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Mitogen-activated protein (MAP) kinases belong to evolutionary conserved cell-cell communication systems that are involved in the coupling of extracellular signaling to changes in gene expression. In Caenorhabditis elegans, one MAP kinase gene has been linked to the vulval induction pathway, which is very similar to numerous mammalian growth factor signaling pathways. Although the exact genetics of the C. elegans vulval induction pathway is not yet clear, the current findings indicate that MAP kinases play an essential role in receptor tyrosine kinase-mediated signal transduction pathways.

Abstract Background Shared-usage high throughput screening (HTS) facilities are becoming more common in academe as large-scale small molecule and genome-scale RNAi screening strategies are adopted for basic research purposes. These shared facilities require a unique informatics infrastructure that must not only provide access to and analysis of screening data, but must also manage the administrative and technical challenges associated with conducting numerous, interleaved screening efforts run by multiple independent research groups. Results We have developed Screensaver, a free, open source, web-based lab information management system (LIMS), to address the informatics needs of our small molecule and RNAi screening facility. Screensaver supports the storage and comparison of screening data sets, as well as the management of information about screens, screeners, libraries, and laboratory work requests. To our knowledge, Screensaver is one of the first applications to support the storage and analysis of data from both genome-scale RNAi screening projects and small molecule screening projects. Conclusions The informatics and administrative needs of an HTS facility may be best managed by a single, integrated, web-accessible application such as Screensaver. Screensaver has proven useful in meeting the requirements of the ICCB-Longwood/NSRB Screening Facility at Harvard Medical School, and has provided similar benefits to other HTS facilities.

Our previous pre-clinical work defined BCL-2 induction as a critical component of the adaptive response to lapatinib-mediated inhibition of HER2. To determine whether a similar BCL-2 upregulation occurs in lapatinib-treated patients, we evaluated gene expression within tumor biopsies, collected before and after lapatinib or trastuzumab treatment, from the TRIO-B-07 clinical trial (NCT#00769470). We detected BCL2 mRNA upregulation in both HER2+/ER- as well as HER2+/ER+ patient tumors treated with lapatinib or trastuzumab. To address whether mRNA expression correlated with protein expression, we evaluated pre- and post-treatment tumors for BCL-2 via immunohistochemistry. Despite BCL2 mRNA upregulation within HER2+/ER- tumors, BCL-2 protein levels were undetectable in most of the lapatinib- or trastuzumab-treated HER2+/ER- tumors. BCL-2 upregulation was evident within the majority of lapatinib-treated HER2+/ER+ tumors and was often coupled with increased ER expression and decreased proliferation. Comparable BCL-2 upregulation was not observed within the trastuzumab-treated HER2+/ER+ tumors. Together, these results provide clinical validation of the BCL-2 induction associated with the adaptive response to lapatinib and support evaluation of BCL-2 inhibitors within the context of lapatinib and other HER2-targeted receptor tyrosine kinase inhibitors. [ABSTRACT FROM AUTHOR]

Background: Shared-usage high throughput screening (HTS) facilities are becoming more common in academe as large-scale small molecule and genome-scale RNAi screening strategies are adopted for basic research purposes. These shared facilities require a unique informatics infrastructure that must not only provide access to and analysis of screening data, but must also manage the administrative and technical challenges associated with conducting numerous, interleaved screening efforts run by multiple independent research groups. Results: We have developed Screensaver, a free, open source, web-based lab information management system (LIMS), to address the informatics needs of our small molecule and RNAi screening facility. Screensaver supports the storage and comparison of screening data sets, as well as the management of information about screens, screeners, libraries, and laboratory work requests. To our knowledge, Screensaver is one of the first applications to support the storage and analysis of data from both genome-scale RNAi screening projects and small molecule screening projects. Conclusions: The informatics and administrative needs of an HTS facility may be best managed by a single, integrated, web-accessible application such as Screensaver. Screensaver has proven useful in meeting the requirements of the ICCB-Longwood/NSRB Screening Facility at Harvard Medical School, and has provided similar benefits to other HTS facilities. [ABSTRACT FROM AUTHOR]

In pancreatic ductal adenocarcinoma cancer (PDAC) drug resistance is a severe clinical problem and patients relapse within a few months after receiving the standard-of-care chemotherapy. One contributing factor to treatment resistance is the desmoplastic nature of PDAC; the tumours are surrounded by thick layers of stroma composing up to 90% of the tumour mass. This stroma, which is mostly comprised of extracellular matrix (ECM) proteins, is secreted by cancer-associated fibroblasts (CAFs) residing in the tumour microenvironment. However, the mechanistic basis by which the tumour stroma directly contributes to chemoresistance remains unclear. Here, we show that CAF-secreted ECM proteins induce chemoresistance by blunting chemotherapy-induced DNA damage. Mechanistically, we identify N-myc downstream regulated gene 1 (NDRG1) as a key protein required for stroma-induced chemoresistance that responds to signals from the ECM and adhesion receptors. We further show that NDRG1 is a novel DNA repair protein that physically interacts with replication forks, maintains DNA replication and functions to resolve stalled forks caused by chemotherapy. More specifically, NDRG1 reduces R-loops, RNA-DNA hybrids that are known to cause genomic instability. R-loops occur during replication-transcription conflicts in S-phase and after chemotherapy treatments, thus posing a major threat to normal replication fork homeostasis. We identify NDRG1 as highly expressed in PDAC tumours, and its high expression correlates with chemoresistance and poor disease-specific survival. Importantly, knock-out of NDRG1 or inhibition of its phosphorylation restores chemotherapy-induced DNA damage and resensitizes tumour cells to treatment. In conclusion, our data reveal an unexpected role for CAF-secreted ECM proteins in enhancing DNA repair via NDRG1, a novel DNA repair protein, directly linking tumour stroma to replication fork homeostasis and R-loop biology, with important therapeutic implications for restoring DNA damage response pathways in pancreatic cancer., Summary Paragraph: Drug resistance is a severe clinical problem in stroma-rich tumours, such as pancreatic ductal adenocarcinoma (PDAC), and patients often relapse within a few months on chemotherapy 1-9 . The stroma, comprised of extracellular matrix (ECM) proteins, is secreted by cancer-associated fibroblasts (CAFs) residing in the tumour microenvironment 10-13 . Prior work show that ECM proteins provide survival benefits to cancer cells 14,15 . However, the precise role of CAF-secreted ECM in resistance to DNA damaging chemotherapies remains poorly understood. Here, we link ECM proteins to chemoresistance by enhanced DNA damage repair (DDR). Mechanistically, we identify N-myc downstream-regulated gene 1 (NDRG1) as a key effector downstream of ECM and the integrin-Src-SGK1-signalling axis that mediates enhanced DDR. We show that NDRG1 loss, mutation of conserved His194, or inhibition of NDRG1 phosphorylation by SGK1 lead to replication fork stalling, increased R-loops, and higher transcription-replication conflicts, resulting in genomic instability and sensitivity to chemotherapies. Our analysis of PDAC patient cohorts 16 found that high NDRG1 expression correlates with chemoresistance and poor patient survival. In conclusion, we uncover an unexpected role for CAF-secreted ECM proteins in promoting therapeutic resistance by enhancing DDR and establish NDRG1 as a novel DNA repair protein directly linking tumour stroma to DDR.

Cells rely on antioxidants to survive. The most abundant antioxidant is glutathione (GSH). The synthesis of GSH is non-redundantly controlled by the glutamate-cysteine ligase catalytic subunit (GCLC). GSH imbalance is implicated in many diseases, but the requirement for GSH in adult tissues is unclear. To interrogate this, we developed a series of in vivo models to induce Gclc deletion in adult animals. We find that GSH is essential to lipid abundance in vivo . GSH levels are reported to be highest in liver tissue, which is also a hub for lipid production. While the loss of GSH did not cause liver failure, it decreased lipogenic enzyme expression, circulating triglyceride levels, and fat stores. Mechanistically, we found that GSH promotes lipid abundance by repressing NRF2, a transcription factor induced by oxidative stress. These studies identify GSH as a fulcrum in the liver's balance of redox buffering and triglyceride production., Competing Interests: DECLARATION OF INTERESTS All other authors declare no competing interests.

Most patients with late-stage high-grade serous ovarian cancer (HGSOC) initially respond to chemotherapy but inevitably relapse and develop resistance, highlighting the need for novel therapies to improve patient outcomes. The MEK/ERK pathway is activated in a large subset of HGSOC, making it an attractive therapeutic target. Here, we systematically evaluated the extent of MEK/ERK pathway activation and efficacy of pathway inhibition in a large panel of well-annotated HGSOC patient-derived xenograft models. The vast majority of models were nonresponsive to the MEK inhibitor cobimetinib (GDC-0973) despite effective pathway inhibition. Proteomic analyses of adaptive responses to GDC-0973 revealed that GDC-0973 upregulated the proapoptotic protein BIM, thus priming the cells for apoptosis regulated by BCL2-family proteins. Indeed, combination of both MEK inhibitor and dual BCL-2/X L inhibitor (ABT-263) significantly reduced cell number, increased cell death, and displayed synergy in vitro in most models. In vivo , GDC-0973 and ABT-263 combination was well tolerated and resulted in greater tumor growth inhibition than single agents. Detailed proteomic and correlation analyses identified two subsets of responsive models-those with high BIM at baseline that was increased with MEK inhibition and those with low basal BIM and high pERK levels. Models with low BIM and low pERK were nonresponsive. Our findings demonstrate that combined MEK and BCL-2/X L inhibition has therapeutic activity in HGSOC models and provide a mechanistic rationale for the clinical evaluation of this drug combination as well as the assessment of the extent to which BIM and/or pERK levels predict drug combination effectiveness in chemoresistant HGSOC., (©2019 American Association for Cancer Research.)