Recent published findings have demonstrated the effectiveness of combining molecules from traditional Chinese medicine with chemotherapeutic drugs to treat cancer. Combined administration of these agents can overcome drug-mitigating responses as well as reduce adverse side effects, thereby enhancing the efficacy of the therapy. Tetramethylpyrazine (TMP), an alkaloid monomer from the medicinal herb Ligusticum chuanxiong hort, is known to exert a variety of antitumor effects including inhibition of tumor cell proliferation, metastasis, and drug resistance. In this research, we investigated antitumor effects of TMP combined with paclitaxel (PTX), a frontline chemotherapeutic drug, in vitro and in vivo. Our results indicate that TMP enhances the antitumor effects of PTX in ovarian cancer A2780 and SKOV3 cells. Furthermore, we found that combined treatment of TMP and PTX suppressed angiogenesis by inhibiting both ERK1/2 and Akt pathways and promoted apoptosis of tumor cells compared to TMP or PTX treatment alone. Moreover, TMP augmented the antitumor effects of PTX in ovarian cancer A2780 xenograft mouse models by significantly decreasing tumor burden and partially decreasing the toxicity of PTX, as evidenced by the decreased expression of proliferation and angiogenesis markers as well as the hematoxylin and eosin (H&E) staining and biochemical indexes assay. Overall, our findings provide novel mechanistic insight into the efficacy of combining of potent molecules present in traditional Chinese medicine with chemotherapeutic drugs for therapeutic intervention in cancer.

Vascularization is an essential step in successful bone tissue engineering. The induction of angiogenesis in bone tissue engineering can be enhanced through the delivery of therapeutic agents that stimulate vessel and bone formation. In this study, we show that cucurbitacin B (CuB), a tetracyclic terpene derived from Cucurbitaceae family plants, facilitates the induction of angiogenesis in vitro.

The redox dye, DCPIP, has recently shown to exhibit anti-melanoma activity in vitro and in vivo. On the other hand, there is increasing evidence that synthetic nanoparticles can serve as highly efficient carriers of drugs and vaccines for treatment of various diseases. These nanoparticles have shown to serve as potent tools that can increase the bioavailability of the drug/vaccine by facilitating absorption or conferring sustained and improved release. Here, we describe results on the effects of free- and nanoparticle-enclosed DCPIP as anti-angiogenesis and anti-inflammation agents in a human colon cancer HCT116 cell line in vitro, and in induced angiogenesis in ovo.

Topoisomerases II are ubiquitous enzymes with significant genotoxic effects in many critical DNA processes. Additionally, epidermal growth factor receptor (EGFR) plays pivotal role in tumour growth and angiogenesis. A novel series of naphtho[2',3':4,5]thiazolo[3,2-a]pyrimidine hybrids have been designed, synthesised and evaluated for their topo IIα/EGFR inhibitory and apoptotic inducer activities. Cytotoxicity of the synthesised hybrids was evaluated against MCF-7, A549 and HCT-116 cell lines. Of the synthesised hybrids, 6i, 6a and 6c experienced superior cytotoxic activity compared to doxorubicin and erlotinib against the tested cancer cells. The molecular mechanism of these hybrids revealed their ability to successfully inhibit topo IIα and EGFR activities in micromolar concentration and may serve as topo II catalytic inhibitor. Moreover, these hybrids significantly arrested cell cycle at G2/M phase together with increased p53, caspae-7, caspase-9 levels and Bax/Bcl-2 ratio. The synthesised hybrids showed efficient binding pattern in molecular docking study and have acceptable drug likeness characters.

Metronomic chemotherapy with cytotoxic agents has been shown to inhibit angiogenesis and, consequently, tumor growth by targeting vascular endothelial cells (ECs). In these regimens, anti-tumor activities additional to anti-angiogenesis may operate. Moreover, chemotherapy typically generates reactive oxygen species in targeted ECs, which can affect angiogenesis. The aim of the present study was to assess the systemic effect of low-dosage metronomic treatment with either irinotecan or mitoxantrone on angiogenesis induced by VEGF-A. Angiogenesis was induced in normal adult rat mesentery by intraperitoneal injection of a low dosage of VEGF-A. Thereafter, irinotecan and mitoxantrone were infused separately continuously at minimally toxic dosages for 14 consecutive days via a subcutaneous osmotic minipump. Angiogenesis was assessed in terms of objective and quantitative variables using morphologic and computerized image analyses. Irinotecan or mitoxantrone significantly stimulated angiogenesis, with ironotecan increasing angiogenesis by 104%, when compared with the vehicle-treated animals. Low-dosage metronomic chemotherapy with irinotecan or mitoxantrone stimulates angiogenesis in the normal mesentery of rats, probably by inducing low-level oxidative stress in the targeted ECs. Whether or not this pertains to tumor angiogenesis may be difficult to confirm, as several anti-tumor modes may operate during low-dosage metronomic chemotherapy.

Hepatocyte growth factor (HGF) and its receptor Met are responsible for a wide variety of cellular responses, both physiologically during embryo development and tissue homeostasis, and pathologically, particularly during tumor growth and dissemination. In cancer, Met can act as an oncogene on tumor cells, as well as a pro-angiogenic factor activating endothelial cells and inducing new vessel formation. Molecules interfering with Met activity could be valuable therapeutic agents. Here we have investigated the antiangiogenic properties of a synthetic peptide mimicking the docking site of the Met carboxyl-terminal tail, which was delivered into the cells by fusion with the internalization sequences from Antennapedia or HIV-Tat. We showed that these peptides inhibit ligand-dependent endothelial cell proliferation, motility, invasiveness and morphogenesis in vitro to an even greater extent and with much less toxicity than the Met inhibitor PHA-665752, which correlated with interference of HGF-dependent downstream signaling. In vivo, the peptides inhibited HGF-induced angiogenesis in the matrigel sponge assay and impaired xenograft tumor growth and vascularization in Kaposi's sarcoma. These data show that interference with the Met receptor intracellular sequence impairs HGF-induced angiogenesis, suggesting the use of antidocking site compounds as a therapeutic strategy to counteract angiogenesis in cancer as well as in other diseases.

Taxanes target microtubules and are clinically established chemotherapeutic agents with proven efficacy in human cancers. Cabazitaxel (XRP-6258, Jevtana®) is a second generation semisynthetic taxane with high chemotherapeutic potential in prostate cancer. There, cabazitaxel can overcome docetaxel-resistant prostate cancer. Here, we tested the effects of cabazitaxel on glioma cells, and non-transformed cells such as neurons and astrocytes. Cabazitaxel operates highly toxic in various human glioma cells at nanomolar concentrations. In contrast, primary astrocytes and neurons are not affected by this agent. Cabazitaxel disrupts cytoskeletal F-actin fibers and induces apoptotic cell death in gliomas. Moreover, cabazitaxel displayed highest efficacy in inhibiting glioma cell migration and invasion. Here we demonstrate that cabazitaxel inhibited tumor migration already at 1 nM. We also tested cabazitaxel in the ex vivo VOGiM assay. Cabazitaxel stalled glioma growth and at the same time inhibited tumor-induced angiogenesis. In summary, we found that cabazitaxel operates as an apoptosis-inducing gliomatoxic agent with strongest effects on migration and invasive growth. Thus, our report uncovered cabazitaxel actions on gliomas and on the brain tumor microenvironment. These data reveal novel aspects for adjuvant approaches when applied to brain tumor patients.

Lipocalin 2 (Lcn2) is a promising therapeutic target as well as a potential diagnostic biomarker for breast cancer. It has been previously shown to promote breast cancer progression by inducing the epithelial to mesenchymal transition in breast cancer cells as well as by enhancing angiogenesis. Lcn2 levels in urine and tissue samples of breast cancer patients has also been correlated with breast cancer status and poor patient prognosis. In this study, we have engineered a novel liposomal small interfering RNA (siRNA) delivery system to target triple negative breast cancer (TNBC) via a recently identified molecular target, intercellular adhesion molecule-1 (ICAM-1). This ICAM-1-targeted, Lcn2 siRNA- encapsulating liposome (ICAM-Lcn2-LP) binds human TNBC MDA-MB-231cells significantly stronger than non-neoplastic MCF-10A cells. Efficient Lcn2 knockdown by ICAM-Lcn2-LPs led to a significant reduction in the production of vascular endothelial growth factor (VEGF) from MDA-MB-231 cells, which, in turn, led to reduced angiogenesis both in vitro and in vivo. Angiogenesis (neovascularization) is a requirement for solid tumor growth and progression, and its inhibition is an important therapeutic strategy for human cancers. Our results indicate that a tumor-specific strategy such as the TNBC-targeted, anti-angiogenic therapeutic approach developed here, may be clinically useful in inhibiting TNBC progression.

BACKGROUND Natural compounds have been utilized in inhibiting metastasis alone or in combination with other anti-tumor agents. Dehydrocostus lactone (DHC), a natural sesquiterpene lactone, was used to investigate its effect on proliferation of lung cancer cells and on the anti-angiogenic efficacy of doxorubicin. MATERIAL AND METHODS Cell proliferation was assessed by MTT assay and clonogenic assay. Apoptosis and migration were assessed by flow cytometry and wound-healing assay, respectively. Western blotting and qPCR were performed for gene and protein expression analysis. Matrigel plug assay was performed for angiogenesis assessment. RESULTS Results of the study show that DHC inhibited the survival and proliferation of lung cancer cells (A549 and H460) and enhanced the growth-inhibitory properties of DOX. Cotreatment of DHC enhanced the apoptosis-inducing effects of DOX by activating caspase-9 and caspase-3 followed by cleavage of PARP. Treatment of A549 and H460 cells with DHC caused suppression of HIF-1α, Akt and pAkt, GSK-3β and pGSK-3β, as well as ERK, pERK, mTOR, and p-mTOR. DHC enhanced the effect of DOX by inhibiting migration of A549 cells as observed by wound-healing assay. DHC caused synergistic inhibition of MMP-2 and MMP-9 genes when treated in combination with DOX. DHC further enhanced the anti-angiogenic properties of DOX in mice implanted with Matrigel plugs. DHC suppressed the proliferation of lung cancer cells and enhanced the anti-angiogenic properties of DOX. CONCLUSIONS The putative mechanism behind the metastasis-limiting effects of DHC may involve the suppression of Akt/GSK-3β and inhibition of MMP-2 and MMP-9 in lung cancer cells.

Biomaterials that promote angiogenesis are required for repair and regeneration of bone. In-situ formed injectable hydrogels functionalised with bioactive agents, facilitating angiogenesis have high demand for bone regeneration. In this study, pH and thermosensitive hydrogels based on chitosan (CS) and hydroxyapatite (HA) composite materials loaded with heparin (Hep) were investigated for their pro-angiogenic potential. Hydrogel formulations with varying Hep concentrations were prepared by sol-gel technique for these homogeneous solutions were neutralised with sodium bicarbonate (NaHCO3) at 4 °C. Solutions (CS/HA/Hep) constituted hydrogels setting at 37 °C which was initiated from surface in 5-10 minutes. Hydrogels were characterised by performing injectability, gelation, rheology, morphology, chemical and biological analyses. Hydrogel solutions facilitated manual dropwise injection from 21 Gauge which is highly used for orthopaedic and dental administrations, and the maximum injection force measured through 19 G needle (17.191 ± 2.296N) was convenient for manual injections. Angiogenesis tests were performed by an ex-ovo chick chorioallantoic membrane (CAM) assay by applying injectable solutions on CAM, which produced in situ hydrogels. Hydrogels induced microvascularity in CAM assay this was confirmed by histology analyses. Hydrogels with lower concentration of Hep showed more efficiency in pro-angiogenic response. Thereof, novel injectable hydrogels inducing angiogenesis (CS/HA/Hep) are potential candidates for bone regeneration and drug delivery applications.

Ovarian cancer is the seventh most common cancer and the second most common cause of cancer-associated mortality among gynecological malignancies worldwide. The combination of antimitotic agents, such as taxanes, and the DNA-damaging agents, such as platinum compounds, is the standard treatment for ovarian cancer. However, due to chemoresistance, development of novel therapeutic strategies for the treatment of ovarian cancer remains critical. Amentoflavone (AMF) is a biflavonoid derived from the extracts of Selaginella tamariscina, which has been used as a Chinese herb for thousands of years. A previous study demonstrated that AMF inhibits angiogenesis of endothelial cells and induces apoptosis in hypertrophic scar fibroblasts. In order to check the influence of AMF on cell proliferation, the effects of AMF on cell cycle and DNA damage were measured by cell viability, flow cytometry, immunofluorescence and western blotting assays in SKOV3 cells, an ovarian cell line. In the present study, treatment with AMF inhibited ovarian cell proliferation, increased P21 expression, decreased CDK1/2 expression, interrupted the balance of microtubule dynamics and arrested cells at the G2 phase. Furthermore, treatment with AMF increased the expression levels of phospho-Histone H2AX (γ-H2AX; a variant of histone 2A, that belongs to the histone 2A family member X) and the DNA repair protein RAD51 homolog 1 (Rad51), indicating the occurrence of DNA damage since γ-H2AX and Rad51 are both key markers of DNA damage. Consistent with previous findings, the results of the present study suggest that AMF is a potential therapeutic agent for the treatment of ovarian cancer. In addition, the effects of AMF on cell cycle arrest and DNA damage induction may be the molecular mechanisms by which AMF might exert its potential therapeutic benefits in ovarian cancer.

Schistosoma haematobium, the helminth causing urogenital schistosomiasis, is a known bladder carcinogen. Despite the causal link between S. haematobium and bladder cancer, the underlying mechanisms are poorly understood. S. haematobium oviposition in the bladder is associated with angiogenesis and urothelial hyperplasia. These changes may be pre-carcinogenic events in the bladder. We hypothesized that the Interleukin-4-inducing principle of Schistosoma mansoni eggs (IPSE), an S. haematobium egg-secreted "infiltrin" protein that enters host cell nuclei to alter cellular activity, is sufficient to induce angiogenesis and urothelial hyperplasia. Methods: Mouse bladders injected with S. haematobium eggs were analyzed via microscopy for angiogenesis and urothelial hyperplasia. Endothelial and urothelial cell lines were incubated with recombinant IPSE protein or an IPSE mutant protein that lacks the native nuclear localization sequence (NLS-) and proliferation measured using CFSE staining and real-time monitoring of cell growth. IPSE's effects on urothelial cell cycle status was assayed through propidium iodide staining. Endothelial and urothelial cell uptake of fluorophore-labeled IPSE was measured. Findings: Injection of S. haematobium eggs into the bladder triggers angiogenesis, enhances leakiness of bladder blood vessels, and drives urothelial hyperplasia. Wild type IPSE, but not NLS-, increases proliferation of endothelial and urothelial cells and skews urothelial cells towards S phase. Finally, IPSE is internalized by both endothelial and urothelial cells. Interpretation: IPSE drives endothelial and urothelial proliferation, which may depend on internalization of the molecule. The urothelial effects of IPSE depend upon its NLS. Thus, IPSE is a candidate pro-carcinogenic molecule of S. haematobium.

Tissue inhibitors of metalloproteinase 3 (TIMP3) are a major endogenous inhibitor of matrix metalloproteinase (MMPs) that inhibit tumor growth, invasion, metastasis and angiogenesis. In this study, we found that TIMP3 expression is associated with positive prognosis of colorectal cancer (CRC) clinicopathologically. Therefore, we developed a series of arylsulfonamide derivatives as TIMP3 inducers in order to define potential colorectal cancer therapeutic agent. Among these, MPT0B390 was selected for anti-tumor, anti-metastasis, and anti-angiogenesis property determination. Methods: The relationship between TIMP3 expression and clinical pathological features in colorectal patients and cell lines were determined by immunohistochemistry, bioinformatics analysis and western blotting. The anti-tumor function was validated by using MTT, apoptosis pathway detection and in vivo xenograft model for tumor growth inhibition determination. The anti-metastatic function was validated using a transwell migration assay, and using in vivo lung metastasis and liver metastasis models. The mechanism of MPT0B390-induced TIMP3 expression was further tested using qPCR and Chromatin IP assay. The anti-angiogenesis function was examined by using transwell migration assay, and in vivo Matrigel plug assay. Results: After screening candidate compounds, we identified MPT0B390 as an effective inducer of TIMP3. We showed that MPT0B390 induces TIMP3 expression significantly and inhibits CRC cell growth in vitro and in vivo. By inducing TIMP3 expression, MPT0B390 can also exert its anti-metastasis effect to inhibit CRC cell migration and invasion and downregulates migration markers such as uPA, uPAR, and c-Met. Subsequent Chromatin immunoprecipitation assay revealed that MPT0B390 can significantly inhibit EZH2 expression as well as its binding to TIMP3 promoter region to regulate TIMP3 induction. In addition to the anti-tumor and anti-metastasis capability, MPT0B390 can also induce TIMP3 expression in endothelial cells to inhibit tumor angiogenesis. Conclusion: These data suggest the potential therapeutic applications of the TIMP3 inducer, MPT0B390, for colorectal cancer treatment.

Organoselenium compounds have attracted growing interests as promising antitumor agents over recent years. Herein, four series of novel selenium-containing chiral 1,4-diarylazetidin-2-ones were asymmetrically synthesized and biologically evaluated for antitumor activities. Among them, compound 7 was found to be about 10-fold more potent than its prototype compound 1a, and compound 9a exhibited the most potent cytotoxicity against five human cancer cell lines, including a paclitaxel-resistant human ovarian cancer cell line A2780T, with IC50 values ranging from 1 to 3 nM. Mechanistic studies revealed that compound 9a worked by disrupting tubulin polymerization, inducing reactive oxygen species (ROS) production, decreasing mitochondrial membrane potential, blocking the cell cycle in the G2/M phase, inducing cellular apoptosis and suppressing angiogenesis. Additionally, compound 9a exhibited appropriate human-microsomal metabolic stability and physicochemical properties. Importantly, compound 9a was found to inhibit tumor growth effectively in a xenograft mice model with low toxicity profile, which rendered 9a a highly promising candidate for further pre-clinical development.

Hepatocellular carcinoma (HCC) is one of the most aggressive cancers with high mortality worldwide. Research and development of novel agents for HCC therapy is in demand, urgently. Morusin has been reported to exhibit potential cytotoxic activity in several cancer cell lines. However, whether it has potential antiangiogenic activity especially in HCC remains unclear. In the current study, we found that morusin exerted growth inhibition effects on human HCC cells (HepG2 and Hep3B) in vitro and human HCC cell (HepG2) xenografts in vivo. Moreover, apoptosis induction was observed in a dose-dependent manner after morusin treatment along with an increase in the expression of active caspase-3 and the Bax/Bcl-2 expression ratio. More importantly, morusin inhibited proliferation, migration, and tube formation of human umbilical vein endothelial cells (HUVECs) in vitro and downregulated angiogenic proteins in HCC cells and HUVECs. In vivo, tumor angiogenesis was also attenuated after morusin treatment. In addition, morusin suppressed constitutive as well as IL-6-induced STAT3 phosphorylation in HCC cells and corresponding tumor tissues. Overall, morusin has a potential anticancer effect on human HCC cells in vitro and in vivo by inducing apoptosis and inhibiting anti-angiogenesis. The corresponding mechanism might be associated with the attenuation of the IL-6/STAT3 signaling pathway. Morusin might serve as a promising novel anticancer agent in HCC therapy, and requires further study.

The edible Rosa hybrida (RH) petal is utilized in functional foods and cosmetics. Although the biological function of RH petal extract is known, mechanism of action studies involving tumor-associated angiogenesis have not yet been reported. Herein, we investigated the regulatory effect of the ethanol extract of RH petal (EERH) on tumor growth and tumor angiogenesis against bladder cancer. EERH treatment inhibited the bladder carcinoma T24 cell and 5637 cell proliferation because of G1-phase cell cycle arrest by inducing p21WAF1 expression and reducing cyclins/CDKs level. EERH regulated signaling pathways differently in both cells. EERH-stimulated suppression of T24 and 5637 cell migration and invasion was associated with the decline in transcription factor-mediated MMP-9 expression. EERH oral administration to xenograft mice reduced tumor growth. Furthermore, no obvious toxicity was observed in acute toxicity test. Decreased CD31 levels in EERH-treated tumor tissues led to examine the angiogenic response. EERH alleviated VEGF-stimulated tube formation and proliferation by downregulating the VEGFR2/eNOS/AKT/ERK1/2 cascade in HUVECs. EERH impeded migration and invasion of VEGF-induced HUVECs, which is attributed to the repressed MMP-2 expression. Suppression of neo-microvessel sprouting, induced by VEGF, was verified by treatment with EERH using the ex vivo aortic ring assay. Finally, kaempferol was identified as the main active compound of EERH. The present study demonstrated that EERH may aid the development of antitumor agents against bladder cancer.

Triple-negative breast cancer (TNBC) is an aggressive subset of breast cancer and currently lacks effective therapeutic targets. As two main phototherapeutic methods, photothermal therapy (PTT) and photodynamic therapy (PDT) show many advantages in TNBC treatment, and their combination with chemotherapy can achieve synergistic therapeutic effects. In the present study, a biomimetic nanoplatform was developed based on leukocyte/platelet hybrid membrane (LPHM) and dendritic large pore mesoporous silicon nanoparticles (DLMSNs). A near infrared (NIR) fluorescent dye IR780 and a chemotherapeutic drug doxorubicin (DOX) were co-loaded into the large pores of DLMSNs to prepare DLMSN@DOX/IR780 (DDI) nanoparticles (NPs), followed by camouflage with LPHM to obtain LPHM@DDI NPs. Through the mediation of LPHM, LPHM@DDI NPs showed an excellent TNBC-targeting ability and very high PTT/PDT performances in vitro and in vivo. Upon NIR laser irradiation, LPHM@DDI NPs exhibited synergistic cytotoxicity and apoptosis-inducing activity in TNBC cells, and effectively suppressed tumor growth and recurrence in TNBC mice through tumor ablation and anti-angiogenesis. These synergistic effects were sourced from the combination of PTT/PDT and chemotherapy. Altogether, this study offers a promising biomimetic nanoplatform for efficient co-loading and targeted delivery of photo/chemotherapeutic agents for TNBC combination treatment.

The aim of the present study was to analyze the molecular mechanisms involved in blocking the signaling pathway and the effects of this on the progression of prostate cancer (CaP) cells in vitro. LNCaP human CaP cell line was stimulated with interleukin-6 (IL-6) in the presence/absence of Janus kinase (JAK) 2 (AG490), signal transducer and activator of transcription 3 [(STAT3) S3I-201] inhibitors and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Cytotoxic activity, the activation of phosphorylated (p)-STAT3 protein, caspase (CASP) 3 activity at protein level, vascular endothelial growth factor (VEGF) A, VEGFC, vascular endothelial growth factor receptor 2, STAT3, matrix metalloproteinase-2, myeloid cell leukemia sequence 1 (MCL-1), CASP8 and CASP9 messenger RNA (mRNA) levels were determined. Morphology and apoptosis were confirmed by DAPI staining and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay. IL-6 rapidly induced the phosphorylation of STAT3 in a dose- and time-dependent manner with a peak expression at 3 h at a concentration of 25 ng/ml. In addition, AG490 (50 μM) and S3I-201 (300 μM) inhibited STAT3 activation. Western blotting results revealed that p-STAT3 protein expression decreased significantly with AG490 and S3I-201 treatment in LNCaP cells. AG490 and S3I-201 induced the downregulation of VEGFA, MCL-1 and STAT3 and the upregulation of CASP8 and CASP9 mRNA transcription levels. In addition, the inhibitors increased the level of CASP3 protein. Combinations of AG490- and S3I-201-TRAIL did not result in an increase in this effect. Parallel results were found by DAPI staining and TUNEL assay. To the best of our knowledge, this is the first study to investigate the possible clinical use of AG490 or S3I-201, together with the reduced use of chemotherapeutic agents with high cytotoxicity, for their ability to exert an apoptotic effect, targeting the JAK/STAT3 pathway.

A major goal of the field of systems biology is to translate genome-wide profiling data (e.g., mRNAs, miRNAs) into interpretable functional networks. However, employing a systems biology approach to better understand the complexities underlying drug resistance phenotypes in cancer continues to represent a significant challenge to the field. Previously, we derived two drug-resistant breast cancer sublines (tamoxifen- and fulvestrant-resistant cell lines) from the MCF7 breast cancer cell line and performed genome-wide mRNA and microRNA profiling to identify differential molecular pathways underlying acquired resistance to these important antiestrogens. In the current study, to further define molecular characteristics of acquired antiestrogen resistance we constructed an "integrative network". We combined joint miRNA-mRNA expression profiles, cancer contexts, miRNA-target mRNA relationships, and miRNA upstream regulators. In particular, to reduce the probability of false positive connections in the network, experimentally validated, rather than prediction-oriented, databases were utilized to obtain connectivity. Also, to improve biological interpretation, cancer contexts were incorporated into the network connectivity.

New strategies for the treatment of hepatocellular carcinoma (HCC) are needed, given that currently available chemotherapeutics are inefficient. Since tumor growth reflects the net balance between pro-proliferative and death signaling, agents shifting the equilibrium toward the latter are of considerable interest. The TWEAK:Fn14 signaling axis promotes tumor cell proliferation and tumor angiogenesis, while TRAIL:TRAIL-receptor (TRAIL-R) interactions selectively induce apoptosis in malignant cells. Fn14•TRAIL, a fusion protein bridging these two pathways, has the potential to inhibit tumor growth, by interfering with TWEAK:Fn14 signaling, while at the same time enforcing TRAIL:TRAIL-R-mediated apoptosis. Consequently, Fn14•TRAIL's capacity to inhibit HCC growth was tested.