Molecular Oncology

Editorial Board

2012-02-01

Exploitation of the propulsive force of chemotherapy for improving the response to cancer immunotherapy

Enrico Proietti, Federica Moschella, Imerio Capone, Filippo Belardelli — 2011-12-09

Abstract: Since the early clinical studies of cancer immunotherapy, the question arose as to whether it was possible to combine it with standard cancer treatments, mostly chemotherapy. The answer, now, is past history. The combined use of immunotherapy and chemotherapy is not only possible but, in certain cases, can be advantageous, depending on the drug, the dose and the combination modalities. In order to find the best synergisms between the two treatments and to turn weak immunotherapeutic interventions into potent anticancer instruments, it is mandatory to understand the complex mechanisms responsible for the positive interactions between chemotherapy and immunotherapy. In this article, we review the current knowledge on mechanisms involved in the immunostimulating activity of chemotherapy and summarize the main studies in both mouse models and patients aimed at exploiting such mechanisms for enhancing the response to cancer immunotherapy.Highlights: ► Chemotherapy strongly enhances anticancer efficacy of immunotherapeutic strategies. ► Three mechanisms of synergism have been hypothesized: “subtractive”, “immunogenic” and “propulsive”. ► Chemo-immunotherapy can effectively be used as secondary prevention against tumor relapse.

Drug development to overcome resistance to EGFR inhibitors in lung and colorectal cancer

Rodrigo Dienstmann, Sara De Dosso, Enriqueta Felip, Josep Tabernero — 2011-12-14

Abstract: Epidermal growth factor receptor (EGFR) is a validated target in different human malignancies. EGFR tyrosine kinase inhibitors (TKIs) are known to contribute considerably to the extension of progression-free survival in EGFR-mutant non-small cell lung cancer and monoclonal antibodies (mAbs) targeting EGFR have also improved the efficacy outcomes in KRAS wild-type colorectal cancer. Nevertheless, a significant percentage of lung and colorectal cancer patients do not respond to anti-EGFR agents and secondary resistance after initial benefit is a challenging reality faced by clinicians. Extensive preclinical work on the potential mechanisms of resistance to EGFR inhibitors in different disease settings has guided the development of second-generation irreversible EGFR TKIs, more efficient anti-EGFR mAbs, and combination strategies with agents targeting other receptors and downstream effectors. In order to elucidate the role of the multiple therapeutic strategies under investigation to overcome EGFR inhibitors-resistance, rational drug development based on stringent preclinical data, biomarker validation and proper selection of patients in the ongoing clinical trials are of paramount importance. Preliminary results of clinical trials evaluating these approaches will be discussed in this manuscript, with emphasis on TKIs in lung cancer and mAbs in advanced colorectal cancer.Graphical abstract: CRC: colorectal cancer; EGFR: epidermal growth factor receptor; TKI: tyrosine kinase inhibitor; mAb: monoclonal antibody; IGF-1R: insulin-like growth factor 1 receptor; MET: mesenchymal–epithelial transition receptor; NSCLC: non-small cell lung cancer.Highlights: ► KRAS and secondary EGFR mutations confer resistance to EGFR inhibitors. ► Upregulation of parallel signaling pathways is a frequent adoptive response. ► Irreversible EGFR TKIs may overtake resistance in lung cancer. ► More efficient anti-EGFR mAbs are promising agents in colorectal cancer. ► Combination of targeted agents is a straightforward approach under investigation.

Systems pathology: A critical review

Jose Costa — 2011-12-05

Abstract: The technological advances of the last twenty years together with the dramatic increase in computational power have injected new life into systems-level thinking in Medicine. This review emphasizes the close relationship of Systems Pathology to Systems Biology and delineates the differences between Systems Pathology and Clinical Systems Pathology. It also suggests an algorithm to support the application of systems-level thinking to clinical research, proposes applying systems-level thinking to the health care systems and forecasts an acceleration of preventive medicine as a result of the coupling of personal genomics with systems pathology.Highlights: ► This review defines Systems Pathology. ► Relates it to Systems Biology. ► Defines Clinical System Biology. ► Discusses difficulties for systems translational medicine.

uPA and uPAR shRNA inhibit angiogenesis via enhanced secretion of SVEGFR1 independent of GM-CSF but dependent on TIMP-1 in endothelial and glioblastoma cells

2011-12-08

Abstract: The uPA/uPAR system is known to play a critical role in angiogenesis of glioblastoma. Previously, we have shown that shRNA against uPA and uPAR attenuates angiogenesis by blocking nuclear translocation of angiogenin, inhibition of angiopoietin/Tie2 signaling, and regulating several other pro-angiogenic, angiostatic and anti-angiogenic molecules. Further analysis revealed that GM-CSF, a pleiotropic cytokine, was significantly inhibited in U87MG and 4910 co-cultures with endothelial cells transfected with shRNA against uPA and uPAR. The role of the uPA/uPAR system in this process is not completely understood. Analysis of tumor conditioned medium of U87MG, 4910 and HMECs transfected with shRNA against uPA or uPAR alone or in combination (pU2) revealed inhibition of GM-CSF-enhanced secretion of SVEGFR1 as shown by Western blotting and ELISA. Moreover, phosphorylation of JAK2 and STAT5, the downstream effectors of GM-CSF signaling, was also inhibited in all three cell lines. Phosphorylation at Tyr 166 position of the GM-CSFRβ subunit, the signal activating subunit of the GM-CSF receptor, was inhibited in HMEC, U87MG and 4910 cells. Further analysis revealed that shRNA against uPA and/or uPAR increased secretion of TIMP-1, which is known to enhance SVEGFR1 secretion in endothelial cells. Moreover, addition of purified uPA (with and without GM-CSF) activated JAK2/STAT5 signaling in HMEC. Exogenous addition of SVEGFR1 to pU2 tumor conditioned medium enhanced inhibition of VEGF-induced endothelial capillary tube formation as assessed by an in vitro angiogenesis assay. To determine the significance of these events in vivo, nude mice with pre-established tumors treated with shRNA against uPA and/or uPAR showed decreased levels of GM-CSF and increased levels of SVEGFR1 and TIMP-1 when compared with controls. Enhanced secretion of SVEGFR1 by puPA, puPAR and pU2 in endothelial and GBM cells was mediated indirectly by MMP-7 and augmented by ectodomain shedding of VEGFr1 by tyrosine phosphorylation at the 1213 position. Taken together, these results suggest that the uPA/uPAR system could prove beneficial as an indirect target for inhibition of angiogenesis in glioblastoma.Highlights: ► First report highlighting the role and mechanism by which shRNA against uPA/uPAR inhibits angiogenesis in glioblastoma. ► shRNA against uPA/uPAR enhances secretion of sVEGFr1, a scavenger for VEGF. ► sVEGFr1 secretion is independent of GM-CSF unlike monocytes but dependent on TIMP-1. ► TIMP-1 secretion is mediated by MMP-7 which is further regulated by uPA and uPAR.

CRABP-II methylation: A critical determinant of retinoic acid resistance of medulloblastoma cells

2011-12-02

Abstract: Medulloblastoma cells exhibit varied responses to therapy by all-trans retinoic acid (RA). The underlying mechanism for such diverse effects however remains largely unclear. In this study, we attempted to elucidate the molecular basis of RA resistance through the study of RA signaling components in both RA-sensitive (Med-3) and RA-resistant (UW228-2 and UW228-3) medulloblastoma cells. The results revealed that RARα/β/γ and RXRα/β/γ were found in the three cell lines. Expression of CRABP-I and CRABP-II was seen in Med-3 cells, up-regulated when treated with RA, but was absent in UW228-2 and UW228-3 cells regardless of RA treatment. Bisulfite sequencing revealed 8 methylated CG sites at the promoter region of CRABP-II in UW228-2 and UW228-3 but not in Med-3 cells. Demethylation by 5-aza-2′-deoxycytidine recovered CRABP-II expression. Upon restoration of CRABP-II expression, both UW228-2 and UW228-3 cells responded to RA treatment by forming neuronal-like differentiation, synaptophysin expression, β-III tubulin upregulation, and apoptosis. Furthermore, CRABP-II specific siRNA reduced RA sensitivity in Med-3 cells. Tissue microarray-based immunohistochemical staining showed variable CRABP-II expression patterns among 104 medulloblastoma cases, ranging from negative (42.3%), partly positive (14.4%) to positive (43.3%). CRABP-II expression was positively correlated with synaptophysin (rs = 0.317; p = 0.001) but not with CRABP-I expression (p > 0.05). In conclusion, aberrant methylation in CRABP-II reduces the expression of CRABP-II that in turn confers RA resistance in medulloblastoma cells. Determination of CRABP-II expression or methylation status may enable a personalized RA therapy in patients with medulloblastomas and other types of cancers.Highlights: ► CRABP-II is silenced in RA-resistant medulloblastoma/MB cells due to promoter methylation. ► Restoration of CRABP-II expression overcomes RA-resistant property of MB cells. ► Inhibition of CRABP-II expression reduces RA sensitivity of MB cells. ► CRABP-II expression patterns are highly variable in medulloblastoma tissues. ► Evaluation of CRABP-II expression would be of values in personalized RA therapy for MB patients.

Atorvastatin reduces vascular endothelial growth factor (VEGF) expression in human non-small cell lung carcinomas (NSCLCs) via inhibition of reactive oxygen species (ROS) production

2011-11-30

Abstract: The high metastatic potential of non-small cell lung cancers (NSCLCs) is closely correlated with the elevated expression of vascular endothelial growth factor (VEGF) and resultant tumor angiogenesis. However, no effective strategies against VEGF expression have been available in NSCLCs therapy. This study demonstrated that elevated reactive oxygen species (ROS) levels derived from both mitochondria and NADPH oxidase were required for VEGF expression in NSCLC cells. Atorvastatin administration could significantly inhibit VEGF expression both in vitro and in vivo via inhibition of ROS production. Atorvastatin inhibited ROS generation partly through suppression of Rac1/NADPH oxidase activity. Specifically, atorvastatin could upregulate the activity of glutathione peroxidase (GPx) and catalase, which are responsible for elimination of hydrogen peroxide (H2O2) in the mitochondria and peroxisomes, respectively. Thus, inhibition of ROS production by concomitant suppression of Rac1/NADPH oxidase activity and upregulation of the activity of GPx and catalase contributes critically to atorvastatin-reduced VEGF expression in NSCLCs. Atorvastatin may be a potential alternative against VEGF expression and angiogenesis in NSCLCs therapy.Highlights: ► Both mitochondria and NADPH oxidase are required for VEGF expression in NSCLCs. ► Atorvastatin suppresses VEGF expression in NSCLCs via inhibition of ROS production. ► Atorvastatin inhibits ROS production mostly via suppression of NADPH oxidase. ► Atorvastatin can upregulate the activity of GPx and catalase.

Down-regulation of tumor suppressor MTUS1/ATIP is associated with enhanced proliferation, poor differentiation and poor prognosis in oral tongue squamous cell carcinoma

2011-11-28

Abstract: Microtubule-associated tumor suppressor gene (MTUS1, also known as mitochondrial tumor suppressor) is a recently identified tumor suppressor gene that has been implicated in several cancer types. The expression of MTUS1 gene leads to 5 known transcript variants and codes for 5 isoforms of Angiotensin II AT2 receptor interacting protein (ATIP). In this study, we first confirmed that the down-regulation of MTUS1/ATIP was a frequent event in oral tongue squamous cell carcinoma (OTSCC) and the premalignant lesion (leukoplakia). We further demonstrated that the down-regulation of MTUS1/ATIP was correlated with poor differentiation and enhanced proliferation (Ki67 proliferation index). Statistical analysis suggests that the down-regulation of MTUS1/ATIP was associated with reduced overall survival. Isoform specific quantitative RT-PCR assays revealed that ATIP1, ATIP3a and ATIP3b were the major isoforms of the MTUS1 gene products in oral tongue epithelial cells. Significant down-regulations were observed for all 3 ATIP isoforms in OTSCC as compared to matching normal tissues. In vitro functional study showed that the restoration of ATIP1 expression led to G1 arrest, apoptosis and reduction of cell proliferation in OTSCC cell lines. These ATIP1-induced cellular changes were accompanied by reduced phosphorylation of ERK1/2 and up-regulation of p53. Taken together, these data suggest that MTUS1 plays major roles in the progression of OTSCC, and may serve as a biomarker or therapeutic target for patients with OTSCC.Highlights: ► Down-regulation of tumor suppressor MTUS1/ATIP is a frequent event in OTSCC. ► MTUS1/ATIP correlates with differentiation, proliferation, and overall survival. ► Restoration of MTUS1/ATIP led to G1 arrest, apoptosis and reduced cell proliferation.

Aquaporins mediate the chemoresistance of human melanoma cells to arsenite

2011-11-21

Abstract: The integral membrane channel protein aquaporin (AQP) is aberrantly expressed with oncogenic characteristics in various human cancers. In this study, we analyzed the expression pattern of all subtypes of AQPs, and found that 8 out of 13 AQPs expressed in melanoma cells. To understand the role of aberrant expression of AQP in this disease, we over-expressed AQP3 and AQP9 in human melanoma WM266.4 cells and found that both AQPs significantly increased the chemoresistance of WM266.4 cells to arsenite. Functional studies showed that AQP3 and AQP9 can inhibit cell apoptosis induced by arsenite through down-regulating p53 and up-regulating Bcl-2 and XIAP. Our data suggest the implication of APQ in melanoma progression and that the over-expression of AQP3 and AQP9 contributes to the chemoresistance of melanoma to arsenite.Highlights: ► We identify the expression pattern of all subtypes of AQP in melanoma cells. ► Over-expression of AQP3 and AQP9 increases the chemoresistance of melanoma cells to arsenite. ► AQP has no effect on melanoma cell growth. ► The mechanism of AQP affecting melanoma chemoresistance is relevant to apoptosis regulated by p53, Bax, Bcl-2, and XIAP.

Lack of independent prognostic and predictive value of centromere 17 copy number changes in breast cancer patients with known HER2 and TOP2A status

2011-12-05

Abstract: The clinical benefit of anthracyclines has been connected to HER2 status, TOP2A status and centromere 17 copy numbers (CEN-17). Data from a clinical trial randomizing patients to anthracyclines was used to assess whether the number of CEN-17 in breast cancers may predict incremental responsiveness to anthracyclines besides what is obtained when used relatively to TOP2A and HER2. As cut sections of paraffin-embedded tissue are prone to truncation of nuclei, strict definition of ploidy levels is lacking. We therefore used normal breast tissue to assist define ploidy levels in cut sections. Fluorescence in situ hybridization (FISH) with centromere 17 (CEN-17) and TOP2A was performed on 120 normal breast specimens. The diploid CEN-17 copy number was reduced from the expected two signals in whole nuclei to an average of 1.68 signals per nucleus in cut sections of normal breast. Ploidy levels determined in normal breast were applied to data on 767 patients with known HER2 and TOP2A status randomized to anthracyclines in the DBCG 89D trial. CEN-17 ploidy levels were in cut sections from the 767 breast cancer patients established as: Haploid: ≤1.25 (10%), diploid: 1.26–2.09 (60%), triploid: 2.10–2.93 (21%), tetraploid: 2.94–3.77 (5%) or higher ploidy: ≥3.78 (4%). Amplification of HER2 and deletion of TOP2A were frequently observed in tumors with a high ploidy level. In univariate analyses increasing ploidy was associated with decreased disease-free survival (DFS) (P=0.0001) and overall survival (OS) (P<0.0001). However, in multivariate analysis CEN-17 was not established as an independent prognostic factor and was neither a statistically significant predictor of benefit from CEF (Cyclophosphamide/Epirubicin/5-Fluorouracil) compared to CMF (Cyclophosphamide/Methotrexate/5-Fluorouracil) (PInteraction 0.39 for DFS and 0.67 for OS). In conclusion, CEN-17 levels do not independently from TOP2A/CEN-17 ratio identify breast cancer patients who achieve an incremental benefit from adjuvant anthracyclines.Highlights: ► Overall, breast cancer patients benefit from treatment with anthracyclines. ► Predictive biomarkers are needed for stratifying patients to anthracyclines. ► TOP2A, HER2 or centromere 17 status may serve as biomarkers. ► TOP2A but not HER2 has predictive value in the DBCG 89D trial. ► CEN-17 status does not independently from TOP2A predict anthracycline benefit.

Genomic imbalances in endometrial adenocarcinomas – Comparison of DNA ploidy, karyotyping and comparative genomic hybridization

2011-10-28

Abstract: DNA ploidy analysis is useful for prognostication in cancer patients, but the genomic details underlying ploidy changes are not fully understood. To improve this understanding, we compared DNA ploidy status with karyotypic and comparative genomic hybridization data on 51 endometrial adenocarcinomas. Out of 34 DNA diploid tumors evaluated by CGH, 16 (47%) showed imbalances, though only two had more than four copy number changes. Ten (29%) had aberrations involving chromosome 1, seven (21%) involving chromosome 10, while one tumor had a chromosome 8 aberration. Four of the seven DNA tetraploid tumors (57%) had imbalances detected by CGH with two (29%) having more than four. Six out of eight DNA aneuploid tumors showed imbalances by CGH, with five (63%) having more than four. The aberrations were observed on chromosomes 1 and 8 in five/eight (63%) cases while four imbalances (50%) involved chromosomes 5, 7 and X. Not surprisingly, we observed a significant correlation between increasing DNA ploidy complexity and increasing number of copy alterations. Gains of material from chromosomes 8 and 7 might be specifically correlated to DNA aneuploidy in endometrial adenocarcinomas since 63% and 50% of the aneuploid compared to 3% of the diploid tumors showed imbalances involving these chromosomes.Highlights: ► Reveal specific CGH aberrations related to DNA aneuploidy in endometrial carcinomas. ► Correlation between increasing DNA ploidy complexity and copy alterations. ► 63% of the DNA aneuploid lesions had aberrations on chromosome 8. ► 50% of the DNA aneuploid lesions had aberrations on chromosome 7. ► Only 3% of the DNA diploid lesions had aberrations on chromosome 7 or 8.