Retinoid chemoprevention trials: cyclin D1 in the crosshairs.

Sarah J. Freemantle, Yongli Guo and Ethan Dmitrovsky Successful clinical cancer chemoprevention with retinoids was promised by compelling preclinical and early clinical evidence. Indeed, the concept of cancer chemoprevention came largely from studies showing retinoid suppression of epithelial carcinogenesis (1–4). Yet, randomized phase III intergroup chemoprevention trials with the classic retinoid isotretinoin did not reduce second primary cancers in patients with early stages of non–small cell lung cancer (5) or head and neck squamous cell carcinoma (6). Absence of preventive activity in these trials may be attributed to frequent silencing of the retinoic acid receptor-β, which can confer resistance to classic retinoids (7). Several hypothesis-driven approaches for circumventing loss of retinoic acid receptor-β signaling have been developed, including the use of nonclassic retinoids that act through the retinoid X receptors or that have receptorindependent activities. The nonclassic retinoid N-(4-hydroxyphenyl)retinamide (or fenretinide) was investigated in two studies by the same group, William and colleagues (8) and Papadimitrakopoulou and colleagues (9), reported in this issue of the journal. The authors are commended for completing hypothesis-driven trials with a nonclassic retinoid. Fenretinide at elevated concentrations can induce apoptosis (10). William and colleagues (8) describe a trial of high-dose fenretinide in oral leukoplakia patients with the rationale of achieving serum levels sufficient to induce apoptosis anticipated from preclinical studies. Yet, this study was closed early by design because of an insufficient response rate in the first cohort of patients. Papadimitrakopoulou and colleagues (9) treated moderate-to-severe laryngeal dysplasia patients with a regimen that combined daily 13-cis-retinoic acid and α-tocopherol with twice weekly α-IFN for 1 year. Nonprogressing patients were randomized to maintenance low-dose fenretinide or placebo for 2 years. Despite a previous positive trial of a regimen similar to that used in this induction phase (11), the present trial showed no apparent effect of either induction or maintenance therapy on laryngeal cancer development (9). A prior study from this group found an intriguing association between shorter cancer-free survival and presence of the G/A870 cyclin D1 polymorphism (12). In the current study, these investigators confirmed this earlier finding and found additionally that high intralesional cyclin D1 protein expression (measured immunohistochemically) was also linked to shorter overall cancer-free survival (9). This work implicates cyclin D1 genotype and protein expression as molecular cancer risk factors and highlights cyclin D1 as a molecular pharmacologic target. Although many interesting aspects of these chemoprevention trials warrant comment, this perspective focuses on the therapeutic and preventive potential of targeting cyclin D1. Cyclin D1 has been associated with carcinogenesis since it was cloned in 1991 as the gene rearranged in a subset of parathyroid tumors (13). Also known as bcl1, the cyclin D1 genomic locus at chromosome 11q13 is altered in multiple cancer types. This region is amplified in a subset of human breast cancers and squamous cell cancers and at the breakpoint of the chromosomal translocation t(11;14)(q13;q32) in certain B-cell neoplasms. Murine studies also identified the cyclin D1 locus as a frequent hotspot for proviral insertions resulting in lymphoma (14–17). The D-type cyclins are activating regulators of cyclindependent kinases (CDK) 4 and 6 (18). These CDKs phosphorylate the retinoblastoma protein, which releases E2F transcription factors allowing expression of genes required for S-phase entry and cell cycle progression. Cyclin D1 levels are elevated in many cancers through diverse mechanisms (19). In both engineered H-ras and c-neu/erbB-2 murine breast cancer models, tumors do not form in the absence of cyclin D1. This finding elegantly shows the key role cyclin D1 plays in carcinogenesis (20). Considerable attention has turned to targeting both CDK activity and associated mitogenic pathways that deregulate cyclin D1 levels (21). Mitogenic signaling pathways maintain sufficient cyclin D1 levels to permit cell cycle progression predominantly through transcriptional regulation of cyclin D1. Signals that activate transcription from the cyclin D1 promoter include those mediated by ErbB2, Ras, Wnt, and signal transducer and activator of transcription (22). Cyclin D1 levels are regulated by multiple mechanisms. When a rapid decrease in cyclin D1 is required, a pathway of accelerated ubiquitin-mediated proteolysis is engaged. This pathway is activated at the end of S phase and in response to cellular stress. Cyclin D1 is phosphorylated at residue Thr, is exported from the nucleus, and is recognized by a specific ubiquitin ligase that targets it for degradation by the proteasome (21, 23). Glycogen synthase kinase-3β (GSK-3β) is the most extensively studied kinase that phosphorylates cyclin D1 at Thr (24), although p38SAPK2 and extracellular signal-regulated kinase 2 can also target this site (23). Cyclin D1b is a naturally occurring alternatively spliced cyclin D1 species, which lacks the COOH terminus, including residue Thr, present in wild-type cyclin D1. It is localized predominantly in the nucleus and exhibits oncogenic activity (25). A specific polymorphism of cyclin D1 (G/A870) at the Authors' Affiliations: Department of Pharmacology and Toxicology, Department of Medicine, and Norris Cotton Cancer Center, Dartmouth Medical School, Hanover, New Hampshire and Dartmouth-Hitchcock Medical Center, Lebanon, New Hampshire Received 10/27/2008; accepted 11/24/2008. Requests for reprints: Sarah J. Freemantle, Department of Pharmacology and Toxicology, Dartmouth Medical School, 7650 Remsen, Hanover, NH 03755. Phone: 603-650-1667; Fax: 603-650-1129; E-mail: sarah.freemantle@ dartmouth.edu. ©2009 American Association for Cancer Research. doi:10.1158/1940-6207.CAPR-08-0218