Agonistic anti-CD148 monoclonal antibody attenuates diabetic nephropathy in mice.

CD148 is a transmembrane protein tyrosine phosphatase (PTP) that is expressed in the renal vasculature, including the glomerulus. Previous studies have shown that CD148 plays a role in the negative regulation of growth factor signals (including epidermal growth factor and vascular endothelial growth factor), suppressing cell proliferation and transformation. However, the role of CD148 in kidney disease remains unknown. Here, we generated an agonistic anti-CD148 antibody and evaluated its effects in murine diabetic nephropathy (DN). Monoclonal antibodies (mAbs) against the mouse CD148 ectodomain sequence were generated by immunizing CD148 knockout (CD148KO) mice. The mAbs that increased CD148 activity were selected by biological (proliferation) and biochemical (PTP activity) assays. The mAb (18E1) that showed strong agonistic activity was injected (10 mg/kg ip) in streptozotocin-induced wild-type and CD148KO diabetic mice for 6 wk, and the renal phenotype was then assessed. The effects of 18E1 mAb in podocyte growth factor signals were also assessed in culture. Compared with control IgG, 18E1 mAb significantly decreased albuminuria and mesangial expansion without altering hyperglycemia and blood pressure in wild-type diabetic mice. Immunohistochemical evaluation showed that 18E1 mAb significantly prevented the reduction of podocyte number and nephrin expression and decreased glomerular fibronectin expression and renal macrophage infiltration. The 18E1 mAb showed no effects in CD148KO diabetic mice. Furthermore, we demonstrated that 18E1 mAb reduces podocyte epidermal growth factor receptor signals in culture and in diabetic mice. These findings suggest that agonistic anti-CD148 mAb attenuates DN in mice, in part by reducing epidermal growth factor receptor signals in podocytes. This antibody may be used for the treatment of early DN.

Sustained activation of the HER1-ERK1/2-RSK signaling pathway controls myoepithelial cell fate in human mammary tissue.

Human mammary glands arise from multipotent progenitor cells, which likely respond both to cell-autonomous and to extrinsic cues. However, the identity of these cues and how they might act remain unclear. We analyzed HER1 ligand effects on mammary morphogenesis using a three-dimensional organoid model generated from human breast tissue that recapitulates both qualitatively and quantitatively the normal ductal network in situ. Strikingly, different HER1 ligands generate distinct patterns of cell fate. Epidermal growth factor (EGF) causes a massive expansion of the myoepithelial lineage. Amphiregulin, in contrast, enables normal ductal development. These differences cannot be ascribed to preferential apoptosis or proliferation of differentiated cell populations, but are dependent on HER1 signal intensity. Inhibition of the extracellular signal-regulated kinase 1/2 (ERK1/2) effector RSK prevents the EGF-induced myoepithelial expansion. Notably, mouse mammary organoids are much less responsive to HER1 ligands. Little is known about the myoepithelial lineage or about growth factor effects on mammary progenitor differentiation, and our studies provide an important window into human mammary development that reveals unexpected differences from the mouse model.

Transcriptome analysis of Haloquadratum walsbyi: vanity is but the surface.

BACKGROUND: Haloquadratum walsbyi dominates saturated thalassic lakes worldwide where they can constitute up to 80-90% of the total prokaryotic community. Despite the abundance of the enigmatic square-flattened cells, only 7 isolates are currently known with 2 genomes fully sequenced and annotated due to difficulties to grow them under laboratory conditions. We have performed a transcriptomic analysis of one of these isolates, the Spanish strain HBSQ001 in order to investigate gene transcription under light and dark conditions. RESULTS: Despite a potential advantage for light as additional source of energy, no significant differences were found between light and dark expressed genes. Constitutive high gene expression was observed in genes encoding surface glycoproteins, light mediated proton pumping by bacteriorhodopsin, several nutrient uptake systems, buoyancy and storage of excess carbon. Two low expressed regions of the genome were characterized by a lower codon adaptation index, low GC content and high incidence of hypothetical genes. CONCLUSIONS: Under the extant cultivation conditions, the square hyperhalophile devoted most of its transcriptome towards processes maintaining cell integrity and exploiting solar energy. Surface glycoproteins are essential for maintaining the large surface to volume ratio that facilitates light and organic nutrient harvesting whereas constitutive expression of bacteriorhodopsin warrants an immediate source of energy when light becomes available.

Codependent functions of RSK2 and the apoptosis-promoting factor TIA-1 in stress granule assembly and cell survival.

Stress granules aid cell survival in response to environmental stressors by acting as sites of translational repression. We report an unanticipated link between stress granules and the serine/threonine kinase RSK2. In stressed breast cells, endogenous RSK2 colocalizes in granules with TIA-1 and poly(A)-binding protein 1, and the sequestration of RSK2 and TIA-1 exhibits codependency. The RSK2 N-terminal kinase domain controls the direct interaction with the prion-related domain of TIA-1. Silencing RSK2 decreases cell survival in response to stress. Mitogen releases RSK2 from the stress granules and permits its nuclear import via a nucleocytoplasmic shuttling sequence in the C-terminal domain. Nuclear accumulation is dependent on TIA-1. Surprisingly, nuclear localization of RSK2 is sufficient to enhance proliferation through induction of cyclin D1, in the absence of other active signaling pathways. Hence, RSK2 is a pivotal factor linking the stress response to survival and proliferation.

The contribution of endophytic bacteria to Albizia lebbeck-mediated phytoremediation of tannery effluent contaminated soil.

Toxicity of chromium often impairs the remediation capacity of plants used in phytoremediation of polluted soils. In this study, we have identified Albizia lebbeck as a prospective chromium hyperaccumulator and examined cultivable diversity of endophytes present in chromium-treated and control saplings. High numbers (22-100%) of endophytic bacteria, isolated from root, stem, and leaf tissues, could tolerate elevated (1-3 mM) concentrations of K2CrO7. 16S rRNA gene sequence-based phylogenetic analysis showed that the 118 isolates obtained comprised of 17 operational taxonomic units affiliated with the proteobacterial genera Rhizobium (18%), Marinomonas (1%), Pseudomonas (16%), and Xanthomonas (7%) but also with members of Firmicutes genera, such as Bacillus (35%) and Salinococcus (3%). The novel isolates belonging to Salinococcus and Bacillus could tolerate high K2CrO7 concentrations (3 mM) and also showed elevated activity of chromate reductase. In addition, majority (%) of the endophytic isolates also showed production of indole-3-acetic acid. Taken together, our results indicate that the innate endophytic bacterial community assists plants in reducing heavy metal toxicity.

Diversity of the cell-wall associated genomic island of the archaeon Haloquadratum walsbyi.

BACKGROUND: Haloquadratum walsbyi represents up to 80% of cells in NaCl-saturated brines worldwide, but is notoriously difficult to maintain under laboratory conditions. In order to establish the extent of genetic diversity in a natural population of this microbe, we screened a H. walsbyi enriched metagenomic fosmid library and recovered seven novel version of its cell-wall associated genomic island. The fosmid inserts were sequenced and analysed. RESULTS: The novel cell-wall associated islands delineated two major clades within H. walsbyi. The islands predominantly contained genes putatively involved in biosynthesis of surface layer, genes encoding cell surface glycoproteins and genes involved in envelope formation. We further found that these genes are maintained in the population and that the diversity of this region arises through homologous recombination but also through the action of mobile genetic elements, including viruses. CONCLUSIONS: The population of H. walsbyi in the studied saltern brine is composed of numerous clonal lineages that differ in surface structures including the cell wall. This type of variation probably reflects a number of mechanisms that minimize the infection rate of predating viruses.

Low Bacterial Diversity and Nitrate Levels in Cores from Deep Boreholes in Pristine Karst.

This study investigates the nitrate gradients within the deep biosphere of karst carbonate rocks and their resident microbiota. Samples were taken from borehole cores at depths down to 350 m below the surface, collected during geological site investigations for proposed railway tunnels and analysed using 16S rRNA amplicon sequencing. 16S rRNA amplicon sequencing analysis revealed relatively low microbial diversity, which can serve as a reliable indicator of the pristine nature of deep karst. However, some local hotspots of diversity are independent of depth. Pseudomonadota dominated the samples, with Gammaproteobacteria dominating at the class level. The low nitrate content in deep karst, in contrast to higher values closer to the surface, serves as an additional marker of its undisturbed and unpolluted status. Based on the prediction of functional profiles from 16S rRNA sequencing data, nitrates remain low due to indigenous microbial denitrification and assimilatory nitrate reduction. Pathways related to nitrogen fixation, ammonia assimilation, and nitrification were not confirmed. When elevated nitrate levels are observed in karst, they are most probably related to anthropogenic activities. Environmental factors other than depth and nitrate content play an important role in shaping bacterial communities.

Structure and community composition of sprout-like bacterial aggregates in a Dinaric Karst subterranean stream.

The Vjetrenica cave in the Dinaric Karst hosts a worldwide extraordinarily high cave biodiversity. Beside a diverse and specialized cave fauna, sprout-like formations attached to the bed of the cave stream were observed and described, but not further characterized, almost a century ago. Here we investigated these sprout-like microbial aggregates by the rRNA approach and detailed microscopy. Based on fluorescence in situ hybridization and ultrastructural analysis, the sprout-like formations are morphologically highly organized, and their core consists of a member of a novel deep-branching lineage in the bacterial phylum Nitrospirae. This organism displays an interesting cellular ultrastructure with different kinds of cytoplasmic inclusions and is embedded in a thick extracellular matrix, which contributes to the stability and shape of the aggregates. This novel bacterium has been provisionally classified as "Candidatus Troglogloea absoloni." The surface of the sprout-like aggregates is more diverse than the core. It is colonized by a bacterial biofilm consisting primarily of filamentous Betaproteobacteria, whereas other microbial populations present in the crust include members of the Bacteriodetes, Gammaproteobacteria, Actinombacteria, Alphaproteobacteria, and Planctomycetes, which are intermingled with mineral inclusions. This study represents the first thorough molecular and ultrastructural characterization of the elusive sprout-like bacterial aggregates, which are also found in other cave systems of the Dinaric Karst. The discovery of Ca. Troglogloea absoloni contributes to the known biodiversity of subterranean ecosystems and especially of macroscopic structures formed in caves by microorganisms, whose composition and ecological function often remain enigmatic.

A toolbox of machine learning software to support microbiome analysis.

The human microbiome has become an area of intense research due to its potential impact on human health. However, the analysis and interpretation of this data have proven to be challenging due to its complexity and high dimensionality. Machine learning (ML) algorithms can process vast amounts of data to uncover informative patterns and relationships within the data, even with limited prior knowledge. Therefore, there has been a rapid growth in the development of software specifically designed for the analysis and interpretation of microbiome data using ML techniques. These software incorporate a wide range of ML algorithms for clustering, classification, regression, or feature selection, to identify microbial patterns and relationships within the data and generate predictive models. This rapid development with a constant need for new developments and integration of new features require efforts into compile, catalog and classify these tools to create infrastructures and services with easy, transparent, and trustable standards. Here we review the state-of-the-art for ML tools applied in human microbiome studies, performed as part of the COST Action ML4Microbiome activities. This scoping review focuses on ML based software and framework resources currently available for the analysis of microbiome data in humans. The aim is to support microbiologists and biomedical scientists to go deeper into specialized resources that integrate ML techniques and facilitate future benchmarking to create standards for the analysis of microbiome data. The software resources are organized based on the type of analysis they were developed for and the ML techniques they implement. A description of each software with examples of usage is provided including comments about pitfalls and lacks in the usage of software based on ML methods in relation to microbiome data that need to be considered by developers and users. This review represents an extensive compilation to date, offering valuable insights and guidance for researchers interested in leveraging ML approaches for microbiome analysis.

The effects of CD148 Q276P/R326Q polymorphisms in A431D epidermoid cancer cell proliferation and epidermal growth factor receptor signaling.

BACKGROUND: CD148 is a transmembrane protein tyrosine phosphatase that is expressed in multiple cell types. Previous studies have shown that CD148 dephosphorylates growth factor receptors and their signaling molecules, including EGFR and ERK1/2, and negatively regulates cancer cell growth. Furthermore, research of clinical patients has shown that highly linked CD148 gene polymorphisms, Gln276Pro (Q276P) and Arg326Gln (R326Q), are associated with an increased risk of several types of cancer. However, the biological effects of these missense mutations have not been studied. AIM: We aimed to determine the biological effects of CD148 Q276P/R326Q mutations in cancer cell proliferation and growth factor signaling, with emphasis on EGFR signaling. METHODS: CD148 forms, wild-type (WT) or Q276P/R326Q, were retrovirally introduced into A431D epidermoid carcinoma cells that lacks CD148 expression. The stable cells that express comparable levels of CD148 were sorted by flow cytometry. A431D cells infected with empty retrovirus was used as a control. CD148 localization, cell proliferation rate, EGFR signaling, and the response to thrombospondin-1 (TSP1), a CD148 ligand, were assessed by immunostaining, cell proliferation assay, enzyme-linked immunosorbent assay, and Western blotting. RESULTS: Both CD148 forms (WT, Q276P/R326Q) were distributed to cell surface and all three cell lines expressed same level of EGFR. Compared to control cells, the A431D cells that express CD148 forms showed significantly lower cell proliferation rates. EGF-induced EGFR and ERK1/2 phosphorylation as well as cell proliferation were also significantly reduced in these cells. Furthermore, TSP1 inhibited cell proliferation in CD148 (WT, Q276P/R326Q)-expressing A431D cells, while it showed no effects in control cells. However, significant differences were not observed between CD148 WT and Q276P/R326Q cells. CONCLUSION: Our data demonstrates that Q276P/R326Q mutations do not have major effects on TSP1-CD148 interaction as well as on CD148's cellular localization and activity to inhibit EGFR signaling and cell proliferation.

Adapting to environmental changes using specialized paralogs.

When a bacterial species survives under changing environmental circumstances (e.g. salinity or temperature), its proteins might not function in all physicochemical conditions. We propose that prokaryotes cope with this problem by having two or more copies of the genes affected by environmental fluctuations, each one performing the same function under different conditions (i.e. ecoparalog). We identify potential examples in the bacterium Salinibacter ruber and in other species that experience wide environmental variations.

Extraction, purification and characterization of a protease from Micrococcus sp. VKMM 037.

The haloalkaliphilic bacterium Micrococcus sp. VKMM 037, isolated from an effluent of the caustic soda industry, was found to produce a protease. Maximal proteolytic activity was observed in cell culture grown at 40 degrees C using 2% (w/v) glycerol, 2% (w/v) beef extract and 2% (w/v) peptone as nutrients in medium also containing 0.85 M NaCl with a pH of 10.0. An efficient purification procedure combining ammonium sulphate precipitation and Q-Sepharose ion-exchange chromatography was developed. The purified 41 kDa protease was stable in a temperature range between 20 degrees C and 60 degrees C. The protease remained active over a wide range of pH values (4.0-12.0) and NaCl concentrations (0-3.42 M) with an optimum at pH 10.0 and 0.85 M NaCl, respectively. Furthermore, the enzyme remained stable or was only marginally inhibited in the presence of various organic solvents, surfactants and reducing agents. The purified protease of Micrococcus sp. VKMM 037 efficiently removed blood stains within 40 minutes of treatment. Given the biochemical characteristics determined, this novel protease could be exploited as an additive in the detergent industry and also for the synthesis of biomolecules and the degradation of protein.

Optimization of growth medium for protease production by Haloferax Lucentensis VKMM 007 by response surface methodology.

The production of halophilic thermostable protease by Haloferax lucentensis VKMM 007 was optimized using a statistical approach. In accordance with factorial design, soluble starch, gelatin, KCl and MgSO4 were selected among 27 variables tested. Next, a second-order quadratic model was estimated and optimal medium concentrations were determined based on quadratic regression equation generated by model. These were 5.14 g L(-1) of KCl, 6.57 g L(-1)of MgSO4, 9.05 g L(-1)of gelatin and 5.27 g L(-1)of soluble starch in high salts media supplemented with 0.5% (w/v) of beef extract and peptone, respectively. In these optimal conditions, the obtained protease concentration of 6.80 U mL(-1) was in agreement with the predicted protease concentration and was further improved to 7.02 U mL(-1) by increasing the concentration of NaCl in the medium to 25% (w/v). An overall 4.0-fold increase in protease production was achieved in the optimized medium compared to activity obtained in initial medium.

Direct Detection of Isolevuglandins in Tissues using a D11 scFv-Alkaline Phosphatase Fusion Protein and Immunofluorescence.

Isolevuglandins (IsoLGs) are highly reactive gamma ketoaldehydes formed from H2-isoprostanes through lipid peroxidation and crosslink proteins leading to inflammation and various diseases including hypertension. Detection of IsoLG accumulation in tissues is crucial in shedding light on their involvement in the disease processes. However, measurement of IsoLGs in tissues is extremely difficult, and currently available tools, including mass spectrometry analysis, are laborious and extremely expensive. Here we describe a novel method for in situ detection of IsoLGs in tissues using alkaline phosphatase-conjugated D11 ScFv and a recombinant phage-display antibody produced in E. coli by immunofluorescent microscopy. Four controls were used for validating the staining: (1) staining with and without D11, (2) staining with bacterial periplasmic extract with the alkaline phosphatase linker, (3) irrelevant scFV antibody staining, and (4) competitive control with IsoLG prior to the staining. We demonstrate the effectiveness of the alkaline phosphatase-conjugated D11 in both human and mouse tissues with or without hypertension. This method will likely serve as an important tool to study the role of IsoLGs in a wide variety of disease processes.

Microbiota entrapped in recently-formed ice: Paradana Ice Cave, Slovenia.

Paradana is one of the biggest ice caves in Slovenia, with an estimated ice volume of 8,000 m3. Reflecting climatological conditions, the cave ice undergoes repeated freeze-thaw cycles and regular yearly deposition of fresh ice. Three distinct ice block samples, collected from the frozen lake in May 2016, were analysed to obtain data on ice physicochemical properties and the composition of associated microbiota. Isotopic composition of the ice samples (18O, 2H) and a local meteoric water line (LMWL) constructed for monthly precipitation at Postojna were used to estimate the isotopic composition of the water that formed the ice, which had high values of deuterium excess and low concentrations of chloride, sulphate and nitrate. The values of total organic carbon (1.93-3.95 mg/l) within the ice blocks fall within the range of those measured in karst streams. Total cell count in the ice was high and the proportion of cell viability increased along the depth gradient and ranged from 4.67 × 104 to 1.52 × 105 cells/ml and from 51.0 to 85.4%, respectively. Proteobacteria represented the core of the cave-ice microbiome (55.9-79.1%), and probably play an essential role in this ecosystem. Actinobacteria was the second most abundant phylum (12.0-31.4%), followed in abundance by Bacteroidetes (2.8-4.3%). Ice phylotypes recorded amounted to 442 genera, but only 43 genera had abundances greater than 0.5%. Most abundant were Pseudomonas, a well-known ice dweller, and Lysobacter, which previously was not reported in this context. Finally, two xanthophytes, Chloridella glacialis and Ellipsoidion perminimum, known from polar environments, were cultured from the ice. This indicates that the abundance and ecological role of phototrophs in such environments might be greater than previously deduced.

Statistical and Machine Learning Techniques in Human Microbiome Studies: Contemporary Challenges and Solutions.

The human microbiome has emerged as a central research topic in human biology and biomedicine. Current microbiome studies generate high-throughput omics data across different body sites, populations, and life stages. Many of the challenges in microbiome research are similar to other high-throughput studies, the quantitative analyses need to address the heterogeneity of data, specific statistical properties, and the remarkable variation in microbiome composition across individuals and body sites. This has led to a broad spectrum of statistical and machine learning challenges that range from study design, data processing, and standardization to analysis, modeling, cross-study comparison, prediction, data science ecosystems, and reproducible reporting. Nevertheless, although many statistics and machine learning approaches and tools have been developed, new techniques are needed to deal with emerging applications and the vast heterogeneity of microbiome data. We review and discuss emerging applications of statistical and machine learning techniques in human microbiome studies and introduce the COST Action CA18131 "ML4Microbiome" that brings together microbiome researchers and machine learning experts to address current challenges such as standardization of analysis pipelines for reproducibility of data analysis results, benchmarking, improvement, or development of existing and new tools and ontologies.

Metagenomic islands of hyperhalophiles: the case of Salinibacter ruber.

BACKGROUND: Saturated brines are extreme environments of low diversity. Salinibacter ruber is the only bacterium that inhabits this environment in significant numbers. In order to establish the extent of genetic diversity in natural populations of this microbe, the genomic sequence of reference strain DSM 13855 was compared to metagenomic fragments recovered from climax saltern crystallizers and obtained with 454 sequencing technology. This kind of analysis reveals the presence of metagenomic islands, i.e. highly variable regions among the different lineages in the population. RESULTS: Three regions of the sequenced isolate were scarcely represented in the metagenome thus appearing to vary among co-occurring S. ruber cells. These metagenomic islands showed evidence of extensive genomic corruption with atypically low GC content, low coding density, high numbers of pseudogenes and short hypothetical proteins. A detailed analysis of island gene content showed that the genes in metagenomic island 1 code for cell surface polysaccharides. The strain-specific genes of metagenomic island 2 were found to be involved in biosynthesis of cell wall polysaccharide components. Finally, metagenomic island 3 was rich in DNA related enzymes. CONCLUSION: The genomic organisation of S. ruber variable genomic regions showed a number of convergences with genomic islands of marine microbes studied, being largely involved in variable cell surface traits. This variation at the level of cell envelopes in an environment devoid of grazing pressure probably reflects a global strategy of bacteria to escape phage predation.

High dietary salt-induced dendritic cell activation underlies microbial dysbiosis-associated hypertension.

Excess dietary salt contributes to inflammation and hypertension via poorly understood mechanisms. Antigen presenting cells including dendritic cells (DCs) play a key role in regulating intestinal immune homeostasis in part by surveying the gut epithelial surface for pathogens. Previously, we found that highly reactive γ-ketoaldehydes or isolevuglandins (IsoLGs) accumulate in DCs and act as neoantigens, promoting an autoimmune-like state and hypertension. We hypothesized that excess dietary salt alters the gut microbiome leading to hypertension and this is associated with increased immunogenic IsoLG-adduct formation in myeloid antigen presenting cells. To test this hypothesis, we performed fecal microbiome analysis and measured blood pressure of healthy human volunteers with salt intake above or below the American Heart Association recommendations. We also performed 16S rRNA analysis on cecal samples of mice fed normal or high salt diets. In humans and mice, high salt intake was associated with changes in the gut microbiome reflecting an increase in Firmicutes, Proteobacteria and genus Prevotella bacteria. These alterations were associated with higher blood pressure in humans and predisposed mice to vascular inflammation and hypertension in response to a sub-pressor dose of angiotensin II. Mice fed a high salt diet exhibited increased intestinal inflammation including the mesenteric arterial arcade and aorta, with a marked increase in the B7 ligand CD86 and formation of IsoLG-protein adducts in CD11c+ myeloid cells. Adoptive transfer of fecal material from conventionally housed high salt-fed mice to germ-free mice predisposed them to increased intestinal inflammation and hypertension. These findings provide novel insight into the mechanisms underlying inflammation and hypertension associated with excess dietary salt and may lead to interventions targeting the microbiome to prevent and treat this important disease.

"Deciphering Archaeal Communities" Omics Tools in the Study of Archaeal Communities.

Archaea constitute one of the three recognized phylogenetic groups of organisms living on the planet, and the latest to be discovered. Most Archaea resist cultivation and are studied using molecular methods. High-throughput amplicon sequencing and metagenomic approaches have been key in uncovering hitherto unknown archaeal diversity, their metabolic potential, and have even provided an insight into genomes of a number of uncultivated members of this group. Here, we summarize protocols describing sampling, molecular, metagenomic, and metatranscriptomic analyses as well as bioinformatics approaches that have proved useful for the study of archaea in natural samples.

Development of a RSK Inhibitor as a Novel Therapy for Triple-Negative Breast Cancer.

Metastatic breast cancer is an incurable disease and identification of novel therapeutic opportunities is vital. Triple-negative breast cancer (TNBC) frequently metastasizes and high levels of activated p90RSK (RSK), a downstream MEK-ERK1/2 effector, are found in TNBC. We demonstrate, using direct pharmacologic and genetic inhibition of RSK1/2, that these kinases contribute to the TNBC metastatic process in vivo Kinase profiling showed that RSK1 and RSK2 are the predominant kinases targeted by the new inhibitor, which is based on the natural product SL0101. Further evidence for selectivity was provided by the observations that silencing RSK1 and RSK2 eliminated the ability of the analogue to further inhibit survival or proliferation of a TNBC cell line. In vivo, the new derivative was as effective as the FDA-approved MEK inhibitor trametinib in reducing the establishment of metastatic foci. Importantly, inhibition of RSK1/2 did not result in activation of AKT, which is known to limit the efficacy of MEK inhibitors in the clinic. Our results demonstrate that RSK is a major contributor to the TNBC metastatic program and provide preclinical proof-of-concept for the efficacy of the novel SL0101 analogue in vivo Mol Cancer Ther; 15(11); 2598-608. ©2016 AACR.