Histone H3.3G34-Mutant Interneuron Progenitors Co-opt PDGFRA for Gliomagenesis.

Histone H3.3 glycine 34 to arginine/valine (G34R/V) mutations drive deadly gliomas and show exquisite regional and temporal specificity, suggesting a developmental context permissive to their effects. Here we show that 50% of G34R/V tumors (n = 95) bear activating PDGFRA mutations that display strong selection pressure at recurrence. Although considered gliomas, G34R/V tumors actually arise in GSX2/DLX-expressing interneuron progenitors, where G34R/V mutations impair neuronal differentiation. The lineage of origin may facilitate PDGFRA co-option through a chromatin loop connecting PDGFRA to GSX2 regulatory elements, promoting PDGFRA overexpression and mutation. At the single-cell level, G34R/V tumors harbor dual neuronal/astroglial identity and lack oligodendroglial programs, actively repressed by GSX2/DLX-mediated cell fate specification. G34R/V may become dispensable for tumor maintenance, whereas mutant-PDGFRA is potently oncogenic. Collectively, our results open novel research avenues in deadly tumors. G34R/V gliomas are neuronal malignancies where interneuron progenitors are stalled in differentiation by G34R/V mutations and malignant gliogenesis is promoted by co-option of a potentially targetable pathway, PDGFRA signaling.

13q12.2 deletions in acute lymphoblastic leukemia lead to upregulation of FLT3 through enhancer hijacking.

Mutations in the FMS-like tyrosine kinase 3 (FLT3) gene in 13q12.2 are among the most common driver events in acute leukemia, leading to increased cell proliferation and survival through activation of the phosphatidylinositol 3-kinase/AKT-, RAS/MAPK-, and STAT5-signaling pathways. In this study, we examine the pathogenetic impact of somatic hemizygous 13q12.2 microdeletions in B-cell precursor (BCP) acute lymphoblastic leukemia (ALL) using 5 different patient cohorts (in total including 1418 cases). The 13q12.2 deletions occur immediately 5' of FLT3 and involve the PAN3 locus. By detailed analysis of the 13q12.2 segment, we show that the deletions lead to loss of a topologically associating domain border and an enhancer of FLT3. This results in increased cis interactions between the FLT3 promoter and another enhancer located distally to the deletion breakpoints, with subsequent allele-specific upregulation of FLT3 expression, expected to lead to ligand-independent activation of the receptor and downstream signaling. The 13q12.2 deletions are highly enriched in the high-hyperdiploid BCP ALL subtype (frequency 3.9% vs 0.5% in other BCP ALL) and in cases that subsequently relapsed. Taken together, our study describes a novel mechanism of FLT3 involvement in leukemogenesis by upregulation via chromatin remodeling and enhancer hijacking. These data further emphasize the role of FLT3 as a driver gene in BCP ALL.

Stable Control of Physiological Parameters, But Not Infection, in Preterm Lambs Maintained on Ex Vivo Uterine Environment Therapy.

Ex vivo uterine environment (EVE) therapy is an experimental neonatal intensive care strategy wherein gas exchange is performed by membranous oxygenators attached to the umbilical vessels. Our aim was to assess the ability of a newly refined EVE system to maintain key physiological parameters in preterm lambs within optimal ranges for 48 h. EVE group; n = 6: Preterm lambs were delivered under general anesthesia at 115 ± 2 days of gestational age. Animals were submerged in a bath of artificial amniotic fluid on EVE therapy for 48 h. Physiological parameters were monitored in real-time over the length of the experiment. Control group; n = 11: Ewes carrying a single fetus (115 ± 2 days of gestational age) underwent recovery surgery to allow placement of a fetal carotid artery catheter. Fetuses received an infusion of sterile saline only. After euthanasia, EVE and Control group fetuses underwent necroscopy to perform static pressure-volume curves and for sampling of lung and cord blood plasma for molecular analyses. Five out of six fetuses in the EVE group completed the study period with key physiological variables remaining within their respective reference ranges for the duration of the 48 h study. Bacteremia was identified in four out of five EVE fetuses, and was associated with a systemic inflammatory response. Using our refined EVE therapy platform, preterm lambs were maintained in a stable physiological condition for 48 h. These findings represent a significant advance over earlier work with this system; however, the identification of bacteremia and a fetal inflammatory response suggests that further refinement to the EVE therapy platform is required.

The uptake of transdermal fentanyl in a pregnant sheep model.

OBJECTIVE: To evaluate the maternal and foetal uptake of transdermal fentanyl patch applied to the groin of pregnant sheep following surgery. STUDY DESIGN: Prospective series. ANIMALS: A group of 16 singleton pregnant sheep underwent anaesthesia for laparotomy, hysterotomy and instrumentation of the foetus. Of these ewes 10 (101 ± 12 days of gestation) were used to evaluate the maternal uptake of transdermal fentanyl, and the efficacy of the drug in the postoperative period (n = 10). To determine the extent of transplacental transfer of fentanyl, six ewes from the group of 10, and six other ewes (92 ± 1 days' gestation) were studied. METHODS: A 75 µg hour-1 fentanyl patch was placed onto the woolless skin of the medial thigh close to the groin at the end of surgery. Maternal blood samples were collected from the cephalic or jugular vein, and pain and sedation scores were determined, prior to application of the patch (time 0) and at 3, 6, 12, 24, 36 and 48 hours after. A commercial Fentanyl ELISA kit was used to determine the concentration of fentanyl. Paired maternal and foetal blood samples were collected 48 hours after surgery. Animals were euthanized at the end of the study. Data were tested for normality and compared with Student t test or one-way anova and are expressed as mean ± standard deviation or median (range). RESULTS: Recovery from anaesthesia and surgery was uneventful in all ewes. The dose of fentanyl was 1.4 ± 0.2 µg kg-1 hour-1. The maximum maternal plasma concentration of fentanyl was 0.547 ng mL-1 (range, 0.349-0.738 ng mL-1) at 12 hours. After 48 hours, the concentration of fentanyl was 0.381 ng mL-1 (range, 0.211-0.487 ng mL-1; maternal) and 0.295 ng mL-1 (range, 0.185-0.377 ng mL-1; foetal; p = 0.175). The placental transfer rate of fentanyl was 77%. CONCLUSIONS AND CLINICAL RELEVANCE: The uptake of fentanyl varied between animals. The placental transfer rate of fentanyl was 77%.

Outside-in? Acute fetal systemic inflammation in very preterm chronically catheterized sheep fetuses is not driven by cells in the fetal blood.

BACKGROUND: The preterm birth syndrome (delivery before 37 weeks gestation) is a major contributor to the global burden of perinatal morbidity and death. The cause of preterm birth is complex, multifactorial, and likely dependent, at least in part, on the gestational age of the fetus. Intrauterine infection is frequent in preterm deliveries that occur at <32 weeks gestation; understanding how the fetus responds to proinflammatory insult will be an important step towards early preterm birth prevention. However, animal studies of infection and inflammation in prematurity commonly use older fetuses that possess comparatively mature immune systems. OBJECTIVE: Aiming to characterize acute fetal responses to microbial agonist at a clinically relevant gestation, we used 92-day-old fetuses (62% of term) to develop a chronically catheterized sheep model of very preterm pregnancy. We hypothesized that any acute fetal systemic inflammatory responses would be driven by signaling from the tissues exposed to Escherichia coli lipopolysaccharide that is introduced into the amniotic fluid. STUDY DESIGN: Eighteen ewes that were carrying a single fetus at 92 days of gestation had recovery surgery to place fetal tracheal, jugular, and intraamniotic catheters. Animals were recovered for 24 hours before being administered either intraamniotic E coli lipopolysaccharide (n = 9) or sterile saline solution (n = 9). Samples were collected for 48 hours before euthanasia and necroscopy. Fetal inflammatory responses were characterized by microarray analysis, quantitative polymerase chain reaction, and enzyme-linked immunosorbent assay. RESULTS: Intraamniotic lipopolysaccharide reached the distal trachea within 2 hours. Lipopolysaccharide increased tracheal fluid interleukin-8 within 2 hours and generated a robust inflammatory response that was characterized by interleukin-6 signaling pathway activation and up-regulation of cell proliferation but no increases in inflammatory mediator expression in cord blood RNA. CONCLUSIONS: In very preterm sheep fetuses, lipopolysaccharide stimulates inflammation in the fetal lung and fetal skin and stimulates a systemic inflammatory response that is not generated by fetal blood cells. These data argue for amniotic fluid-exposed tissues that play a key role in driving acute fetal and intrauterine inflammatory responses.

Clinical and Genomic Predictors of Adverse Events in Newly Diagnosed Glioblastoma.

PURPOSE: Adverse clinical events cause significant morbidity in patients with GBM (GBM). We examined whether genomic alterations were associated with AE (AE) in patients with GBM. EXPERIMENTAL DESIGN: We identified adults with histologically confirmed IDH-wild-type GBM with targeted next-generation sequencing (OncoPanel) at Dana Farber Cancer Institute from 2013 to 2019. Seizure at presentation, lymphopenia, thromboembolic events, pseudoprogression, and early progression (within 6 months of diagnosis) were identified as AE. The biologic function of genetic variants was categorized as loss-of-function (LoF), no change in function, or gain-of-function (GoF) using a somatic tumor mutation knowledge base (OncoKB) and consensus protein function predictions. Associations between functional genomic alterations and AE were examined using univariate logistic regressions and multivariable regressions adjusted for additional clinical predictors. RESULTS: Our study included 470 patients diagnosed with GBM who met the study criteria. We focused on 105 genes that had sequencing data available for ≥ 90% of the patients and were altered in ≥10% of the cohort. Following false-discovery rate (FDR) correction and multivariable adjustment, the TP53, RB1, IGF1R, and DIS3 LoF alterations were associated with lower odds of seizures, while EGFR, SMARCA4, GNA11, BRD4, and TCF3 GoF and SETD2 LoF alterations were associated with higher odds of seizures. For all other AE of interest, no significant associations were found with genomic alterations following FDR correction. CONCLUSIONS: Genomic biomarkers based on functional variant analysis of a routine clinical panel may help identify AE in GBM, particularly seizures. Identifying these risk factors could improve the management of patients through better supportive care and consideration of prophylactic therapies.

Integrated molecular and clinical analysis of BRAF-mutant glioma in adults.

BRAF mutations are a significant driver of disease in pediatric low-grade glioma, but the implications of BRAF alterations on the clinical course and treatment response in adult glioma remain unclear. Here, we characterize a multi-institutional cohort of more than 300 patients (>200 adults) with BRAF-mutated glioma using clinical, pathological/molecular, and outcome data. We observed that adult and pediatric BRAF-mutant gliomas harbor distinct clinical and molecular features, with a higher prevalence of BRAFV600E (Class I) and BRAF fusions in pediatric tumors. BRAFV600E alterations were associated with improved survival in adults with glioma overall, though not in glioblastoma. Other genomic alterations observed within functional classes were consistent with the putative roles of those BRAF mutation classes in glioma pathogenesis. In our adult cohort, BRAFV600E alterations conferred sensitivity to targeted therapies. Overall, this large cohort of BRAF-altered adult gliomas demonstrates a broad range of molecular alterations with implications for treatment sensitivity and survival.

Epigenetic reactivation of tumor suppressor genes with CRISPRa technologies as precision therapy for hepatocellular carcinoma.

BACKGROUND: Epigenetic silencing of tumor suppressor genes (TSGs) is a key feature of oncogenesis in hepatocellular carcinoma (HCC). Liver-targeted delivery of CRISPR-activation (CRISPRa) systems makes it possible to exploit chromatin plasticity, by reprogramming transcriptional dysregulation. RESULTS: Using The Cancer Genome Atlas HCC data, we identify 12 putative TSGs with negative associations between promoter DNA methylation and transcript abundance, with limited genetic alterations. All HCC samples harbor at least one silenced TSG, suggesting that combining a specific panel of genomic targets could maximize efficacy, and potentially improve outcomes as a personalized treatment strategy for HCC patients. Unlike epigenetic modifying drugs lacking locus selectivity, CRISPRa systems enable potent and precise reactivation of at least 4 TSGs tailored to representative HCC lines. Concerted reactivation of HHIP, MT1M, PZP, and TTC36 in Hep3B cells inhibits multiple facets of HCC pathogenesis, such as cell viability, proliferation, and migration. CONCLUSIONS: By combining multiple effector domains, we demonstrate the utility of a CRISPRa toolbox of epigenetic effectors and gRNAs for patient-specific treatment of aggressive HCC.

Clonal origin and development of high hyperdiploidy in childhood acute lymphoblastic leukaemia.

High hyperdiploid acute lymphoblastic leukemia (HeH ALL), one of the most common childhood malignancies, is driven by nonrandom aneuploidy (abnormal chromosome numbers) mainly comprising chromosomal gains. In this study, we investigate how aneuploidy in HeH ALL arises. Single cell whole genome sequencing of 2847 cells from nine primary cases and one normal bone marrow reveals that HeH ALL generally display low chromosomal heterogeneity, indicating that they are not characterized by chromosomal instability and showing that aneuploidy-driven malignancies are not necessarily chromosomally heterogeneous. Furthermore, most chromosomal gains are present in all leukemic cells, suggesting that they arose early during leukemogenesis. Copy number data from 577 primary cases reveals selective pressures that were used for in silico modeling of aneuploidy development. This shows that the aneuploidy in HeH ALL likely arises by an initial tripolar mitosis in a diploid cell followed by clonal evolution, in line with a punctuated evolution model.

Synthetic Epigenetic Reprogramming of Mesenchymal to Epithelial States Using the CRISPR/dCas9 Platform in Triple Negative Breast Cancer.

Epithelial-mesenchymal transition (EMT) is a reversible transcriptional program invoked by cancer cells to drive cancer progression. Transcription factor ZEB1 is a master regulator of EMT, driving disease recurrence in poor-outcome triple negative breast cancers (TNBCs). Here, this work silences ZEB1 in TNBC models by CRISPR/dCas9-mediated epigenetic editing, resulting in highly-specific and nearly complete suppression of ZEB1 in vivo, accompanied by long-lasting tumor inhibition. Integrated "omic" changes promoted by dCas9 linked to the KRAB domain (dCas9-KRAB) enabled the discovery of a ZEB1-dependent-signature of 26 genes differentially-expressed and -methylated, including the reactivation and enhanced chromatin accessibility in cell adhesion loci, outlining epigenetic reprogramming toward a more epithelial state. In the ZEB1 locus transcriptional silencing is associated with induction of locally-spread heterochromatin, significant changes in DNA methylation at specific CpGs, gain of H3K9me3, and a near complete erasure of H3K4me3 in the ZEB1 promoter. Epigenetic shifts induced by ZEB1-silencing are enriched in a subset of human breast tumors, illuminating a clinically-relevant hybrid-like state. Thus, the synthetic epi-silencing of ZEB1 induces stable "lock-in" epigenetic reprogramming of mesenchymal tumors associated with a distinct and stable epigenetic landscape. This work outlines epigenome-engineering approaches for reversing EMT and customizable precision molecular oncology approaches for targeting poor outcome breast cancers.

The anti-inflammatory actions of IL-4 in human monocytes are not mediated by IL-10, RP105 or the kinase activity of RIPK2.

The anti-inflammatory actions of IL-4 in activated human monocytes may reflect transcriptional regulation of genes involved in TLR signaling pathways. Tailored gene arrays were conducted to profile the expression of 84 genes central to TLR-mediated signal transduction in human monocytes treated with the TLR4 ligand, LPS, with or without IL-4. In the first 3h, IL-4 down-regulated mRNA levels of LPS-induced inflammatory cytokines and chemokines, without altering mRNA levels of TLRs, TLR-related signaling molecules or multiple transcription factors. The down-regulation of inflammatory genes by IL-4 was preceded by an early up-regulation of IL-10 mRNA and protein and mRNA for receptor-interacting serine-threonine kinase 2 (RIPK2), the TLR homolog, RP105, and c-Maf, a transcription factor required for IL-10 gene expression. However, IL-4 still suppressed LPS-induced TNFα production in bone-marrow derived macrophages from IL10(-/-) mice, and in the presence of a neutralizing antibody to IL-10 in human monocytes. The up-regulation of RIPK2 and RP105 mRNA by IL-4 occurred independently of IL-10. IL-4 maintained the ability to suppress LPS-induced TNFα and enhance IL-10 production in the presence of RIPK2 kinase inhibitors. Further, IL-4 failed to up-regulate expression of RP105 at the cell surface. In conclusion, the anti-inflammatory actions of IL-4 occur independently of IL-10, RP105, and the kinase activity of RIPK2.

Posttraumatic stress disorder symptoms associated with protective and risky behaviors for coronavirus disease 2019.

OBJECTIVE: Psychiatric disorders increase risk for contracting coronavirus disease 2019 (COVID-19), but we know little about relationships between psychiatric symptoms and COVID-19 risky and protective behaviors. Posttraumatic stress disorder (PTSD) has been associated with increased propensity to engage in risky behaviors, but may also be associated with increased COVID-19 protective behaviors due to increased threat sensitivity and social isolation. METHOD: We examined associations of PTSD symptoms with COVID-19-related protective and risky behaviors using data from a cross-sectional online United States study among 845 US adults in August through September 2020. PTSD symptoms (PTSD Checklist-5), sociodemographics, COVID-19-related experiences and vulnerabilities, and past 30-day engagement in 10 protective and eight risky behaviors for COVID-19 were assessed via self-report. We examined associations between PTSD symptoms and COVID-19 protective and risky behaviors with linear regressions, adjusting for covariates. RESULTS: Probable PTSD and higher PTSD symptom severity were associated with greater engagement in protective behaviors, but also greater engagement in risky behaviors. Associations were only slightly attenuated by adjustment for COVID-19 exposures and perceived likelihood and severity of COVID-19. Associations varied by PTSD clusters: intrusions and arousal were associated with both more protective and more risky behaviors, whereas negative cognitions or mood was associated only with more risky, and avoidance only with more protective, behaviors. CONCLUSION: Higher PTSD symptoms were associated with engagement in more protective but also more risky behaviors for COVID-19. Mental health should be considered in the design of public health campaigns dedicated to limiting infectious disease spread. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Identifying correlates of suicide ideation during the COVID-19 pandemic: A cross-sectional analysis of 148 sociodemographic and pandemic-specific factors.

The coronavirus disease 2019 (COVID-19) pandemic has created a global health crisis, with disproportionate effects on vulnerable sociodemographic groups. Although the pandemic is showing potential to increase suicide ideation (SI), we know little about which sociodemographic characteristics or COVID-19 experiences are associated with SI. Our United States-based sample (n = 837 adults [mean age = 37.1 years]) completed an online survey during August-September 2020. The study utilized an online convenience sample from a prior study, which was enriched for exposure to trauma and experiences of posttraumatic stress symptoms. We assessed SI using the Beck Depression Inventory-II. Traditional (i.e., logistic regression) and machine learning (i.e., LASSO, random forest) methods evaluated associations of 148 self-reported COVID-19 factors and sociodemographic characteristics with current SI. 234 participants (28.0%) reported SI. Twenty items were significantly associated with SI from logistic regression. Of these 20 items, LASSO identified seven sociodemographic characteristics (younger age, lower income, single relationship status, sexual orientation other than heterosexual as well as specifically identifying as bisexual, non-full-time employment, and living in a town) and six COVID-19 factors (not engaging in protective COVID-19 behaviors, receiving mental health treatment (medication and/or psychotherapy) due to the COVID-19 pandemic, socializing during the pandemic, losing one's job due to COVID-19, having a friend with COVID-19, and having an acquaintance with COVID-19) associated with SI. Random forest findings were largely consistent with LASSO. These findings may inform multidisciplinary research and intervention work focused on understanding and preventing adverse mental health outcomes such as SI during and in the aftermath of the pandemic.

Clusters of COVID-19 protective and risky behaviors and their associations with pandemic, socio-demographic, and mental health factors in the United States.

Individual behaviors are critical for preventing the spread of coronavirus disease 2019 (COVID-19) infection. Given that both protective and risky behaviors influence risk of infection, it is critical that we understand how such behaviors cluster together and in whom. Using a data-driven approach, we identified clusters of COVID-19-related protective and risky behaviors and examined associations with socio-demographic, pandemic, and mental health factors. Data came from a cross-sectional online U.S. nationwide study of 832 adults with high levels of pre-pandemic trauma. Latent class analysis was performed with ten protective (e.g., washing hands, wearing masks) and eight risky (e.g., attending indoor restaurants, taking a flight) behaviors for COVID-19. Then, we examined distributions of socio-demographic and pandemic factors across behavior classes using ANOVA or Chi-square tests, and associations between mental health factors (depressive, anxiety, posttraumatic stress symptoms) and behavior classes using multinomial logistic regression. We identified four classes, including three classes with relatively low risky but high (28.8%), moderate (33.5%) and minimal (25.5%) protective behaviors and one high risky behaviors class with associated moderate protective behaviors (12.1%). Age, sexual orientation, political preference, and most pandemic factors differed significantly across behavior classes. Anxiety and posttraumatic stress symptoms, but not depression, were higher in the High Risk, but also Highly and Moderately Protective classes, relative to Minimally Protective. Prevention and intervention efforts should examine constellations of protective and risky behaviors to comprehensively understand risk, and consider current anxiety and posttraumatic stress symptoms as potential risk indicators.

A molecularly integrated grade for meningioma.

BACKGROUND: Meningiomas are the most common primary intracranial tumor in adults. Clinical care is currently guided by the World Health Organization (WHO) grade assigned to meningiomas, a 3-tiered grading system based on histopathology features, as well as extent of surgical resection. Clinical behavior, however, often fails to conform to the WHO grade. Additional prognostic information is needed to optimize patient management. METHODS: We evaluated whether chromosomal copy-number data improved prediction of time-to-recurrence for patients with meningioma who were treated with surgery, relative to the WHO schema. The models were developed using Cox proportional hazards, random survival forest, and gradient boosting in a discovery cohort of 527 meningioma patients and validated in 2 independent cohorts of 172 meningioma patients characterized by orthogonal genomic platforms. RESULTS: We developed a 3-tiered grading scheme (Integrated Grades 1-3), which incorporated mitotic count and loss of chromosome 1p, 3p, 4, 6, 10, 14q, 18, 19, or CDKN2A. 32% of meningiomas reclassified to either a lower-risk or higher-risk Integrated Grade compared to their assigned WHO grade. The Integrated Grade more accurately identified meningioma patients at risk for recurrence, relative to the WHO grade, as determined by time-dependent area under the curve, average precision, and the Brier score. CONCLUSION: We propose a molecularly integrated grading scheme for meningiomas that significantly improves upon the current WHO grading system in prediction of progression-free survival. This framework can be broadly adopted by clinicians with relative ease using widely available genomic technologies and presents an advance in the care of meningioma patients.

Genomic landscape of gliosarcoma: distinguishing features and targetable alterations.

Gliosarcoma is an aggressive brain tumor with histologic features of glioblastoma (GBM) and soft tissue sarcoma. Despite its poor prognosis, its rarity has precluded analysis of its underlying biology. We used a multi-center database to characterize the genomic landscape of gliosarcoma. Sequencing data was obtained from 35 gliosarcoma patients from Genomics Evidence Neoplasia Information Exchange (GENIE) 5.0, a database curated by the American Association of Cancer Research (AACR). We analyzed genomic alterations in gliosarcomas and compared them to GBM (n = 1,449) and soft tissue sarcoma (n = 1,042). 30 samples were included (37% female, median age 59 [IQR: 49-64]). Nineteen common genes were identified in gliosarcoma, defined as those altered in > 5% of samples, including TERT Promoter (92%), PTEN (66%), and TP53 (60%). Of the 19 common genes in gliosarcoma, 6 were also common in both GBM and soft tissue sarcoma, 4 in GBM alone, 0 in soft tissue sarcoma alone, and 9 were more distinct to gliosarcoma. Of these, BRAF harbored an OncoKB level 1 designation, indicating its status as a predictive biomarker of response to an FDA-approved drug in certain cancers. EGFR, CDKN2A, NF1, and PTEN harbored level 4 designations in solid tumors, indicating biological evidence of these biomarkers predicting a drug-response. Gliosarcoma contains molecular features that overlap GBM and soft tissue sarcoma, as well as its own distinct genomic signatures. This may play a role in disease classification and inclusion criteria for clinical trials. Gliosarcoma mutations with potential therapeutic indications include BRAF, EGFR, CDKN2A, NF1, and PTEN.

The oncogene AAMDC links PI3K-AKT-mTOR signaling with metabolic reprograming in estrogen receptor-positive breast cancer.

Adipogenesis associated Mth938 domain containing (AAMDC) represents an uncharacterized oncogene amplified in aggressive estrogen receptor-positive breast cancers. We uncover that AAMDC regulates the expression of several metabolic enzymes involved in the one-carbon folate and methionine cycles, and lipid metabolism. We show that AAMDC controls PI3K-AKT-mTOR signaling, regulating the translation of ATF4 and MYC and modulating the transcriptional activity of AAMDC-dependent promoters. High AAMDC expression is associated with sensitization to dactolisib and everolimus, and these PI3K-mTOR inhibitors exhibit synergistic interactions with anti-estrogens in IntClust2 models. Ectopic AAMDC expression is sufficient to activate AKT signaling, resulting in estrogen-independent tumor growth. Thus, AAMDC-overexpressing tumors may be sensitive to PI3K-mTORC1 blockers in combination with anti-estrogens. Lastly, we provide evidence that AAMDC can interact with the RabGTPase-activating protein RabGAP1L, and that AAMDC, RabGAP1L, and Rab7a colocalize in endolysosomes. The discovery of the RabGAP1L-AAMDC assembly platform provides insights for the design of selective blockers to target malignancies having the AAMDC amplification.

The anti-inflammatory effects of interleukin-4 are not mediated by suppressor of cytokine signalling-1 (SOCS1).

While it is known that the anti-inflammatory effects of interleukin (IL)-4 require new protein synthesis, the exact mechanisms by which IL-4 suppresses the production of pro-inflammatory cytokines by human monocytes and macrophages is unclear. IL-4 rapidly induced suppressor of cytokine signalling-1 (SOCS1) mRNA and protein, which peaked at 60 min, much earlier than lipopolysaccharide (LPS)-induced SOCS1 mRNA and protein which were consistently maximal 4 hr post-exposure. SOCS1 is a molecule generally considered to be induced for negative feedback of inflammatory processes. We investigated whether the early induction of SOCS1 by IL-4 was responsible for the suppression of LPS-induced tumour necrosis factor (TNF)-alpha production by IL-4. IL-4 suppressed LPS-induced TNF-alpha in freshly isolated monocytes at the level of transcription but acted by a different, possibly translational, mechanism in monocytes cultured overnight in macrophage colony-stimulating factor (M-CSF). Despite different modes of regulation by IL-4, the kinetics and magnitude of induction of SOCS1 mRNA and protein by IL-4 in the two cell types were identical. There was no significant difference in the suppression by IL-4 of LPS-induced TNF-alpha production by bone-marrow derived macrophages from wild-type mice, Ifngamma(-/-) mice and mice lacking SOCS1 (Socs1(-/-)Ifngamma(-/-)). These data suggest that SOCS1 is not involved in the suppression of LPS-induced TNF-alpha production by IL-4.

SOCS1 regulates the IFN but not NFkappaB pathway in TLR-stimulated human monocytes and macrophages.

SOCS1 can regulate TLR-mediated signal transduction, yet mechanistic studies in murine macrophages have been confusing and contradictory. This study has used an adenoviral transfection system to determine the role of SOCS1 in the regulation of TNF-alpha production by activated human monocytes. Monocytes were infected with AdV-SOCS1 or with an empty vector control, AdV-GFP, for 24 h before activation with the TLR4 ligand, LPS. SOCS1 did not regulate TNF-alpha mRNA or protein production within the first two hours of TLR4 activation. However, SOCS1 suppressed the sustained production of TNF-alpha by primary human monocytes and synovial fluid macrophages ex vivo. In addition, SOCS1 regulated the production of IL-6, but not IL-10, by monocytes. Analysis of the early signaling pathway downstream of TLR4 demonstrated that SOCS1 had no regulatory effect on the activation or on the DNA binding capacity of NFkappaB. The late effects of LPS are mediated in part through the MyD88-independent pathway activating IRF3 and initiating the production of IFN-beta. In response to adenoviral infection and before LPS exposure, monocytes expressed enhanced levels of IFN-beta and Myxovirus A mRNA, an anti-viral molecule characterizing IFN-beta activity. These two genes were reduced in AdV-SOCS1-infected cells. Further, SOCS1 regulated IFN-dependent pathways in LPS-activated cells as evidenced by reduced IFN-beta production and STAT1 phosphorylation. Using AdV-infection to dissect SOCS1 control of IFN-dependent pathways, this study suggests that SOCS1-regulation of the IFN-dependent component of the LPS-induced TLR4 signaling pathway may contribute to the down-regulation of inflammatory cytokine production by AdV-SOCS1-infected human monocytes.

Rab GTPases: Emerging Oncogenes and Tumor Suppressive Regulators for the Editing of Survival Pathways in Cancer.

The Rab GTPase family of proteins are mediators of membrane trafficking, conferring identity to the cell membranes. Recently, Rab and Rab-associated factors have been recognized as major regulators of the intracellular positioning and activity of signaling pathways regulating cell growth, survival and programmed cell death or apoptosis. Membrane trafficking mediated by Rab proteins is controlled by intracellular localization of Rab proteins, Rab-membrane interactions and GTP-activation processes. Aberrant expression of Rab proteins has been reported in multiple cancers such as lung, brain and breast malignancies. Mutations in Rab-coding genes and/or post-translational modifications in their protein products disrupt the cellular vesicle trafficking network modulating tumorigenic potential, cellular migration and metastatic behavior. Conversely, Rabs also act as tumor suppressive factors inducing apoptosis and inhibiting angiogenesis. Deconstructing the signaling mechanisms modulated by Rab proteins during apoptosis could unveil underlying molecular mechanisms that may be exploited therapeutically to selectively target malignant cells.