alvocidib and Carcinoma--Hepatocellular

To clarify the effects of cylcin E1 expression on HCC tumor progression, we studied the expression of cyclin E1 and inhibitory efficacy of regorafenib and sorafenib in HCC cells, and investigated a potential therapy that combines regorafenib treatment with cyclin E1 inhibition.. Western blotting for caspase-3 and Hoechst 33225 staining was used to measure the expression level of apoptosis-related proteins under drug treatment.. Our results showed that enhanced expression of cyclin E1 after transfection compromised apoptosis in HCC cells induced by regorafenib or sorafenib. Conversely, down-regulation of cyclin E1 gene expression or inhibition of cyclin E1 by the cyclin-dependent kinase (CDK) inhibitors dinaciclib (DIN) or flavopiridol sensitized HCC cells to regorafenib and sorafenib by inducing apoptosis. The expression of Mcl-1, which is modulated by STAT3, plays a key role in regulating the therapeutic effects of CDK inhibitors. Xenograft experiments conducted to test the efficacy of regorafenib combined with DIN showed dramatic tumor inhibitory effects due to induction of apoptosis. Our results suggested that the level of cyclin E1 expression in HCCs may be used as a pharmacodynamic biomarker to assess the antitumor effects of regorafenib or sorafenib.. Combining regorafenib and CDK inhibitors may enhance the clinical efficiency of the treatment of HCCs.

Topics: Animals; Apoptosis; Bridged Bicyclo Compounds, Heterocyclic; Carcinoma, Hepatocellular; Cell Line, Tumor; Cyclic N-Oxides; Cyclin E; Drug Synergism; Flavonoids; Gene Expression Regulation, Neoplastic; Humans; Indolizines; Liver Neoplasms; Male; Mice; Myeloid Cell Leukemia Sequence 1 Protein; Oncogene Proteins; Phenylurea Compounds; Piperidines; Prognosis; Pyridines; Pyridinium Compounds; Sorafenib; STAT3 Transcription Factor; Xenograft Model Antitumor Assays

A previous study showed that flavopiridol increased doxorubicin sensitivity in hypoxic hepatocellular carcinoma (HCC) cells by increasing apoptosis through suppressing hypoxia-inducible N-myc downstream-regulated gene-1 (NDRG1) expression. However, this has not been investigated in an in vivo HCC model. Therefore, we aimed to elucidate whether the combination of doxorubicin and flavopiridol has a synergistic anti-tumor effect in an in vivo HCC model.. An HCC mouse model was established by implanting C3H/He mouse with MH134 cells. Then, doxorubicin with or without flavopiridol was injected. The anti-tumor efficacy was assessed by evaluating tumor volumes, and the underlying mechanism was investigated by quantifying apoptotic cells, the Ki-67 proliferation index, and microvessel densities (MVDs). Immunohistochemistry of NDRG1 was performed to determine the underlying mechanism.. Tumor growth was significantly suppressed in the doxorubicin + flavopiridol combination group compared to the other three groups. The percentage of apoptotic cells was significantly higher, and Ki-67-positive proliferating cells were significantly lower in the combination group compared to the other groups; however, MVDs were not significantly different across the groups. Increased apoptosis by flavopiridol occurred by suppressing hypoxia-inducible NDRG1 expression.. These results show that a combination of doxorubicin and flavopiridol has a synergistic anti-tumor effect in an in vivo HCC model. This synergistic effect of combination therapy was attributed to increased apoptosis and decreased proliferation of tumor cells rather than decreased angiogenesis. These findings suggest that flavopiridol might be an effective adjuvant therapy to doxorubicin-resistant HCC cells by inducing apoptosis through suppression of NDRG1 expression.

Topics: Animals; Antibiotics, Antineoplastic; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Carcinoma, Hepatocellular; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Doxorubicin; Drug Synergism; Flavonoids; Hexokinase; Hypoxia-Inducible Factor 1, alpha Subunit; Intracellular Signaling Peptides and Proteins; Liver Neoplasms; Male; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Piperidines; Protein Kinase Inhibitors; Tumor Burden

Flavopiridol was one of the first cyclin-dependent kinase inhibitors demonstrated to have an antitumor effect in several cancer types. Here, we investigated the effects of flavopiridol on TNF-related apoptosis-inducing ligand (TRAIL) in the human hepatocellular carcinoma (HCC) cell lines HLE and HepG2, and evaluated the role of flavopiridol in apoptosis. To better understand the mechanism of increased TRAIL sensitivity in HCC cells, we determined the effect of flavopiridol on cell surface expression of TRAIL and TRAIL receptors using flow cytometry analysis. The levels of survivin, FLIP, Bcl-xL and X-chromosome-linked IAP (XIAP) in treated and untreated cells was also determined. Flavopiridol decreased cell viability in a dose-dependent manner in the two HCC cell lines tested. The pan-caspase inhibitor z-VAD-FMK did not inhibit the effect. However, subtoxic levels of flavopiridol dramatically enhanced TRAIL-induced apoptosis in both cells. Flavopiridol up-regulated TRAIL, TRAIL-R1 and TRAIL-R2 in both cell lines. In addition, flavopiridol down-regulated expression of survivin in both cell lines, and expression of FLIP and Bcl-xL were down-regulated in HLE cells. In summary, flavopiridol augmented TRAIL sensitivity by up-regulation of TRAIL receptors and down-regulation of survivin, FLIP and Bcl-xL. Thus, combining flavopiridol with a TRAIL agonist may prove to be an effective new strategy for treatment of HCC.

Topics: Antineoplastic Agents; Apoptosis; bcl-X Protein; Carcinoma, Hepatocellular; CASP8 and FADD-Like Apoptosis Regulating Protein; Cell Line, Tumor; Cell Survival; Cyclin-Dependent Kinases; fas Receptor; Flavonoids; Humans; Inhibitor of Apoptosis Proteins; Liver Neoplasms; Microtubule-Associated Proteins; Neoplasm Proteins; Piperidines; Protein Kinase Inhibitors; Receptors, TNF-Related Apoptosis-Inducing Ligand; Survivin; TNF-Related Apoptosis-Inducing Ligand; Tumor Necrosis Factor-alpha