Scutellaria baicalensis Induces Cell Apoptosis and Elicits Mesenchymal-Epithelial Transition to Alleviate Metastatic Hepatocellular Carcinoma via Modulating HSP90ß.

Hepatocellular carcinoma is one of the most common malignant tumors in the world and shows strong metastatic potential. Current medicine for hepatocellular carcinoma therapy is invalid, while Scutellaria baicalensis Georgi exhibits the pharmaceutical potential to treat liver diseases and liver cancer. Herein, we verified the inhibitory properties and the pivotal molecules regimented by Scutellaria baicalensis on advanced hepatocellular carcinoma. At first, the viability of SK-Hep-1 cells was significantly reduced under treatment of Scutellaria baicalensis extract in a dose-dependent manner without affecting the growth of normal hepatocyte. Scutellaria baicalensis extract application could remarkably cause apoptosis of SK-Hep-1 cells through p53/cytochrome C/poly-ADP ribose polymerase cascades and arrest the cell cycle at the G1/S phase by downregulating cyclin-dependent kinases. Meanwhile, administration of Scutellaria baicalensis extract remarkably attenuated the migration capability as well as suppressed matrix metalloproteinase activity of advanced hepatocellular carcinoma cells. The proteome profiles and network analysis particularly implied that exposure to Scutellaria baicalensis extract downregulated the expression of HSP90ß, and the clinical stage of hepatocellular carcinoma is also positively correlated with the HSP90ß level. Combined treatment of Scutellaria baicalensis extract and HSP90ß siRNAs could markedly enhance the ubiquitination activity and the degradation of vimentin to subsequently inhibit the metastatic property of SK-Hep-1 cells. Moreover, application of Scutellaria baicalensis extract and HSP90ß siRNAs depleted phosphorylation of AKT, which stimulated the expression of p53 and consecutively triggered cell apoptosis. These findings suggest that HSP90ß may be a prospective target for the effective therapy of advanced hepatocellular carcinoma via accelerating apoptosis of hepatocellular carcinoma cells and eliciting mesenchymal-epithelial transition with the administration of Scutellaria baicalensis extract.

Inhibition of CDK9 exhibits anticancer activity in hepatocellular carcinoma cells via targeting ribonucleotide reductase.

Cyclin-dependent kinase 9 (CDK9) inhibitors are a novel category of anticancer treatment for cancers. However, their effects on hepatocellular carcinoma (HCC) are rarely investigated. Human ribonucleotide reductase (RR, which consists of RRM1 and RRM2 subunits) catalyzes the conversion of ribonucleoside diphosphate into 2'-deoxyribonucleoside diphosphate to maintain the homeostasis of nucleotide pools, which play essential roles in DNA synthesis and DNA repair. In this study, we identified that CDK9 protein expression in adjacent non-tumor tissues predicted HCC patients' overall and progression-free survivals. The anticancer activity of a CDK9-selective inhibitor, LDC000067, on HCC cells was positively associated with its ability to inhibit the expression of RRM1 and RRM2. LDC000067 downregulated RRM1 and RRM2 expression through post-transcriptional pathway. Specifically, LDC000067 triggered RRM2 protein degradation via multiple pathways, including proteasome-, lysosome-, and calcium-dependent pathways. Furthermore, CDK9 positively correlates with RRM1 or RRM2 expression in HCC patients, and the expressions of these three genes were associated with the higher infiltration of immune cells in HCC. Taken together, this study identified the prognostic relevance of CDK9 in HCC and the molecular mechanism for the anticancer effect of CDK9 inhibitors on HCC.

Repurposing of ingenol mebutate for treating human colorectal cancer by targeting S100 calcium-binding protein A4 (S100A4).

Colorectal cancer (CRC) is the world's second most common cause of cancer-related death. Novel treatments are still urgently needed. S100 calcium-binding protein A4 (S100A4) was demonstrated to be an anticancer therapeutic target. Herein, we found that higher S100A4 expression was associated with a poorer prognosis in publicly available cohorts and a Taiwanese CRC patient cohort. To identify repurposed S100A4 inhibitors, we mined the Connectivity Map (CMap) database for clinical drugs mimicking the S100A4-knockdown gene signature. Ingenol mebutate, derived from the sap of the plant Euphorbia peplus, is approved as a topical treatment for actinic keratosis. The CMap analysis predicted ingenol mebutate as a potent S100A4 inhibitor. Indeed, both messenger RNA and protein levels of S100A4 were attenuated by ingenol mebutate in human CRC cells. In addition, CRC cells with higher S100A4 expressions and/or the wild-type p53 gene were more sensitive to ingenol mebutate, and their migration and invasion were inhibited by ingenol mebutate. Therefore, our results suggest the repurposing of ingenol mebutate for treating CRC by targeting S100A4.

A chemical probe inhibitor targeting STAT1 restricts cancer stem cell traits and angiogenesis in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is a worldwide cancer with rising annual incidence. New medications for patients with CRC are still needed. Recently, fluorescent chemical probes have been developed for cancer imaging and therapy. Signal transducer and activator of transcription 1 (STAT1) has complex functions in tumorigenesis and its role in CRC still needs further investigation. METHODS: RNA sequencing datasets in the NCBI GEO repository were analyzed to investigate the expression of STAT1 in patients with CRC. Xenograft mouse models, tail vein injection mouse models, and azoxymethane/dextran sodium sulfate (AOM/DSS) mouse models were generated to study the roles of STAT1 in CRC. A ligand-based high-throughput virtual screening approach combined with SWEETLEAD chemical database analysis was used to discover new STAT1 inhibitors. A newly designed and synthesized fluorescently labeled 4',5,7-trihydroxyisoflavone (THIF) probe (BODIPY-THIF) elucidated the mechanistic actions of STAT1 and THIF in vitro and in vivo. Colonosphere formation assay and chick chorioallantoic membrane assay were used to evaluate stemness and angiogenesis, respectively. RESULTS: Upregulation of STAT1 was observed in patients with CRC and in mouse models of AOM/DSS-induced CRC and metastatic CRC. Knockout of STAT1 in CRC cells reduced tumor growth in vivo. We then combined a high-throughput virtual screening approach and analysis of the SWEETLEAD chemical database and found that THIF, a flavonoid abundant in soybeans, was a novel STAT1 inhibitor. THIF inhibited STAT1 phosphorylation and might bind to the STAT1 SH2 domain, leading to blockade of STAT1-STAT1 dimerization. The results of in vitro and in vivo binding studies of THIF and STAT1 were validated. The pharmacological treatment with BODIPY-THIF or ablation of STAT1 via a CRISPR/Cas9-based strategy abolished stemness and angiogenesis in CRC. Oral administration of BODIPY-THIF attenuated colitis symptoms and tumor growth in the mouse model of AOM/DSS-induced CRC. CONCLUSIONS: This study demonstrates that STAT1 plays an oncogenic role in CRC. BODIPY-THIF is a new chemical probe inhibitor of STAT1 that reduces stemness and angiogenesis in CRC. BODIPY-THIF can be a potential tool for CRC therapy as well as cancer cell imaging.

Differential effect of herbal tea extracts on free fatty acids-, ethanol- and acetaminophen-induced hepatotoxicity in FL83B hepatocytes.

In recent years, herbal tea consumption becomes popular because of the potential health benefits and attractive flavors. However, there is also a growing concern that herbal supplements contribute to the drug-drug/drug-herb interactions and hepatotoxicity. In this study, FL83B mouse hepatocytes were used as an in vitro mode of hepatotoxicity induced by free fatty acids, including palmitic acid (PA) and oleic acid (OA), ethanol, and acetaminophen. Herbal tea extracts were obtained from eight common herbal plants, including Verbena officinalis L., Hyssopus officinalis L., Salvia officinalis L., Urtica dioica L., Hemerocallis fulva (L.) L., Citrus maxima (Burm.) Merr., Citrus limon (L.) Osbeck, and Ficus formosana Maxim. MTT assay was used to evaluate the impact of these herbal tea extracts on hepatoxocitity. We found that these herbal tea extracts per se did not exhibit hepatotoxicity, and had no effect on OA-induced hepatotoxicity. However, extracts from Verbena officinalis L., Hyssopus officinalis L., Salvia officinalis L., and Hemerocallis fulva (L.) L. exhibited protective effect against PA-induced hepatotoxicity. In addition, herbal tea extracts from Verbena officinalis L., Hyssopus officinalis L., Salvia officinalis L., Urtica dioica L., Hemerocallis fulva (L.) L., and Ficus formosana Maxim. exhibited protective effect against acetaminophen-induced hepatotoxicity. Interestingly, all these herbal tea extracts enhanced ethanol-induced hepatotoxicity. Our results suggest that herbal tea extracts have differential effects on different modes of hepatotoxicity.

Long Non-Coding RNA MIR31HG Promotes the Transforming Growth Factor ß-Induced Epithelial-Mesenchymal Transition in Pancreatic Ductal Adenocarcinoma Cells.

The epithelial-to-mesenchymal transition (EMT) describes a biological process in which polarized epithelial cells are converted into highly motile mesenchymal cells. It promotes cancer cell dissemination, allowing them to form distal metastases, and also involves drug resistance in metastatic cancers. Transforming growth factor ß (TGFß) is a multifunctional cytokine that plays essential roles in development and carcinogenesis. It is a major inducer of the EMT. The MIR31 host gene (MIR31HG) is a newly identified long non-coding (lnc)RNA that exhibits ambiguous roles in cancer. In this study, a cancer genomics analysis predicted that MIR31HG overexpression was positively correlated with poorer disease-free survival of pancreatic ductal adenocarcinoma (PDAC) patients, which was associated with upregulation of genes related to TGFß signaling and the EMT. In vitro evidence demonstrated that TGFß induced MIR31HG expression in PDAC cells, and knockdown of MIR31HG expression reversed TGFß-induced EMT phenotypes and cancer cell migration. Therefore, MIR31HG has an oncogenic role in PDAC by promoting the EMT.

Hepatic Stellate Cell Modulates the Immune Microenvironment in the Progression of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a major cause of increases in the mortality rate due to cancer that usually develops in patients with liver fibrosis and impaired hepatic immunity. Hepatic stellate cells (HSCs) may directly or indirectly crosstalk with various hepatic cells and subsequently modulate extracellular remodeling, cell invasion, macrophage conversion, and cancer deterioration. In this regard, the tumor microenvironment created by activated HSC plays a critical role in mediating pathogenesis and immune escape during HCC progression. Herein, intermediately differentiated human liver cancer cell line (J5) cells were co-cultured with HSC-conditioned medium (HSC-CM); changes in cell phenotype and cytokine profiles were analyzed to assess the impact of HSCs on the development of hepatoma. The stage of liver fibrosis correlated significantly with tumor grade, and the administration of conditioned medium secreted by activated HSC (aHSC-CM) could induce the expression of N-cadherin, cell migration, and invasive potential, as well as the activity of matrix metalloproteinases in J5 cells, implying that aHSC-CM could trigger the epithelial-mesenchymal transition (EMT). Next, the HSC-CM was further investigated and network analysis indicated that specific cytokines and soluble proteins, such as activin A, released from activated HSCs could remarkably affect the tumor-associated immune microenvironment involved in macrophage polarization, which would, in turn, diminish a host's immune surveillance and drive hepatoma cells into a more malignant phenotype. Together, our findings provide a novel insight into the integral roles of HSCs to enhance hepatocarcinogenesis through their immune-modulatory properties and suggest that HSC may serve as a potent target for the treatment of advanced HCC.

PERK/ATF4-Dependent ZFAS1 Upregulation Is Associated with Sorafenib Resistance in Hepatocellular Carcinoma Cells.

Sorafenib, a multi-kinase inhibitor, is the first-line treatment for advanced hepatocellular carcinoma (HCC) patients. However, this drug only provides a short improvement of patients' overall survival, and drug resistance is commonly developed. Thus, the identification of resistant factor(s) or biomarker(s) is needed to develop more efficient therapeutic strategies. Long, non-coding RNAs (lncRNAs) have recently been viewed as attractive cancer biomarkers and drive many important cancer phenotypes. A lncRNA, ZFAS1 (ZNFX1 antisense RNA 1) has been found to promote HCC metastasis. This study found that sorafenib induced ZFAS1 expression specifically in sorafenib-resistant HCC cells. Although ZFAS1 knockdown did not restore the sensitivity of HCC cells to sorafenib, its expression may act as a resistant biomarker for sorafenib therapy. Bioinformatics analysis predicted that sorafenib tended to induce pathways related to endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) in sorafenib-resistant HCC cells. In vitro experimental evidence suggested that sorafenib induced protein kinase RNA-like ER kinase (PERK)/activating transcription factor 4 (ATF4)-dependent ZFAS1 expression, and sorafenib resistance could be overcome by PERK/ATF inhibitors. Therefore, PERK/ATF4/ZFAS1 signaling axis might be an attractive therapeutic and prognostic biomarker for sorafenib therapy in HCC.

Dicoumarol suppresses HMGA2-mediated oncogenic capacities and inhibits cell proliferation by inducing apoptosis in colon cancer.

Colon cancer is one of the leading causes of cancer-related deaths and its five-year survival rate remains low in locally advanced or metastatic stages of colon cancer. Overexpression of high mobility group protein AT-hook2 (HMGA2) is associated with cancer progression, metastasis, and poor prognosis in many malignancies. Oxidative stress regulates cellular mechanisms and provides an environment that favors the cancer cells to survive and progress, yet, at the same time, oxidative stress can also be utilized as a cancer-damaging strategy. The molecular regulatory roles of HMGA2 in oxidative stress and their involvement in cancer progression are largely unknown. In this study, we investigated the involvement of HMGA2 in regulation of oxidative stress responses by luciferase reporter assays. Moreover, we utilized dicoumarol (DIC), a derivative of coumarin which has been suggested to be involved in oxidation regulation with anticancer effects, and demonstrated that DIC could induce apoptosis and inhibit cell migration of HMGA2 overexpressing colon cancer cells. Further investigation also evidenced that DIC can enhance the cancer inhibition effect of 5-FU in colony formation assays. Taken together, our data revealed novel insights into the molecular mechanisms underlying HMGA2 and highlighted the possibility of targeting the cellular antioxidant system for treating patients and preventing from cancer progression in HMGA2 overexpressing colon cancer cells.

Characterizing the Relapse Potential in Different Luminal Subtypes of Breast Cancers with Functional Proteomics.

Poor prognosis due to the high relapse and metastasis rates of breast cancer has been particularly linked to the luminal B subtype. The current study utilized MCF-7 and ZR-75-1 to investigate various luminal subtypes of breast cancers that have discrepant expressions in the estrogen receptor (ER) and human epidermal growth factor receptor 2 (HER2). Understanding of the differential protein profiles and the associated pathways could help alleviate the malignance and promote the long-term survival rate of breast cancer patients. Functional proteome tools were applied to comprehensively delineate the global protein alterations that reflect the varieties of biological features between the two subtypes. In this study, a total of 11 proteins with significant and meaningful changes were identified. These protein targets including PRX2, CK19, nucleophosmin and cathepsin D were mostly involved in cell differentiation or proliferation. Particularly, cathepsin D was highly expressed in the luminal B subtype. Moreover, the level of cathepsin-D was also upregulated in the clinical metastatic tissues. Accordingly, the RNA interference-mediated silencing of cathepsin D stimulated ER expression but suppressed the level of HER2. The knockdown of cathepsin D enhanced the level of ZO-1 and a remarkable decrease in N-cadherin was also detected. Again, the matrix metalloproteinases (MMP) activity was impaired under the cathepsin D abolishment. Collectively, this study represented a modality to explore novel relationships in a proteome complex and highlighted the functional roles of cathepsin D in treatment options for different subtypes of breast cancer.

Characterization of Fibrinogen as a Key Modulator in Patients with Wilson's Diseases with Functional Proteomic Tools.

Wilson's disease (WD) is an autosomal recessive disorder of copper metabolism caused by defects in the ATPase gene (ATP7B). The various clinical features result from the massive accumulation of copper in the liver, cornea and basal ganglia. Although WD can be effectively treated with proper medicine, this disease is difficult to clearly diagnose due to its indefinite symptoms. In the current study, we achieved a positive correlation between clinical symptoms and the enzymatic activity of ceruloplasmin in WD patients. Furthermore, proteome profiles of plasma as well as network analysis demonstrated that fibrinogen is a critical indicator which is significantly unregulated in WD subjects in comparison to healthy donors and closely linked to pathogenesis of WD. Here, we applied 2DE-immunoblots and immunohistochemistry to verify the protein level and localization in situ. The enhanced expression of fibrinogen in the plasma of WD subjects with respect to that of healthy controls and patients with distinct disorders was also confirmed by utilizing clinical samples. As expected, application of high dose of copper induced expression of fibrinogen, while knockdown of ceruloplasmin also resulted in upregulation of fibrinogen as well as elimination of superoxide dismutase (SOD), leading to increased oxidative stress in cells. In summary, the liver injury or oxidative stress induced by the progression of WD may account for the obvious increase of fibrinogen, which in turn triggers inflammatory responses and interferes coagulation cascades; this finding sheds light on the early detection and diagnosis of WD.

[Research advance in chemical compounds,pharmacological action and clinical application of Dengzhan Shengmai Capsules].

Dengzhan Shengmai Capsules( DZSMC),a well-known traditional Chinese medicine( TCM) formula,is comprised of the main drug of Erigeron breviscapus,and supplemented with Panax ginseng,Ophiopogon japonicus and Schisandra chinensis,with functions of supplementing Qi and nourishing Yin,promoting blood circulation and strengthening brain. DZSMC is the only Chinese patent drug with A-level evidence-based medicine in secondary prevention for stroke and ranks first among TCMs for neurological treatment. Modern studies indicate that the chemical constituents of DZSMC mainly include flavonoids,phenolic acids,lignans,saponins and so on. Pharmacological experimental studies have shown that DZSMC has such pharmacological effects as anti-oxidation,anti-inflammatory and anti-myocardial ischemia. DZSMC is mainly used in the convalescent care of ischemic cardiovascular and cerebrovascular diseases,and is often used in combination with various conventional therapeutic drugs to exert clinical efficacy through brain protection,neuroprotection,etc.,and improve clinical symptoms in patients. In this review,according to domestic and international related literature combined with research results obtained by our project,the research advances in the chemical constituents,pharmacological effects and clinical application of DZSMC have been systematically reviewed and summarized,providing reference and support for further study and secondary development of the formula.

Insights into the hepatocellular carcinoma patient journey: results of the first global quality of life survey.

AIM: To better understand the hepatocellular carcinoma (HCC) patient journey, we conducted a patient survey across 13 countries. METHODS: The survey included closed- and open-ended questions developed using an iterative process to gather information on demographics, diagnosis and treatment. Patients self-selected or were directed to the online survey by their doctor. RESULTS: A total of 256 patients completed the survey. More than two-thirds (68%) felt they did not receive enough information about HCC at diagnosis. Treatments included oral anticancer therapy, transarterial chemoembolization (TACE), and selective internal radiation therapy (SIRT). A total of 81% receiving sorafenib, 45% receiving SIRT and 32% receiving TACE reported impaired quality-of-life (QoL). A total of 42, 19 and 0% of patients using sorafenib rated their current QoL as 'poor', 'good' and 'excellent', respectively; compared with SIRT (22, 33 and 6%) or TACE (11, 37 and 13%). CONCLUSION: Most patients with HCC require additional accessible information. People with incurable HCC require treatments that preserve QoL.

Laser galvanometric scanning system for improved average power uniformity and larger scanning area.

A new laser galvanometric scanning optical system incorporating a dynamic-tilt focusing lens is proposed to improve the laser spot performance in adaptive manufacturing applications. The simulations focus specifically on the laser spot size, the spot profile, the spot position, the spot energy distribution, and the size of the scanning working field. It is shown that for a designed spot size of 50 µm, the proposed system achieves an average spot size of 50.5 µm. Moreover, the maximum position deviation of the laser beam is reduced from (x=-3.02%, y=1.30%) in a traditional scanning system to (x=-0.055%, y=0.162%) in the proposed system. Finally, the maximum working field area is increased by around 240% compared to that of a traditional system. Overall, the results show that the proposed laser galvanometric scanning system achieves a small spot size, a symmetrical and round spot profile, a uniform spot energy distribution, and a large working area. As a result, it is ideally suited to rapid prototyping applications.

Two new phenolic compounds from the tuber of Sparganium stoloniferum.

Two new phenolic compounds, 2-(2-hydroxyphenyl)-4-methoxycarbonyl-5-hydroxybenzofuran (1) and 1-methoxycarbonyl-2, 3-dihydroxydibenzo[b, f]oxepine (2), were isolated from the tuber of Sparganium stoloniferum. The structures of both new compounds were determined on basis of spectroscopic means including HR-ESI-MS, 1D and 2D NMR experiments.

S-Adenosyl-L-methionine-competitive inhibitors of the histone methyltransferase EZH2 induce autophagy and enhance drug sensitivity in cancer cells.

The enhancer of zeste homolog 2 (EZH2) has emerged as a novel anticancer target. Various EZH2 inhibitors have been developed in recent years. Among these, 3-deazaneplanocin A (DZNep) is known to deplete EZH2 protein expression through an indirect pathway. In contrast, GSK343 directly inhibits enzyme activity through an S-adenosyl-L-methionine-competitive pathway. Therefore, we proposed that DZNep and GSK343 may exert differential effects against cancer cells. In this study, we found that GSK343 but not DZNep induced autophagic cell death of cancer cells. Inhibition of EZH2 expression was not required for GSK343-induced autophagy. In addition, GSK343 enhanced the anticancer activity of a multikinase inhibitor, sorafenib, in human hepatocellular carcinoma cells. Our results show that GSK343 is a more potent anticancer agent than DZNep, and for the first time, we show that it acts as an autophagy inducer.

Berberine inhibits the metastatic ability of prostate cancer cells by suppressing epithelial-to-mesenchymal transition (EMT)-associated genes with predictive and prognostic relevance.

BACKGROUND: Over 70% of cancer metastasis from prostate cancer develops bone metastases that are not sensitive to hormonal therapy, radiation therapy, or chemotherapy. The epithelial-to-mesenchymal transition (EMT) genetic program is implicated as a significant contributor to prostate cancer progression. As such, targeting the EMT represents an important therapeutic strategy for preventing or treating prostate cancer metastasis. Berberine is a natural alkaloid with significant antitumor activities against many types of cancer cells. In this study, we investigated the molecular mechanism by which berberine represses the metastatic potential of prostate cancer. METHODS: The effects of berberine on cell migration and invasion were determined by transwell migration assay and Matrigel invasion assay. Expressions of EMT-related genes were determined by an EMT PCR Array and a quantitative RT-PCR. The prognostic relevance of berberine's modulation of EMT-related genes in prostate cancer was evaluated using Kaplan-Meier survival analysis. RESULTS: Berberine exerted inhibitory effects on the migratory and invasive abilities of highly metastatic prostate cancer cells. These inhibitory effects of berberine resulted in significant repression of a panel of mesenchymal genes that regulate the developmental EMT. Among EMT-related genes downregulated by berberine, high BMP7, NODAL and Snail gene expressions of metastatic prostate cancer tissues were associated with shorter survival of prostate cancer patients and provide potential therapeutic interventions. CONCLUSIONS: We concluded that berberine should be developed as a pharmacological agent for use in combination with other anticancer drug for treating metastatic prostate cancer.

Extraction, preliminary characterization and evaluation of in vitro antitumor and antioxidant activities of polysaccharides from Mentha piperita.

This study describes the extraction, preliminary characterization and evaluation of the in vitro antitumor and antioxidant activities of polysaccharides extracted from Mentha piperita (MPP). The optimal parameters for the extraction of MPP were obtained by Box-Behnken experimental design and response surface methodology (RSM) at the ratio of water to raw material of 20, extraction time of 1.5 h and extraction temperature at 80 °C. Chemical composition analysis showed that MPP was mainly composed of glucuronic acid, galacturonic acid, glucose, galactose and arabinose, and the molecular weight of its two major fractions were estimated to be about 2.843 and 1.139 kDa, respectively. In vitro bioactivity experiments showed that MPP not only inhibited the growth of A549 cells but possessed potent inhibitory action against DNA topoisomerase I (topo I), and an appreciative antioxidant action as well. These results indicate that MPP may be useful for developing safe natural health products.

Repositioning VU-0365114 as a novel microtubule-destabilizing agent for treating cancer and overcoming drug resistance.

Microtubule-targeting agents represent one of the most successful classes of anticancer agents. However, the development of drug resistance and the appearance of adverse effects hamper their clinical implementation. Novel microtubule-targeting agents without such limitations are urgently needed. By employing a gene expression-based drug repositioning strategy, this study identifies VU-0365114, originally synthesized as a positive allosteric modulator of human muscarinic acetylcholine receptor M5 (M5 mAChR), as a novel type of tubulin inhibitor by destabilizing microtubules. VU-0365114 exhibits a broad-spectrum in vitro anticancer activity, especially in colorectal cancer cells. A tumor xenograft study in nude mice shows that VU-0365114 slowed the in vivo colorectal tumor growth. The anticancer activity of VU-0365114 is not related to its original target, M5 mAChR. In addition, VU-0365114 does not serve as a substrate of multidrug resistance (MDR) proteins, and thus, it can overcome MDR. Furthermore, a kinome analysis shows that VU-0365114 did not exhibit other significant off-target effects. Taken together, our study suggests that VU-0365114 primarily targets microtubules, offering potential for repurposing in cancer treatment, although more studies are needed before further drug development.

ASPM stabilizes the NOTCH intracellular domain 1 and promotes oncogenesis by blocking FBXW7 binding in hepatocellular carcinoma cells.

Notch signaling is aberrantly activated in approximately 30% of hepatocellular carcinoma (HCC), significantly contributing to tumorigenesis and disease progression. Expression of the major Notch receptor, NOTCH1, is upregulated in HCC cells and correlates with advanced disease stages, although the molecular mechanisms underlying its overexpression remain unclear. Here, we report that expression of the intracellular domain of NOTCH1 (NICD1) is upregulated in HCC cells due to antagonism between the E3-ubiquitin ligase F-box/WD repeat-containing protein 7 (FBXW7) and the large scaffold protein abnormal spindle-like microcephaly-associated protein (ASPM) isoform 1 (ASPM-i1). Mechanistically, FBXW7-mediated polyubiquitination and the subsequent proteasomal degradation of NICD1 are hampered by the interaction of NICD1 with ASPM-i1, thereby stabilizing NICD1 and rendering HCC cells responsive to stimulation by Notch ligands. Consistently, downregulating ASPM-i1 expression reduced the protein abundance of NICD1 but not its FBXW7-binding-deficient mutant. Reinforcing the oncogenic function of this regulatory module, the forced expression of NICD1 significantly restored the tumorigenic potential of ASPM-i1-deficient HCC cells. Echoing these findings, NICD1 was found to be strongly co-expressed with ASPM-i1 in cancer cells in human HCC tissues (P < 0.001). In conclusion, our study identifies a novel Notch signaling regulatory mechanism mediated by protein-protein interaction between NICD1, FBXW7, and ASPM-i1 in HCC cells, representing a targetable vulnerability in human HCC.