SUMO2 Inhibition Reverses Aberrant Epigenetic Rewiring Driven by Synovial Sarcoma Fusion Oncoproteins and Impairs Sarcomagenesis.

Synovial Sarcoma (SySa) is an aggressive soft tissue sarcoma that accounts for 5 - 10% of all soft tissue sarcomas. Current treatment involves radiation and radical surgery including limb amputation, highlighting the urgent need to develop targeted therapies. We reasoned that transcriptional rewiring by the fusion protein SS18-SSX, the sole oncogenic driver in SySa, creates specific vulnerabilities that can be exploited for treatment. To uncover genes that are selectively essential for SySa, we mined The Cancer Dependency Map (DepMap) data to identify genes that specifically impact the fitness of SySa compared to other tumor cell lines. Targeted CRISPR library screening of SySa-selective candidates revealed that the small ubiquitin-like modifier 2 (SUMO2) was one of the strongest dependencies both in vitro as well as in vivo. TAK-981, a clinical-stage small molecule SUMO2 inhibitor potently inhibited growth and colony-forming ability. Strikingly, transcriptomic studies showed that pharmacological SUMO2 inhibition with TAK-981 treatment elicited a profound reversal of a gene expression program orchestrated by SS18-SSX fusions. Of note, genetic or pharmacological SUMO2 inhibition reduced global and chromatin levels of the SS18-SSX fusion protein with a concomitant reduction in histone 2A lysine 119 ubiquitination (H2AK119ub), an epigenetic mark that plays an important role in SySa pathogenesis. Taken together, our studies identify SUMO2 as a novel, selective vulnerability in SySa. Since SUMO2 inhibitors are currently in Phase 1/2 clinical trials for other cancers, our findings present a novel avenue for targeted treatment of synovial sarcoma.

Detection of Genomic Structural Variations Associated with Drug Sensitivity and Resistance in Acute Leukemia.

Acute leukemia is a particularly problematic collection of hematological cancers, and, while somewhat rare, the survival rate of patients is typically abysmal without bone marrow transplantation. Furthermore, traditional chemotherapies used as standard-of-care for patients cause significant side effects. Understanding the evolution of leukemia to identify novel targets and, therefore, drug treatment regimens is a significant medical need. Genomic rearrangements and other structural variations (SVs) have long been known to be causative and pathogenic in multiple types of cancer, including leukemia. These SVs may be involved in cancer initiation, progression, clonal evolution, and drug resistance, and a better understanding of SVs from individual patients may help guide therapeutic options. Here, we show the utilization of optical genome mapping (OGM) to detect known and novel SVs in the samples of patients with leukemia. Importantly, this technology provides an unprecedented level of granularity and quantitation unavailable to other current techniques and allows for the unbiased detection of novel SVs, which may be relevant to disease pathogenesis and/or drug resistance. Coupled with the chemosensitivities of these samples to FDA-approved oncology drugs, we show how an impartial integrative analysis of these diverse datasets can be used to associate the detected genomic rearrangements with multiple drug sensitivity profiles. Indeed, an insertion in the gene MUSK is shown to be associated with increased sensitivity to the clinically relevant agent Idarubicin, while partial tandem duplication events in the KMT2A gene are related to the efficacy of another frontline treatment, Cytarabine.

Early life programming by diet can play a role in risk reduction of otitis in dogs.

Introduction: Otitis in dogs is often chronic while local treatment primarily consists of flushing, antibiotics, and/or antifungals. We were interested in finding early life variables that associate with otitis later in life, preferably some that could be modified. Methods: A cross-sectional hypothesis-driven study with longitudinal data was performed to search for associations between pre- and postnatal exposures, and the incidence of owner-reported otitis in dogs at over 1 year of age. The multivariate logistic regression analysis study included data from 3,064 dogs and explored 26 different early life variables at four early life stages: prenatal, neonatal, postnatal, and puppyhood. We compared two feeding patterns, a non-processed meat-based diet (NPMD, raw) and an ultra-processed carbohydrate-based diet (UPCD, dry). Results: We report that eating a NPMD diet significantly decreased the risk of otitis later in life, while eating a UPCD diet significantly increased the risk. This was seen in different life stages of mother or puppy: The maternal diet during pregnancy (p=0.011) and the puppies' diet from 2 to 6 months of age (p=0.019) were both significantly associated with otitis incidence later in life, whereas the puppies' first solid diet, was associated in the same way, but did not reach significance (p=0.072). Also, analyzing food ratios showed that when puppies were consuming >25% of their food as NPMD it significantly decreased their incidence of otitis later in life, while a ratio of >75% UPCD in their diet significantly increased their risk of otitis. Also, if the dog was born in the current family, was exposed to sunlight for more than 1 hour daily, and was raised on a dirt floor during puppyhood, there was a lower risk of otitis development later in life. Discussion: The findings only suggest causality, and further studies are required. However, we propose that veterinarians, breeders, and owners can impact otitis risk by modifying factors such as diet and environment.

Assessing the association between supplemented puppyhood dietary fat sources and owner-reported epilepsy in adulthood, among Finnish companion dogs.

Introduction: Epilepsy is a serious and common neurological condition in dogs, despite the wide number of antiepileptic drugs available, in approximately one third of the patients, epilepsy remains unsatisfactorily controlled. We aim to analyze whether feeding dietary fat sources during puppyhood was associated with canine epilepsy in adulthood. Methods: A nested case-control study was compiled from the validated DogRisk food frequency questionnaire (DogRisk FFQ). DogRisk FFQ collected feeding, disease, and background data about the dog. The study sample consisted of 108 owner-reported epileptic cases and 397 non-epileptic controls. Each case was matched with up to four controls for the key confounding factors of sex, breed, and age. We analyzed associations between feeding as a puppy and owner-reported epilepsy as an adult dog using Cox regression. We tested 55 different food variables. Results: We found that feeding fish fat from dietary sources at least once a week during puppyhood was inversely associated with epilepsy in later life in the unadjusted analysis [OR 0.46 (95% CI 0.25-0.83), p=0.01], while when adjusting for keeping conditions and dog characteristics the association was [OR 0.45 (95% CI 0.23-0.88), p=0.02]. When adjusted for keeping conditions, dog characteristics, and other feeding factors, the association was of similar magnitude but not significance [OR 0.56 (95% CI 0.27-1.15), p=0.12]. Discussion: The study indicates possible protective associations of feeding the dog with dietary sources of fish fat against epilepsy, although the result could be confounded by other feeding factors. Findings are compatible with current knowledge regarding the role of omega-3 fatty acids and ketogenic diet, a low carbohydrate, high fat diet as supportive treatments of epilepsy. As our findings are based on observations, we suggest the possibility of causality but do not prove it. Dietary intervention studies should now be conducted to confirm our findings.

Multi-level assessment of the origin, feeding area and organohalogen contamination on salmon from the Baltic Sea.

The Atlantic salmon (Salmo salar) population in the Baltic Sea consists of wild and hatchery-reared fish that have been released into the sea to support salmon stocks. During feeding migration, salmon migrate to different parts of the Baltic Sea and are exposed to various biotic and abiotic stressors, such as organohalogen compounds (OHCs). The effects of salmon origin (wild or hatchery-reared), feeding area (Baltic Main Basin, Bothnian Sea, and Gulf of Finland), and OHC concentration on the differences in hepatic proteome of salmon were investigated. Multi-level analysis of the OHC concentration, transcriptome, proteome, and oxidative stress biomarkers measured from the same salmon individuals were performed to find the key variables (origin, feeding area, OHC concentrations, and oxidative stress) that best account for the differences in the transcriptome and proteome between the salmon groups. When comparing wild and hatchery-reared salmon, differences were found in xenobiotic and amino acid metabolism-related pathways. When comparing salmon from different feeding areas, the amino acid and carbohydrate metabolic pathways were notably different. Several proteins found in these pathways are correlated with the concentrations of polychlorinated biphenyls (PCBs). The multi-level analysis also revealed amino acid metabolic pathways in connection with PCBs and oxidative stress variables related to glutathione metabolism. Other pathways found in the multi-level analysis included genetic information processes related to ribosomes, signaling and cellular processes related to the cytoskeleton, and the immune system, which were connected mainly to the concentrations of Polychlorinated biphenyls and Dichlorodiphenyltrichloroethane and their metabolites. These results suggest that the hepatic proteome of salmon in the Baltic Sea, together with the transcriptome, is more affected by the OHC concentrations and oxidative stress of the feeding area than the origin of the salmon.

Mineral, trace element, and toxic metal concentration in hair from dogs with idiopathic epilepsy compared to healthy controls.

BACKGROUND: Altered trace element status is associated with epilepsy in humans and dogs with idiopathic epilepsy (IE). OBJECTIVES: Compare hair element concentrations in epileptic and healthy dogs. ANIMALS: Sixty-three dogs with IE (53 treated, 10 untreated) and 42 controls. METHODS: Case-control study using ICP-MS to determine hair calcium, magnesium, phosphorus, sodium, potassium, iron, copper, manganese, zinc, selenium, chromium, lead, mercury, cadmium, arsenic, aluminum, and nickel concentration. Groups were compared using nonparametric tests. Results were controlled for diet, sex, age, and hair color using generalized linear mixed models. RESULTS: Compared to healthy controls, dogs with IE had lower hair phosphorus (mean ± SD; IE: 286.19 ± 69.62 µg/g, healthy: 324.52 ± 58.69 µg/g; P = .001), higher hair copper (IE: 10.97 ± 3.51 µg/g, healthy: 8.41 ± 1.27 µg/g; P < .001), zinc (IE: 158.25 ± 19.64 µg/g, healthy: 144.76 ± 32.18 µg/g; P < .001), copper/zinc ratio (IE: 0.07 ± 0.02, healthy: 0.06 ± 0.01; P = .003), selenium (IE: 1.65 ± 0.43 µg/g, healthy: 0.94 ± 0.73 µg/g; P < .001), and arsenic (IE: 0.40 ± 0.78 µg/g, healthy: 0.05 ± 0.08 µg/g; P < .001). When comparing treated and untreated epileptic dogs with healthy dogs, the differences in phosphorus and selenium remained significant for both groups, whereas the differences in copper, zinc, and arsenic were significant only for treated dogs. Potassium bromide treatment was strongly associated with high hair arsenic (P = .000). CONCLUSIONS AND CLINICAL IMPORTANCE: Altered trace element status could be involved in the pathophysiology of IE in dogs. Antiseizure drugs might affect trace element and arsenic metabolism.

The effect of puppyhood and adolescent diet on the incidence of chronic enteropathy in dogs later in life.

Diet has a key role in the homeostasis of the gut microenvironment, influencing the microbiome, the gut barrier, host immunity and gut physiology. Yet, there is little information on the role of early diet in the onset of inflammatory gastrointestinal disorders later in life, especially in dogs. Therefore, the aim of the present cross-sectional, epidemiological study with longitudinal data, was to explore associations of companion dogs' early life diet style and food items with owner-reported chronic enteropathy (CE) incidence in later life. Food frequency questionnaire data from Finnish companion dogs was analyzed using principal component analysis and logistic regression. We found that feeding a non-processed meat-based diet and giving the dog human meal leftovers and table scraps during puppyhood (2-6 months) and adolescence (6-18 months) were protective against CE later in life. Especially raw bones and cartilage as well as leftovers and table scraps during puppyhood and adolescence, and berries during puppyhood were associated with less CE. In contrast, feeding an ultra-processed carbohydrate-based diet, namely dry dog food or "kibble" during puppyhood and adolescence, and rawhides during puppyhood were significant risk factors for CE later in life.

Single-cell multiomics reveals the complexity of TGFß signalling to chromatin in iPSC-derived kidney organoids.

TGFß1 plays a regulatory role in the determination of renal cell fate and the progression of renal fibrosis. Here we show an association between SMAD3 and the histone methyltransferase, EZH2, during cell differentiation; ChIP-seq revealed that SMAD3 and EZH2 co-occupy the genome in iPSCs and in iPSC-derived nephron progenitors. Through integration of single cell gene expression and epigenome profiling, we identified de novo ACTA2+ve/POSTN+ve myofibroblasts in kidney organoids treated with TGFß1, characterised by increased SMAD3-dependent cis chromatin accessibility and gene expression associated with fibroblast activation. We have identified fibrosis-associated regulons characterised by enrichment of SMAD3, AP1, the ETS family of transcription factors, and NUAK1, CREB3L1, and RARG, corresponding to enriched motifs at accessible loci identified by scATACseq. Treatment with the EZH2 specific inhibitor GSK343, blocked SMAD3-dependent cis co-accessibility and inhibited myofibroblast activation. This mechanism, through which TGFß signals directly to chromatin, represents a critical determinant of fibrotic, differentiated states.

SRC plays a specific role in the cross-talk between apoptosis and autophagy via phosphorylation of a novel regulatory site on AMPK.

Cell detachment from the extracellular matrix (ECM) typically promotes cell death via a form of apoptosis known as anoikis. However, in tumor cells, detachment can also induce cell survival, utilizing a process known as macroautophagy/autophagy, which involves degradation and removal of apoptotic proteins as well as rewiring of metabolic pathways so that cells can survive under stress. The crosstalk between the competing processes of anoikis and autophagy is only partially understood but may be critical for the design of multi-drug therapeutic strategies. Here, we summarize our recent studies, which reveal a direct regulatory link between a major mediator of cell survival in adherent cells, the ECM-integrin-activated dual tyrosine kinase complex of SRC and PTK2/FAK, and a major regulator of cell metabolism and autophagy, AMP-activated protein kinase (AMPK). We identify a novel SRC phosphorylation site on AMPK and demonstrate that this phosphorylation event plays key roles in AMPK regulation, autophagy induction, and cell survival.

Copepod reproductive effort and oxidative status as responses to warming in the marine environment.

The marine ecosystems are under severe climate change-induced stress globally. The Baltic Sea is especially vulnerable to ongoing changes, such as warming. The aim of this study was to measure eco-physiological responses of a key copepod species to elevated temperature in an experiment, and by collecting field samples in the western Gulf of Finland. The potential trade-off between reproductive output and oxidative balance in copepods during thermal stress was studied by incubating female Acartia sp. for reproduction rate and oxidative stress measurements in ambient and elevated temperatures. Our field observations show that the glutathione cycle had a clear response in increasing stress and possibly had an important role in preventing oxidative damage: Lipid peroxidation and ratio of reduced and oxidized glutathione were negatively correlated throughout the study. Moreover, glutathione-s-transferase activated in late July when the sea water temperature was exceptionally high and Acartia sp. experienced high oxidative stress. The combined effect of a heatwave, increased cyanobacteria, and decreased dinoflagellate abundance may have caused larger variability in reproductive output in the field. An increase of 7°C had a negative effect on egg production rate in the experiment. However, the effect on reproduction was relatively small, implying that Acartia sp. can tolerate warming at least within the temperature range of 9-16°C. However, our data from the experiment suggest a link between reproductive success and oxidative stress during warming, shown as a significant combined effect of temperature and catalase on egg production rate.

Discovery of novel furanylbenzamide inhibitors that target oncogenic tyrosine phosphatase SHP2 in leukemia cells.

Disturbance of the dynamic balance between tyrosine phosphorylation and dephosphorylation of signaling molecules, controlled by protein tyrosine kinases and protein tyrosine phosphatases (PTPs), is known to lead to the development of cancer. While most approved targeted cancer therapies are tyrosine kinase inhibitors, PTPs have long been stigmatized as undruggable and have only recently gained renewed attention in drug discovery. One PTP target is the Src-homology 2 domain-containing phosphatase 2 (SHP2). SHP2 is implicated in tumor initiation, progression, metastasis, and treatment resistance, primarily because of its role as a signaling nexus of the extracellular signal-regulated kinase pathway, acting upstream of the small GTPase Ras. Efforts to develop small molecules that target SHP2 are ongoing, and several SHP2 allosteric inhibitors are currently in clinical trials for the treatment of solid tumors. However, while the reported allosteric inhibitors are highly effective against cells expressing WT SHP2, none have significant activity against the most frequent oncogenic SHP2 variants that drive leukemogenesis in several juvenile and acute leukemias. Here, we report the discovery of novel furanylbenzamide molecules as inhibitors of both WT and oncogenic SHP2. Importantly, these inhibitors readily cross cell membranes, bind and inhibit SHP2 under physiological conditions, and effectively decrease the growth of cancer cells, including triple-negative breast cancer cells, acute myeloid leukemia cells expressing either WT or oncogenic SHP2, and patient-derived acute myeloid leukemia cells. These novel compounds are effective chemical probes of active SHP2 and may serve as starting points for therapeutics targeting WT or mutant SHP2 in cancer.

Whole blood trace element and toxic metal concentration in dogs with idiopathic epilepsy and healthy dogs: A case-control study.

Background: Idiopathic epilepsy (IE) is the most common neurological disease in dogs. Multiple genes and environmental factors interact to cause clinical signs, although the pathogenesis remains poorly understood. Extensive evidence from recent decades shows that trace elements play a role in epilepsy in humans, and recently it was shown for the first time that also dogs with IE have altered trace element status. On the other hand, toxic metals may cause seizures but research on their role in canine IE is lacking. Therefore, we aimed to investigate trace element and toxic metal concentrations in whole blood from dogs that had been diagnosed with IE and compare them to those of healthy dogs. Materials and methods: Whole blood concentrations of trace elements (selenium, zinc, copper, manganese, iron, and chromium) and toxic metals (arsenic, cadmium, mercury, and lead) were analyzed from 19 dogs that had been diagnosed with IE by board-certified neurologists and 19 healthy control dogs using inductively coupled plasma mass spectrometry. The concentrations in study and control group were compared using the Mann-Whitney U test. Results: Dogs diagnosed with IE had significantly higher blood copper concentration (P = 0.007), higher copper/zinc ratio (P = 0.04), and higher selenium concentration (P < 0.001), as well as lower chromium concentration (P = 0.01) when compared to healthy dogs. Treatment of IE with potassium bromide was associated with a significant elevation in blood arsenic concentration (P = 0.01). Conclusion: In conclusion, the present results support the role of altered trace element status in dogs diagnosed with IE and suggest that copper, selenium, and chromium may be involved in the pathogenesis of canine epilepsy or seizures. The results also suggest that potassium bromide may alter arsenic metabolism in dogs.

Cell adhesion suppresses autophagy via Src/FAK-mediated phosphorylation and inhibition of AMPK.

Autophagy is a multi-step process regulated in part by AMP-activated protein kinase (AMPK). Phosphorylation of threonine 172 on the AMPK α-subunit enhances AMPK kinase activity, resulting in activation of downstream signaling. Integrin-mediated cell adhesion activates Src/ Focal Adhesion Kinase (FAK) signaling complex, which regulates multiple cellular processes including cell survival. We show here that Src signaling leads to direct phosphorylation of the AMPK-α subunit on a novel site, tyrosine 179, resulting in suppression of AMPK-T172 phosphorylation and autophagy upon integrin-mediated cell adhesion. By using chemical inhibitors, genetic cell models and targeted mutagenesis, we confirm an important role for Src and FAK in suppressing AMPK signaling and autophagy induced by various additional stimuli, including glucose starvation. Furthermore, we found that autophagy suppression by hydroxychloroquine promotes apoptosis in a cancer cell model that had been treated with Src inhibitors. Our findings reveal a link between the Src/ FAK complex and AMPK/ autophagy regulation, which may play an important role in the maintenance of normal cellular homeostasis and tumor progression.

Diet and dog characteristics affect major and trace elements in hair and blood of healthy dogs.

Obtaining correct amounts of essential elements, and avoiding toxic metals are key factors in dog health. Through analyzing major and trace elements in hair and blood of 50 healthy companion dogs using ICP-MS, we study their associations with dog characteristics and diet, hypothesizing that eating the same diet long-term results in strong correlations between hair and blood element concentrations, and that dog characteristics and diet affect element status. The correlation between hair and blood was significant for Hg (R = 0.601, p = 0.000) and Pb (R = 0.384, p = 0.010). The following associations were significant (p < 0.05): Dark hair had higher Ca and Mg compared to light hair. Females had higher hair Zn, blood Mn, and blood As compared to males. Blood Mn and Se increased, while blood Pb decreased with age. Raw diet fed dogs had higher hair Zn and Se compared to dry or mixed diet fed dogs, and lower blood Mn compared to dry diet fed dogs. Dry and mixed diet fed dogs had higher blood Cd compared to raw diet fed dogs. Mixed diet fed dogs had higher hair Ca and Mg compared to raw or dry diet fed dogs, and higher hair Pb compared to dry diet fed dogs. Wild game consumption was associated with higher blood Pb, and rice consumption with higher blood As. In conclusion, hair provides an alternative for assessing Hg and Pb exposure, and major and trace elements status is affected by hair color, sex, age, and diet.

The ubiquitin ligase RNF5 determines acute myeloid leukemia growth and susceptibility to histone deacetylase inhibitors.

Acute myeloid leukemia (AML) remains incurable, largely due to its resistance to conventional treatments. Here, we find that increased abundance of the ubiquitin ligase RNF5 contributes to AML development and survival. High RNF5 expression in AML patient specimens correlates with poor prognosis. RNF5 inhibition decreases AML cell growth in culture, in patient-derived xenograft (PDX) samples and in vivo, and delays development of MLL-AF9-driven leukemogenesis in mice, prolonging their survival. RNF5 inhibition causes transcriptional changes that overlap with those seen upon histone deacetylase (HDAC)1 inhibition. RNF5 induces the formation of K29 ubiquitin chains on the histone-binding protein RBBP4, promoting its recruitment to and subsequent epigenetic regulation of genes involved in AML maintenance. Correspondingly, RNF5 or RBBP4 knockdown enhances AML cell sensitivity to HDAC inhibitors. Notably, low expression of both RNF5 and HDAC coincides with a favorable prognosis. Our studies identify an ERAD-independent role for RNF5, demonstrating that its control of RBBP4 constitutes an epigenetic pathway that drives AML, and highlight RNF5/RBBP4 as markers useful to stratify patients for treatment with HDAC inhibitors.

Puppyhood diet as a factor in the development of owner-reported allergy/atopy skin signs in adult dogs in Finland.

BACKGROUND: The increased prevalence of atopic dermatitis (AD) in dogs necessitates research in its disease etiology. OBJECTIVES: To explore the association between puppyhood dietary exposures and prevalence of owner-reported allergy/atopy skin signs (AASS) after the age of 1 year. ANIMALS: Four thousand and twenty-two dogs were eligible, 1158 cases, and 2864 controls. METHODS: This cross-sectional hypothesis-driven observational study was extracted from the DogRisk food frequency questionnaire. Forty-six food items and the ratio of 4 major diet types were tested for their association with AASS incidence later in life. Potential puppyhood dietary risk factors for AASS incidence were specified using binary multivariable logistic regression. The model was adjusted for age and sex. RESULTS: Eating raw tripe (odds ratio, 95% confidence intervals OR, 95% CI = 0.36, 0.16-0.79; P = .01), raw organ meats (OR, 95% CI = 0.23, 0.08-0.67; P = .007), human meal leftovers, and fish oil supplements as well as eating more that 20% of the diet as raw and/or <80% of the diet as dry, in general, were associated with significantly lower AASS incidence in adulthood. In contrast, dogs fed fruits (OR, 95% CI = 2.01, 1.31-3.07; P = .001), mixed-oil supplements, dried animal parts, and dogs that drank from puddles showed significantly higher AASS incidence in adulthood. CONCLUSIONS AND CLINICAL IMPORTANCE: Puppyhood exposure to raw animal-based foods might have a protective influence on AASS incidence in adulthood, while puppyhood exposure to mixed oils, heat processed foods and sugary fruits might be a potential risk factor of AASS incidence later. The study suggests a causal relationship but does not prove it.

Early Life Modifiable Exposures and Their Association With Owner Reported Inflammatory Bowel Disease Symptoms in Adult Dogs.

Background: Inflammatory bowel disease (IBD) is an idiopathic multifactorial disease in humans and dogs, usually assigned to the interactions between genes, gut microbiota, diet, environment, and the immune system. We aimed to investigate the modifiable early life exposures associated with IBD in dogs. Materials and Methods: The study data was extracted from the validated owner-reported DogRisk food frequency questionnaire. This was a cross-sectional questionnaire-based study that tested 21 different early life dietary and environmental, demographic and genetic variables for their association with IBD or not, in adult dogs. A total of 7,015 dogs participated in this study. The study covered early life periods; prenatal, neonatal, early, and late postnatal periods. Two feeding patterns, a non-processed meat-based diet (NPMD) and an ultra-processed carbohydrate-based diet (UPCD) were studied. Data was analyzed using logistic regression analysis with a backward stepwise deletion. Results: From the final models we found that the NPMD during early and late postnatal periods were significantly associated with lower IBD risk later in life. The UPCD during the same periods was associated with a higher risk of IBD incidence. Also, the maternal diet during the neonatal period showed a non-significant trend of lower IBD risk in the offspring with the NPMD and a higher IBD risk with the UPCD. Additionally, the normal body weight of puppies during the first 6 months of age was associated with a lower risk of IBD in adulthood while, slim puppies associated significantly with IBD in adulthood. From the non-modifiable background variables, we identified the maternal history of IBD as the strongest risk factor for later incidence of IBD. Furthermore, male dogs were twice as likely to develop IBD as female dogs were. Conclusions: It is reassuring for owners to know that they themselves can have an impact on their dog's health. A high-fat, low-carbohydrate NPMD exposure during early life, and a normal body condition in puppyhood were significantly associated with less IBD in adult dogs. The opposite was true for UPCD exposure and abnormal body condition score in 6 month old puppies.

PTEN deficiency leads to proteasome addiction: a novel vulnerability in glioblastoma.

BACKGROUND: Glioblastoma (GBM) is the most common primary brain tumor in adults with a median survival of approximately 15 months; therefore, more effective treatment options for GBM are required. To identify new drugs targeting GBMs, we performed a high-throughput drug screen using patient-derived neurospheres cultured to preferentially retain their glioblastoma stem cell (GSC) phenotype. METHODS: High-throughput drug screening was performed on GSCs followed by a dose-response assay of the 5 identified original "hits." A PI3K/mTOR dependency to a proteasome inhibitor (carfilzomib), was confirmed by genetic and pharmacologic experiments. Proteasome Inhibition Response Signatures were derived from proteomic and bioinformatic analysis. Molecular mechanism of action was determined using three-dimensional (3D) GBM-organoids and preclinical orthotopic models. RESULTS: We found that GSCs were highly sensitive to proteasome inhibition due to an underlying dependency on an increased protein synthesis rate, and loss of autophagy, associated with PTEN loss and activation of the PI3K/mTOR pathway. In contrast, combinatory inhibition of autophagy and the proteasome resulted in enhanced cytotoxicity specifically in GSCs that did express PTEN. Finally, proteasome inhibition specifically increased cell death markers in 3D GBM-organoids, suppressed tumor growth, and increased survival of mice orthotopically engrafted with GSCs. As perturbations of the PI3K/mTOR pathway occur in nearly 50% of GBMs, these findings suggest that a significant fraction of these tumors could be vulnerable to proteasome inhibition. CONCLUSIONS: Proteasome inhibition is a potential synthetic lethal therapeutic strategy for GBM with proteasome addiction due to a high protein synthesis rate and autophagy deficiency.

MicroRNA-211 Modulates the DUSP6-ERK5 Signaling Axis to Promote BRAFV600E-Driven Melanoma Growth In Vivo and BRAF/MEK Inhibitor Resistance.

MicroRNAs (miRs) are important posttranscriptional regulators of cell fate in both normal and disease states. miR-211 has previously been shown to be a direct regulator of metabolism in BRAFV600E-mutant melanoma cells in vitro. Here, we report that miR-211 expression promotes the aggressive growth of BRAFV600E-mutant melanoma xenografts in vivo. miR-211 promoted proliferation through the posttranscriptional activation of extracellular signal-regulated kinase (ERK) 5 signaling, which has recently been implicated in the resistance to BRAF and MAPK/ERK kinase inhibitors. We therefore examined whether miR-211 similarly modulated melanoma resistance to the BRAF inhibitor vemurafenib and the MAPK/ERK kinase inhibitor cobimetinib. Consistent with this model, miR-211 expression increased melanoma cell resistance to both the inhibitors, and this resistance was associated with an increased ERK5 phosphorylation. miR-211 mediates these effects by directly inhibiting the expression of DUSP6, an ERK5 pathway-specific phosphatase and now shown to be an miR-211 target gene. These results dissect the role of the miR-211-DUSP6-ERK5 axis in melanoma tumor growth and suggest a mechanism for the development of drug-resistant tumors and a target for overcoming resistance.