MEK2 Is Sufficient but Not Necessary for Proliferation and Anchorage-Independent Growth of SK-MEL-28 Melanoma Cells

Mitogen-activated protein kinase kinases (MKK or MEK) 1 and 2 are usually treated as redundant kinases. However, in assessing their relative contribution towards ERK-mediated biologic response investigators have relied on tests of necessity, not sufficiency. In response we developed a novel experimental model using lethal toxin (LeTx), an anthrax toxin-derived pan-MKK protease, and genetically engineered protease resistant MKK mutants (MKKcr) to test the sufficiency of MEK signaling in melanoma SK-MEL-28 cells. Surprisingly, ERK activity persisted in LeTx-treated cells expressing MEK2cr but not MEK1cr. Microarray analysis revealed non-overlapping downstream transcriptional targets of MEK1 and MEK2, and indicated a substantial rescue effect of MEK2cr on proliferation pathways. Furthermore, LeTx efficiently inhibited the cell proliferation and anchorage-independent growth of SK-MEL-28 cells expressing MKK1cr but not MEK2cr. These results indicate in SK-MEL-28 cells MEK1 and MEK2 signaling pathways are not redundant and interchangeable for cell proliferation. We conclude that in the absence of other MKK, MEK2 is sufficient for SK-MEL-28 cell proliferation. MEK1 conditionally compensates for loss of MEK2 only in the presence of other MKK

Preclinical Evaluation of Genomic-Based Therapies in Pancreatic Cancer and Glioblastoma

INTRODUCTION: The focus of this study is the testing of biomarker-driven analytical methods to identify targeted therapies in pancreatic cancer and glioblastoma, which are highly invasive and metastatic cancers with poor outcomes and few treatment options. The objective was to make treatment predictions based on the molecular signatures of pancreatic cancer and glioblastoma samples, then to evaluate the efficacy of these therapies using preclinical models. METHODS AND MATERIALS: XenoBase Bio-Integration Suite (XB-BIS) in an informatics platform for the analysis of molecular data using Personalized Medicine (PMED) algorithms. PMED applies four independent methods (Drug Target Expression, Connectivity Map, Parametric Gene Set Enrichment, and GeneGo Network Topological Enrichment Analysis) to a genomic dataset to identify targeted therapies. Affymetrix data was collected from panels of pancreatic cancer cell lines and human glioblastoma specimens and analyzed in XB-BIS to predict therapies, which were evaluated in vivo. RESULTS: Treatment of mice with subcutaneous pancreatic tumors with Chlorpromazine, predicted by CMAP, resulted in a decrease in tumor volume and extended survival compared to control animals. Predictive algorithms identified BCNU, Doxorubicin, and Marimastat as potential treatments for glioblastoma. Combination treatment of mice implanted intracranially with U251 glioblastoma cells showed extended survival compared to control mice and similar survival to standard-of-care treatment, Temozolomide. CONCLUSIONS: We have demonstrated efficacy of therapies identified by the PMED approach in relevant models of pancreatic cancer and glioblastoma. While further investigation is needed, these therapies could prove to be a great resource against two devastating human diseases

Precision Medicine in Rhabdomyosarcoma: Using Patient Derived Xenografts as models of drug efficacy and acquired resistance

Background. Precision (Personalized) medicine has the potential to revolutionize patient health care and whilst there have been huge advances for a few cancers of known etiology, for many cancers, the fundamental causes of the disease process remain either elusive or have no available therapy. Here we outline a study in alveolar rhabdomyosarcoma, in which we use gene expression profiling and a series of drug prediction algorithms combined with a matched patient derived xenograft model (PDX) to test predicted therapies. Procedure. A PDX model was developed from a patient biopsy and a number of drugs identified using gene expression analysis in combination with drug prediction algorithms. Drugs chosen from each of the predictive methodologies, along with the patient’s standard-of-care (ICE-T), were tested in vivo in the PDX tumor. A second study was initiated using the tumors that re-grew following the ICE-T treatment. Further expression analysis identified additional therapies with potential anti-tumor efficacy. Results. A number of the predicted therapies were found to be active against the tumors in particular BGJ398 (FGFR2) and ICE-T. Re-transplantation of the ICE-T treated tumorgrafts demonstrated a decrease in response to ICE-T recapitulating the patient’s refractory disease. Gene expression profiling of the ICE-T treated tumorgrafts identified cytarabine (SLC29A1) as a potential therapy, which was shown, along with BGJ398, to be highly active in vivo. Conclusions. This study illustrates that tumorgrafts are ideal surrogates for testing potential therapeutic strategies based on gene expression analysis, modeling clinical drug resistance and hold the potential to assist in guiding prospective patient care

Sox2 Promotes Malignancy in Glioblastoma by Regulating Plasticity and Astrocytic Differentiation

The high-mobility group–box transcription factor sex-determining region Y–box 2 (Sox2) is essential for the maintenance of stem cells from early development to adult tissues. Sox2 can reprogram differentiated cells into pluripotent cells in concert with other factors and is overexpressed in various cancers. In glioblastoma (GBM), Sox2 is a marker of cancer stemlike cells (CSCs) in neurosphere cultures and is associated with the proneural molecular subtype. Here, we report that Sox2 expression pattern in GBM tumors and patient-derived mouse xenografts is not restricted to a small percentage of cells and is coexpressed with various lineage markers, suggesting that its expression extends beyond CSCs to encompass more differentiated neoplastic cells across molecular subtypes. Employing a CSC derived from a patient with GBM and isogenic differentiated cell model, we show that Sox2 knockdown in the differentiated state abolished dedifferentiation and acquisition of CSC phenotype. Furthermore, Sox2 deficiency specifically impaired the astrocytic component of a biphasic gliosarcoma xenograft model while allowing the formation of tumors with sarcomatous phenotype. The expression of genes associated with stem cells and malignancy were commonly downregulated in both CSCs and serum-differentiated cells on Sox2 knockdown. Genes previously shown to be associated with pluripontency and CSCs were only affected in the CSC state, whereas embryonic stem cell self-renewal genes and cytokine signaling were downregulated, and the Wnt pathway activated in differentiated Sox2-deficient cells. Our results indicate that Sox2 regulates the expression of key genes and pathways involved in GBM malignancy, in both cancer stemlike and differentiated cells, and maintains plasticity for bidirectional conversion between the two states, with significant clinical implications