Perspective Too Much Cleavage of Cyclin E Promotes Breast Tumorigenesis Keith R. Loeb1,2,3*, Xueyan Chen1 1 Department of Laboratory Medicine, University of Washington School of Medicine, Seattle, Washington, United States of America, 2 Department of Pathology, University of Washington School of Medicine, Seattle, Washington, United States of America, 3 Division of Clinical Research, Fred Hutchinson Cancer Research Center, Seattle, Washington, United States of America Cyclin E, together with cyclin-depen- dent kinase 2 (CDK2), functions as a gatekeeper to promote G1/S transitions and the initiation of DNA replication. In normal cells, cyclin E–associated kinase activity is exquisitely regulated, with ac- tivity being limited to a brief time interval between late G1 and early S phase. Human cancers frequently exhibit dereg- ulated cyclin E–associated kinase activity resulting from overexpression of cyclin E and loss of cyclin-dependent kinase inhi- bition (via p53 mutations) promoting genetic instability and cell proliferation [1]. Increased levels of cyclin E correlate with tumorigenesis and are a poor prog- nostic indicator independent of prolifera- tion rate, suggesting that cyclin E’s role in tumorigenesis is not limited to promoting increased cell proliferation [2,3]. By elim- inating regulatory constraints using p53 cells, we and others have shown that overexpression or endogenous expression of stabilizing mutant forms of cyclin E can lead to hyperproliferation, genetic insta- bility, and malignancy in cell culture and murine models [4,5]. Normal cells sup- press the effects of excess/stabilized cyclin E via the G1/S checkpoint involving the p53/p21 pathway. Five isoforms of cyclin E, ranging in size from 33 to 44 kDa, have been identified in tumors over-expressing cyclin E. These low molecular weight forms of cyclin E (LMW-E) are generated through post- translational cleavage of full-length cyclin E by the elastase family of serine proteases in tumor cells [6,7]. In comparison to full- length cyclin E (50 kDa), LMW-E forms are uniquely expressed in tumor cells, exhibit enhanced CDK2-associated kinase activity, have increased affinity for CDK2 [7–9], and exhibit decreased inhibition by CDK2 inhibitors, p21 and p27 (Figure 1) [10,11]. Ectopic expression of LMW-E isoforms promotes cell proliferation, ge- netic instability, centrosome amplification, and malignancy [12,13]. In addition, clinical studies have shown that high LMW-E is strongly associated with poor survival in breast cancer [2], colorectal cancer [14,15], ovarian cancer [16], and melanomas [17]. Given its unique prop- erties and distinct function in human cancers, targeting LMW-E could have important therapeutic implications. The study by Duong et al. in this issue of PLoS Genetics [18] convincingly uncovers the tumorigenic potential of LMW-E. The authors used three different model sys- tems—3D acinar cultures, xenograft trans- plantation, and transgenic mice—to show that overexpression of LMW-E is sufficient to induce aberrant acinar morphogenesis in culture and mammary tumors in mice (Figure 1). When grown on Matrigel, immortalized human mammary epithelial cells (hMECs) expressing LMW-E exhibit large misshapen multiacinar structures resulting from defective growth arrest and apoptosis that mimic morphologic features of breast carcinomas. Further, ectopic expression of LMW-E in immor- talized hMECs promotes tumorigenesis in xenografts and transgenic mice to a much greater extent than full-length cyclin E. Consistent with the reports by Akli et al. and Nanos-Webb et al., tumorigenesis associated with LMW-E is dependent on CDK2 [19,20]. Furthermore, in vivo passaging of tumor cells increases the expression of LMW-E, suggesting that LMW-E provides a selective growth ad- vantage to the tumor. Duong et al. also took advantage of a proteomic analysis termed reverse-phase protein array assay (RPPA) to examine protein expression patterns in cultured tumor cells and in breast tumors expressing high LMW-E levels. Their analyses revealed that multi- ple components of the b-RAF-ERK1/2- mTOR pathway are elevated in these cells. Activation of the b-RAF-ERK1/2- mTOR pathway normally promotes cell division and cell survival. Consistent with this, the authors observed that endogenous cyclin E levels are also increased in cells expressing high LMW-E, indicative of the existence of a positive feedback loop that promotes cell proliferation. Both high LMW-E levels and up-regulation of the b-RAF-ERK1/2-mTOR signaling path- way are associated with poor survival, suggesting functional correlation of these events in aggressive tumors. Importantly, the authors demonstrated that combina- tion therapy targeting LMW-E/CDK2 and the b-RAF-ERK1/2-mTOR pathway has a synergistic effect in abrogating the tumorigenic effect of LMW-E. Thus, the identification of these downstream regula- tors may provide novel biomarkers and/or potential therapeutic targets for LMW-E– expressing tumors. The report that LMW-E potentiates tumorigenesis in three independent model systems associated with activation of the b- RAF-ERK1/2-mTOR pathway is intrigu- ing. However, there are many important questions about the role of LMW-E in tumorigenesis that need to be addressed. 1) What is the functional relationship be- tween LMW-E and full-length cyclin E? In each tumor model reported by Duong et al., the effect was examined by over- expressing LMW-E in a background of endogenous full-length cyclin E. Further, the authors show that ectopic expression of LMW-E in transplanted xenografts trig- gers tumor evolution and results in increased levels of endogenous cyclin E. Thus, the contribution of endogenous full- Citation: Loeb KR, Chen X (2012) Too Much Cleavage of Cyclin E Promotes Breast Tumorigenesis. PLoS Genet 8(3): e1002623. doi:10.1371/journal.pgen.1002623 Editor: Marshall S. Horwitz, University of Washington, United States of America Published March 29, 2012 Copyright: 2012 Loeb, Chen. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The authors received no specific funding for this article. Competing Interests: The authors have declared that no competing interests exist. * E-mail: kloeb@fhcrc.org PLoS Genetics | www.plosgenetics.org 1 March 2012 | Volume 8 | Issue 3 | e1002623