Cancer cells exhibit alterations in histone modification patterns at individual genes and globally at the level of single nuclei in individual cells. We demonstrated previously that lower global/cellular levels of histone H3 lysine 4 dimethylation (H3K4me2) and H3K18 acetylation (ac) predict a higher risk of prostate cancer recurrence. Here we show that the cellular levels of both H3K4me2 and H3K18ac also predict clinical outcome in both lung and kidney cancer patients, with lower levels predicting significantly poorer survival probabilities in both cancer groups. We also show that lower cellular levels of H3K9me2, a modification associated with both gene activity and repression, is also prognostic of poorer outcome for individuals with either prostate or kidney cancers. The predictive power of these histone modifications was independent of tissue-specific clinicopathological variables, the proliferation marker Ki-67, or a p53 tumor suppressor mutation. Chromatin immunoprecipitation experiments indicated that the lower cellular levels of histone modifications in more aggressive cancer cell lines correlated with lower levels of modifications at DNA repetitive elements but not with gene promoters across the genome. Our results suggest that lower global levels of histone modifications are predictive of a more aggressive cancer phenotype, revealing a surprising commonality in prognostic epigenetic patterns of adenocarcinomas of different tissue origins.

BACKGROUND New biomarkers are needed to detect pleural mesothelioma at an earlier stage and to individualize treatment strategies. We investigated whether fibulin-3 in plasma and pleural effusions could meet sensitivity and specificity criteria for a robust biomarker. METHODS We measured fibulin-3 levels in plasma (from 92 patients with mesothelioma, 136 asbestos-exposed persons without cancer, 93 patients with effusions not due to mesothelioma, and 43 healthy controls), effusions (from 74 patients with mesothelioma, 39 with benign effusions, and 54 with malignant effusions not due to mesothelioma), or both. A blinded validation was subsequently performed. Tumor tissue was examined for fibulin-3 by immunohistochemical analysis, and levels of fibulin-3 in plasma and effusions were measured with an enzyme-linked immunosorbent assay. RESULTS Plasma fibulin-3 levels did not vary according to age, sex, duration of asbestos exposure, or degree of radiographic changes and were significantly higher in patients with pleural mesothelioma (105±7 ng per milliliter in the Detroit cohort and 113±8 ng per milliliter in the New York cohort) than in asbestos-exposed persons without mesothelioma (14±1 ng per milliliter and 24±1 ng per milliliter, respectively; P<0.001). Effusion fibulin-3 levels were significantly higher in patients with pleural mesothelioma (694±37 ng per milliliter in the Detroit cohort and 636±92 ng per milliliter in the New York cohort) than in patients with effusions not due to mesothelioma (212±25 and 151±23 ng per milliliter, respectively; P<0.001). Fibulin-3 preferentially stained tumor cells in 26 of 26 samples. In an overall comparison of patients with and those without mesothelioma, the receiver-operating-characteristic curve for plasma fibulin-3 levels had a sensitivity of 96.7% and a specificity of 95.5% at a cutoff value of 52.8 ng of fibulin-3 per milliliter. In a comparison of patients with early-stage mesothelioma with asbestos-exposed persons, the sensitivity was 100% and the specificity was 94.1% at a cutoff value of 46.0 ng of fibulin-3 per milliliter. Blinded validation revealed an area under the curve of 0.87 for plasma specimens from 96 asbestos-exposed persons as compared with 48 patients with mesothelioma. CONCLUSIONS Plasma fibulin-3 levels can distinguish healthy persons with exposure to asbestos from patients with mesothelioma. In conjunction with effusion fibulin-3 levels, plasma fibulin-3 levels can further differentiate mesothelioma effusions from other malignant and benign effusions. (Funded by the Early Detection Research Network, National Institutes of Health, and others.)

Elevated tumor cyclooxygenase-2 (COX-2) expression is associated with tumor invasion, metastasis, and poor prognosis in non-small cell lung cancer (NSCLC). Here, we report that COX-2-dependent pathways contribute to the modulation of E-cadherin expression in NSCLC. First, whereas genetically modified COX-2-sense (COX-2-S) NSCLC cells expressed low E-cadherin and showed diminished capacity for cellular aggregation, genetic or pharmacologic inhibition of tumor COX-2 led to increased E-cadherin expression and resulted in augmented homotypic cellular aggregation among NSCLC cells in vitro. An inverse relationship between COX-2 and E-cadherin was shown in situ by double immunohistochemical staining of human lung adenocarcinoma tissue sections. Second, treatment of NSCLC cells with exogenous prostaglandin E 2 (PGE 2 ) significantly decreased the expression of Ecadherin, whereas treatment of COX-2-S cells with celecoxib (1 Amol/L) led to increased E-cadherin expression. Third, the transcriptional suppressors of E-cadherin, ZEB1 and Snail, were up-regulated in COX-2-S cells or PGE 2 -treated NSCLC cells but decreased in COX-2-antisense cells. PGE 2 exposure led to enhanced ZEB1 and Snail binding at the chromatin level as determined by chromatin immunoprecipitation assays. Small interfering RNA-mediated knockdown of ZEB1 or Snail interrupted the capacity of PGE 2 to downregulate E-cadherin. Fourth, an inverse relationship between E-cadherin and ZEB1 and a direct relationship between COX-2 and ZEB1 were shown by immunohistochemical staining of human lung adenocarcinoma tissue sections. These findings indicate that PGE 2 , in autocrine or paracrine fashion, modulates transcriptional repressors of E-cadherin and thereby regulates COX-2-dependent E-cadherin expression in NSCLC. Thus, blocking PGE 2 production or activity may contribute to both prevention and treatment of NSCLC. (Cancer Res 2006; 66(10): 5338-45)

Lung cancer is the most common cause of cancer mortality in male and female patients in the US. The etiology of non-small cell lung cancer (NSCLC) is not fully defined, but new data suggest that estrogens and growth factors promote tumor progression. In this work, we confirm that estrogen receptors (ER), both ERalpha and ERbeta, occur in significant proportions of archival NSCLC specimens from the clinic, with receptor expression in tumor cell nuclei and in extranuclear sites. Further, ERalpha in tumor nuclei was present in activated forms as assessed by detection of ER phosphorylation at serines-118 and -167, residues commonly modulated by growth factor receptor as well as steroid signaling. In experiments using small interfering RNA (siRNA) constructs, we find that suppressing expression of either ERalpha or ERbeta elicits a significant reduction in NSCLC cell proliferation in vitro. Estrogen signaling in NSCLC cells may also include steroid receptor coactivators (SRC), as SRC-3 and MNAR/PELP1 are both expressed in several lung cell lines, and both EGF and estradiol elicit serine phosphorylation of SRC-3 in vitro. EGFR and ER also cooperate in promoting early activation of p42/p44 MAP kinase in NSCLC cells. To assess new strategies to block NSCLC growth, we used Faslodex alone and with erlotinib, an EGFR kinase inhibitor. The drug tandem elicited enhanced blockade of the growth of NSCLC xenografts in vivo, and antitumor activity exceeded that of either agent given alone. The potential for use of antiestrogens alone and with growth factor receptor antagonists is now being pursued further in clinical trials.

SUMMARY Monocarboxylate Transporter 1 (MCT1) inhibition is thought to block tumor growth through disruption of lactate transport and glycolysis. Here we show MCT1 inhibition impairs proliferation of glycolytic breast cancer cells co-expressing MCT1 and MCT4 via disruption of pyruvate rather than lactate export. MCT1 expression is elevated in glycolytic breast tumors, and high MCT1 expression predicts poor prognosis in breast and lung cancer patients. Acute MCT1 inhibition reduces pyruvate export but does not consistently alter lactate transport or glycolytic flux in breast cancer cells that co-express MCT1 and MCT4. Despite the lack of glycolysis impairment, MCT1 loss-of-function decreases breast cancer cell proliferation and blocks growth of mammary fat pad xenograft tumors. Our data suggest MCT1 expression is elevated in glycolytic cancers to promote pyruvate export, which when inhibited enhances oxidative metabolism and reduces proliferation. This study presents an alternative molecular consequence of MCT1 inhibitors further supporting their use as anti-cancer therapeutics.