Cyclin B1 is the regulatory subunit of M-phase promoting factor, and proper regulation of cyclin B1 is essential for the initiation of mitosis. Increasing evidence indicates that the deregulation of cyclin B1 is involved in neoplastic transformation, suggesting the suppression of cyclin B1 could be an attractive strategy for antiproliferative therapy. In the present work, we analysed the impact of small interfering RNAs (siRNAs) targeted to cyclin B1 on different human tumor cell lines. Cyclin B1 siRNAs reduced the protein level of cyclin B1 in HeLa, MCF-7, BT-474 and MDA-MB-435 tumor cells and efficiently reduced the kinase activity of Cdc2/cyclin B1 in HeLa cells. siRNA-treated cells were arrested in G2/M phase in all tumor cell lines tested. Proliferation of tumor cells from different origins was suppressed by 50-80% 48 h after transfection and apoptosis was increased from 5 to 40-50%. Furthermore, tumor cells showed less colonyforming ability after siRNA treatment. In contrast, primary human umbilical vein endothelial cells exhibited only a slight change in cell cycle, and neither apoptosis nor clear inhibition of proliferation was observed after cyclin B1 siRNA treatment for 48 h. These results indicate that siRNAs against cyclin B1 could become a powerful antiproliferative tool in future antitumor therapy.

Aldo-keto reductase family 1 member B10 (AKR1B10) is primarily expressed in the normal human colon and small intestine but overexpressed in liver and lung cancer. Our previous studies have shown that AKR1B10 mediates the ubiquitin-dependent degradation of acetyl-CoA carboxylase-␣. In this study, we demonstrate that AKR1B10 is critical to cell survival. In human colon carcinoma cells (HCT-8) and lung carcinoma cells (NCI-H460), small-interfering RNA-induced AKR1B10 silencing resulted in caspase-3-mediated apoptosis. In these cells, the total and subspecies of cellular lipids, particularly of phospholipids, were decreased by more than 50%, concomitant with 2-3-fold increase in reactive oxygen species, mitochondrial cytochrome c efflux, and caspase-3 cleavage. AKR1B10 silencing also increased the levels of ␣,␤-unsaturated carbonyls, leading to the 2-3-fold increase of cellular lipid peroxides. Supplementing the HCT-8 cells with palmitic acid (80 M), the end product of fatty acid synthesis, partially rescued the apoptosis induced by AKR1B10 silencing, whereas exposing the HCT-8 cells to epalrestat, an AKR1B10 inhibitor, led to more than 2-fold elevation of the intracellular lipid peroxides, resulting in apoptosis. These data suggest that AKR1B10 affects cell survival through modulating lipid synthesis, mitochondrial function, and oxidative status, as well as carbonyl levels, being an important cell survival protein.Aldo-keto reductase family 1 member B10 (AKR1B10, 2 also designated aldose reductase-like-1, ARL-1) is primarily expressed in the human colon, small intestine, and adrenal gland, with a low level in the liver (1-3). However, this protein is overexpressed in hepatocellular carcinoma, cervical cancer, lung squamous cell carcinoma, and lung adenocarcinoma in smokers, being a potential diagnostic and/or prognostic marker (1, 2, 4 -6).The biological function of AKR1B10 in the intestine and adrenal gland, as well as its role in tumor development and progression, remains unclear. AKR1B10 is a monomeric enzyme that efficiently catalyzes the reduction to corresponding alcohols of a range of aromatic and aliphatic aldehydes and ketones, including highly electrophilic ␣,␤-unsaturated carbonyls and antitumor drugs containing carbonyl groups, with NADPH as a co-enzyme (1, 7-12). The electrophilic carbonyls are constantly produced by lipid peroxidation, particularly in oxidative conditions, and are highly cytotoxic; through interaction with proteins, peptides, and DNA, the carbonyls cause protein dysfunction and DNA damage (breaks and mutations), resulting in mutagenesis, carcinogenesis, or apoptosis (10, 13-19). AKR1B10 also shows strong enzymatic activity toward all-trans-retinal, 9-cis-retinal, and 13-cis-retinal, reducing them to the corresponding retinols, which may regulate the intracellular retinoic acid, a signaling molecule modulating cell proliferation and differentiation (6, 20 -23). In lung cancer, AKR1B10 expression is correlated with the patient smoking history and activates procarcinogens in cigarette sm...

Recent studies have demonstrated that aldo-keto reductase family 1 B10 (AKR1B10), a novel protein overexpressed in human hepatocellular carcinoma and non-small cell lung carcinoma, may facilitate cancer cell growth by detoxifying intracellular reactive carbonyls. This study presents a novel function of AKR1B10 in tumorigenic mammary epithelial cells (RAO-3), regulating fatty acid synthesis. In RAO-3 cells, Sephacryl-S 300 gel filtration and DEAE-Sepharose ion exchange chromatography demonstrated that AKR1B10 exists in two distinct forms, monomers (ϳ40 kDa) bound to DEAE-Sepharose column and protein complexes (ϳ300 kDa) remaining in flow-through. Co-immunoprecipitation with AKR1B10 antibody and protein mass spectrometry analysis identified that AKR1B10 associates with acetyl-CoA carboxylase-␣ (ACCA), a rate-limiting enzyme of de novo fatty acid synthesis. This association between AKR1B10 and ACCA proteins was further confirmed by co-immunoprecipitation with ACCA antibody and pulldown assays with recombinant AKR1B10 protein. Intracellular fluorescent studies showed that AKR1B10 and ACCA proteins colocalize in the cytoplasm of RAO-3 cells. More interestingly, small interfering RNA-mediated AKR1B10 knock down increased ACCA degradation through ubiquitination-proteasome pathway and resulted in >50% decrease of fatty acid synthesis in RAO-3 cells. These data suggest that AKR1B10 is a novel regulator of the biosynthesis of fatty acid, an essential component of the cell membrane, in breast cancer cells.Aldo-keto reductase family 1 B10 (AKR1B10, 2 also designated aldose reductase-like-1, ARL-1) is a novel protein identified from human hepatocellular carcinoma (1). This protein belongs to the aldo-keto reductase superfamily, a group of proteins implicated in intracellular detoxification, cell carcinogenesis, and cancer therapeutics (2-5). AKR1B10 is primarily expressed in the colon and small intestine with low levels in the liver, thymus, prostate, and testis (1). However, this gene is overexpressed in 54% of human hepatocellular carcinoma, 84.4% of lung squamous cell carcinoma, and 29.2% of lung adenocarcinoma in smokers, making it a potential diagnostic and/or prognostic marker (1, 6, 7). AKR1B10 is an enzyme that efficiently catalyzes the reduction of carbonyls to corresponding alcohols with NADPH as a co-enzyme (1). Recent studies demonstrate that AKR1B10 expression facilitates growth of cancer cells, enhances their clonogenic capability, and reduces their susceptibility to reactive carbonyls such as acrolein and crotonaldehyde (8, 9). In vitro, AKR1B10 also shows strong enzymatic activity toward all-trans-retinal, 9-cis-retinal, and 13-cis-retinal, reducing them to the corresponding retinols. The diversity of retinal metabolism may diminish intracellular retinoic acid, a signaling molecule regulating cell proliferation and differentiation (4, 10).The current study presents a novel biological function of AKR1B10, regulating long chain fatty acid synthesis, in human breast cancer cells. During tumorigenic transformatio...

Mitochondria are dynamic organelles that undergo constant fission and fusion cycles. In response to cellular damage, this balance is shifted dramatically toward fission and apoptosis. This work describes the role of the transcription factor cyclin C in mediating both responses.

Cyclin B1 is the regulatory subunit of cyclin-dependent kinase 1 (Cdk1) and is critical for the initiation of mitosis. Accumulating data indicate that the deregulation of cyclin B1 is tightly linked to neoplastic transformation. To study the phenotype and the potential preclinical relevance, we generated HeLa cell lines stably transfected with the plasmids encompassing short hairpin RNA (shRNA) targeting cyclin B1. We demonstrate that the reduction of cyclin B1 caused inhibition of proliferation by arresting cells in G2 phase and by inducing apoptosis. Cells, entering mitosis, were impaired in chromosome condensation and alignment. Importantly, HeLa cells with reduced cyclin B1 were more susceptible to the treatment of small interfering RNA targeting Polo-like kinase 1 (Plk1) and to the administration of the chemotherapeutic agent taxol. Finally, HeLa cells with reduced cyclin B1 showed inhibited tumor growth in nude mice compared to that of control cells. In summary, our data indicate that cyclin B1 is an essential molecule for tumor cell survival and aggressive proliferation, suggesting that the downregulation of cyclin B1, especially in combination with other molecular targets, might become an interesting strategy for antitumor intervention.