Molecular Oncology - Articles in Press

Heterogeneity among RIP-Tag2 insulinomas allows vascular endothelial growth factor-A independent tumor expansion as revealed by studies in Shb mutant mice: Implications for tumor angiogenesis - Uncorrected Proof

2012-02-06

Abstract: The Shb adapter protein is a signaling intermediate that operates downstream of vascular endothelial growth factor receptor-2 (VEGFR-2) in endothelial cells. The Shb knockout mouse displays a dysfunctional microvasculature and impaired growth of subcutaneously implanted tumor cells. We decided to investigate tumor growth and angiogenesis in the absence of Shb in an inheritable tumor model, the RIP-Tag2 mouse, which produces insulinomas in a manner highly dependent on de novo angiogenesis. We observed a reduced tumor incidence and burden in both RIP-Tag2 Shb−/− and RIP-Tag2 Shb+/− mice. This correlated with a reduced microvascular density, measured as a percentage of insulinoma area positive for CD31 staining, and altered vascular morphology. However, treatment with a VEGF-A blocking antibody was without effect on the Shb mutant tumor volume whereas it significantly inhibited tumor volume in the wild-type mice, suggesting that in mice with reduced Shb expression tumor angiogenesis was primarily sustained by VEGF-A independent pathway(s). This notion was further substantiated by gene expression analysis of angiogenic markers showing reduced VEGF-A expression in Shb-deficient tumors. Considerable heterogeneity with respect to the gene expression profiles of other angiogenic markers and the signal-transduction characteristics was observed between different tumors, suggesting that multiple “rescue” pathways could be operating. The numbers of invasive tumors or metastases were unchanged in the Shb mutant.It is concluded that the Shb mutant background reduces tumor frequency by chronically suppressing VEGF-A dependent angiogenesis. However, VEGF-A independent angiogenesis supports a significant degree of tumor expansion in Shb-deficient mice, indicating heterogeneity in the mechanisms by which tumor expansion is promoted. Interference with Shb signaling may provide novel means for future cancer therapy.Highlights: ► Shb is an adapter protein operating downstream of VEGFR-2. ► Shb was presently investigated in the context of RIP-Tag2 angiogenesis. ► Absence of Shb reduces RIP-Tag2 tumor burden and VEGF-dependent tumor angiogenesis. ► A fraction of tumors escape the angiogenic inhibition imposed by the absence of Shb. ► Tumor heterogeneity allows escape from angiogenic restriction by various means.

Inhibition of NEDD8-conjugation pathway by novel molecules: Potential approaches to anticancer therapy - Uncorrected Proof

Tomoaki Tanaka, Tatsuya Nakatani, Tetsu Kamitani — 2012-01-27

Abstract: Cancer cells can survive through the upregulation of cell cycle and the escape from apoptosis induced by numerous cellular stresses. In the normal cells, these biological cascades depend on scheduled proteolytic degradation of regulatory proteins via the ubiquitin–proteasome pathway. Therefore, interruption of regulated proteolytic pathways leads to abnormal cell-proliferation. Ubiquitin ligases called SCF complex (consisting of Skp-1, cullin, and F-box protein) or CRL (cullin-RING ubiquitin ligase) are predominant in a family of E3 ubiquitin ligases that control a final step in ubiquitination of diverse substrates. To a great extent, the ubiquitin ligase activity of the SCF complex requires the conjugation of NEDD8 to cullins, i.e. scaffold proteins. This review is anticipated to review the downregulation system of NEDD8 conjugation by several factors including a chemical compound such as MLN4924 and protein molecules (e.g. COP9 signalosome, inactive mutant of Ubc12, and NUB1/NUB1L). Since the downregulation of NEDD8 conjugation affects cell-cycle progression by inhibiting the ligase activity of SCF complexes, such knowledge in the NEDD8-conjugation pathway will contribute to the more magnificent therapies that selectively suppress tumorigenesis.Highlights: ► We have provided recent insights in the NEDD8-associated researches. ► We have revealed NEDD8-associated molecules regulating the SCF ubiquitin E3 ligases. ► Deneddylation-related proteins are candidates in inhibiting the SCF ligase. ► MLN4924 is a compound involved in initial inhibition of the neddylation cascade.

The toxin component of targeted anti-tumor toxins determines their efficacy increase by saponins - Uncorrected Proof

2012-01-27

Abstract: Tumor-targeting protein toxins are composed of a toxin enzyme coupled to a specific cell binding domain that targets cancer-associated antigens. The anti-tumor treatment by targeted toxins is accompanied by dose-limiting side effects. The future prospects of targeted toxins for therapeutic use in humans will be determined by reduce side effects. Certain plant secondary metabolites (saponins) were shown to increase the efficacy of a particular epidermal growth factor receptor (EGFR)-targeted toxin, paralleled by a tremendous decrease of side effects.This study was conducted in order to investigate the effects of substituting different toxin moieties fused to an EGF ligand binding domain on the augmentative ability of saponins for each against therapeutic potential of the saponin-mediated efficacy increase for different anti-tumor toxins targeting the EGFR.We designed several EGFR-targeted toxins varying in the toxic moiety. Each targeted toxin was used in combination with a purified saponin (SA1641), isolated from the ornamental plant Gypsophila paniculata L. SA1641 was characterized and the SA1641-mediated efficacy increase was investigated on EGFR-transfected NIH-3T3 cells.We observed a high dependency of the SA1641-mediated efficacy increase on the nature of toxin used for the construction of the targeted toxin, indicating high specificity.Structural alignments revealed a high homology between saporin and dianthin-30, the two toxic moieties that benefit most from the combination with SA1641.We further demonstrate that SA1641 did not influence the plasma membrane permeability, indicating an intracellular interaction of SA1641 and the toxin components of targeted toxins. Surface plasmon resonance measurements point to a transient binding of SA1641 to the toxin components of targeted toxins.Graphical abstract: Highlights: ► Combination therapy. ► Targeted toxins. ► Efficacy increase by saponins. ► Binding of saponins to toxins.

Germline pharmacogenomics in oncology: Decoding the patient for targeting therapy - Uncorrected Proof

Peter H. O'Donnell, Mark J. Ratain — 2012-01-25

Abstract: Pharmacogenomics is the study of genetic factors determining drug response or toxicity. The use of pharmacogenomics is especially desirable in oncology because the therapeutic index of oncology drugs is often narrow, the need for favorable drug response is often acute, and the consequences of drug toxicity can be life-threatening. In this review, we examine the state of pharmacogenomics in oncology, focusing only on germline pharmacogenomic variants. We consider several critical points when assessing the quality of pharmacogenomic findings and their relevance to clinical use, and discuss potential confounding factors limiting interpretation and implementation. Several of the most extensively studied drug–gene pairs (irinotecan and UGT1A1; tamoxifen and CYP2D6; 5-fluorouracil and DPYD) are inspected in depth as illustrations of both the state of advancement—and the current limitations of—present knowledge. We argue that there will likely soon be a critical mass of important germline pharmacogenomic biomarkers in oncology which deserve clinical implementation to provide optimal, personalized oncologic care. We conclude with a vision of how routine clinical testing of such germline markers could one day change the paradigm for cancer care.

Structural and genic characterization of stable genomic regions in breast cancer: Relevance to chemotherapy - Uncorrected Proof

Nicole I. Park, Peter K. Rogan, Heather E. Tarnowski, Joan H.M. Knoll — 2012-01-24

Abstract: Background: Cancer genomes accumulate frequent and diverse chromosomal abnormalities as well as gene mutations but must maintain the ability to survive in vivo. We hypothesize that genetic selection acts to maintain tumour survival by preserving copy number of specific genes and genomic regions. Genomic regions and genes that remain unaltered in copy number and expression, respectively, may be essential for maintaining tumour survival.Methods: We analyzed copy number data of 243 previously reported breast tumours and computationally derived stable copy number regions. To identify genes in stable copy number regions with nominal changes in expression, datasets for tumour and normal samples were compared. Results were replicated by analysis of a series of independent copy number, expression and genomic sequencing studies. A subset of stable regions, including stable paralogous regions, were confirmed by quantitative PCR and fluorescence in situ hybridization (FISH) in 5 breast cancer cell lines. We deduced a comprehensive set of dually stable genes (i.e. maintaining nominal copy number and expression) which were categorized according to pathway and ontology assignments. The stability of genes encoding therapeutic drug targets was also assessed.Results and Conclusion: Tumour genome analysis revealed 766 unstable (amplified and/or deleted) and 812 stable contiguous genomic regions. Replication analysis of an independent set of 171 breast tumours confirmed copy number stability of 1.3 Gb of the genome. We found that 5804 of these genes were dually stable. The composition of this gene set remained essentially unchanged (<2% reduction) after accounting for commonly mutated breast cancer genes found by sequencing and differential expression. The stable breast cancer genome is enriched for cellular metabolism, regulation of gene expression, DNA packaging (chromatin and nucleosome assembly), and regulation of apoptosis functions. Stable genes participating in multiple essential pathways were consistently found to be targets of chemotherapies. Preservation of stable, essential genes may be related to the effectiveness of certain chemotherapeutic agents that act on multiple gene products in this set.Graphical abstract: Highlights: ► A portion of the genome is stable in a large number of breast tumours. ► Breast cancer genome contains stable paralogous gene-rich sequence families. ► Genes with stable copy number and expression are consistent between independent studies. ► Stable genes define essential biochemical pathways/functions in tumours. ► Stable genes encode targets of systemic breast cancer chemotherapies.

Cancer immunotherapy - Corrected Proof

Thomas F. Gajewski — 2012-01-13

Abstract: The remarkable specificity of the immune system through antigen recognition has long attracted investigators to the possibility of immune-based therapy for cancer. Previous cancer immunotherapeutics had been restricted to non-specific immunomodulatory agents, such as the cytokines IL-2 or IFN-α. However, the molecular definition of cancer-associated antigens introduced the possibility of specific vaccines and adoptive T cell approaches aiming to target the tumor cells more specifically. The recent introduction of total exome sequencing has enabled the identification of patient tumor-specific epitopes generated through somatic point mutations, raising the possibility of targeting tumor antigens in individual patients which are even more tumor-specific. Transcriptional profiling and immunohistochemistry analyses have revealed a subset of patients with a pre-existing T cell-inflamed tumor microenvironment. This phenotype may be predictive of clinical outcome to immunotherapies and offers the possibility of a predictive biomarker. Further analysis of these tumors has identified a set of defined immune suppressive factors which themselves are being targeted with new immunotherapeutics, already with interesting early phase clinical trial results. Understanding not only the expression of tumor antigens but also the dynamic between a growing tumor and the host immune response is thus generating a rich set of opportunities for the specific immunotherapy of cancer.

Adseverin: A novel cisplatin-resistant marker in the human bladder cancer cell line HT1376 identified by quantitative proteomic analysis - Corrected Proof

2012-01-12

Abstract: Cisplatin is currently the most effective antitumor agent available against bladder cancer. However, a majority of patients eventually relapse with cisplatin-resistant disease. Chemoresistance thus remains a major obstacle in bladder cancer therapy. To clarify the molecular mechanisms underlying cisplatin resistance in bladder cancer, we established a cisplatin-resistant subline from the human bladder cancer cell line HT1376 (HT1376-CisR), and conducted large-scale analyses of the expressed proteins using two-dimensional (2D) gel electrophoresis coupled with mass spectrometry (MS). Comparative proteomic analysis of HT1376 and HT1376-CisR cells revealed 36 differentially expressed proteins, wherein 21 proteins were upregulated and 15 were downregulated in HT1376-CisR cells. Among the differentially regulated proteins, adseverin (SCIN), a calcium-dependent actin-binding protein, was overexpressed (4-fold upregulation) in HT1376-CisR, with the increase being more prominent in the mitochondrial fraction than in the cytosol fraction. SCIN mRNA knockdown significantly reduced cell proliferation with mitochondria-mediated apoptosis in HT1376-CisR cells. Immunoprecipitation analysis revealed voltage-dependent anion channels (VDACs) to be bound to SCIN in the mitochondrial fraction. Our results suggest that the VDAC-SCIN interaction may inhibit mitochondria-mediated apoptosis in cisplatin-resistant cells. Targeting the VDAC-SCIN interaction may offer a new therapeutic strategy for cisplatin-resistant bladder cancer.Highlights: ► We established a cisplatin-resistant subline (HT1376-CisR). ► We used quantitative proteomic analysis to compare between HT1376-CisR and HT1376. ► Adseverin, an actin-binding protein, was overexpressed in HT1376-CisR. ► It affected behaviors of voltage-dependent anion channel, interfering with apoptosis. ► Targeting the VDAC-adseverin interaction may offer a new therapeutic strategy.

The interaction of PKN3 with RhoC promotes malignant growth - Corrected Proof

2012-01-03

Abstract: PKN3 is an AGC-family protein kinase implicated in growth of metastatic prostate cancer cells with phosphoinositide 3-kinase pathway deregulation. The molecular mechanism, however, by which PKN3 contributes to malignant growth and tumorigenesis is not well understood. Using orthotopic mouse tumor models, we now show that inducible knockdown of PKN3 protein not only blocks metastasis, but also impairs primary prostate and breast tumor growth. Correspondingly, overexpression of exogenous PKN3 in breast cancer cells further increases their malignant behavior and invasiveness in-vitro. Mechanistically, we demonstrate that PKN3 physically interacts with Rho-family GTPases, and preferentially with RhoC, a known mediator of tumor invasion and metastasis in epithelial cancers. Likewise, RhoC predominantly associates with PKN3 compared to its closely related PKN family members. Unlike the majority of Rho GTPases and PKN molecules, which are ubiquitously expressed, both PKN3 and RhoC show limited expression in normal tissues and become upregulated in late-stage malignancies. Since PKN3 catalytic activity is increased in the presence of Rho GTPases, the co-expression and preferential interaction of PKN3 and RhoC in tumor cells are functionally relevant. Our findings provide novel insight into the regulation and function of PKN3 and suggest that the PKN3–RhoC complex represents an attractive therapeutic target in late-stage malignancies.Highlights: ► Inducible PKN3 knockdown inhibits primary prostate and breast tumor growth. ► PKN3 associates with Rho-family GTPases, and preferentially with RhoC. ► RhoC predominantly associates with PKN3 compared to other PKN family members. ► PKN3 catalytic activity is stimulated in the presence of RhoA and RhoC. ► Turn-motif phosphorylation at T860 closely correlates with PKN3 activity.

Serous ovarian carcinoma patients with high alpha-folate receptor had reducing survival and cytotoxic chemo-response - Corrected Proof

2011-12-29

Abstract: The alpha-folate receptor (α-FR) is highly-expressed in various non-mucinous tumors of epithelial origin, including ovarian carcinoma. The aim of this study was to investigate the relationship between alpha-folate receptor (α-FR) and the clinico-pathologic features and outcomes of serous ovarian carcinoma patients and the possible mechanism of α-FR to chemo-resistance. Therefore, semi-quantitative reverse-transcription polymerase chain reactions for α-FR expression were performed in the 91 specimens of serous ovarian carcinomas. The expression of α-FR in each ovarian cancer tissue specimen was defined as the ratio of density of α-FR to density of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In vitro apoptotic experiments were tested in the original OVCAR-3 tumor cells and various OVCAR-3 α-FR-transfectants. Patients with an increased α-FR expression level had poorer responses to chemotherapy (per α-FR expression level increase: odds ratio (OR): 8.97 (95% confidence interval (CI): 1.40–57.36), p = 0.021). An increased α-FR expression level was an independently poor prognostic factor for disease free interval (DFI) (per α-FR expression level increase: hazard ratio (HR): 2.45 (95% CI: 1.16–5.18), p = 0.02) and had a negative impact on overall survival (OS) of these serous ovarian cancer patients (per α-FR expression level increase: HR: 3.6 (95% CI: 0.93–13.29), p = 0.03) by multivariate analyses. α-FR inhibited cytotoxic drug-induced apoptosis in our in vitro apoptotic assays. α-FR could induce chemo-resistance via regulating the expression of apoptosis-related molecules, Bcl-2 and Bax. Therefore, α-FR can be a potential biomarker for the prediction of chemotherapeutic responses and clinical prognosis. It also could be the target of ovarian cancer treatment.Highlights: ► Patients with an increased α-FR expression level had poorer responses to chemotherapy. ► Increased α-FR expression level was a poor prognostic factor for overall survival of the patients. ► Ovarian serous cancer patients with high levels of α-FR had poor chemo-response. ► α-FR can be a biomarker for the outcome of ovarian serous cancer patients.