Noncanonical Roles of tRNAs: tRNA Fragments and Beyond.

As one of the most abundant and conserved RNA species, transfer RNAs (tRNAs) are well known for their role in reading the codons on messenger RNAs and translating them into proteins. In this review, we discuss the noncanonical functions of tRNAs. These include tRNAs as precursors to novel small RNA molecules derived from tRNAs, also called tRNA-derived fragments, that are abundant across species and have diverse functions in different biological processes, including regulating protein translation, Argonaute-dependent gene silencing, and more. Furthermore, the role of tRNAs in biosynthesis and other regulatory pathways, including nutrient sensing, splicing, transcription, retroelement regulation, immune response, and apoptosis, is reviewed. Genome organization and sequence variation of tRNA genes are also discussed in light of their noncanonical functions. Lastly, we discuss the recent applications of tRNAs in genome editing and microbiome sequencing.

XRN2 suppresses aberrant entry of tRNA trailers into argonaute in humans and Arabidopsis.

MicroRNAs (miRNAs) are a well-characterized class of small RNAs (sRNAs) that regulate gene expression post-transcriptionally. miRNAs function within a complex milieu of other sRNAs of similar size and abundance, with the best characterized being tRNA fragments or tRFs. The mechanism by which the RNA-induced silencing complex (RISC) selects for specific sRNAs over others is not entirely understood in human cells. Several highly expressed tRNA trailers (tRF-1s) are strikingly similar to microRNAs in length but are generally excluded from the microRNA effector pathway. This exclusion provides a paradigm for identifying mechanisms of RISC selectivity. Here, we show that 5' to 3' exoribonuclease XRN2 contributes to human RISC selectivity. Although highly abundant, tRF-1s are highly unstable and degraded by XRN2 which blocks tRF-1 accumulation in RISC. We also find that XRN mediated degradation of tRF-1s and subsequent exclusion from RISC is conserved in plants. Our findings reveal a conserved mechanism that prevents aberrant entry of a class of highly produced sRNAs into Ago2.

The tumor-suppressive long noncoding RNA DRAIC inhibits protein translation and induces autophagy by activating AMPK.

Long noncoding RNAs (lncRNAs) are long RNA transcripts that do not code for proteins and have been shown to play a major role in cellular processes through diverse mechanisms. DRAIC, a lncRNA that is downregulated in castration-resistant advanced prostate cancer, inhibits the NF-κB pathway by inhibiting the IκBα kinase. Decreased DRAIC expression predicted poor patient outcome in gliomas and seven other cancers. We now report that DRAIC suppresses invasion, migration, colony formation and xenograft growth of glioblastoma-derived cell lines. DRAIC activates AMP-activated protein kinase (AMPK) by downregulating the NF-κB target gene GLUT1, and thus represses mTOR, leading to downstream effects, such as a decrease in protein translation and increase in autophagy. DRAIC, therefore, has an effect on multiple signal transduction pathways that are important for oncogenesis, namely, the NF-κB pathway and AMPK-mTOR-S6K/ULK1 pathway. The regulation of NF-κB, protein translation and autophagy by the same lncRNA explains the tumor-suppressive role of DRAIC in different cancers and reinforces the importance of lncRNAs as emerging regulators of signal transduction pathways. This article has an associated First Person interview with the first author of the paper.

MicroDNA levels are dependent on MMEJ, repressed by c-NHEJ pathway, and stimulated by DNA damage.

Extrachromosomal circular DNA (eccDNA) are present within all eukaryotic organisms and actively contribute to gene expression changes. MicroDNA (200-1000bp) are the most abundant type of eccDNA and can amplify tRNA, microRNA, and novel si-like RNA sequences. Due to the heterogeneity of microDNA and the limited technology to directly quantify circular DNA molecules, the specific DNA repair pathways that contribute to microDNA formation have not been fully elucidated. Using a sensitive and quantitative assay that quantifies eight known abundant microDNA, we report that microDNA levels are dependent on resection after double-strand DNA break (DSB) and repair by Microhomology Mediated End Joining (MMEJ). Further, repair of DSB without resection by canonical Non-Homologous End Joining (c-NHEJ) diminishes microDNA formation. MicroDNA levels are induced locally even by a single site-directed DSB, suggesting that excision of genomic DNA by two closely spaced DSB is not necessary for microDNA formation. Consistent with all this, microDNA levels accumulate as cells undergo replication in S-phase, when DNA breaks and repair are elevated, and microDNA levels are decreased if DNA synthesis is prevented. Thus, formation of microDNA occurs during the repair of endogenous or induced DNA breaks by resection-based DNA repair pathways.

Promoting parent-child relationships and preventing violence via home-visiting: a pre-post cluster randomised trial among Rwandan families linked to social protection programmes.

BACKGROUND: Sugira Muryango is a father-engaged early child development and violence-prevention home-visiting programme delivered by trained lay workers. This cluster-randomised trial evaluates whether families living in extreme poverty (Ubudehe 1, the poorest category in the Government of Rwanda's wealth ranking) who receive Sugira Muryango in combination with a government-provided social protection programme demonstrate greater responsive, positive caregiving, nutrition, care seeking, hygiene, and father involvement compared with control families receiving usual care (UC). METHODS: Using detailed maps, we grouped closely spaced villages into 284 geographic clusters stratified by the type of social protection programmes operating in the village clusters; 198 clusters met all enrolment criteria. Sugira Muryango was delivered to n = 541 families in 100 treatment clusters with children aged 6-36 months living in extreme poverty. We assessed changes in outcomes in intervention and n = 508 UC control families using structured surveys and observation. Analyses were intent to treat using mixed models to accommodate clustering. RESULTS: Families receiving Sugira Muryango improved on core outcomes of parent-child relationships assessed using the Home Observation for Measurement of the Environment (Cohen's d = 0.87, 95% CI: 0.74, 0.99) and the Observation of Mother-Child Interaction (Cohen's d = 0.29, 95% CI: 0.17, 0.41). We also saw reductions in harsh discipline on items from the UNICEF MICS (OR = 0.30: 95% CI: 0.19, 0.47) and in violent victimisation of female caregivers by their partners (OR = 0.49, 95% CI: 0.24, 1.00) compared with UC. Moreover, children in families receiving SM had a 0.45 higher increase in food groups consumed in the past 24 h (Cohen's d = 0.35, 95% CI: 0.22, 0.47), increased care seeking for diarrhoea (OR = 4.43, 95% CI: 1.95, 10.10) and fever (OR = 3.28, 95% CI: 1.82, 5.89), and improved hygiene behaviours such as proper treatment of water (OR = 3.39, 95% CI: 2.16, 5.30) compared with UC. Finally, Sugira Muryango was associated with decreased caregiver depression and anxiety (OR = 0.58, 95% CI: 0.38, 0.88). CONCLUSIONS: Sugira Muryango led to improvements in caregiver behaviours linked to child development and health as well as reductions in violence. TRIAL REGISTRATION: ClinicalTrials.gov number NCT02510313.

sRNA-Effector: A tool to expedite discovery of small RNA regulators.

microRNAs (miRNAs) are small regulatory RNAs that repress target mRNA transcripts through base pairing. Although the mechanisms of miRNA production and function are clearly established, new insights into miRNA regulation or miRNA-mediated gene silencing are still emerging. In order to facilitate the discovery of miRNA regulators or effectors, we have developed sRNA-Effector, a machine learning algorithm trained on enhanced crosslinking and immunoprecipitation sequencing and RNA sequencing data following knockdown of specific genes. sRNA-Effector can accurately identify known miRNA biogenesis and effector proteins and identifies 9 putative regulators of miRNA function, including serine/threonine kinase STK33, splicing factor SFPQ, and proto-oncogene BMI1. We validated the role of STK33, SFPQ, and BMI1 in miRNA regulation, showing that sRNA-Effector is useful for identifying new players in small RNA biology. sRNA-Effector will be a web tool available for all researchers to identify potential miRNA regulators in any cell line of interest.

Tumor necrosis factor α antagonism improves neurological recovery in murine intracerebral hemorrhage.

BACKGROUND: Intracerebral hemorrhage (ICH) is a devastating stroke subtype characterized by a prominent neuroinflammatory response. Antagonism of pro-inflammatory cytokines by specific antibodies represents a compelling therapeutic strategy to improve neurological outcome in patients after ICH. To test this hypothesis, the tumor necrosis factor alpha (TNF-α) antibody CNTO5048 was administered to mice after ICH induction, and histological and functional endpoints were assessed. METHODS: Using 10 to 12-week-old C57BL/6J male mice, ICH was induced by collagenase injection into the left basal ganglia. Brain TNF-α concentration, microglia activation/macrophage recruitment, hematoma volume, cerebral edema, and rotorod latency were assessed in mice treated with the TNF-α antibody, CNTO5048, or vehicle. RESULTS: After ICH induction, mice treated with CNTO5048 demonstrated reduction in microglial activation/macrophage recruitment compared to vehicle-treated animals, as assessed by unbiased stereology (P = 0.049). This reduction in F4/80-positive cells was associated with a reduction in cleaved caspase-3 (P = 0.046) and cerebral edema (P = 0.026) despite similar hematoma volumes, when compared to mice treated with vehicle control. Treatment with CNTO5048 after ICH induction was associated with a reduction in functional deficit when compared to mice treated with vehicle control, as assessed by rotorod latencies (P = 0.024). CONCLUSIONS: Post-injury treatment with the TNF-α antibody CNTO5048 results in less neuroinflammation and improved functional outcomes in a murine model of ICH.

Measuring the cost-effectiveness of a home-visiting intervention to promote early child development among rural families linked to the Rwandan social protection system.

Early childhood development (ECD) programmes are heralded as a way to improve children's health and educational outcomes. However, few studies in developing countries calculate the effectiveness of quality early childhood interventions. This study estimates the cost and cost-effectiveness of the Sugira Muryango (SM) trial, a home-visiting intervention to improve ECD outcomes through positive parent-child relationships. Cost-effectiveness analysis of ECD interventions is challenging given their potential to have multiple benefits. We propose a cost-effectiveness method using a single outcome, in this case the improvement in cognitive development per home-visit session, as an indication of efficiency comparable across similar interventions. The trial intervention cost US$456 per family. This cost will likely fall below US$200 if the intervention is scaled through government systems. The cost-effectiveness analysis suggests that while SM generated a relatively small impact on markers of early development, it did so efficiently. The observed improvements in cognitive development per home-visit are similar to other home-visiting interventions of longer duration. SM by focusing on the family had benefits beyond ECD, including reductions in violence against children and intermate partner violence, further analysis is needed to include these returns in the economic evaluation.

Function and Therapeutic Implications of tRNA Derived Small RNAs.

tRNA derived small RNAs are mainly composed of tRNA fragments (tRFs) and tRNA halves (tiRs). Several functions have been attributed to tRFs and tiRs since their initial characterizations, spanning all aspects of regulation of the Central Dogma: from nascent RNA silencing, to post-transcriptional gene silencing, and finally, to translational regulation. The length distribution, sequence diversity, and multifaceted functions of tRFs and tiRs positions them as attractive new models for small RNA therapeutics. In this review, we will discuss the principles of tRF biogenesis and function in order to highlight their therapeutic potential.

tRForest: a novel random forest-based algorithm for tRNA-derived fragment target prediction.

tRNA fragments (tRFs) are small RNAs comparable to the size and function of miRNAs. tRFs are generally Dicer independent, are found associated with Ago, and can repress expression of genes post-transcriptionally. Given that this expands the repertoire of small RNAs capable of post-transcriptional gene expression, it is important to predict tRF targets with confidence. Some attempts have been made to predict tRF targets, but are limited in the scope of tRF classes used in prediction or limited in feature selection. We hypothesized that established miRNA target prediction features applied to tRFs through a random forest machine learning algorithm will immensely improve tRF target prediction. Using this approach, we show significant improvements in tRF target prediction for all classes of tRFs and validate our predictions in two independent cell lines. Finally, Gene Ontology analysis suggests that among the tRFs conserved between mice and humans, the predicted targets are enriched significantly in neuronal function, and we show this specifically for tRF-3009a. These improvements to tRF target prediction further our understanding of tRF function broadly across species and provide avenues for testing novel roles for tRFs in biology. We have created a publicly available website for the targets of tRFs predicted by tRForest.

TRMT6/61A-dependent base methylation of tRNA-derived fragments regulates gene-silencing activity and the unfolded protein response in bladder cancer.

RNA modifications are important regulatory elements of RNA functions. However, most genome-wide mapping of RNA modifications has focused on messenger RNAs and transfer RNAs, but such datasets have been lacking for small RNAs. Here we mapped N1-methyladenosine (m1A) in the cellular small RNA space. Benchmarked with synthetic m1A RNAs, our workflow identified specific groups of m1A-containing small RNAs, which are otherwise disproportionally under-represented. In particular, 22-nucleotides long 3' tRNA-fragments are highly enriched for TRMT6/61A-dependent m1A located within the seed region. TRMT6/61A-dependent m1A negatively affects gene silencing by tRF-3s. In urothelial carcinoma of the bladder, where TRMT6/61A is over-expressed, higher m1A modification on tRFs is detected, correlated with a dysregulation of tRF targetome. Lastly, TRMT6/61A regulates tRF-3 targets involved in unfolded protein response. Together, our results reveal a mechanism of regulating gene expression via base modification of small RNA.

Prophylactic low-dose, bi-weekly benznidazole treatment fails to prevent Trypanosoma cruzi infection in dogs under intense transmission pressure.

Trypanosoma cruzi naturally infects a wide variety of wild and domesticated mammals, in addition to humans. Depending on the infection dose and other factors, the acute infection can be life-threatening, and in all cases, the risk of chagasic heart disease is high in persistently infected hosts. Domestic, working, and semi-feral dogs in the Americas are at significant risk of T. cruzi infection and in certain settings in the southern United States, the risk of new infections can exceed 30% per year, even with the use of vector control protocols. In this study, we explored whether intermittent low-dose treatment with the trypanocidal compound benznidazole (BNZ) during the transmission season, could alter the number of new infections in dogs in an area of known, intense transmission pressure. Preliminary studies in mice suggested that twice-weekly administration of BNZ could prevent or truncate infections when parasites were delivered at the mid-point between BNZ doses. Pre-transmission season screening of 126 dogs identified 53 dogs (42.1%) as T. cruzi infection positive, based upon blood PCR and Luminex-based serology. Serial monitoring of the 67 uninfected dogs during the high transmission season (May to October) revealed 15 (22.4%) new infections, 6 in the untreated control group and 9 in the group receiving BNZ prophylaxis, indicating no impact of this prophylaxis regimen on the incidence of new infections. Although these studies suggest that rigorously timed and more potent dosing regimen may be needed to achieve an immediate benefit of prophylaxis, additional studies would be needed to determine if drug prophylaxis reduced disease severity despite this failure to prevent new infections.

Developing a curriculum framework for global health in family medicine: emerging principles, competencies, and educational approaches.

BACKGROUND: Recognizing the growing demand from medical students and residents for more comprehensive global health training, and the paucity of explicit curricula on such issues, global health and curriculum experts from the six Ontario Family Medicine Residency Programs worked together to design a framework for global health curricula in family medicine training programs. METHODS: A working group comprised of global health educators from Ontario's six medical schools conducted a scoping review of global health curricula, competencies, and pedagogical approaches. The working group then hosted a full day meeting, inviting experts in education, clinical care, family medicine and public health, and developed a consensus process and draft framework to design global health curricula. Through a series of weekly teleconferences over the next six months, the framework was revised and used to guide the identification of enabling global health competencies (behaviours, skills and attitudes) for Canadian Family Medicine training. RESULTS: The main outcome was an evidence-informed interactive framework http://globalhealth.ennovativesolution.com/ to provide a shared foundation to guide the design, delivery and evaluation of global health education programs for Ontario's family medicine residency programs. The curriculum framework blended a definition and mission for global health training, core values and principles, global health competencies aligning with the Canadian Medical Education Directives for Specialists (CanMEDS) competencies, and key learning approaches. The framework guided the development of subsequent enabling competencies. CONCLUSIONS: The shared curriculum framework can support the design, delivery and evaluation of global health curriculum in Canada and around the world, lay the foundation for research and development, provide consistency across programmes, and support the creation of learning and evaluation tools to align with the framework. The process used to develop this framework can be applied to other aspects of residency curriculum development.

HPVE6-USP46 Mediated Cdt2 Stabilization Reduces Set8 Mediated H4K20-Methylation to Induce Gene Expression Changes.

E6 from high-risk strains of HPV is well known to transform cells by deregulating p53. We reported that in HPV transformed cell-lines E6 from high-risk HPV can recruit the USP46 deubiquitinase to substrates such as Cdt2 and stabilize the latter, and that USP46 is important for growth of HPV induced tumors in xenografts. Here we show that in cervical cancer biopsies the stabilization of Cdt2 in the HPV-induced cancers leads to the decrease of a CRL4-Cdt2 substrate, the histone H4K20 mono-methyltransferase Set8, and decrease in H4K20me1 or H4K20me3 that can be detected by immunohistochemistry. In HPV-transformed cancer cell lines in vitro, knockdown of E6 decreases Cdt2 and increases Set8. Co-knockdown of Set8 shows that some of the gene expression changes produced by E6 knockdown is due to the increase of Set8. EGFR and EGFR regulated genes were identified in this set of genes. Turning to the mechanism by which E6 stabilizes Cdt2, we find that a purified E6:USP46 complex has significantly more de-ubiquitinase activity in vitro than USP46 alone, demonstrating that E6 can directly interact with USP46 in the absence of other proteins and that it can substitute for the known activators of USP46, UAF1 and WDR20. Deletion mapping of Cdt2 shows that there are three discrete, but redundant, parts of the substrate that are essential for stabilization by E6: USP46. The helix-loop-helix region or the WD40 repeat driven beta-propeller structure of Cdt2 are dispensable for the stabilization implying that interaction with DDB1 (and the rest of the CRL4 complex) or with the substrate of the CRL4-Cdt2 E3 ligase is not necessary for E6:USP46 to interact with and stabilize Cdt2. The identification of 50 amino acid stretches in the 731 amino acid Cdt2 protein as being important for the stabilization by E6 underlines the specificity of the process. In summary, E6 activates the deubiquitinase activity of USP46, stabilizes Cdt2 utilizing multiple sites on Cdt2, and leads to degradation of Set8 and changes in gene-expression in HPV-transformed cells.

Effect of a home-visiting parenting program to promote early childhood development and prevent violence: a cluster-randomized trial in Rwanda.

INTRODUCTION: Families living in extreme poverty require interventions to support early-childhood development (ECD) due to broad risks. This longitudinal cluster randomised trial examines the effectiveness of Sugira Muryango (SM), a home-visiting intervention linked to Rwanda's social protection system to promote ECD and reduce violence compared with usual care (UC). METHODS: Families with children aged 6-36 months were recruited in 284 geographical clusters across three districts. Cluster-level randomisation (allocated 1:1 SM:UC) was used to prevent diffusion. SM was hypothesised to improve child development, reduce violence and increase father engagement. Developmental outcomes were assessed using the Ages and Stages Questionnaire (ASQ-3) and the Malawi Development Assessment Tool (MDAT) and anthropometric assessments of growth. Violence was assessed using questions from UNICEF Multiple Indicators Cluster Survey (MICS) and Rwanda Demographic and Health Surveys (DHS). Father engagement was assessed using the Home Observation for Measurement of the Environment. Blinded enumerators conducted interviews and developmental assessments. RESULTS: A total of 541 SM families and 508 UC families were enrolled and included in the analyses. Study attrition (2.0% children; 9.6% caregivers) was addressed by hot deck imputation. Children in SM families improved more on gross motor (d=0.162, 95% CI 0.065 to 0.260), communication (d=0.081, 95% CI 0.005 to 0.156), problem solving (d=0.101, 95% CI 0.002 to 0.179) and personal-social development (d=0.096, 95% CI -0.015 to 0.177) on the ASQ-3. SM families showed increased father engagement (OR=1.592, 95% CI 1.069 to 2.368), decreased harsh discipline (incidence rate ratio, IRR=0.741, 95% CI 0.657 to 0.835) and intimate partner violence (IRR=0.616, 95% CI:0.458 to 0.828). There were no intervention-related improvements on MDAT or child growth. CONCLUSION: Social protection programmes provide a means to deliver ECD intervention. TRIAL REGISTRATION NUMBER: NCT02510313.

Extrinsic interactions in the microenvironment in vivo activate an antiapoptotic multidrug-resistant phenotype in CLL.

The Bcl-2 inhibitor venetoclax has yielded exceptional clinical responses in chronic lymphocytic leukemia (CLL). However, de novo resistance can result in failure to achieve negative minimal residual disease and predicts poor treatment outcomes. Consequently, additional proapoptotic drugs, such as inhibitors of Mcl-1 and Bcl-xL, are in development. By profiling antiapoptotic proteins using flow cytometry, we find that leukemic B cells that recently emigrated from the lymph node (CD69+/CXCR4Low) in vivo are enriched for cell clusters simultaneously overexpressing multiple antiapoptotic proteins (Mcl-1High/Bcl-xLHigh/Bcl-2High) in both treated and treatment-naive CLL patients. These cells exhibited antiapoptotic resistance to multiple BH-domain antagonists, including inhibitors of Bcl-2, Mcl-1, and Bcl-xL, when tested as single agents in a flow cytometry-based functional assay. Antiapoptotic multidrug resistance declines ex vivo, consistent with resistance being generated in vivo by extrinsic microenvironmental interactions. Surviving "persister" cells in patients undergoing venetoclax treatment are enriched for CLL cells displaying the functional and molecular properties of microenvironmentally induced multidrug resistance. Overcoming this resistance required simultaneous inhibition of multiple antiapoptotic proteins, with potential for unwanted toxicities. Using a drug screen performed using patient peripheral blood mononuclear cells cultured in an ex vivo microenvironment model, we identify novel venetoclax drug combinations that induce selective cytotoxicity in multidrug-resistant CLL cells. Thus, we demonstrate that antiapoptotic multidrug-resistant CLL cells exist in patients de novo and show that these cells persist during proapoptotic treatment, such as venetoclax. We validate clinically actionable approaches to selectively deplete this reservoir in patients.

The pan-cancer landscape of prognostic germline variants in 10,582 patients.

BACKGROUND: While clinical factors such as age, grade, stage, and histological subtype provide physicians with information about patient prognosis, genomic data can further improve these predictions. Previous studies have shown that germline variants in known cancer driver genes are predictive of patient outcome, but no study has systematically analyzed multiple cancers in an unbiased way to identify genetic loci that can improve patient outcome predictions made using clinical factors. METHODS: We analyzed sequencing data from the over 10,000 cancer patients available through The Cancer Genome Atlas to identify germline variants associated with patient outcome using multivariate Cox regression models. RESULTS: We identified 79 prognostic germline variants in individual cancers and 112 prognostic germline variants in groups of cancers. The germline variants identified in individual cancers provide additional predictive power about patient outcomes beyond clinical information currently in use and may therefore augment clinical decisions based on expected tumor aggressiveness. Molecularly, at least 12 of the germline variants are likely associated with patient outcome through perturbation of protein structure and at least five through association with gene expression differences. Almost half of these germline variants are in previously reported tumor suppressors, oncogenes or cancer driver genes with the other half pointing to genomic loci that should be further investigated for their roles in cancers. CONCLUSIONS: Germline variants are predictive of outcome in cancer patients and specific germline variants can improve patient outcome predictions beyond predictions made using clinical factors alone. The germline variants also implicate new means by which known oncogenes, tumor suppressor genes, and driver genes are perturbed in cancer and suggest roles in cancer for other genes that have not been extensively studied in oncology. Further studies in other cancer cohorts are necessary to confirm that germline variation is associated with outcome in cancer patients as this is a proof-of-principle study.

Siderophores in Iron Metabolism: From Mechanism to Therapy Potential.

Iron is an essential nutrient for life. During infection, a fierce battle of iron acquisition occurs between the host and bacterial pathogens. Bacteria acquire iron by secreting siderophores, small ferric iron-binding molecules. In response, host immune cells secrete lipocalin 2 (also known as siderocalin), a siderophore-binding protein, to prevent bacterial reuptake of iron-loaded siderophores. To counter this threat, some bacteria can produce lipocalin 2-resistant siderophores. This review discusses the recently described molecular mechanisms of siderophore iron trafficking between host and bacteria, highlighting the therapeutic potential of exploiting pathogen siderophore machinery for the treatment of antibiotic-resistant bacterial infections. Because the latter reflect a persistent problem in hospital settings, siderophore-targeting or siderophore-based compounds represent a promising avenue to combat such infections.

Gender and age interact to affect early outcome after intracerebral hemorrhage.

BACKGROUND: Intracerebral hemorrhage (ICH) is a common and devastating form of cerebrovascular disease. In ICH, gender differences in outcomes remain relatively understudied but have been examined in other neurological emergencies. Further, a potential effect of age and gender on outcomes after ICH has not been explored. This study was designed to test the hypothesis that age and gender interact to modify neurological outcomes after ICH. METHODS: Adult patients admitted with spontaneous primary supratentorial ICH from July 2007 through April 2010 were assessed via retrospective analysis of an existing stroke database at Duke University. Univariate analysis of collected variables was used to compare gender and outcome. Unfavorable outcome was defined as discharge to hospice or death. Using multivariate regression, the combined effect of age and gender on outcome after ICH was analyzed. RESULTS: In this study population, women were younger (61.1+14.5 versus 65.8+17.3 years, p=0.03) and more likely to have a history of substance abuse (35% versus 8.9%, p<0.0001) compared to men. Multivariable models demonstrated that advancing age had a greater effect on predicting discharge outcome in women compared to men (p=0.02). For younger patients, female sex was protective; however, at ages greater than 60 years, female sex was a risk factor for discharge to hospice or death. CONCLUSION: While independently associated with discharge to hospice or death after ICH, the interaction effect between gender and age demonstrated significantly stronger correlation with early outcome after ICH in a single center cohort. Prospective study is required to verify these findings.