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Driver genes with a mutually exclusive mutation pattern across tumor genomes are thought to have overlapping roles in tumorigenesis. In contrast, we show here that mutually exclusive prostate cancer driver alterations involving the ERG transcription factor and the ubiquitin ligase adaptor SPOP are synthetic sick. At the molecular level, the incompatible cancer pathways are driven by opposing functions in SPOP. ERG upregulates wild type SPOP to dampen androgen receptor (AR) signaling and sustain ERG activity through degradation of the bromodomain histone reader ZMYND11. Conversely, SPOP-mutant tumors stabilize ZMYND11 to repress ERG-function and enable oncogenic androgen receptor signaling. This dichotomy regulates the response to therapeutic interventions in the AR pathway. While mutant SPOP renders tumor cells susceptible to androgen deprivation therapies, ERG promotes sensitivity to high-dose androgen therapy and pharmacological inhibition of wild type SPOP. More generally, these results define a distinct class of antagonistic cancer drivers and a blueprint toward their therapeutic exploitation.

It is generally assumed that recurrent mutations within a given cancer driver gene elicit similar drug responses. Cancer genome studies have identified recurrent but divergent missense mutations affecting the substrate-recognition domain of the ubiquitin ligase adaptor SPOP in endometrial and prostate cancers. The therapeutic implications of these mutations remain incompletely understood. Here we analyzed changes in the ubiquitin landscape induced by endometrial cancer-associated SPOP mutations and identified BRD2, BRD3 and BRD4 proteins (BETs) as SPOP-CUL3 substrates that are preferentially degraded by endometrial cancer-associated SPOP mutants. The resulting reduction of BET protein levels sensitized cancer cells to BET inhibitors. Conversely, prostate cancer-specific SPOP mutations resulted in impaired degradation of BETs, promoting their resistance to pharmacologic inhibition. These results uncover an oncogenomics paradox, whereby mutations mapping to the same domain evoke opposing drug susceptibilities. Specifically, we provide a molecular rationale for the use of BET inhibitors to treat patients with endometrial but not prostate cancer who harbor SPOP mutations.

Androgen receptor (AR) signaling is a key driver of prostate cancer (PC). While androgen-deprivation therapy is transiently effective in advanced disease, tumors often progress to a lethal castration-resistant state (CRPC). We show that recurrent PC-driver mutations in speckle-type POZ protein (SPOP) stabilize the TRIM24 protein, which promotes proliferation under low androgen conditions. TRIM24 augments AR signaling, and AR and TRIM24 co-activated genes are significantly upregulated in CRPC. Expression of TRIM24 protein increases from primary PC to CRPC, and both TRIM24 protein levels and the AR/TRIM24 gene signature predict disease recurrence. Analyses in CRPC cells reveal that the TRIM24 bromodomain and the AR-interacting motif are essential to support proliferation. These data provide a rationale for therapeutic TRIM24 targeting in SPOP mutant and CRPC patients., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Integrins are extracellular matrix receptors involved in several pathologies. Despite homologies between the RGD-binding α5β1 and αvβ3 integrins, selective small antagonists for each heterodimer have been proposed. Herein, we evaluated the effects of such small antagonists in a cellular context, the U87MG cell line, which express both integrins. The aim of the study was to determine if fibronectin-binding integrin antagonists are able to impact on cell adhesion and migration in relationships with their defined affinity and selectivity for α5β1 and αvβ3/β5 purified integrins., Methods: Small antagonists were either selective for α5β1 integrin, for αvβ3/β5 integrin or non-selective. U87MG cell adhesion was evaluated on fibronectin or vitronectin. Migration assays included wound healing recovery and single cell tracking experiments. U87MG cells stably manipulated for the expression of α5 integrin subunit were used to explore the impact of α5β1 integrin in the biological assays., Results: U87MG cell adhesion on fibronectin or vitronectin was respectively dependent on α5β1 or αvβ3/β5 integrin. Wound healing migration was dependent on both integrins. However U87MG single cell migration was highly dependent on α5β1 integrin and was inhibited selectively by α5β1 integrin antagonists but increased by αvβ3/β5 integrin antagonists., Conclusions: We provide a rationale for testing new integrin ligands in a cell-based assay to characterize more directly their potential inhibitory effects on integrin cellular functions., General Significance: Our data highlight a single cell tracking assay as a powerful cell-based test which may help to characterize true functional integrin antagonists that block α5β1 integrin-dependent cell migration., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Integrins emerge nowadays as crucial actors of tumor aggressiveness and resistance to therapies. Integrin α5β1, the fibronectin receptor, determines malignant properties of colon carcinoma which is one of the most important causes of cancer-related deaths in the world. Here we show that inhibition of α5 integrin subunit expression by siRNA or α5β1 integrin function by specific antagonist affects the survival of HCT116 colon cancer cells. We also evidence that pharmacological reactivation of the tumor suppressor p53 by Nutlin-3a inhibits specifically the expression of the α5 integrin subunit both at the transcriptional and protein level. Inversely repression of α5 integrin modulates p53 activity. A clear relationship between p53 activation by Nutlin-3a, α5 repression and cell survival is shown. No such effects are obtained in cells lacking p53 or when another non-genotoxic activator of p53, RITA, is used. Our results emphasize the crucial role of α5β1 integrin in colon tumors. Data also suggest that interfering with the integrin α5β1 through the reactivation of p53 by Nutlin-3a may be of valuable interest as a new therapeutic option for colon tumors expressing high level of the integrin and a wild type p53., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

The transient receptor potential (TRP) protein superfamily consists of seven major groups, among them the "canonical TRP" family. The TRPC proteins are calcium-permeable nonselective cation channels activated after the emptying of intracellular calcium stores and appear to be gated by various types of messengers. The TRPC6 channel has been shown to be expressed in various tissues and cells, where it modulates the calcium level in response to external signals. Calcium binding proteins such as Calmodulin or the family of S100A proteins are regulators of TRPC channels. Here we characterized the overlapping integrative binding site for S100A1 at the C-tail of TRPC6, which is also able to accomodate various ligands such as Calmodulin and phosphatidyl-inositol-(4,5)-bisphosphate. Several positively charged amino acid residues (Arg852, Lys856, Lys859, Arg860 and Arg864) were determined by fluorescence anisotropy measurements for their participation in the calcium-dependent binding of S100A1 to the C terminus of TRPC6. The triple mutation Arg852/Lys859/Arg860 exhibited significant disruption of the binding of S100A1 to TRPC6. This indicates a unique involvement of these three basic residues in the integrative overlapping binding site for S100A1 on the C tail of TRPC6.

Glioblastoma is the most common malignant primary brain tumor. Surgical resection, postoperative radiotherapy plus concomitant and adjuvant chemotherapy with temozolomide (TMZ) is the standard of care for newly diagnosed glioblastoma. In the past decade, efforts have been made to decipher genomic and core pathway alterations to identify clinically relevant glioblastoma subtypes. Based on these studies and more academic explorations, new potential therapeutic targets were found and several targeting agents were developed. Such molecules should hopefully overcome the resistance of glioblastoma to the current therapy. One of the hallmarks of glioblastoma subtypes was the enrichment of extracellular matrix/invasion-related genes. Integrins, which are cell adhesion molecules important in glioma cell migration/invasion and angiogenesis were one of those genes. Integrins seem to be pertinent therapeutic targets and antagonists recently reached the clinic. Although the p53 pathway appears often altered in glioblastoma, conflicting results can be found in the literature about the clinically relevant impact of the p53 status in the resistance to TMZ. Here, we will summarize the current knowledge on (1) integrin expression, (2) p53 status, and (3) relationship between integrins and p53 to discuss their potential impact on the resistance of glioblastoma to temozolomide.

Integrins play a role in the resistance of advanced cancers to radiotherapy and chemotherapy. In this study, we show that high expression of the α5 integrin subunit compromises temozolomide-induced tumor suppressor p53 activity in human glioblastoma cells. We found that depletion of the α5 integrin subunit increased p53 activity and temozolomide sensitivity. However, when cells were treated with the p53 activator nutlin-3a, the protective effect of α5 integrin on p53 activation and cell survival was lost. In a functional p53 background, nutlin-3a downregulated the α5 integrin subunit, thereby increasing the cytotoxic effect of temozolomide. Clinically, α5β1 integrin expression was associated with a more aggressive phenotype in brain tumors, and high α5 integrin gene expression was associated with decreased survival of patients with high-grade glioma. Taken together, our findings indicate that negative cross-talk between α5β1 integrin and p53 supports glioma resistance to temozolomide, providing preclinical proof-of-concept that α5β1 integrin represents a therapeutic target for high-grade brain tumors. Direct activation of p53 may remain a therapeutic option in the subset of patients with high-grade gliomas that express both functional p53 and a high level of α5β1 integrin.

The transient receptor potential channel TRPC6 is a non-selective cation channel which modulates the calcium level in eukaryotic cells (including sensory receptor cells) in response to external signals. Calmodulin (CaM) is a ubiquitously expressed Ca(2+) binding protein that is an important mediator of Ca(2+)-dependent regulation of the TRPC6 channel. One CaM binding site was identified within the C-tail of TRPC6. The aim of this study is to map in detail the CaM and inositol (1,4,5)-triphosphate receptor binding (CIRB) domain in the C-terminal region of mouse TRPC6 that is capable of interacting with CaM using in vitro binding assays. Besides the set of positively charged amino acid residues Arg852, Lys856, Arg864, Lys859/Arg860, a hydrophobic Ile857, at the position 1 in 1-5-10 motif, was located and the effect of replacing it with a neutral residue was tested using fluorescence anisotropy measurement. Participation of Ile857 could indicate a strong role of this conserved CaM binding motif., (Copyright 2009 Elsevier Ltd. All rights reserved.)

Calmodulin (CaM) is known to play an important role in the regulation of TRP channels activity. Although it has been reported that CaM binds to the C-terminus of TRPV1 (TRPV1-CT), no classic CaM-binding motif was found in this region. In this work, we explored this unusual TRPV1 CaM-binding motif in detail and found that five residues from a putative CaM-binding motif are important for TRPV1-CT's binding to CaM, with arginine R785 being the most essential residue. The homology modelling suggests that a CaM-binding motif of TRPV1-CT forms an alpha helix that docks into the central cavity of CaM.