PTPN11 Plays Oncogenic Roles and Is a Therapeutic Target for BRAF Wild-Type Melanomas.

Melanoma is one of the most highly mutated cancer types. To identify functional drivers of melanoma, we searched for cross-species conserved mutations utilizing a mouse melanoma model driven by loss of PTEN and CDKN2A, and identified mutations in Kras, Erbb3, and Ptpn11. PTPN11 encodes the SHP2 protein tyrosine phosphatase that activates the RAS/RAF/MAPK pathway. Although PTPN11 is an oncogene in leukemia, lung, and breast cancers, its roles in melanoma are not clear. In this study, we found that PTPN11 is frequently activated in human melanoma specimens and cell lines and is required for full RAS/RAF/MAPK signaling activation in BRAF wild-type (either NRAS mutant or wild-type) melanoma cells. PTPN11 played oncogenic roles in melanoma by driving anchorage-independent colony formation and tumor growth. In Pten- and Cdkn2a-null mice, tet-inducible and melanocyte-specific PTPN11E76K expression significantly enhanced melanoma tumorigenesis. Melanoma cells derived from this mouse model showed doxycycline-dependent tumor growth in nude mice. Silencing PTPN11E76K expression by doxycycline withdrawal caused regression of established tumors by induction of apoptosis and senescence, and suppression of proliferation. Moreover, the PTPN11 inhibitor (SHP099) also caused regression of NRASQ61K -mutant melanoma. Using a quantitative tyrosine phosphoproteomics approach, we identified GSK3α/ß as one of the key substrates that were differentially tyrosine-phosphorylated in these experiments modulating PTPN11. This study demonstrates that PTPN11 plays oncogenic roles in melanoma and regulates RAS and GSK3ß signaling pathways. IMPLICATIONS: This study identifies PTPN11 as an oncogenic driver and a novel and actionable therapeutic target for BRAF wild-type melanoma.

Epidermal growth factor expression as a predictor of chemotherapeutic resistance in muscle-invasive bladder cancer.

BACKGROUND: Epidermal growth factor receptor (EGFR) overexpression is believed to be associated with bladder cancer (BC) progression and poor clinical outcomes. In vivo studies have linked EGFR subcellular trafficking and chemo-resistance to cisplatin-based chemotherapies. This has not been studied in the clinical adjuvant setting. We aimed to investigate the prognostic significance of EGFR expression in patients receiving cisplatin-based adjuvant chemotherapy following radical cystectomy for advanced BC. METHODS: The database from the Urology and Nephrology Center at Mansoura University was reviewed. BC patients who were treated with radical cystectomy and adjuvant chemotherapy for adverse pathological features or node positive disease were identified. Patients who underwent palliative cystectomy, had histological diagnoses other than pure urothelial carcinoma, or received adjuvant radiotherapy were excluded from the study. Immunohistochemical staining for EGFR expression was performed on archived bladder specimens. The following in vitro functional analyses were performed to study the relationship of EGFR expression and chemoresponse. RESULTS: The study included 58 patients, among which the mean age was 57 years old. Majority of patients had node positive disease (n = 53, 91%). Mean follow up was 26.61 months. EGFR was overexpressed in 25 cystectomy specimens (43%). Kaplan-Meier analysis revealed that EGFR over-expression significantly correlated with disease recurrence (p = 0.021). Cox proportional hazard modeling identified EGFR overexpression as an independent predictor for disease recurrence (p = 0.04). Furthermore, in vitro experiments demonstrated that inhibition of EGFR may sensitize cellular responses to cisplatin. CONCLUSIONS: Our findings suggest that EGFR overexpression is associated with disease recurrence following adjuvant chemotherapy for advanced BC. This may aid in patient prognostication and selection prior to chemotherapeutic treatment for BC.

Integrated proteomic and phosphoproteomic analyses of cisplatin-sensitive and resistant bladder cancer cells reveal CDK2 network as a key therapeutic target.

BACKGROUND: Cisplatin-based chemotherapy is currently part of the standard of care for bladder cancer (BC). Unfortunately, some patients respond poorly to chemotherapy and have acquired or developed resistance. The molecular mechanisms underlying this resistance remain unclear. Here, we introduce a multidimensional proteomic analysis of a cisplatin-resistant BC model that provides different levels of protein information, including that of the global proteome and phosphoproteome. METHODS: To characterize the global proteome and phosphoproteome in cisplatin-resistant BC cells, liquid chromatography-mass spectrometry/mass spectrometry experiments combined with comprehensive bioinformatics analysis were performed. Perturbed expression and phosphorylation levels of key kinases associated with cisplatin resistance were further studied using various cell biology assays, including western blot analysis. RESULTS: Analyses of protein expression and phosphorylation identified significantly altered proteins, which were also EGF-dependent and independent. This suggests that protein phosphorylation plays a significant role in cisplatin-resistant BC. Additional network analysis of significantly altered proteins revealed CDK2, CHEK1, and ERBB2 as central regulators mediating cisplatin resistance. In addition to this, we identified the CDK2 network, which consists of CDK2 and its 5 substrates, as being significantly associated with poor survival after cisplatin chemotherapy. CONCLUSIONS: Collectively, these findings potentially provide a novel way of classifying higher-risk patients and may guide future research in developing therapeutic targets.

Differential perturbation of the interstitial cystitis-associated genes of bladder and urethra in rat model.

Interstitial cystitis (IC) is a chronic bladder dysfunction characterized as urinary frequency, urgency, nocturia, and pelvic pain. The changes in urethra may wind up with the bladder changes in structure and functions, however, the functions of the urethra in IC remains elusive. The aim of this study was to understand the perturbed gene expression in urethra, compared with urinary bladder, associated with the defected urodynamics. Using female IC mimic rats, a comprehensive RNA-sequencing combined with a bioinformatics analysis was performed and revealed that IC-specific genes in bladder or urethra. Gene ontology analysis suggested that the cell adhesion or extracellular matrix regulation, intracellular signaling cascade, cardiac muscle tissue development, and second messenger-mediated signaling might be the most enriched cellular processes in IC context. Further study of the effects of these bladder- or urethra-specific genes may suggest underlying mechanism of lower urinary tract function and novel therapeutic strategies against IC.

Interstitial Cystitis-Associated Urinary Metabolites Identified by Mass-Spectrometry Based Metabolomics Analysis.

This study on interstitial cystitis (IC) aims to identify a unique urine metabolomic profile associated with IC, which can be defined as an unpleasant sensation including pain and discomfort related to the urinary bladder, without infection or other identifiable causes. Although the burden of IC on the American public is immense in both human and financial terms, there is no clear diagnostic test for IC, but rather it is a disease of exclusion. Very little is known about the clinically useful urinary biomarkers of IC, which are desperately needed. Untargeted comprehensive metabolomic profiling was performed using gas-chromatography/mass-spectrometry to compare urine specimens of IC patients or health donors. The study profiled 200 known and 290 unknown metabolites. The majority of the thirty significantly changed metabolites before false discovery rate correction were unknown compounds. Partial least square discriminant analysis clearly separated IC patients from controls. The high number of unknown compounds hinders useful biological interpretation of such predictive models. Given that urine analyses have great potential to be adapted in clinical practice, research has to be focused on the identification of unknown compounds to uncover important clues about underlying disease mechanisms.