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PURPOSESeveral groups and resources provide information that pertains to the validity of gene-disease relationships used in genomic medicine and research; however, universal standards and terminologies to define the evidence base for the role of a gene in disease, and a single harmonized resource were lacking. To tackle this issue, the Gene Curation Coalition (GenCC) was formed.METHODSThe GenCC drafted harmonized definitions for differing levels of gene-disease validity based on existing resources, and performed a modified Delphi survey with three rounds to narrow the list of terms. The GenCC also developed a unified database to display curated gene-disease validity assertions from its members.RESULTSBased on 241 survey responses from the genetics community, a consensus term set was chosen for grading gene-disease validity and database submissions. As of December 2021, the database contains 15,241 gene-disease assertions on 4,569 unique genes from 12 submitters. When comparing submissions to the database from distinct sources, conflicts in assertions of gene-disease validity ranged from 5.3% to 13.4%.CONCLUSIONTerminology standardization, sharing of gene-disease validity classifications, and resolution of curation conflicts will facilitate collaborations across international curation efforts and in turn, improve consistency in genetic testing and variant interpretation.

Cardiovascular disease (CVD) is a major cause of mortality among people with diabetic kidney disease (DKD). The pathophysiology is inadequately explained by traditional CVD risk factors. The uremic solutes trimethylamine-N-oxide (TMAO) and asymmetric and symmetric dimethylarginine (ADMA, SDMA) have been linked to CVD in kidney failure with replacement therapy (KFRT), but data are limited in populations with diabetes and less severe kidney disease.Observational cohort.Random subcohort of 555 REGARDS (Reasons for Geographic and Racial Differences in Stroke) study participants with diabetes and estimated glomerular filtration rate (eGFR) 60 mL/min/1.73 mADMA, SDMA, and TMAO assayed by liquid chromatography-mass spectrometry in plasma and urine.Cardiovascular mortality (primary outcome); all-cause mortality and incident KFRT (secondary outcomes).Plasma concentrations and ratios of urine to plasma concentrations of ADMA, SDMA, and TMAO were tested for association with outcomes. Adjusted Cox regression models were fitted and hazard ratios of outcomes calculated per standard deviation and per doubling, and as interquartile comparisons.The mean baseline eGFR was 44 mL/min/1.73 mSingle cohort, restricted to patients with diabetes and eGFR 60 mL/min/1.73 mHigher plasma concentrations and lower ratios of urine to plasma concentrations of uremic solutes were independently associated with cardiovascular and all-cause mortality in DKD. Associations of ratios of urine to plasma concentrations with mortality suggest a connection between renal uremic solute clearance and CVD pathogenesis.

The objective of this study was to explore nurses' perceptions of end-of-life care following the withdrawal of the Liverpool Care Pathway (LCP). Thirteen semi-structured interviews were conducted with nurses working in palliative care. Data were analysed using thematic analysis. Three themes emerged: perceptions of the LCP, prevailing issues, and patients' and families' experiences. This study suggested that the removal of the pathway has not remedied the issues attributed to it. Further, the way in which the LCP was removed indicates that the non-expert media can play a negative role in how palliative care is perceived, which inhibits the care process. In this respect it is important that ‘insider’ voices are also heard, in order to educate and also redress disinformation. Similarly, broader, persisting, contextual challenges facing staff need addressing in order to prevent a repeat of the issues leading to the removal of the LCP.

High-throughput sequencing technologies facilitate in-depth sequencing of T-cell receptor (TCR) repertoires to gain insights into human adaptive immunity. These studies can help decipher the diversity of T-cell receptors, relevant to physiological and disease conditions, and potential therapy responses. A variety of TCR kits and programs have been developed for the identification of V, D and J gene segments, complementarity-determining region 3 (CDR3) sequence extraction and clonality analysis. However, there is still a deficit of systematic comparative studies to assist in the selection of an optimal platform. Here, we present a detailed comparison of 4 state-of-the-art TCR platform on samples with different complexities (ranging from cancers to immune disorders) by taking into account aspects such as clonotype detection [unique V(D)J combination] and CDR3 identification. Our results provide new insights into the effect of method selection on analysis results, and will consequently inform users in the selection of an appropriate analysis method for the desired application(s). Citation Format: Mukta Dutta, Mariya Smit, Heather Collins, Anjali Malge, Steve Anderson, Corey Braastad. Comparative study of methods and platforms for T-cell receptor repertoire [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3178.

A Polycomb group protein, Posterior sex combs (Psc), was identified in a genetic screen designed to find factors that can specifically induce morphological defects in rbf1 mutant eyes. We discovered that rbf1 mutations enhance developmental phenotypes caused by Psc overexpression such as ectopic cell death and disorganized ommatidia. Our genetic analysis revealed that Psc-induced developmental defects are strongly influenced by CAF1p55, which is a shared component of several chromatin-associated complexes including a histone chaperone complex, chromatin assembly factor-1 (dCAF-1). Interestingly, the expression levels of dCAF-1 components, CAF1p105 and CAF1p180, are increased in rbf1 mutants, whereas the expression level of CAF1p55 itself remains relatively unchanged. We demonstrated that the increased levels of CAF1p105 and CAF1p180 are required for the hypersensitivity of rbf1 mutant cells to Psc-induced cell death and for the developmentally regulated cell death normally observed in rbf1 mutant eyes. We propose that Caf1p105 and Caf1p180 are important determinants of cell death sensitivity in rbf1 mutant cells and contribute to the genetic interaction between Psc and rbf1.

Cross-sectional.To evaluate the accuracy with which physical therapists identify fear-avoidance beliefs in patients with low back pain by comparing therapist ratings of perceived patient fear-avoidance to the Fear-Avoidance Beliefs Questionnaire (FABQ), Tampa Scale of Kinesiophobia 11-item (TSK-11), and Pain Catastrophizing Scale (PCS). To compare the concurrent validity of therapist ratings of perceived patient fear-avoidance and a 2-item questionnaire on fear of physical activity and harm, with clinical measures of fear-avoidance (FABQ, TSK-11, PCS), pain intensity as assessed with a numeric pain rating scale (NPRS), and disability as assessed with the Oswestry Disability Questionnaire (ODQ).The need to consider psychosocial factors for identifying patients at risk for disability and chronic low back pain has been well documented. Yet the ability of physical therapists to identify fear-avoidance beliefs using direct observation has not been studied.Eight physical therapists and 80 patients with low back pain from 3 physical therapy clinics participated in the study. Patients completed the FABQ, TSK-11, PCS, ODQ, NPRS, and a dichotomous 2-item fear-avoidance screening questionnaire. Following the initial evaluation, physical therapists rated perceived patient fear-avoidance on a 0-to-10 scale and recorded 2 influences on their ratings. Spearman correlation and independent t tests determined the level of association of therapist 0-to-10 ratings and 2-item screening with fear-avoidance and clinical measures.Therapist ratings of perceived patient fear-avoidance had fair to moderate interrater reliability (ICC2,1 = 0.663). Therapist ratings did not strongly correlate with FABQ or TSK-11 scores. Instead, they unexpectedly had stronger associations with ODQ and PCS scores. Both 2-item screening questions were associated with FABQ-physical activity scores, while the fear of physical activity question was also associated with FABQ-work, TSK-11, PCS, and ODQ scores.Therapists' ratings of perceived patient fear-avoidance were not associated with self-reported fear-avoidance scores, showing a potential disconnect between therapist judgments and commonly used fear-avoidance measures. Instead, therapist ratings had small but statistically significant correlations with pain catastrophizing and disability, findings that may support therapists' inability to discriminate fear-avoidance from these other factors. The 2-item screening questions based on fear of physical activity and harm showed potential to identify elevated FABQ physical activity scores.Differential diagnosis, level 2b.

Pedagogical agent research seeks to exploit Reeves and Nass's media equation theory, which holds that users respond to interactive media as if they were social actors. Investigations have tended to focus on the media used to realize the pedagogical agent, e.g., the use of animated talking heads and voices, and the results have been mixed. This paper focuses instead on the manner in which a pedagogical agent communicates with learners, i.e., on the extent to which it exhibits social intelligence. A model of socially intelligent tutorial dialog was developed based on politeness theory, and implemented in an agent interface within an online learning system called virtual factory teaching system. A series of Wizard-of-Oz studies was conducted in which subjects either received polite tutorial feedback that promotes learner face and mitigates face threat, or received direct feedback that disregards learner face. The polite version yielded better learning outcomes, and the effect was amplified in learners who expressed a preference for indirect feedback, who had less computer experience, and who lacked engineering backgrounds. These results confirm the hypothesis that learners tend to respond to pedagogical agents as social actors, and suggest that research should focus less on the media in which agents are realized, and place more emphasis on the agent's social intelligence.

Objective Recognising overweight and obesity is critical to prompting action, and consequently preventing and treating obesity. The present study examined the association between parental perceptions of child weight status and child’s diet. Methods Participants were members of the Gateshead Millennium Study. Parental perception of their child’s weight status was assessed using a questionnaire and compared against International Obesity Task Force cut-offs for childhood overweight and obesity when the children were aged 6–8 years old. Diet was assessed at age 6-8years old using the FAST (Food Assessment in Schools Tool) food diary method. The association between parental perception and dietary patterns as defined by Principal Components Analysis, was assessed using multivariate regression after adjustment for child’s gender, child’s weight status, maternal body mass index (BMI), maternal education and deprivation status. Results Of the 361 parents who provided complete data on confounders and on their perception of their child’s weight status, 63 (17%) parents perceived their child as being of ‘normal’ weight or ‘overweight’ when they were actually ‘overweight’ or ‘obese’, respectively. After adjustment for confounders, parents who misperceived their child’s weight had children with a lower ‘healthy’ dietary pattern score compared to children whose parents correctly perceived their weight (β = -0.88; 95% CI: -1.7, -0.1; P-value = 0.028). This association was found despite higher consumption of reduced sugar carbonated drinks amongst children whose parents incorrectly perceived their weight status compared to children whose parents perceived their weight correctly (52.4% vs. 33.6%; P-value = 0.005). Conclusions In conclusion, children whose parents did not correctly perceive their weight status scored lower on the ‘healthy’ dietary pattern. Further research is required to define parents’ diets based on their perception status and to examine if a child’s or parent’s diet mediates the association between parental perception and child weight.

This study surveyed current education majors (n=70) in two Kansas universities to gain a perspective on their understanding of Information Literacy (IL) concepts and skills, and to learn whether they anticipated teaching such concepts to their future K-12 students. School media specialists in the state were also surveyed (n=85) and asked to share their observations of teachers new to the profession as to their understanding and practice of IL. Results indicate many education students were not familiar with IL concept terminology and at least some new teachers in the state do not have a clear understanding or priority for teaching such skills in K-12 classrooms.

Simulation-based training has been shown to be a useful adjunct to standard didactic lecture in teaching residents appropriate management of adverse contrast reactions. In addition, it has been suggested that a biannual refresher is needed; however, the type of refresher education has not been assessed.This was a prospective study involving 31 radiology residents across all years in a university program. All residents underwent standard didactic lecture followed by high-fidelity simulation-based training. At approximately 6 months, residents were randomized into a didactic versus simulation group for a refresher. At approximately 9 months, all residents returned to the simulation center for performance testing. Knowledge and confidence assessments were obtained from all participants before and after each phase. Performance testing was obtained at each simulation session and scored based on predefined critical actions.There was significant improvement in knowledge (P .002) and confidence (P .001) after baseline education of combined didactic and simulation-based training. There was no statistical difference between the simulation and didactic groups in knowledge or confidence at any phase of the study. There was no significant difference in tested performance between the groups in either performance testing session.This study suggests that a curriculum consisting of an annual didactic lecture combined with simulation-based training followed by a didactic refresher at 6 months is an effective and efficient (both cost-effective and time-effective) method of educating radiology residents in the management of adverse contrast reactions.

Donor lymphocyte infusions (DLI) can induce remissions in patients who have relapsed after allogeneic bone marrow transplantation (BMT). However, DLI frequently also result in significant acute and/or chronic graft-versus-host disease (GVHD). Several clinical and experimental lines of evidence have suggested that CD8+ T cells play a critical role in the pathogenesis of GVHD. To develop methods to reduce the incidence of GVHD associated with DLI, we administered defined numbers of CD4+ donor T cells after ex vivo depletion of CD8+ lymphocytes to 40 patients with relapsed hematologic malignancies after allogeneic BMT. Cohorts of patients received 0.3, 1.0, or 1.5 × 108CD4+ cells/kg. Overall, 12 of 38 patients (32%) evaluable for toxicity developed acute or chronic GVHD. However, 6 of 27 patients (22%) receiving 0.3 × 108 CD4 cells/kg developed GVHD compared with 6 of 11 patients (55%) who received ≥1.0 × 108 CD4 cells/kg (P = .07). Treatment-related mortality was low (3%), with 1 death related to infection in the setting of immunosuppression for GVHD. Disease responses after CD4+ DLI were documented in 15 of 19 patients (79%) with early-phase chronic myelogenous leukemia (CML) relapse, 5 of 6 patients (83%) with relapsed multiple myeloma, and 1 patient with myelodysplasia. For patients with early-phase CML relapse, the Kaplan-Meier probability of achieving complete cytogenetic remission was 87% and the probability of complete molecular response was 78% at 1 year after DLI. The median time to complete cytogenetic response and molecular response in patients with CML was 13 weeks (range, 9 to 30 weeks) and 34 weeks (range, 10 to 56 weeks), respectively. The median time to response in patients with multiple myeloma was 26 weeks (range, 15 to 62 weeks). All patients in this trial who developed GVHD demonstrated tumor regression, but the presence of GVHD was not required for patients to achieve a response, because 48% of responding patients never developed evidence of GVHD. Two patients with CML who did not respond at dose level 1 subsequently achieved complete cytogenetic remission after a second infusion of CD8-depleted cells at dose level 2. In patients with evidence of mixed hematopoietic chimerism who achieved a complete remission after DLI, cytogenetic analysis of marrow cells also demonstrated conversion to complete donor hematopoiesis in all evaluable patients. These studies suggest that relatively low numbers of CD8-depleted donor lymphocytes are effective in inducing complete remissions in patients with stable-phase CML and multiple myeloma who have relapsed after allogeneic BMT. Because of the relatively low risk of toxicity associated with the infusion of defined numbers of CD4+donor cells, further studies can be undertaken in the setting of persistent minimal residual disease to prevent relapse after allogeneic BMT.

The clinical efficacy of donor lymphocyte infusions (DLI) in patients with relapsed chronic myelocytic leukemia after allogeneic bone marrow transplantation has been demonstrated in several recent studies. Although it is presumed that allogeneic T cells mediate this graft-versus-leukemia (GVL) effect, the influence of DLI on the T cell compartment of recipients has not been determined. To characterize the immunologic effects of DLI and to identify T cell changes selectively associated with the GVL response, we analyzed the T cell receptor (TCR) repertoire in four patients with relapsed chronic myelocytic leukemia who achieved a complete remission after infusion of CD4+ lymphocytes from HLA-identical sibling donors. Only one of the four patients developed clinically significant graft-versus-host disease (GVHD) after infusion of donor lymphocytes. TCR repertoire was examined after PCR amplification of 24 Vbeta gene subfamilies in serial samples obtained over a 1-yr period before and after DLI. Results were compared to 10 normal donors. Before DLI, all four patients were found to have abnormal TCR Vbeta repertoire in peripheral T cells, associated with a large number of clonal and oligoclonal patterns. Abnormal TCR patterns persisted for at least 3 mo after DLI, but thereafter gradually began to normalize. By 1 yr after DLI, all patients demonstrated almost complete normalization of Vbeta repertoire with polyclonal representation within almost all Vbeta gene subfamilies. We also examined changes in the TCR Vbeta repertoire associated with the disappearance of Ph+ cells. In each patient, we were able to identify the expansion of at least 1 Vbeta gene subfamily that coincided with the time of the cytogenetic response. In one patient who was studied in greater detail, CDR3 size analysis of serial samples after DLI indicated that these changes were associated with the appearance of clonal T cells. This finding was confirmed through CDR3 sequence analysis and use of CDR3 clone-specific oligonucleotide probes. A putative GVL clone identified by this technique was not detectable in either donor or patient T cells before DLI, but persisted in peripheral T cells for approximately 1 yr. These experiments therefore provide evidence for the clonal expansion of allogeneic T cells that may be selective mediators of antileukemia activity without also mediating graft-versus-host disease.

R24 is a monoclonal antibody directed against the cell surface ganglioside GD3. It can detect GD3 on the surface of a subset of T lymphocytes and can stimulate proliferation and secretion of cytokines in vitro. In the present report, we examined the effects of the R24 antibody upon antigen-specific T cell response, employing an HLA-DR7-specific T cell clonal model. As previously shown, primary stimulation of HLA-DR7-specific alloreactive T cell clones by transfectants expressing HLA-DR7 alone (t-DR7) in the absence of B7 co-stimulation resulted in anergy. Binding of cell surface GD3 on HLA-DR7-specific alloreactive T cell clones with R24 under these anergizing conditions resulted in interleukin-2 (IL-2) accumulation and prevented the induction of alloantigen-specific T cell clonal anergy. Binding of GD3 by R24 also prevented anergy under conditions where B7:CD28 interactions were blocked by CTLA4-Ig. The effect of R24 was abrogated in the presence of a combination of monoclonal antibodies for the alpha and beta chains of the IL-2 receptor (IL-2R) or a neutralizing anti-IL-2 antibody. R24 does not appear to interact directly with the IL-2R since incubation of T cell clones with R24 did not induce early activation of IL-2R associated Jak kinases, Jak1 and Jak3, as was induced following incubation with IL-2. In contrast, incubation of HLA-DR7-specific clones with t-DR7 in the presence of R24 did result in phosphorylation of IL-2R related Jak kinases after 24 h. Our data indicate that the membrane ganglioside GD3 structure recognized by R24 may play an important role in antigen-specific T cell clonal response.

Colorectal cancer (CRC) is the third most common type of cancer in the United States. Although chemotherapy, radiation and targeted therapies can improve survival rates, recent studies have shown the potential benefit of immunotherapies to improve outcomes for patients with advanced CRC. Targeted therapies that use monoclonal antibodies (mAbs) to EGFR have been shown to benefit some CRC patients. Until recently, KRAS has been the only predictive biomarker for anti-EGFR therapy for metastatic CRC. However, 40% to 60% of patients with wild-type KRAS do not respond to anti-EGFR therapy. Therefore, to accurately predict patients’ response to treatments and improve clinical outcomes, additional prediction and treatment methods are imperative. One of the many efforts to improve prediction for CRC patient's response to the anti-EGFR therapy is the development of gene expression based RAS signature scores for identification of RAS activated tumors independent of mutations in the KRAS gene. Recently there have been major advances in immunotherapeutic approaches in a wide variety of cancers. In solid tumors such as melanoma and colon cancers, immune checkpoints have been shown to improve clinical outcomes. There is considerable effort being placed on combinations of targeted therapy and immunotherapies to improve responses for these cancers. Similarly, since no single treatment can apply to all CRC patients, we aim to stratify patients using a combination of three methods: 1. RAS signature score based on the expression profile of 18 genes. This RAS signature score enables measurements of mitogen-activated protein/extracellular signal-regulated kinase (MEK) pathway functional output independent of tumor genotype. 2. Expression profile of immune checkpoint inhibitor target genes, such as PD1 and PDL1, and 3. In-silico prediction of neo-antigens and peptide binding affinity between tumor antigens derived from mutations and human HLA alleles. 55 FFPE samples were selected from a cohort of 468 samples with matching FF samples. These 55 samples have about 1:1:1 ratio of high, medium and low RAS scores. Here we showed our ability to obtain RAS signature scores with concordant results using different platforms including RNA-seq, targeted RNA-seq, Nanostring and Affymetrix microarray. Samples that have RAS activating mutations such as KRAS and BRAF have significant higher RAS scores (p Citation Format: FangYin Lo, Sharon Austin, Kellie Howard, Mollie McWhorter, Heather Collins, Amanda Leonti, Lindsey Maassel, Christopher Subia, Tuuli Saloranta, Nicole Christopherson, Kathryn Shiji, Shradha Patil, Saman Tahir, Sally Dow, Evan Anderson, Jon Oblad, Kerry Deutsch, Timothy Yeatman, Steven Anderson, Anup Madan. Stratification of metastatic colorectal cancer patients using DNA and RNA sequencing and in-silico prediction of tumor antigens for consideration in immunotherapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3946.

As the practice of genetically profiling patient tumors is considered for making clinical treatment decisions, recent methodologies for screening of genomic aberrations in circulating tumor cells (CTCs) and cell-free plasma DNA (cfDNA) may provide non-invasive tools for such applications. Genomic analysis of DNA from CTCs and plasma can also provide useful insight into tumor heterogeneity and thus disease progression by revealing sub-populations of tumor cells that evolve during treatment, have novel drug-resistant genotypes, or carry alternate cancer driver mutations not identified by the sequencing of primary tumors. Comprehensive evaluation of DNA isolated from CTCs and cfDNA from a breast cancer patient by whole exome sequencing was performed to better understand the role of liquid biopsies in investigating the etiology of tumor progression. The patient was diagnosed with metastatic triple negative breast cancer (TNBC) six years after remission from estrogen receptor (ER-3+), progesterone receptor (PR-1+), human epidermal receptor growth factor 2 negative (Her2-), grade 3 intra-ductal carcinoma of the right breast. Metastatic lesions were found in the spine, pelvis and sacrum and bone-marrow. The patient was enrolled in the Intensive Trial of OMics in Cancer clinical Trial (ITOMIC-001; ClinicalTrials.gov ID NCT01957514) and initially received weekly cisplatin infusions followed by additional targeted therapy. Peripheral blood was obtained during regular clinic visits over the 272 days the patient was enrolled in the clinical trial. CTCs were identified and enumerated from each blood draw using the AccuCyte® -CyteFinder® (AC/CF) system (RareCyte, Seattle WA). Multiple CTCs along with white blood cells (WBCs) were picked from various time points throughout the treatment regimen. The selected CTCs and WBCs were whole genome amplified and whole exome sequencing was performed to identify tumor specific variants. A comparative analysis with variants present in genomic DNA isolated from the bone-marrow metastasis tissue biopsy samples and cfDNA revealed the evolution of tumor-specific variants during therapy. Each CTC had somatic alterations in genes associated with therapies in current use or those in the clinical trials setting. Sequencing analysis of cfDNA provided similar information on potential therapeutic approaches. The monitoring of disease over time through genomic analysis of CTCs and cfDNA can identify novel sub-populations related to disease progression for the tailoring of cancer treatment regimens. Further analysis is being performed to better understand the evolution of the genomic heterogeneity among CTCs at the same time point and across different time points and therefore better understand the etiology of progression of metastatic breast cancer in this patient. Citation Format: Howard K, Austin S, Ramirez AB, Ritter L, Boles D, Pruitt J, Collins H, Mahen E, Leonti A, Maassel L, Subia C, Tuuli S, Heying N, Deutsch K, Cox J, Lo FY, Stilwell JL, Kaldjian EP, Dorschner M, Blau S, Blau A, Eisenberg M, Anderson S, Madan A. Longitudinal analysis of circulating tumor cells and cell free tumor DNA by next generation sequencing in triple negative breast cancer. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P2-02-21.

Imagine a simple clinical test that can not only diagnose a disease, but that can also identify the exact, personal therapeutic regime to cure it. Not only that, imagine tests that can accurately predict the potential of developing a disease and provide an individualized roadmap on how it will progress. Now imagine that all you had to do was to spit in a vial, or have a few hairs plucked for the analysis. While the promise of “personalized medicine” is technologically a reality, it relies on the development of disease and progression biomarkers.

Background Genome-wide gene expression profiling of whole blood is an attractive method for discovery of biomarkers due to its non-invasiveness, simple clinical site processing and rich biological content. Except for a few successes, this technology has not yet matured enough to reach its full potential of identifying biomarkers useful for clinical prognostic and diagnostic applications or in monitoring patient response to therapeutic intervention. A variety of technical problems have hampered efforts to utilize this technology for identification of biomarkers. One significant hurdle has been the high and variable concentrations of globin transcripts in whole blood total RNA potentially resulting in non-specific probe binding and high background. In this study, we investigated and quantified the power of three whole blood profiling approaches to detect meaningful biological expression patterns. Methods To compare and quantify the impact of different mitigation technologies, we used a globin transcript spike-in strategy to synthetically generate a globin-induced signature and then mitigate it with the three different technologies. Biological differences, in globin transcript spiked samples, were modeled by supplementing with either 1% of liver or 1% brain total RNA. In order to demonstrate the biological utility of a robust globin artifact mitigation strategy in biomarker discovery, we treated whole blood ex vivo with suberoylanilide hydroxamic acid (SAHA) and compared the overlap between the obtained signatures and signatures of a known biomarker derived from SAHA-treated cell lines and PBMCs of SAHA-treated patients. Results We found cDNA hybridization targets detect at least 20 times more specific differentially expressed signatures (2597) between 1% liver and 1% brain in globin-supplemented samples than the PNA (117) or no treatment (97) method at FDR = 10% and p-value < 3x10-3. In addition, we found that the ex vivo derived gene expression profile was highly concordant with that of the previously identified SAHA pharmacodynamic biomarkers. Conclusions We conclude that an amplification method for gene expression profiling employing cDNA targets effectively mitigates the negative impact on data of abundant globin transcripts and greatly improves the ability to identify relevant gene expression based pharmacodynamic biomarkers from whole blood.

College campuses are rapidly adopting Course Management Systems (CMS) such as Moodle and Sakai for undergraduate instruction. These online support systems offer the potential to extend student-instructor interaction beyond the classroom, yet little is known about the effects of instructor engagement on student use.This study evaluates the extent to which instructor use of a CMS affects student use and whether increased student use results in improved academic performance. Participants in two intact courses taught by the same instructor used course resources available on Moodle during an academic quarter. Levels of online course support provided by the instructor were compared with levels of student use. Levels of student use were then compared with academic performance. In this study, we hypothesize that instructor engagement with a CMS affects how students use the system and that this use relates to academic performance.

Previously, the mutation status of KRAS was the only validated predictive biomarker for metastatic colorectal cancer (CRC). While KRAS mutated tumors demonstrated resistance to epidermal growth factor (EGFR) inhibitors like cetuximab, KRAS WT and EGFR-expressing tumors were predicted to be responsive. However, KRAS WT metastatic colorectal cancer (CRC) patients have a poor prognosis even with EGFR inhibitor therapy as not all KRAS WT CRCs are responsive to such targeted agents. A gene expression based RAS signature score was developed based on multiple tumor tissue samples to identify RAS activated tumors independent of mutations in the KRAS gene [1, 2]. To further refine this score and define technologies that can be used on FFPE samples isolated in a clinical setting, we analyzed DNA and RNA derived from fifty-five (55) FFPE preserved colorectal cancer tumor biopsies using multiple sequencing, digital and array-based technologies. These samples were selected from a CRC cohort in which the initial gene expression-based RAS signature score was calculated utilizing data compiled from fresh frozen (FF) tumor samples from the same 55 patients. The 55 samples were selected for this study as they had representative samples with high, medium and low RAS signature scores. Transcriptomic analyses (RNA-Seq, Affymetrix microarrays, Nanostring and Targeted RNA-Seq) were performed on all 55 FFPE samples and three new RAS scores were calculated from the gene expression datasets. These RAS scores were based on different gene signatures (1) an 18 gene signature (2) a 13 gene signature, and (3) a 147 gene signature. A significant correlation was identified between RAS scores calculated from the 18 and 13 gene signatures (Correlation coefficient ∼ 0.88 and ∼0.76 respectively, p-value < 0.0001). To further refine gene expression signatures, samples were grouped based upon their mutation status obtained by whole exome sequencing (WES) and targeted DNA sequencing data (Illumina TruSight and LifeTech Cancer Panels). In our sample set, the 18 gene RAS score was found to be dependent on the mutation status of KRAS. Further analysis is being carried out to better understand the relationship between the calculated RAS signature scores and the mutation status of other genes. This analysis will lead to the development of a novel genomic signature for better pharmacodynamic stratification of colorectal carcinoma patients. 1. Loboda A et al. A gene expression signature of RAS pathway dependence predicts response to PI3K and RAS pathway inhibitors and expands the population of RAS pathway activated tumors. 2. BMC Medical Genomics 2010, 3:26Dry JR et al. Transcriptional Pathway Signatures Predict MEK Addiction and Response to Selumetinib (AZD6244). Cancer Res. 2010 Mar 15;70(6):2264-73. Citation Format: Sharon Austin, Fang Yin Lo, Kellie Howard, Mollie McWhorter, Heather Collins, Amanda Leonti, Lindsey Maassel, Christopher Subia, Tuuli Saloranta, Nicole Heying, Leila Ritter, Kerry Deutsch, James Cox, Steven Anderson, Anup Madan, Timothy Yeatman. Pharmacodynamic stratification of metastatic colorectal cancer patients using genomic datasets. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4842. doi:10.1158/1538-7445.AM2015-4842