Enhancing the anti-angiogenic action of histone deacetylase inhibitors

<p>Abstract</p> <p>Background</p> <p>Histone deacetylase inhibitors (HDACIs) have many effects on cancer cells, such as growth inhibition, induction of cell death, differentiation, and anti-angiogenesis, all with a wide therapeutic index. However, clinical trials demonstrate that HDACIs are more likely to be effective when used in combination with other anticancer agents. Moreover, the molecular basis for the anti-cancer action of HDACIs is still unknown. In this study, we compared different combinations of HDACIs and anti-cancer agents with anti-angiogenic effects, and analysed their mechanism of action.</p> <p>Results</p> <p>Trichostatin A (TSA) and α-interferon (IFNα) were the most effective combination across a range of different cancer cell lines, while normal non-malignant cells did not respond in the same manner to the combination therapy. There was a close correlation between absence of basal p21^WAF1expression and response to TSA and IFNα treatment. Moreover, inhibition of p21^WAF1expression in a p21^WAF1-expressing breast cancer cell line by a specific siRNA increased the cytotoxic effects of TSA and IFNα. <it>In vitro </it>assays of endothelial cell function showed that TSA and IFNα decreased endothelial cell migration, invasion, and capillary tubule formation, without affecting endothelial cell viability. TSA and IFNα co-operatively inhibited gene expression of some pro-angiogenic factors: vascular endothelial growth factor, hypoxia-inducible factor 1α and matrix metalloproteinase 9, in neuroblastoma cells under hypoxic conditions. Combination TSA and IFNα therapy markedly reduced tumour angiogenesis in neuroblastoma-bearing transgenic mice.</p> <p>Conclusion</p> <p>Our results indicate that combination TSA and IFNα therapy has potent co-operative cytotoxic and anti-angiogenic activity. High basal p21^WAF1expression appears to be acting as a resistance factor to the combination therapy.</p

ROR1 and ROR2 expression in pancreatic cancer

Background: The Wnt receptors ROR1 and ROR2 are generating increased interest as cancer therapeutic targets but remain understudied in pancreatic ductal adenocarcinoma (PDAC). Compared to canonical Wnt/ β-catenin signalling, the role of noncanonical Wnt signalling in PDAC remains largely unknown. Only one study has investigated the prognostic significance of the noncanonical Wnt signalling receptor, ROR2 in PDAC. No studies have investigated the prognostic role of ROR1 in PDAC. Methods: Here, we performed analysis of ROR1 and ROR2 mRNA expression in three publicly available datasets ICGC-PACA-AU (n = 81), TCGA-PAAD (n = 150) and CPTAC-PDAC (n = 137). ROR1 and ROR2 protein expression from the CPTAC-PDAC discovery cohort were also analysed. Immunohistochemistry (IHC) using the validated anti ROR1 monoclonal antibody (4A5) was performed on the Australian Pancreatic Cancer Genome Initiative (APGI) cohort of PDAC samples (n = 152). Association between ROR1 cytoplasmic staining intensity and clinicopathological parameters including stage, grade and overall survival (OS) was investigated. Results: High ROR1 mRNA expression levels correlated with a favourable OS outcome in all of the ICGC-PACA-AU, TCGA-PAAD and CPTAC-PDAC cohorts. ROR1 protein expression was not associated with stage, grade or OS in the APGI cohort. Conclusion: ROR1 and ROR2 have potential as prognostic markers when measured at the mRNA level in PDAC. Our IHC cohort did not support ROR1 protein expression in predicting OS, and highlighted the discrepancy of prognostic biomarkers when measured by MS, IHC and RNAseq

Genomic and molecular analyses identify molecular subtypes of pancreatic cancer recurrence

Pancreatic cancer (PC) remains a highly lethal malignancy, and most patients with localized disease that undergo surgical resection still succumb to recurrent disease. Pattern of recurrence after pancreatectomy is heterogenous, with some studies illustrating that site of recurrence can be associated with prognosis.1 Another study suggested that tumors that develop local and distant recurrence can be regarded as a homogenous disease with similar outcomes.2 Here we investigate novel molecular determinants of recurrence pattern after pancreatectomy for PC

DNA methylation patterns identify subgroups of pancreatic neuroendocrine tumors with clinical association

Here we report the DNA methylation profile of 84 sporadic pancreatic neuroendocrine tumors (PanNETs) with associated clinical and genomic information. We identified three subgroups of PanNETs, termed T1, T2 and T3, with distinct patterns of methylation. The T1 subgroup was enriched for functional tumors and ATRX, DAXX and MEN1 wild-type genotypes. The T2 subgroup contained tumors with mutations in ATRX, DAXX and MEN1 and recurrent patterns of chromosomal losses in half of the genome with no association between regions with recurrent loss and methylation levels. T2 tumors were larger and had lower methylation in the MGMT gene body, which showed positive correlation with gene expression. The T3 subgroup harboured mutations in MEN1 with recurrent loss of chromosome 11, was enriched for grade G1 tumors and showed histological parameters associated with better prognosis. Our results suggest a role for methylation in both driving tumorigenesis and potentially stratifying prognosis in PanNETs

DNA methylation patterns identify subgroups of pancreatic neuroendocrine tumors with clinical association

Here we report the DNA methylation profile of 84 sporadic pancreatic neuroendocrine tumors (PanNETs) with associated clinical and genomic information. We identified three subgroups of PanNETs, termed T1, T2 and T3, with distinct patterns of methylation. The T1 subgroup was enriched for functional tumors and ATRX, DAXX and MEN1 wild-type genotypes. The T2 subgroup contained tumors with mutations in ATRX, DAXX and MEN1 and recurrent patterns of chromosomal losses in half of the genome with no association between regions with recurrent loss and methylation levels. T2 tumors were larger and had lower methylation in the MGMT gene body, which showed positive correlation with gene expression. The T3 subgroup harboured mutations in MEN1 with recurrent loss of chromosome 11, was enriched for grade G1 tumors and showed histological parameters associated with better prognosis. Our results suggest a role for methylation in both driving tumorigenesis and potentially stratifying prognosis in PanNETs

SAHA and IFNα exerted co-operative cytotoxic effects in cancer cell lines, but not in normal non-malignant cells

<p><b>Copyright information:</b></p><p>Taken from "Enhancing the anti-angiogenic action of histone deacetylase inhibitors"</p><p>http://www.molecular-cancer.com/content/6/1/68</p><p>Molecular Cancer 2007;6():68-68.</p><p>Published online 25 Oct 2007</p><p>PMCID:PMC2173905.</p><p></p> Neuroblastoma BE(2)-C, breast cancer MCF-7, and normal non-malignant lung MRC-5 fibroblasts were treated with control, 0.5 μM SAHA and/or 500 IU/ml IFNα for 72 hours. Cell viability was examined using the Alamar blue assay, measured as optical density (OD) units of absorbance, and expressed as the absorbance of treated over control samples (ie., % viable cells). * p < 0.05, ** p < 0.01, *** p < 0.001

TSA and IFNα co-operatively suppress tumour-driven angiogenesis in neuroblastoma bearing transgenic MYCN mice

<p><b>Copyright information:</b></p><p>Taken from "Enhancing the anti-angiogenic action of histone deacetylase inhibitors"</p><p>http://www.molecular-cancer.com/content/6/1/68</p><p>Molecular Cancer 2007;6():68-68.</p><p>Published online 25 Oct 2007</p><p>PMCID:PMC2173905.</p><p></p> A. Photomicrographs of neuroblastoma tumour tissue sections from homozygous MYCN transgenic mice treated with either control, TSA, IFNα, or TSA and IFNα, which were subject to immunohistochemical studies using an anti-PECAM-1 antibody. Arrows indicate PECAM-1 positive microvessels (brown colored). B. Quantitation of the number of PECAM positive microvessels per 40× high power field in neuroblastoma tumour cross-sections. *** p < 0.001

TSA and IFNα exerted co-operative cytotoxic effects in cancer cell lines from a range of different tissue origins, but not in normal non-malignant cells

<p><b>Copyright information:</b></p><p>Taken from "Enhancing the anti-angiogenic action of histone deacetylase inhibitors"</p><p>http://www.molecular-cancer.com/content/6/1/68</p><p>Molecular Cancer 2007;6():68-68.</p><p>Published online 25 Oct 2007</p><p>PMCID:PMC2173905.</p><p></p> Neuroblastoma [BE(2)-C], breast (MCF-7 and MDA-MB-468), lung (H460 and Calu-6), prostate (DU-145 and LNCaP), and colon (HT-29 and Caco-2) cancer cells were treated with control (Cont), 0.02 μM TSA and/or 500 IU/ml IFNα for 72 hours. Cell viability was examined using the Alamar blue assay, measured as optical density (OD) units of absorbance, and expressed as the absorbance of treated over control samples (ie., % viable cells). ** p < 0.01, *** p < 0.001. B. MRC-5 cells were treated with control, 0.02 μM TSA and/or 500 IU/ml IFNα for 72 hours, and cell viability was assessed as above. Moreover, histone protein was extracted and subject to immunoblot analysis with anti-acetylated histone H3 antibody, after 6 hour exposure to control, TSA and/or IFNα

Absence of p21expression correlated with sensitivity to TSA and IFNα combination therapy

<p><b>Copyright information:</b></p><p>Taken from "Enhancing the anti-angiogenic action of histone deacetylase inhibitors"</p><p>http://www.molecular-cancer.com/content/6/1/68</p><p>Molecular Cancer 2007;6():68-68.</p><p>Published online 25 Oct 2007</p><p>PMCID:PMC2173905.</p><p></p> A. MCF-7, MDA-MB-468, H460, Calu-6, DU-145, LNCaP, HT-29 and Caco-2 cells were treated with control, 0.02 μM TSA, 500 IU/ml IFNα, or TSA and IFNα for 24 hours. Whole cell protein was extracted and subjected to immunoblot with an anti- p21antibody, and, an anti-actin antibody as a loading control. B. MCF-7 cells were transfected with control scrambled or p21siRNA for 8 hours, followed by treatment with control, 0.02 μM TSA and/or 500 IU/ml IFNα for 72 hours. The effect of the siRNAs on p21gene and protein expression was analysed by semi-quantitative RT-PCR with the house-keeping gene β-2-microglobulin (β2M) as a loading control or by immunoblot, with actin as a loading control. C. Cell viability was examined by the Alamar blue assay, measured as optical density (OD) units of absorbance, and expressed as percentage of absorbance for drug-treated samples over control-treated samples (% viable cells). ** p < 0.01