Bortezomib Stabilizes Mitotic Cyclins and Prevents Cell Cycle Progression via Inhibition of UBE2C in Colorectal Carcinoma

Abstract

Substantial evidence implicates the ubiquitin-conjugating enzyme E2C (UBE2C) gene, in several human cancers, including colorectal carcinoma (CRC). We therefore investigated the prognostic value of UBE2C alterations in CRC and UBE2C signaling in CRC cell lines. UBE2C protein expression and UBE2C gene copy number were evaluated on clinical samples by immunohistochemistry and fluorescence in situ hybridization in a TMA format. The effect of the proteasome inhibitor bortezomib and small-interfering RNA knockdown was assessed by apoptotic assays and immunoblotting. UBE2C dysregulation was associated with proliferative marker Ki-67, accumulation of cyclin A and B1, and a poor overall survival. UBE2C expression was an independent prognostic marker in early-stage (I and II) CRC. UBE2C depletion resulted in suppression of cellular growth and accumulation of cyclin A and B1. In vitro, bortezomib treatment of CRC cells caused inhibition of cell viability via down-regulation of UBE2C. UBE2C knockdown by bortezomib or transfection with specific small-interfering RNA against UBE2C also caused cells to be arrested at the G2/M level, leading to accumulation of cyclin A and cyclin B1. In vivo, a significant reduction in tumor volume and weight was noted in mice treated with a combination of subtoxic doses of oxaliplatin and bortezomib compared with treatment with oxaliplatin or bortezomib alone. Altogether, our results suggest that UBE2C and the ubiq- Although colorectal cancer (CRC) is a major cause of mortality and morbidity worldwide, various therapeutic modalities followed in clinical practice are not life saving. More recently, advances in understanding of tumor biology have led to the development of targeted therapies, 1 allowing progress in the treatment of CRC. 2 The ubiquitin proteasome system is important for the orchestration of several important cell cycle events, such as proteolysis of cyclin-dependent kinase and their inhibitors. 3,4 Proper cell cycle progression is orchestrated by the controlled oscillation of a series of cell cycle events. Deregulation of appropriate cell cycle control often results in chromosomal instability, which is a potential trigger for the initiation of cancer. 5 In this system, substrate molecules are regulated for degradation by ubiquitinactivating enzyme (E1), ubiquitin-conjugating enzyme (E2), and E3 ligase. The ubiquitin-conjugating enzyme E2C (UBE2C), also known as UBCH10, along with the E3 ligase of the anaphase-promoting complex catalyze the ubiquitination of mitotic cyclins A and B1, as well as securin. 3,6 UBE2C is essential for cell cycle progression, and mutation of its active site cysteine confers a dominant-negative phenotype. 3,6 Overexpression of UBE2C at the mRNA level has been reported in a number of cancer cell lines and primary tumors, whereas only low levels were found in normal tissues. 7 The oncogenic role of UBE2C is also reported in thyroid, 8 ovarian, 9 esopha-

New targeted approaches against the ubiquitin–proteasome system in gastrointestinal malignancies

“…Despite the fact that several preclinical trials have shown that bortezomib may be useful for the treatment of colorectal cancer, by inhibition of UBE2C [77] and increasing the CDK inhibitor p27 Kip1 [78], up to now, the results in the clinical setting have been disappointing (Table 1).…”

UBE2C induces EMT through Wnt/β-catenin and PI3K/Akt signaling pathways by regulating phosphorylation levels of Aurora-A

“…Interestingly, UBE2C is nearly undetectable in normal tissues, at both the RNA or protein levels. In contrast, increasing evidence indicates that UBE2C protein is overexpressed in many human tumor types, including brain tumors (10), malignant breast carcinomas (11), lung cancer (12,13), anaplastic thyroid carcinomas (14), esophageal adenocarcinomas (15), hepatocellular carcinomas (16) and colorectal cancer (17–19), suggesting that UBE2C may play an important role in tumorigenesis and progression. UBE2C overexpression causes the centrosome duplication instability, and variety of proteins can cause this phenomenon, including the anaphase-promoting complex (APC/C) substrates cyclin B, AURKA and PLK1 (20,21).…”

Therapeutic targeting of cancer cell cycle using proteasome inhibitors

“…Rapid proteasomal degradation of CKIs in subsequent cell cycle phases contribute to the uncontrolled cell division in cancer cells. Recent studies have established the involvement of ubiquitin proteasome pathway (UPP) in degradation and recycling of the above mentioned class of proteins, crucial for cancer onset and progression [29]. The proteasomal degradation of proteins is well complemented by the ubiquitin conjugating enzymes E1, E2 and E3 ligases, which add multiple ubiquitin molecules to the proteasome substrate proteins.…”