Calcitriol (1α, 25(OH)-Vitamin D3) binds to the vitamin D receptor (VDR) and regulates differentiation of the normal mammary gland, may therefore be useful in breast cancer treatment or ...prevention.Many breast cancer cells are, however, resistant to Calcitriol. In this study, we investigated the resistance mechanism and the role of epigenetic silencing of VDR by promoter hypermethylation.Bisulfite sequencing of the VDR promoter region revealed methylated CpG islands at -700 base pairs (bp) upstream and near the transcription start site.VDR CpG islands were demethylated by 5'deoxy-azacytidine treatment, and this was accompanied by a parallel increase in VDR mRNA levels in breast cancer cell lines. Quantitative methylation-specific PCR analyses confirmed hypermethylation of these CpG islands in primary tumors, and its absence in normal breast tissue.VDR transcripts detected in breast cancers were predominantly 5'-truncated, while normal breast tissue expressed full-length transcripts. Consistent with this observation, genes containing the VDR-responsive element (VDRE), such as cytochrome p450 hydroxylases,p21, or C/EBP were underexpressed in breast cancers compared to normal breast samples. Expression of the active longer transcripts of VDR was restored with 5'deoxy-Azacytidine (AZA) treatment, with a concurrent increase in expression of VDRE-containing genes. Thus, promoter methylation-mediated silencing of expression of the functional variants of VDR may contribute to reduced expression of downstream effectors of the VDR pathway and subsequent Calcitriol insensitivity in breast cancer. These data suggest that pharmacological reversal of VDR methylation may re-establish breast cancer cell susceptibility to differentiation therapy using Calcitriol.
Background
Methylated gene markers have shown promise in predicting breast cancer outcomes and treatment response. We evaluated whether baseline and changes in tissue and serum methylation levels ...would predict pathological complete response (pCR) in patients with HER2-negative early breast cancer undergoing preoperative chemotherapy.
Methods
The TBCRC008 trial investigated pCR following 12 weeks of preoperative carboplatin and albumin-bound paclitaxel + vorinostat/placebo (
n
= 62). We measured methylation of a 10-gene panel by quantitative multiplex methylation-specific polymerase chain reaction and expressed results as cumulative methylation index (CMI). We evaluated association between CMI level baseline, day 15 (D15), and change and pCR using univariate and multivariable logistic regression models controlling for treatment and hormone receptor (HR) status, and performed exploratory subgroup analyses.
Results
In univariate analysis, one log unit increase in tissue CMI levels at D15 was associated with 40% lower chance of obtaining pCR (odds ratio, OR 0.60, 95% CI 0.37–0.97;
p
= 0.037). Subgroup analyses suggested a significant association between tissue D15 CMI levels and pCR in vorinostat-treated OR 0.44 (0.20, 0.93),
p
= 0.03, but not placebo-treated patients.
Conclusion
In this study investigating the predictive roles of tissue and serum CMI levels in patients with early breast cancer for the first time, we demonstrate that high D15 tissue CMI levels may predict poor response. Larger studies and improved analytical procedures to detect methylated gene markers in early stage breast cancer are needed. TBCRC008 is registered on ClinicalTrials.gov (NCT00616967).
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OBVAL, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Purpose: Recent studies sought to refine lung cancer classification using gene expression microarrays. We evaluate the extent to which
these studies agree and whether results can be integrated.
...Experimental Design: We developed a practical analysis plan for cross-study comparison, validation, and integration of cancer molecular classification
studies using public data. We evaluated genes for cross-platform consistency of expression patterns, using integrative correlations,
which quantify cross-study reproducibility without relying on direct assimilation of expression measurements across platforms.
We then compared associations of gene expression levels to differential diagnosis of squamous cell carcinoma versus adenocarcinoma via reproducibility of the gene-specific t statistics and to survival via reproducibility of Cox coefficients.
Results: Integrative correlation analysis revealed a large proportion of genes in which the patterns agreed across studies more than
would be expected by chance. Correlation of t statistics for diagnosis of squamous cell carcinoma versus adenocarcinoma is high (0.85) and increases (0.925) when using only the most consistent genes identified by integrative correlation.
Correlations of Cox coefficients ranged from 0.13 to 0.31 (0.33–0.49 with genes selected for consistency). Although we find
genes that are significant in multiple studies but show discordant effects, their number is approximately that expected by
chance. We report genes that are reproducible by integrative analysis, significant in all studies, and concordant in effect.
Conclusions: Cross-study comparison revealed significant, albeit incomplete, agreement of gene expression patterns related to lung cancer
biology and identified genes that reproducibly predict outcomes. This analysis approach is broadly applicable to cross-study
comparisons of gene expression profiling projects.
Purpose: Fatty acid synthase (FAS) is overexpressed in lung cancer, and we have investigated the potential use of FAS inhibitors for
chemoprevention of lung cancer.
Experimental Design: Expression of ...FAS was evaluated in preinvasive human lung lesions (bronchial squamous dysplasia and atypical adenomatous
hyperplasia) and in murine models of lung tumorigenesis 4-(methylnitrosamino)-I-(3-pyridyl)-1-butanone–induced and urethane-induced
lung tumors in A/J mice. Then, the ability of pharmacologic inhibitors of FAS to prevent development of the murine tumors
was investigated. Finally, the effect of the FAS inhibitor treatment of levels of phosphorylated Akt in the murine tumors
was evaluated by immunohistochemistry.
Results: Immunohistochemical studies show that human bronchial dysplasia and atypical adenomatous hyperplasia express high levels
of FAS compared with normal lung tissues, suggesting that FAS might be a target for intervention in lung carcinogenesis. FAS
is also expressed at high levels in chemically induced murine lung tumors, and the numbers and sizes of those murine tumors
are significantly reduced by treating carcinogen-exposed mice with pharmacologic inhibitors of FAS, C75 and C93. C93 treatment
is associated with reduced levels of phosphorylated Akt in tumor tissues, suggesting that inhibition of this signal transduction
pathway might be involved in the chemopreventative activity of this compound.
Conclusions: We conclude that increased levels of FAS are common in human preinvasive neoplasia of the lung. Based on studies in mouse
models, it seems that inhibiting FAS is an effective strategy in preventing and retarding growth of lung tumors that have
high expression of this enzyme.
Background:
Bronchoalveolar lavage (BAL) is a specific type of air-way fluid. It is a commonly used clinical specimen for the diagnosis of benign diseases and cancers of the lung. Although previous ...studies have identified several disease-associated proteins in the BAL, the potential utility of BAL in lung cancer is still not well-studied. Based upon the fact that the majority of secreted proteins are glycoproteins, we have profiled
N
-glycoproteins in BAL collected from lung cancers, and investigated the expression of glycoproteins such as the matrix
N-
glycoprotein, periostin, in lung cancers.
Methods:
BAL specimens (
n
= 16) were collected from lung cancer patients, and analyzed using mass spectrometry-based quantitative
N
-glycoproteomic technique. Additional BAL specimens (
n
= 39) were independently collected to further evaluate the expression of periostin by using an enzyme-linked immunosorbent assay (ELISA).
Results:
A total of 462 glycoproteins were identified in BAL samples using
N
-glycoproteomic technique, including 290 in lung adenocarcinoma (ADC,
n
= 5), 376 in squamous cell carcinoma (SQCC,
n
= 4), 309 in small cell lung carcinoma (SCLC,
n
= 4), and 316 in benign lung disease (
n
= 3). The expressions of several glycoproteins were elevated, including 8 in ADC, 12 in SQCC, and 17 in SCLC, compared to benign BALs. The expression of periostin was detected in all subtypes of lung cancers. To further investigate the expression of periostin, an ELISA assay was performed using additional independently collected BALs (
n
= 39) The normalized levels of periostin in benign disease, ADC, SQCC, and SCLC were 255 ± 104 (mean ± SE) and 4,002 ± 2,181, 3,496 ± 1,765, and 1,772 ± 1,119 ng/mg of total BAL proteins.
Conclusion:
Our findings demonstrate that proteomic analysis of BAL can be used for the study of cancer-associated extracellular proteins in air-way fluid from lung cancer patients.
Cancer cells hijack autophagy pathway to evade anti-cancer therapeutics. Many molecular signaling pathways associated with drug-resistance converge on autophagy induction. Honokiol (HNK), a natural ...phenolic compound purified from
, has recently been shown to impede breast tumorigenesis and, in the present study, we investigated whether breast cancer cells evoke autophagy to modulate therapeutic efficacy and functional networks of HNK. Indeed, breast cancer cells exhibit increased autophagosomes-accumulation, MAP1LC3B-II/LC3B-II-conversion, expression of ATG proteins as well as elevated fusion of autophagosomes and lysosomes upon HNK treatment. Breast cancer cells treated with HNK demonstrate significant growth inhibition and apoptotic induction, and these biological processes are blunted by macroautophagy/autophagy. Consequently, inhibiting autophagosome formation, abrogating autophagosome-lysosome fusion or genetic-knockout of
and
effectively increase HNK-mediated apoptotic induction and growth inhibition. Next, we explored the functional impact of tumor suppressor STK11 in autophagy induction in HNK-treated cells. STK11-silencing abrogates LC3B-II-conversion, and blocks autophagosome/lysosome fusion and lysosomal activity as illustrated by LC3B-Rab7 co-staining and DQ-BSA assay. Our results exemplify the cytoprotective nature of autophagy invoked in HNK-treated breast cancer cells and put forth the notion that a combined strategy of autophagy inhibition with HNK would be more effective. Indeed, HNK and chloroquine (CQ) show synergistic inhibition of breast cancer cells and HNK-CQ combination treatment effectively inhibits breast tumorigenesis and metastatic progression. Tumor-dissociated cells from HNK-CQ treated tumors exhibit abrogated invasion and migration potential. Together, these results implicate that breast cancer cells undergo cytoprotective autophagy to circumvent HNK and a combined treatment with HNK and CQ can be a promising therapeutic strategy for breast cancer.
p16 and p53 are frequently altered intracellular pathways in cancers. We investigated the aberrant expression of p16 and its relationship with p53 and HPV status in primary non-small-cell lung ...carcinoma.
Lung tumor tissue microarray (n = 163), immunohistochemical study of p16 and p53, and HPV
hybridization were analyzed.
p16 and p53 were detected in 50.7 and 57.3% of adenocarcinoma (ADCs; n = 75), and 35.2 and 63.6% of squamous cell carcinoma (n = 88). HPV was detected in 16 and 10.2% of ADC and squamous cell carcinoma. In ADCs, p16 positive tumors demonstrated a favorable median overall survival time of 60.9 months, compared with p16 negative tumors of 46.9 months (p < 0.05). Furthermore, we did not find significant relationships between p16 expression and HPV status, nor with p53 expression.
p16 play an unique role in lung cancer survival. The mechanism of p16 needs to be further studied.
Rapid advances in generating new mouse genetic models for lung neoplasia provide a continuous challenge for pathologists and investigators. Frequently, phenotypes of new models either have no ...precedents or are arbitrarily attributed according to incongruent human and mouse classifications. Thus, comparative characterization and validation of novel models can be difficult. To address these issues, a series of discussions was initiated by a panel of human, veterinary, and experimental pathologists during the Mouse Models of Human Cancers Consortium (NIH/National Cancer Institute) workshop on mouse models of lung cancer held in Boston on June 20-22, 2001. The panel performed a comparative evaluation of 78 cases of mouse and human lung proliferative lesions, and recommended development of a new practical classification scheme that would (a) allow easier comparison between human and mouse lung neoplasms, (b) accommodate newly emerging mouse neoplasms, and (c) address the interpretation of benign and preinvasive lesions of the mouse lung. Subsequent discussions with additional experts in pulmonary pathology resulted in the current proposal of a new classification. It is anticipated that this classification, as well as the complementary digital atlas of virtual histological slides, will help investigators and pathologists in their characterization of new mouse models, as well as stimulate further research aimed at a better understanding of proliferative lesions of the lung.
Purpose: Fatty acid synthase (FAS) is overexpressed in many human cancers and is considered to be a promising target for therapy.
However, in vitro use of previous generations of FAS inhibitors has ...been limited by severe, but reversible, anorexia in treated animals, which
is thought to be related to a parallel stimulation of fatty acid oxidation by these agents. This study investigated pharmacologic
inhibition of FAS using C93, a rationally designed molecule that inhibits FAS activity without affecting fatty acid oxidation
in preclinical models of lung cancer.
Experimental Design: Activity of C93 on FAS and fatty acid oxidation was evaluated in cultured non–small cell lung cancer (NSCLC) cells. Antineoplastic
activity of the compound, given orally or by i.p. injection, was evaluated in s.c. and orthotopic NSCLC xenografts.
Results: Our experiments confirm that C93 effectively inhibits FAS without stimulating fatty acid oxidation in lung cancer cells.
More importantly, C93 significantly inhibits the growth of both s.c. and orthotopic xenograft tumors from human NSCLC cell
lines without causing anorexia and weight loss in the treated animals.
Conclusions: We conclude that inhibition of FAS can be achieved without parallel stimulation of fatty acid oxidation and that inhibition
of tumor growth in vivo can be achieved without anorexia and weight loss. Thus, this therapeutic strategy holds promise for clinical treatment of
cancers, including non–small cell lung cancer, the leading cause of cancer mortality in the United States and Europe.
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