EGFR feedback-inhibition by Ran-binding protein 6 is disrupted in cancer

Transport of macromolecules through the nuclear pore by importins and exportins plays a critical role in the spatial regulation of protein activity. How cancer cells co-opt this process to promote tumorigenesis remains unclear. The epidermal growth factor receptor (EGFR) plays a critical role in normal development and in human cancer. Here we describe a mechanism of EGFR regulation through the importin β family member RAN-binding protein 6 (RanBP6), a protein of hitherto unknown functions. We show that RanBP6 silencing impairs nuclear translocation of signal transducer and activator of transcription 3 (STAT3), reduces STAT3 binding to the EGFR promoter, results in transcriptional derepression of EGFR, and increased EGFR pathway output. Focal deletions of the RanBP6 locus on chromosome 9p were found in a subset of glioblastoma (GBM) and silencing of RanBP6 promoted glioma growth in vivo. Our results provide an example of EGFR deregulation in cancer through silencing of components of the nuclear import pathway.This research was supported by the National Brain Tumor Society (I.K.M.), the National Institutes of Health grants 1R01NS080944-01 (I.K.M.), 1 R35 NS105109 01 (I.K.M.), and P30CA008748 (MSKCC Core Grant), the Geoffrey Beene Cancer Research Foundation (I.K.M.), the Cycle of Survival (I.K.M.), and the Seve Ballesteros Foundation (M.S.). B.O. was supported by an American–Italian Cancer Foundation fellowship and a MSKCC Brain Tumor Center grant. W.-Y.H. is the recipient of a FY15 Horizon Award from the U.S. Department of Defense (W81XWH-15-PRCRP-HA). A.C.-G. is the recipient of the Severo-Ochoa PhD fellowship. Further support was provided by the Sontag Foundation (B.S.T.). We thank all members of the Mellinghoff laboratory for helpful suggestions. We thank Dr. Fiona Ginty (Diagnostic Imaging and Biomedical Technologies, GE Global Research Center, Niskayuna, New York, USA) for assistance with multiplexed immunofluorescence. We thank A.J. Schuhmacher and C.S. Clemente-Troncone for assistance with the in vivo experiments, M. Kaufmann for assistance in the luciferase assays and N. Yannuzzi for assistance in cloning.S

Opposing effects of cancer-type-specific SPOP mutants on BET protein degradation and sensitivity to BET inhibitors.

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

Development and bioorthogonal activation of palladium-labile prodrugs of gemcitabine

Bioorthogonal chemistry has become one of the main driving forces in current chemical biology, inspiring the search for novel biocompatible chemospecific reactions for the past decade. Alongside the well-established labeling strategies that originated the bioorthogonal paradigm, we have recently proposed the use of heterogeneous palladium chemistry and bioorthogonal Pd⁰-labile prodrugs to develop spatially targeted therapies. Herein, we report the generation of biologically inert precursors of cytotoxic gemcitabine by introducing Pd⁰-cleavable groups in positions that are mechanistically relevant for gemcitabine’s pharmacological activity. Cell viability studies in pancreatic cancer cells showed that carbamate functionalization of the 4-amino group of gemcitabine significantly reduced (>23-fold) the prodrugs’ cytotoxicity. The <i>N</i>-propargyloxycarbonyl (<i>N</i>-Poc) promoiety displayed the highest sensitivity to heterogeneous palladium catalysis under biocompatible conditions, with a reaction half-life of less than 6 h. Zebrafish studies with allyl, propargyl, and benzyl carbamate-protected rhodamines confirmed <i>N</i>-Poc as the most suitable masking group for implementing <i>in vivo</i> bioorthogonal organometallic chemistry