastasis and restoration of IL-12b expression could translate into clinical benefits.

Sorafenib down-regulates expression of HTATIP2 to promote invasiveness and metastasis of orthotopic hepatocellular carcinoma tumors in mice.

BACKGROUND & AIMS: Antiangiogenic agents can sometimes promote tumor invasiveness and metastasis, but little is known about the effects of the antiangiogenic drug sorafenib on progression of hepatocellular carcinoma (HCC). METHODS: Sorafenib was administered orally (30 mg · kg(-1) · day(-1)) to mice with orthotopic tumors grown from HCC-LM3, SMMC7721, or HepG2 cells. We analyzed survival times of mice, along with tumor growth, metastasis within liver and to lung, and induction of the epithelial-mesenchymal transition. Polymerase chain reaction arrays were used to determine the effects of sorafenib on gene expression patterns in HCC cells. We analyzed regulation of HIV-1 Tat interactive protein 2 (HTATIP2) by sorafenib and compared levels of this protein in tumor samples from 75 patients with HCC (21 who received sorafenib after resection and 54 who did not). RESULTS: Sorafenib promoted invasiveness and the metastatic potential of orthotopic tumors grown from SMMC7721 and HCC-LM3 cells but not from HepG2 cells. In gene expression analysis, HTATIP2 was down-regulated by sorafenib. HCC-LM3 cells that expressed small hairpin RNAs against HTATIP2 (knockdown) formed less invasive tumors in mice following administration of sorafenib than HCC-LM3 without HTATIP2 knockdown. Alternatively, HepG2 cells that expressed transgenic HTATIP2 formed more invasive tumors in mice following administration of sorafenib. Sorafenib induced the epithelial-mesenchymal transition in HCC cell lines, which was associated with expression of HTATIP2. Sorafenib regulated expression of HTATIP2 via Jun-activated kinase (JAK) and signal transducer and activator of transcription (STAT)3 signaling. Sorafenib therapy prolonged recurrence-free survival in patients who expressed lower levels of HTATIP2 compared with higher levels. CONCLUSIONS: Sorafenib promotes invasiveness and the metastatic potential of orthotopic tumors from HCC cells in mice, down-regulating expression of HTATIP2 via JAK-STAT3 signaling.