BackgroundHistone deacetylase inhibitors (HDACis) are promising anticancer drugs; however, the molecular mechanisms leading to HDACi-induced cell death have not been well understood and no clear mechanism of resistance has been elucidated to explain limited efficacy of HDACis in clinical trials.Methods and FindingsHere, we show that protein levels of checkpoint kinase 1 (Chk1), which has a major role in G2 cell cycle checkpoint regulation, was markedly reduced at the protein and transcriptional levels in lung cancer cells treated with pan-and selective HDACis LBH589, scriptaid, valproic acid, apicidin, and MS-275. In HDACi treated cells Chk1 function was impaired as determined by decreased inhibitory phosphorylation of cdc25c and its downstream target cdc2 and increased expression of cdc25A and phosphorylated histone H3, a marker of mitotic entry. In time course experiments, Chk1 downregulation occurred after HDACi treatment, preceding apoptosis. Ectopic expression of Chk1 overcame HDACi-induced cell death, and pretreating cells with the cdc2 inhibitor purvalanol A blocked entry into mitosis and prevented cell death by HDACis. Finally, pharmacological inhibition of Chk1 showed strong synergistic effect with LBH589 in lung cancer cells.ConclusionsThese results define a pathway through which Chk1 inhibition can mediate HDACi-induced mitotic entry and cell death and suggest that Chk1 could be an early pharmacodynamic marker to assess HDACi efficacy in clinical samples.

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Purpose The goal of this study was to examine the therapeutic potential of the vascular endothelial growth factor (VEGF) signaling inhibitor cediranib in a human model of renal cell carcinoma (Caki-1). Methods and Materials The effects of cediranib treatment on in vitro endothelial cell function (proliferation, migration, and tube formation), as well as in vivo angiogenesis and tumor growth, were determined. Results In vitro, cediranib significantly impaired the proliferation and migration of endothelial cells and their ability to form tubes, but had no effect on the proliferation of Caki-1 tumor cells. In vivo, cediranib significantly reduced Caki-1 tumor cell–induced angiogenesis, reduced tumor perfusion, and inhibited the growth of Caki-1 tumor xenografts. Conclusions The present results are consistent with the notion that inhibition of VEGF signaling leads to an indirect (i.e., antiangiogenic) antitumor effect, rather than a direct effect on tumor cells. These results further suggest that inhibition of VEGF signaling with cediranib may impair the growth of renal cell carcinoma.

Lung cancer remains the leading cause of cancer-related death in the United States. The 5 year survival for stage I-IIIA lung cancer ranges from only 15-50%. For many in the early stages of the disease, especially stage II patients, adjuvant chemotherapy (ACT) following surgical resection is standard treatment. Still, it appears that the benefits for ACT have plateaued at 4%. With these marginal survival rates and the toxicities associated with chemotherapy, there is a need to develop a predictive tool to identify those patients with early-stage disease who would most benefit from receiving ACT. Previously, we developed an E2F-based gene expression signature derived from microarrays of two non-small cell lung cancer cell lines that were transfected with small interfering RNAs targeted against several members of the E2F pathway. This signature was then filtered for genes that were altered in cancer, but not in normal lung tissues, agreeing between two publicly-available lung tumor/adjacent normal datasets. Through principal component analysis (PCA), the E2F signature was shown to be prognostic in the Molecular Classification of Lung Adenocarcinoma (MCLA) from the Director's Challenge Consortium, as well as predictive of early-stage lung adenocarcinoma patients’ response to ACT in the JBR.10 clinical trial. In order to demonstrate efficacy of the signature in a typical clinical setting, we evaluated whether the NanoString platform (a method that involves the use of a novel “barcode” system to measure gene expression) would be as effective using RNA derived from formalin-fixed, paraffin-embedded (FFPE) tumor samples as compared to fresh frozen RNA. NanoString quantification of RNA from fresh frozen samples correlated very well to FFPE samples (average r=0.9070, median r=0.9290, SD=0.0859). Further, NanoString expression for 32 pairs of both the fresh frozen and FFPE RNA were compared to gene expression microarray readings of the genes in the signature. This analysis demonstrated that the NanoString readings corresponded well to microarray expression, regardless of the method of RNA extraction for NanoString (paired NanoString-frozen vs. microarray: average r=0.5585, median r=0.5608, SD=0.0718; paired NanoString-FFPE vs. microarray: average r=0.5384, median r=0.5401, SD=0.0891). Taken together, these results demonstrate that this E2F signature may be a useful tool for predicting which patients would best respond to adjuvant chemotherapy, and NanoString analysis may be a very effective means of applying this signature to patient samples. Citation Format: Courtney A. Kurtyka, Lu Chen, Matthew B. Schabath, Dung-Tsa Chen, William Brazelle, Eric A. Welsh, Anders E. Berglund, Steven A. Eschrich, Jhanelle E. Gray, Eric B. Haura, W. Douglas Cress. Development of a prognostic and predictive E2F signature in formalin-fixed, paraffin-embedded early-stage non-small cell lung cancer samples. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 914. doi:10.1158/1538-7445.AM2014-914

Early-stage non-small cell lung cancer (NSCLC) is primarily treated by surgical resection. Unfortunately, after resection, one-third to one-half of early-stage patients will die of metastatic recurrence. Adjuvant chemotherapy (ACT) improves the survival of patients with early-stage disease and has become the standard treatment for patients with resected stage II-III NSCLC. However, the five-year survival advantage of ACT is only 4%-15% suggesting that many patients do not benefit. Given the morbidity associated with ACT, it is imperative to develop new prognostic tools to identify those patients with a high probability of relapse. Toward this end, we have used small inhibitory RNAs targeting multiple E2F pathway components to derive an E2F gene expression signature in vitro. This signature was refined by filtering for its components that were altered in non-small cell lung cancers compared to normal tissue. Principle component analysis was then used to represent the signature which was tested for correlation to overall survival in two large cohorts. The first of the two cohorts was the Molecular Classification of Lung Adenocarcinoma (MCLA) from the Director's Challenge Consortium and the second was a novel database on 444 lung adenocarcinomas treated as a part of Moffitt's Total Cancer Care Network. The E2F signature is strongly prognostic in both cohorts with P values <0.0001. Additionally, using data in the JBR.10 trial (GSE14814) for patients who either did or did not receive ACT we were able to determine that patients having a high E2F signature benefit from ACT (have increased overall survival), whereas patients with a low E2F signature do not. Overall, these results suggest that this approach could be optimized in the clinical setting to distinguish patients likely to benefit from ACT from those who will not. Citation Format: Courtney A. Kurtyka, Dung-Tsa Chen, Lu Chen, William Brazelle, Eric A. Welsh, Anders E. Berglund, Steven A. Eschrich, Matthew B. Schabath, Eric B. Haura, W. Douglas Cress. An E2F signature predicts benefit of adjuvant chemotherapy in early-stage non-small cell lung cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2354. doi:10.1158/1538-7445.AM2013-2354