17-(dimethylaminoethylamino)-17-demethoxygeldanamycin and Carcinoma--Hepatocellular

Hepatocellular carcinoma (HCC) is the third foremost cause of cancer-related deaths. HCC has a very bad prognosis because it is asymptomatic in the early stages, resulting in a late diagnosis, and it is highly resistant to conventional chemotherapy. Such chemotherapies have been proven disappointing because they provide extremely low survival benefits. This study discloses that the STAT3/HIF-1α is an auspicious therapeutic attack site for conceivable repression of HCC development. A site that can be targeted by simultaneous administration of a STAT3 inhibitor in the context of HSP90 inhibition. 17-DMAG binds to HSP90 and constrains its function, resulting in the degradation of HSP90 client proteins HIF-1α and STAT3. Hypoxia recruits STAT3/HIF-1α complex within the VEGF promoter. Additionally, it was acknowledged that STAT3 is an essential mediator of VEGF transcription by direct binding to its promoter. Furthermore, it induces HIF-1α stability and enhances its transcriptional activity. Herein, we revealed that the combination therapy using 17-DMAG and nifuroxazide, a STAT3 inhibitor, repressed the diethylnitrosamine-induced alterations in the structure of the liver. This effect was mediated via decreasing the levels of the HSP90 client proteins HIF-1α and pSTAT3 resulting in the suppression of the STAT3/HIF-1α complex transcriptional activity. To conclude, 17-DMAG/NFXZD combination therapy-induced disruption in the STAT3/HIF-1α loop led to a potential antiangiogenic activity and showed apoptotic potential by inhibiting autophagy and inducing ROS/apoptosis signaling. Additionally, this combination therapy exhibited promising survival prolongation in mice with HCC. Consequently, the use of 17-DMAG/NFXZD renders an inspirational perspective in managing HCC. However, further investigations are compulsory.

Topics: Animals; Carcinoma, Hepatocellular; Cell Line, Tumor; Hypoxia-Inducible Factor 1, alpha Subunit; Liver Neoplasms; Mice; Vascular Endothelial Growth Factor A

Hypoxia stress plays a pivotal role in tumor formation, proliferation, and invasion. Conventional chemotherapy is less effective in the hypoxia microenvironment of solid tumor. Heat shock protein 90 (Hsp90) is an important molecular chaperone in cancer cells and has been a pharmaceutical target for decades. However, Hsp90 inhibitors demonstrate limited effect on solid tumor and the mechanism underlying is not clear. To determine whether hypoxia impairs the therapeutic effect of Hsp90 N-terminal inhibitor, 17-demethoxygeldanamycin hydrochloride (17-DMAG), in live cancer cells, we measured cell proliferation and cell cycle distribution. Cell proliferation assay indicates that hypoxia obviously promotes the proliferation of HepG2 and Huh7 cells after 24, 48, and 72 h and impairs 17-DMAG-induced G2/M arrest in liver cancer cells. As a client protein of Hsp90, cyclin B1 is critical for the transition from G2 to M phase and is related to the prognosis of the patients. We further checked the cyclin B1 messenger RNA (mRNA) level, protein level, ubiquitination of cyclin B1, nuclear translocation, and degradation of cyclin B1 affected by hypoxia after 17-DMAG treatment. The results demonstrate that hypoxia decreases the transcription of cyclin B1 and accelerates the ubiquitination, nuclear translocation, and degradation of cyclin B1. Taken together, our results suggest that hypoxia attenuates cyclin B1 accumulation induced by 17-DMAG and, hence, alleviates 17-DMAG-induced G2/M arrest.

Topics: Antineoplastic Agents; Apoptosis; Benzoquinones; Carcinoma, Hepatocellular; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cyclin B1; HSP90 Heat-Shock Proteins; Humans; Hypoxia; Lactams, Macrocyclic; Liver; Liver Neoplasms

Due to the lack of effective treatment, hepatocellular carcinoma (HCC) is one of the malignancies with low survival rates worldwide. Combination of hyperthermia and chemotherapy has shown promising results in several abdominal tumours, but high expression of HSP90 in tumours attenuated the efficacy of hyperthermia. Thus a combination of hyperthermia and inhibition of HSP90 might be a feasible therapeutic strategy for HCC. One hepatic cell line (L02) and two HCC cell lines (Huh7 and HepG2) were heated at 42 °C for 0, 0.5 or 4 h with or without 100 nM 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG). HCC cells of the combination group exhibited more G2/M arrest and higher apoptotic rates which might result from suffering from more reactive oxygen species and serious DNA damage. Heat shock/17-DMAG co-treatment of HCC cells also destabilized CDK1, Cyclin B1 and CDC25C with a concomitant decreased proportion of cells in the M phase. Furthermore, co-treatment impaired the interaction of HSP90α with CDC37 and with CDK1, accompanied with decreased soluble CDK1. Combination of 17-DMAG with a 1.5-h whole body hyperthermia treatment attenuated tumour growth in xenograft mice models. These results suggest hyperthermia sensitize HCC to 17-DMAG, and combination of hyperthermia with 17-DMAG might be a potential therapeutic strategy for HCC.

Topics: Animals; Benzoquinones; Carcinoma, Hepatocellular; CDC2 Protein Kinase; Cell Cycle Proteins; Cell Line, Tumor; Cell Survival; Chaperonins; Cyclin B1; DNA Damage; G2 Phase Cell Cycle Checkpoints; Hep G2 Cells; HSP90 Heat-Shock Proteins; Humans; Hyperthermia, Induced; Lactams, Macrocyclic; Liver Neoplasms; Male; Mice, Inbred BALB C; Reactive Oxygen Species; Xenograft Model Antitumor Assays

Genomic analyses promise to improve tumor characterization to optimize personalized treatment for patients with hepatocellular carcinoma (HCC). Exome sequencing analysis of 243 liver tumors identified mutational signatures associated with specific risk factors, mainly combined alcohol and tobacco consumption and exposure to aflatoxin B1. We identified 161 putative driver genes associated with 11 recurrently altered pathways. Associations of mutations defined 3 groups of genes related to risk factors and centered on CTNNB1 (alcohol), TP53 (hepatitis B virus, HBV) and AXIN1. Analyses according to tumor stage progression identified TERT promoter mutation as an early event, whereas FGF3, FGF4, FGF19 or CCND1 amplification and TP53 and CDKN2A alterations appeared at more advanced stages in aggressive tumors. In 28% of the tumors, we identified genetic alterations potentially targetable by US Food and Drug Administration (FDA)-approved drugs. In conclusion, we identified risk factor-specific mutational signatures and defined the extensive landscape of altered genes and pathways in HCC, which will be useful to design clinical trials for targeted therapy.

Topics: Aged; Antineoplastic Agents; Benzoquinones; Carcinoma, Hepatocellular; Cell Line, Tumor; DNA Mutational Analysis; Exome; Female; Genetic Association Studies; HSP90 Heat-Shock Proteins; Humans; Lactams, Macrocyclic; Liver Neoplasms; Male; Molecular Targeted Therapy; NAD(P)H Dehydrogenase (Quinone); Risk Factors; Sequence Deletion

Hepatocellular carcinoma (HCC) is a severe liver malignancy with few drug treatment options. In finding an effective treatment for HCC, screening drugs that are already FDA-approved will fast track the clinical trial and drug approval process. Connectivity Map (CMap), a large repository of chemical-induced gene expression profiles, provides the opportunity to analyze drug properties on the basis of gene expression. Support Vector Machines (SVM) were utilized to classify the effectiveness of drugs against HCC using gene expression profiles in CMap. The results of this classification will help us (1) identify genes that are chemically sensitive, and (2) predict the effectiveness of remaining chemicals in CMap in the treatment of HCC and provide a prioritized list of possible HCC drugs for biological verification. Four HCC cell lines were treated with 146 distinct chemicals, and cell viability was examined. SVM successfully classified the effectiveness of the chemicals with an average Area Under ROC Curve (AUROC) of 0.9. Using reported HCC patient samples, we identified chemically sensitive genes that may be possible HCC therapeutic targets, including MT1E, MYC, and GADD45B. Using SVM, several known HCC inhibitors, such as geldanamycin, alvespimycin (HSP90 inhibitors), and doxorubicin (chemotherapy drug), were predicted. Seven out of the 23 predicted drugs were cardiac glycosides, suggesting a link between this drug category and HCC inhibition. The study demonstrates a strategy of in silico drug screening with SVM using a large repository of microarrays based on initial in vitro drug screening. Verifying these results biologically would help develop a more accurate chemical sensitivity model.

Topics: Antineoplastic Agents; Benzoquinones; Carcinoma, Hepatocellular; Cell Line, Tumor; Computer Simulation; Doxorubicin; Drug Screening Assays, Antitumor; Gene Expression Regulation, Neoplastic; Humans; Lactams, Macrocyclic; Liver Neoplasms; Oligonucleotide Array Sequence Analysis; ROC Curve; Support Vector Machine; Transcriptome

Primary liver cancer is one of the highly malignant tumours. The traditional surgery, chemotherapy and radiation therapy only established 6% of 5-year survival rate in HCC (hepatocellular carcinoma). Therefore there is an urgent need to develop new therapeutic strategies. HSP90 (heat shock protein 90) is one of the important molecular chaperones and was identified with high expression in the primary liver cancer. In this study, we evaluated the therapeutic effect of specific HSP90 inhibitor 17-DMAG (17-dimethylaminoethylamino-17-demethoxy geldanamycin) in HCC cells. The time and concentration effects of 17-DMAG were investigated in HCC cells. Cell proliferation was measured by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay and cell counting. Apoptosis was detected by flow cytometry with staining of Annexin V-FITC/PI (propidium iodide). The protein levels of survivin, cyclin D1, p53 and NF-κB (nuclear factor κB) were measured by Western blotting. 17-DMAG inhibited the proliferation of HCC cells in a time- and concentration-dependent manner. Treatment with 400 nmol/l 17-DMAG for 48 h significantly induced early-stage apoptosis (22.4%). Conversely, it induced less late-stage apoptosis (3.03%). The 5 mg/l of cisplatin induced significantly less early-stage apoptosis (6.5%), but similar proportion of late-stage apoptosis (4.89%) compared with 17-DMAG. Inhibition of HSP90 activity by 400 nmol/l 17-DMAG decreased protein levels of survivin, cyclin D1 and NF-κB protein levels, whereas increased p53 protein level. HSP90 plays a key role in HCC cell growth and survival through regulation of survivin, cyclin D1, p53 and nucleus NF-κB protein levels and the specific HSP90 inhibitor 17-DMAG can play a therapeutic role in HCC treatment.

Topics: Antineoplastic Agents; Apoptosis; Benzoquinones; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Proliferation; Cisplatin; Cyclin D1; HSP90 Heat-Shock Proteins; Humans; Inhibitor of Apoptosis Proteins; Lactams, Macrocyclic; Liver Neoplasms; NF-kappa B; Survivin; Tumor Suppressor Protein p53