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Self-efficacy, the internal belief that one can perform a specific task successfully, influences behavior. To promote critical appraisal of medical literature, rheumatology training programs should foster both competence and self-efficacy for critical appraisal. This study aimed to investigate whether select items from the Clinical Research Appraisal Inventory (CRAI), an instrument measuring clinical research self-efficacy, could be used to measure critical appraisal self-efficacy (CASE).One hundred twenty-five trainees from 33 rheumatology programs were sent a questionnaire that included two sections of the CRAI. Six CRAI items relevant to CASE were identified a priori; responses generated a CASE score (total score range 0-10; higher = greater confidence in one's ability to perform a specific task successfully). CASE scores' internal structure and relation to domain-concordant variables were analyzed.Questionnaires were completed by 112 of 125 (89.6%) trainees. CASE scores ranged from 0.5 to 8.2. The six CRAI items contributing to the CASE score demonstrated high internal consistency (Cronbach's α = 0.95) and unidimensionality. Criterion validity was supported by the findings that participants with higher CASE scores rated their epidemiology and biostatistics understanding higher than that of peers (P 0.0001) and were more likely to report referring to studies to answer clinical questions (odds ratio 2.47, 95% confidence interval 1.41-4.33; P = 0.002). The correlation of CASE scores with percentage of questions answered correctly was only moderate, supporting discriminant validity.The six-item CASE instrument demonstrated content validity, internal consistency, discriminative capability, and criterion validity, including correlation with self-reported behavior, supporting its potential as a useful measure of critical appraisal self-efficacy.

Human Central memory T cells express pcdh18 coincident with in vitro conversion to Effector memory cells

S1. Pcdh 18 is expressed in anti-L. monocytogenes (SIINFEKL-specific) CD8+ T cells. S2. Table of TIL ISR from gene array. S3. Expression of immune cell molecules in wild-type and pcdh 18-/- CD8+ T spleen cells. S4. Antibody blockade of PD-1 or pcdh18 in wild type mice. S5. IFN� secretion by human CD8+ naive T cells is not impacted by anti-pcdh18 amino terminal-specific Ab, but is enhanced in CD8+ effector memory cells. Human central memory T cells express pcdh18 coincident with in vitro conversion to effector memory cells. S6. Human central memory T cells express pcdh18 coincident with in vitro conversion to effector memory cells.

Supplemental Figures 1-6

Antibody blockade of PD-1 or pcdh18 in wild type mice

Table of TIL ISR from gene array

Cancers are infiltrated with antitumor CD8+ T cells that arise during tumor growth, but are defective in effector phase functions because of the suppressive microenvironment. The reactivation of TILs can result in tumor destruction, showing that lytic dysfunction in CD8+ tumor-infiltrating lymphocytes (TIL) permits tumor growth. Like all memory T cells, TILs express inhibitory signaling receptors (aka checkpoint inhibitor molecules) that downregulate TCR-mediated signal transduction upon TIL interaction with cells expressing cognate ligands, thereby restricting cell activation and preventing the effector phase. Previously, we identified a novel murine CD8+ TIL inhibitory signaling receptor, protocadherin-18, and showed that it interacts with p56lck kinase to abrogate proximal TCR signaling. Here, we show that TILs from mice deleted in protocadherin-18 had enhanced antitumor activity and that coblockade of PD-1 and protocadherin-18 in wild-type mice significantly enhanced TIL effector phase function. These results define an important role for protocadherin-18 in antitumor T-cell activity. Cancer Immunol Res; 5(10); 920–8. ©2017 AACR.

Expression of immune cell molecules in wild type and pcdh 18 -/- CD8+ T spleen cells.

IFNg secretion by human CD8+ naive T cells is not impacted by anti-pcdh18 amino terminal-specific Ab but is enhanced in CD8+ effector memory cells.

pcdh 18 is expressed in anti-L. monocytogenes (SIINFEKL-specific) CD8+ T cells.

Supplementary Figure 4 from Suppression of Proximal T Cell Receptor Signaling and Lytic Function in CD8+ Tumor-Infiltrating T Cells

Supplementary Legends 1-6 from Suppression of Proximal T Cell Receptor Signaling and Lytic Function in CD8+ Tumor-Infiltrating T Cells

Supplementary Figure 5 from Suppression of Proximal T Cell Receptor Signaling and Lytic Function in CD8+ Tumor-Infiltrating T Cells

Supplementary Figure 3 from Suppression of Proximal T Cell Receptor Signaling and Lytic Function in CD8+ Tumor-Infiltrating T Cells

Supplementary Figure 2 from Suppression of Proximal T Cell Receptor Signaling and Lytic Function in CD8+ Tumor-Infiltrating T Cells

Supplementary Figure 6 from Suppression of Proximal T Cell Receptor Signaling and Lytic Function in CD8+ Tumor-Infiltrating T Cells

Supplementary Figure 1 from Suppression of Proximal T Cell Receptor Signaling and Lytic Function in CD8+ Tumor-Infiltrating T Cells

CD8+ tumor-infiltrating lymphocytes (TIL) lack in vivo and in vitro lytic function due to a signaling deficit characterized by failure to flux calcium or activate tyrosine kinase activity upon contact with cognate tumor cells. Although CD3ζ is phosphorylated by conjugation in vitro with cognate tumor cells, showing that TIL are triggered, PLCγ-1, LAT, and ZAP70 are not activated and LFA-1 is not affinity-matured, and because p56lck is required for LFA-1 activation, this implies that the signaling blockade is very proximal. Here, we show that TIL signaling defects are transient, being reversed upon purification and brief culture in vitro, implying a fast-acting “switch”. Biochemical analysis of purified nonlytic TIL shows that contact with tumor cells causes transient activation of p56lck (∼10 s) which is rapidly inactivated. In contrast, tumor-induced activation of p56lck in lytic TIL is sustained coincident with downstream TCR signaling and lytic function. Shp-1 is robustly active in nonlytic TIL compared with lytic TIL, colocalizes with p56lck in nonlytic TIL, and inhibition of Shp-1 activity in lytic TIL in vitro blocks tumor-induced defective TIL cytolysis. Collectively, our data support the notion that contact of nonlytic TIL with tumor cells, and not with tumor-infiltrating myeloid-derived suppressor cells, causes activation of Shp-1 that rapidly dephosphorylates the p56lck activation motif (Y394), thus inhibiting effector phase functions. [Cancer Res 2007;67(23):11447–54]

Aiming at improving the first-year engineering experience by engaging, inspiring, and holistically preparing students for the challenges in the future, the College of Engineering at the University of Saskatchewan redesigned and launched a RE-ENGINEERED First Year (REFY) Program. The project has had three phases, and the first two were reported at CEEA conferences in 20181 and 20192. A small-scale pilot of the competency-based assessment (CBA), one of the key features of the REFY program, was presented at CEEA 20213. In this paper, we will report the changes made to the program structure (Phase II), present the systematic procedure followed to develop the course materials (Phase III) and reflect on the first implementation of the REFY program. This paper will also discuss the proposed changes to the program for next year’s implementation. The REFY program consists of 10-12 modular courses with various durations and intensities each term. To better sequence materials and to facilitate just-in-time learning, the structural design of the program was further refined over the past two years via adjusting the start/end dates and contact hours of the modular courses. The development of course materials was directed by the identified graduate attributes (Phase I), integrated curriculum schedule (Phase II), and the philosophy of CBA. With the lessons learned from the pilot of CBA in a single first year engineering course, we defined four types of assessments and the evaluation criteria. We also developed a course preparation procedure template and created a series of policies to support the program. Initiating the REFY program during the COVID pandemic was a victory, although we identified some challenges and problems in Term 1 through teaching, observations, and reflection, as well as anecdotal comments. We will share the lessons learned, our opinions on the potential causes of the challenges and problems, and propose revisions for the next iteration of program refinement.

To provide optimal patient care, rheumatologists must be equipped and motivated to critically appraise the literature. The conceptual frameworks Retrieval Enhanced Learning, Self-Determination Theory, and Communities of Practice can inform the design of educational approaches to promote critical appraisal in practice. HSS CLASS-Rheum® is a learning tool that can be used to help rheumatologists learn skills for critical appraisal through retrieval practice. Combining retrieval practice with opportunities for connection through Peer Instruction, journal clubs, and other forums can help support engagement and internalization of motivation, promoting persistence with critical appraisal in practice.

BackgroundPaucity of T cells in the immune privileged tumor microenvironment (TME) of pancreatic ductal adenocarcinoma (PDAC) is a major reason that PDAC is refractory to immune checkpoint blockade.1 In this study, we show that human PDAC tumors over-express vasoactive intestinal peptide (VIP), an immunosuppressive neuropeptide, that inhibits effector T cell responses and regulates chemokine receptor expression on activated T cells.2 3 We thus hypothesized that pharmacological inhibition of VIP receptor signaling could enhance anti-tumor responses in PDAC.MethodsVIP levels in plasma were determined via VIP-specific enzyme immunoassay and confirmed with immunohistochemistry (IHC) of tissue sections. VIP receptor (VIP-R) signaling in C57BL/6 immunocompetent murine models of KPC, MT5 or Panc02 pancreatic cancer was inhibited by daily sub-cutaneous treatment with ANT008 or ANT308, two novel VIP-R antagonists with predicted high binding affinities to VIP receptors.4–7 Multiplex IHC or flow cytometry detected frequencies and phenotypes of intra-tumoral T cells across treatment groups.ResultsHuman PDAC tumors expressed VIP by immunohistochemistry, and PDAC patients had significantly elevated plasma VIP levels when compared to healthy volunteers (pConclusionsVIP-R antagonists represent a novel approach to treat PDAC. VIP and VIP-R sequences are highly conserved between humans and mice,8 and human T cells are activated in vitro following treatment with VIP-R antagonists. Thus, we predict comparable anti-tumor activity of the combination of VIP-R antagonist and anti-PD-1 MoAb in human PDAC patients. Further clinical development of this novel concept will require appropriate pre-clinical pharmacokinetic and toxicology studies.AcknowledgementsThe authors thank healthy volunteers and patients for blood and/or tissue samples. The authors also thank the shared resources at Emory University, namely the Emory Integrated Genomics Core (EIGC), Emory Flow Cytometry Core (EFCC), Cancer Animal Models Shared Resource (CAMS), Cancer Tissue Pathology Core (CTP), Biostatistics Shared Resource (BSR) and Integrated Cellular Imaging Core (ICI), that provided services or instruments at subsidized cost to conduct some of the reported experiments. BioRender was used to make figure 4A and 5C. This work was supported in part by Katz Foundation funding and Emory School of Medicine Dean's Imagine, Innovate and Impact (I3) venture award to Edmund K. Waller and NIH R01 CA207619 awarded to Susan N. Thomas. Part of the cost for the immunohistochemistry staining of tissues was covered by Winship Cancer Institute Development Discovery and Therapeutic Program Pilot funding to Sruthi Ravindranathan.Abstract 748 Figure 1VIP is over-expressed by PDAC. (A) VIP mRNA expression levels in various solid malignancies, as obtained from TCGA. (B) Representative images of human PDAC tumor stained with antibodies to VIP or CK19, showing VIP co-expression in islets (black arrow) and cancer epithelial cells (red arrow). Levels of VIP in (C) culture supernatants collected from murine and human PDAC cell lines cultured for 24 hours (n=3 per cell line) were compared to culture supernatants from B16F10 and D4M melanoma cells; (D) plasma of mice bearing melanoma or PDAC tumors (n=5) compared to plasma of non-tumor-bearing mice; (E) plasma of PDAC patients (n=19) compared to that from healthy volunteers (n=26). Statistical differences in C and D were performed by ANOVA followed by Dunnett's post-test and in E were performed by student's t-test. Error bars show mean ± SEM. *pAbstract 748 Figure 2VIP-R antagonists improve responses to anti-PD-1. KPC.Luc, MT5 or Panc02 cells were subcutaneously implanted in immunocompetent C57BL/6 mice. About one week after tumor implantation, when the tumors were palpable, mice were randomized into treatment groups and treated with VIP-R antagonist and/or anti-PD-1 as described in methods. (A) KPC.Luc, MT5 and Panc02 tumor volumes as measured by Vernier calipers on day 22 after subcutaneous tumor implantation. (B) Kaplan-Meier survival plots of C57BL/6 mice with subcutaneously implanted KPC.Luc, MT5 or Panc02 tumors stratified by treatment. Kaplan-Meier survival plots of (C) C57BL/6 mice receiving monoclonal CD4 and/or CD8 monoclonal antibodies (D) CD4KO or (E) CD8KO mice compared to wild-type CD57BL/6 mice with subcutaneously implanted KPC.Luc tumors, stratified by treatment. Statistical differences in A were calculated by ANOVA followed by Dunnett's post-test. Solid line shows mean with in each treatment group. Statistical differences in B-E are calculated via Log-rank test. *pAbstract 748 Figure 3Enhanced T cell response with combination therapy. mRNA expression in T cells isolated from subcutaneous KPC.Luc tumors in C57BL/6 mice treated with ANT008 and/or anti-PD-1 (n=3 per treatment group), were analyzed via Nanostring metabolism panel. Volcano plot showing differential expression of genes in T cells from (A) ANT008+ isotype IgG (IgG) vs scrambled peptide (Scram) + isotype IgG, (B) scrambled peptide +anti-PD-1 vs scrambled peptide + isotype IgG and (C) ANT008+anti-PD-1 vs scrambled peptide + isotype IgG (n=3 mice per treatment group). Genes that are associated with TCR activation and co-stimulation and are at levels significantly higher when compared to Scram+ isotype IgG (FDRAbstract 748 Figure 4Increased T cell density with combination therapy. KPC.Luc cells were orthotopically implanted in the tail of the pancreas of C57BL/6 mice and treated with ANT008 and/or anti-PD-1 with n=9, 10, 8 and 11 in scrambled+IgG, ANT008+IgG, scrambled+anti-PD-1 and ANT008+anti-PD-1, respectively. (A) Schematic showing orthotopic implantation of KPC.Luc cells and treatment strategy with ANT008 and/or anti-PD-1. (B) Waterfall plot showing % change in tumor flux on day 22 relative to day 7 prior to start of treatment. (C) Total flux as measured by IVIS bioluminescent imaging in the different treatment groups. Cross symbol represents mice that were euthanized before day 25 due to ulceration of the tumor and circle symbol represent mouse that were imaged on day 26 via MRI imaging shown in supplementary figure S5. (D) Bar graph showing weight of pancreas on day 25 when the mice were euthanized. 'Star' shaped data points indicate tumor free mice and dotted horizontal line represents the average weight of healthy pancreas from naïve mice. (E) Representative multiplex IHC images (right) showing pancreatic tumors stained for DAPI (blue), CD4 (yellow), CD8 (red) and Ki67 (cyan) and trichrome staining (left) with black arrows showing blue collagen stain in the tissue. XY plot showing the correlation between number of (F) CD4+ or (G) CD8+ T cells/mm2; and (H) Ki67+ CD4+ or (I) Ki67+ CD8+ T cells/mm2 with weight of the pancreas with n=4 to 6 mice per group. P values in panel D were calculated using student ANOVA followed by Dunnett's post hoc test (comparing each treatment group with Scram+IgG). Error bars show mean ± SEM. *pAbstract 748 Figure 5Increased T cell homing with combination therapy. KPC.Luc tumors were subcutaneously implanted in C57BL/6 mice and treated with VIP-R antagonist and/or anti-PD-1 checkpoint therapy for 10 days after the tumors were palpable. Tumor draining lymph nodes were then analyzed for percentage of (A) CXCR4+CD69+ and (B) CXCR4+Ki67+ cells in CD4+ (left) and CD8+ (right) subsets of T cells. In a separate experiment, on day 15 after subcutaneous implantation of KPC.Luc tumors, GFP+ T cells from enhanced GFP transgenic mice (C57BL/6 background) were adoptively transferred (via tail vein injections) and treated with ANT308± aPD-1 for 3 days. (C) Schematic showing GFP+ T cell transfer and treatment strategy in mice with subcutaneous KPC.Luc tumors. (D) Representative Hoescht (blue for nucleus) stained tumor tissues from tumors of each treatment group. Two regions of interest (ROI) in ANT308+aPD-1 treated tumors are shown at higher magnification. Statistical differences in A and B were determined via repeated measures ANOVA and Dunnett's post-test with n=4–5 mice per group. *pReferencesSahin IH, et al. Immunotherapy in pancreatic ductal adenocarcinoma: an emerging entity? Ann Oncol 2017;28(12):2950–2961.Gonzalez-Rey E, Anderson P, Delgado M. Emerging roles of vasoactive intestinal peptide: a new approach for autoimmune therapy. Ann Rheum Dis 2007;66(Suppl 3):p. iii70–6.Anderson P, Gonzalez-Rey E. Vasoactive intestinal peptide induces cell cycle arrest and regulatory functions in human T cells at multiple levels. Mol Cell Biol 2010;30(10):2537–51.Li JM, et al. VIPhyb, an antagonist of vasoactive intestinal peptide receptor, enhances cellular antiviral immunity in murine cytomegalovirus infected mice. PLoS One 2013;8(5):e63381.Moody TW, et al., VIP receptor antagonists and chemotherapeutic drugs inhibit the growth of breast cancer cells. Breast Cancer Res Treat 2001;68(1):55–64.Moody TW, et al. A vasoactive-Intestinal-Peptide antagonist inhibits nonsmall cell lung-cancer growth. Proceedings of the National Academy of Sciences of the United States of America 1993;90(10):4345–4349.Zia H, et al. Breast cancer growth is inhibited by vasoactive intestinal peptide (VIP) hybrid, a synthetic VIP receptor antagonist. Cancer Res 1996;56(15):3486–9.Sena M, et al. High conservation of upstream regulatory sequences on the human and mouse vasoactive intestinal peptide (VIP) genes. DNA Seq 1994;5(1):25–9.Ethics ApprovalAll experimental procedures involving mice were approved by the Institutional Animal Care and Use Committee (IACUC) at Emory University. De-identified blood samples from consented patients with PDAC (IRB 00087397) or healthy volunteers (IRB 00046063) were obtained with approval from Institutional Review Boards.

A paucity of effector T cells within tumors renders pancreatic ductal adenocarcinoma (PDAC) resistant to immune checkpoint therapies. While several under-development approaches target immune-suppressive cells in the tumor microenvironment, there is less focus on improving T cell function. Here we show that inhibiting vasoactive intestinal peptide receptor (VIP-R) signaling enhances anti-tumor immunity in murine PDAC models. In silico data mining and immunohistochemistry analysis of primary tumors indicate overexpression of the neuropeptide vasoactive intestinal peptide (VIP) in human PDAC tumors. Elevated VIP levels are also present in PDAC patient plasma and supernatants of cultured PDAC cells. Furthermore, T cells up-regulate VIP receptors after activation, identifying the VIP signaling pathway as a potential target to enhance T cell function. In mouse PDAC models, VIP-R antagonist peptides synergize with anti-PD-1 antibody treatment in improving T cell recruitment into the tumors, activation of tumor-antigen-specific T cells, and inhibition of T cell exhaustion. In contrast to the limited single-agent activity of anti-PD1 antibodies or VIP-R antagonist peptides, combining both therapies eliminate tumors in up to 40% of animals. Furthermore, tumor-free mice resist tumor re-challenge, indicating anti-cancer immunological memory generation. VIP-R signaling thus represents a tumor-protective immune-modulatory pathway that is targetable in PDAC.

Compartmentalization of biochemical processes is essential for cell function. Although membrane-bound organelles are well studied in this context, recent work has shown that phase separation is a key contributor to cellular compartmentalization through the formation of liquid-like membraneless organelles (MLOs). In this Minireview, the key mechanistic concepts that underlie MLO dynamics and function are first briefly discussed, including the relevant noncovalent interaction chemistry and polymer physical chemistry. Next, a few examples of MLOs and relevant proteins are given, along with their functions, which highlight the relevance of the above concepts. The developing area of active matter and non-equilibrium systems, which can give rise to unexpected effects in fluctuating cellular conditions, are also discussed. Finally, our thoughts for emerging and future directions in the field are discussed, including in vitro and in vivo studies of MLO physical chemistry and function.

OBJECTIVE The present study was undertaken to evaluate the pregnancy experiences of women receiving care in the division of rheumatology at a major academic center in New York City during the COVID-19 pandemic. METHODS A web-based COVID-19 survey was emailed to 26,045 patients who were followed in the division of rheumatology at a single center in New York City. Women ages 18-50 years were asked about their pregnancy. We compared the COVID-19 experience between pregnant and nonpregnant women and also explored the impact of the pandemic on prenatal care and perinatal outcomes. RESULTS Among 7,094 of the 26,045 respondents, 1,547 were women ages 18-50 years, with 61 (4%) reporting being pregnant during the pandemic. The prevalence of self-reported COVID-19 was similar in pregnant and nonpregnant women (8% versus 9%, respectively; P = 0.76). Among women with COVID-19, pregnant women had a shorter duration of symptoms (P < 0.01) and were more likely to experience loss of smell or taste (P = 0.02) than nonpregnant women. Approximately three-fourths of women had a systemic rheumatic disease, with no differences when stratified by pregnancy or COVID-19 status. In all, 67% of pregnant women noted changes to prenatal care during the pandemic, and 23% of postpartum women stated that the pandemic affected delivery. CONCLUSION Among women followed in the division of rheumatology at a major center in New York City, pregnancy was not associated with increased self-reported COVID-19. Pregnancy was associated with a shorter duration of COVID-19 symptoms and a higher prevalence of loss of smell or taste. The COVID-19 pandemic impacted prenatal care for the majority of pregnant patients.

Many students entering an engineering program have a strong appreciation of the importance of math- and science-based skills for their future career as an engineer, but often have little grasp of what it means to be entering a professional college. For this reason, many engineering programs in Canada include some form of an “Introduction to the Engineering Profession” in their first-year program. The University of Saskatchewan’s College of Engineering has been working toward the launch of a completely redesigned first year program. This project has afforded the College an opportunity to apply a novel and transferable approach to shaping this “Introduction to the Engineering” experience. The structure of the proposed new first year program has allowed for short and intensive “Introduction to Engineering” modules, which bookend each of the regular session terms. This timing makes them an orientation for the program, allowing for timely deep dives into matters of importance to engineering students: study skills, time-management, teamwork, self-assessment, support services, student well-being, ethics, academic integrity, and health and safety. The timing of the modules also allows for completion of term-long assignments and reflection on both personal and academic growth. This paper describes the process employed to develop the course learning outcomes, schedule of topics and activities, and syllabi. The process focused on over-arching target attitudes, such as “I am on the path to becoming a professional”, and ensured constructive alignment between these attitudes and the learning outcomes, learning activities, and assessment. The nature of the process made it easy to clarify what was essential to include in the courses, and to make a compelling case for the importance of the courses in the context of a myriad of foundational technical topics.

Through the action of two virus-encoded decapping enzymes (D9 and D10) that remove protective caps from mRNA 5′-termini, Vaccinia virus (VACV) accelerates mRNA decay and limits activation of host defenses. D9- or D10-deficient VACV are markedly attenuated in mice and fail to counter cellular double-stranded RNA-responsive innate immune effectors, including PKR. Here, we capitalize upon this phenotype and demonstrate that VACV deficient in either decapping enzyme are effective oncolytic viruses. Significantly, D9- or D10-deficient VACV displayed anti-tumor activity against syngeneic mouse tumors of different genetic backgrounds and human hepatocellular carcinoma xenografts. Furthermore, D9- and D10-deficient VACV hyperactivated the host anti-viral enzyme PKR in non-tumorigenic cells compared to wild-type virus. This establishes a new genetic platform for oncolytic VACV development that is deficient for a major pathogenesis determinant while retaining viral genes that support robust productive replication like those required for nucleotide metabolism. It further demonstrates how VACV mutants unable to execute a fundamental step in virus-induced mRNA decay can be unexpectedly translated into a powerful anti-tumor therapy. Keywords: oncolytic virus, mRNA decay, decapping

Pancreatic ductal adenocarcinoma (PDAC) is unresponsive to immune checkpoint therapy largely due to a paucity of T cells within the tumor microenvironment (TME) and abundant immunosuppressive signaling pathways. In this study we show that human PDAC tumors over-express vasoactive intestinal peptide (VIP), an immunosuppressive neuropeptide that suppresses T cell effector properties and promotes the generation of regulatory T cells (Tregs). Therefore, we treated tumor-bearing mice with VIP receptor peptide antagonists and measured T cell homing, activation, and anti-tumor responses in preclinical murine models of PDAC. Pharmacological inhibition of VIP receptor (VIP-R) signaling using daily subcutaneous injections of peptide antagonists had no discernable toxicity in healthy and tumor-bearing mice. Treatment with VIP-R antagonists in combination with anti-PD-1 checkpoint blockade significantly decreased tumor burden, and improved survival in subcutaneous and orthotopic murine PDAC models. Combination therapy significantly enhanced T cell activation and proliferation and decreased frequencies of Tregs within the TME. Anti-tumor responses were T cell dependent, as the combination therapy failed to improve survival in CD4 or CD8 deficient mice using knock-out strains and antibody depletion. Furthermore, combination therapy significantly increased frequencies of tumor specific T cells (measured with a tetramer reagent) and provided protective immunity against tumor rechallenge. Combination therapy led to significant increases in the infiltration of adoptively transferred GFP+ T cells into PDAC tumors and decreased CXCR4 expression levels on T cells. Encouragingly, peptide-based VIP-R antagonists enhanced the in vitro activation of human T cells isolated from peripheral blood of PDAC patients. Human T cells cultured with VIP-R antagonists had increased proliferation, activation, and decreased proportions of T regs and exhausted T cells co-expressing PD-1, Tim-3 and Lag-3. Taken together, our findings show that VIP is a targetable mechanism of immune escape in PDAC. Inhibiting VIP receptor signaling improves T cell effector properties and synergistically improves anti-tumor responses to checkpoint inhibitors in mouse PDAC models. Additionally, as the VIP sequence is identical between human and mice, and since VIP-R antagonists have similar effects on human and murine T cells in culture, clinical translation is highly feasible. Citation Format: Sruthi Ravindranathan, Passang Tenzin, Jian Ming Li, Rohan Dhamsania, Michael Ware, Mohammad Zaidi, Shuhua Wang, Jingru Zhu, Maria Cardenas, Yuan Liu, Gaurav Joshi, Sanjeev Gumber, Brian Robinson, Anish Sen-Majumdar, Shanmuganathan Chandrakasan, Haydn Kissick, Alan Frey, Susan Thomas, Bassel El-Rayes, Gregory Lesinski, Edmund K. Waller. Inhibiting vasoactive intestinal peptide receptor signaling elicits T cell dependent anti-tumor response of pancreatic ductal adenocarcinoma to immune checkpoint therapy [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-072.

Cancers are infiltrated with antitumor CD8+ T cells that arise during tumor growth, but are defective in effector phase functions because of the suppressive microenvironment. The reactivation of TILs can result in tumor destruction, showing that lytic dysfunction in CD8+ tumor-infiltrating lymphocytes (TIL) permits tumor growth. Like all memory T cells, TILs express inhibitory signaling receptors (aka checkpoint inhibitor molecules) that downregulate TCR-mediated signal transduction upon TIL interaction with cells expressing cognate ligands, thereby restricting cell activation and preventing the effector phase. Previously, we identified a novel murine CD8+ TIL inhibitory signaling receptor, protocadherin-18, and showed that it interacts with p56lck kinase to abrogate proximal TCR signaling. Here, we show that TILs from mice deleted in protocadherin-18 had enhanced antitumor activity and that coblockade of PD-1 and protocadherin-18 in wild-type mice significantly enhanced TIL effector phase function. These results define an important role for protocadherin-18 in antitumor T-cell activity. Cancer Immunol Res; 5(10); 920–8. ©2017 AACR.

The steroid profile changes dramatically from prenatal to postnatal life. Recently a novel backdoor pathway for androgen biosynthesis has been discovered. However its role remains elusive. Therefore we investigated androgen production from birth to one year of life with a focus on minipuberty and on production of androgens through the backdoor pathway. Additionally we assessed the development of the specific steroid enzyme activities in early life. To do so we collected urine specimens from diapers in 43 healthy newborns (22 females) at 13 time points from birth to one year of age in an ambulatory setting and performed in house GC/MS steroid profiling for 67 steroid metabolites. Data were analyzed for androgen production through the classic and backdoor pathway and calculations of diagnostic ratios for steroid enzyme activities were performed. Analysis revealed that during minipuberty androgen production is much higher in boys than in girls (e.g. androsterone (An)) originates largely from the testis (Anboys Angirls) and uses predominantly the alternative backdoor pathway (An/Et; ?5 < ?4 lyase activity). Modelling of steroid enzyme activities showed age related effects for 21 11 17 hydroxylase and P450 oxidoreductase activities as well as 3ß hydroxysteroid dehydrogenase 11ß hydroxylase type 1/2 and 5a reductase activities. Sex related characteristics were found for 21 hydroxylase and 5a reductase activities. Overall our study shows that androgen biosynthesis during minipuberty favors the backdoor pathway over the classic pathway. Calculations of specific diagnostic ratios for enzyme activities seem to allow the diagnosis of specific steroid disorders from the urinary steroid metabolome.

Over the last three years, the University of Saskatchewan’s College of Engineering has embarked upon a complete redesign of its first year common program. This project started from a blank slate and posed the question, "If we could design any first year program that we wanted, what would we create?" The goal is to offer a first year program that excites, engages, inspires, and holistically prepares students for learning in subsequent years. At CEEA 2018, Phase I of this project was reviewed with a focus on the content of the new first year. This year, the focus is on the structure of the proposed program and how it aims to satisfy programmatic design objectives. The proposed first year program is highly modular, allowing for more intentional uses of time during the academic year. Course duration and intensity vary and are selected to best serve student learning, rather than conform to the traditional academic schedule. To provide more timely and targeted feedback, summative assessments occur throughout each term allowing course scheduling to extend into the traditional end-of-term final exam period. This paper presents the current structural design of the new first year and the rationale for its significant features. Some of the program design objectives that have been facilitated by this structure include: • strategic sequencing of learning with opportunities to integrate and reinforce essential skills, • multiple, individualized opportunities for students to stumble and recover, • holistic balancing of content and pacing for better student wellbeing, and • comprehensive, well-timed exposure to wide-ranging programmatic choices for students. Throughout this project, the program structure has evolved continuously. This paper will describe the development process, the challenges faced in that process, and the lessons learned. The paper will conclude by describing the current status of the project, and the focus of work currently being undertaken to prepare for implementation.

This paper describes the progress made over the past year on UVic’s project to redesign their co-op work term requirements. The new requirements are intended to increase the amount of instruction and assessment of higher-level Graduate Attributes in the program, to improve student engagement with and performance on work term requirement, and to replace a final year technical writing course that does not seem to be achieving its intended outcomes. A pilot of the one component of this project will run from May – August 2019. In this paper we will describe the material that has been developed for the pilot. We will also discuss the process by which the material was developed and the details of organizing the pilot.

CD8+ T cells develop increased sensitivity following Ag experience, and differences in sensitivity exist between T cell memory subsets. How differential TCR signaling between memory subsets contributes to sensitivity differences is unclear. We show in mouse effector memory T cells (TEM) that >50% of lymphocyte-specific protein tyrosine kinase (Lck) exists in a constitutively active conformation, compared with

Although counteracting innate defenses allows oncolytic viruses (OVs) to better replicate and spread within tumors, CD8+ T-cells restrict their capacity to trigger systemic anti-tumor immune responses. Herpes simplex virus-1 (HSV-1) evades CD8+ T-cells by producing ICP47, which limits immune recognition of infected cells by inhibiting the transporter associated with antigen processing (TAP). Surprisingly, removing ICP47 was assumed to benefit OV immuno-therapy, but the impact of inhibiting TAP remains unknown because human HSV-1 ICP47 is not effective in rodents. Here, we engineer an HSV-1 OV to produce bovine herpesvirus UL49.5, which unlike ICP47, antagonizes rodent and human TAP. Significantly, UL49.5-expressing OVs showed superior efficacy treating bladder and breast cancer in murine models that was dependent upon CD8+ T-cells. Besides injected subcutaneous tumors, UL49.5-OV reduced untreated, contralateral tumor size and metastases. These findings establish TAP inhibitor-armed OVs that evade CD8+ T-cells as an immunotherapy strategy to elicit potent local and systemic anti-tumor responses., Highlights • Inhibiting TAP (transporter associated with antigen processing) effectively stimulates oncolytic virus (OV) immunotherapy • TAP inhibitor-armed OVs that evade CD8+ T-cells elicit potent local and systemic immunotherapeutic, anti-tumor responses. Oncolytic viruses (OVs) offer advantages over traditional cancer therapies, selectively killing tumor cells and stimulating anti-tumor immune responses. However, sensitivity to CD8+ T-cell killing limits therapeutic responses to OVs. We engineer herpes simplex virus-1 OVs that evade CD8+ T-cells by expressing a TAP (transporter associated with antigen processing) inhibitor and demonstrate their greater efficacy treating bladder and breast cancer in pre-clinical mouse models. TAP inhibitor-armed OVs better stimulated systemic anti-tumor responses at distant, untreated sites and metastases; moreover, their superiority was dependent upon CD8+ T-cell responses. This establishes arming OVs to evade CD8+ T-cells as an effective OV immunotherapy strategy.

The rapidly evolving role of the engineering profession in society requires an engineering graduate with a more diverse and robust skill set than ever before. To answer this challenge, the University of Saskatchewan’s College of Engineering has embarked upon a complete redesign of its first year program. This project essentially started from a “blank slate” and posed the question, “If we could design any first year program that we wanted, what would we create?” The outcome of this endeavor is intended to be an extremely effective first year program that excites, engages and inspires students, and that holistically prepares them for the challenges to come in later years. In this paper, we review the broad learning objectives of our new first year, and the values that we applied to our decision making during its design.The overall project consists of three distinct phases: determination of required first year graduate attributes, development of program structure and delivery methods, and detailed course design. Phase I has been completed. It has left us with a detailed inventory of knowledge, skills, experiences, and attitudes, distributed across 23 content categories, that the College wants students to internalize by the end of their first year of study.We will outline the methods that we used to compile and refine this attribute inventory, including multiple approaches aimed at meaningful stakeholder engagement, surveys of existing first year programs across Canada, and an analysis of gaps and redundancies between the Saskatchewan high school curriculum and our existing first year program. We will also describe the 23 content categories used to organize the graduate attributes of the proposed first year program and how these categories are weighted in relative terms. We share some of our key learnings from Phase I of the project, including which consultation strategies worked most effectively, why we focused on first year graduate attributes and not content, and key elements that will be emphasized in our new program. We will also briefly describe the process by which we are starting to develop the program structure and delivery methods i.e. Phase II.

The function of membrane proteins relies on a defined orientation of protein relative to lipid. In apparent correlation to protein anchoring, tryptophan residues are enriched in the lipid headgroup region. To characterize the thermodynamic and structural basis of this relationship in α-helical membrane proteins, we examined the role of three conserved tryptophans in the folding of the heterodimeric integrin αIIbβ3 transmembrane (TM) complex in phospholipid bicelles and mammalian membranes. In the homogenous lipid environment of bicelles, tryptophan was replaceable by residues of distinct polarities. The appropriate polarity was guided by the electrostatic potential of the tryptophan surrounding, suggesting that tryptophan can complement diverse environments by adjusting the orientation of its anisotropic side chain to achieve site-specific anchoring. As a sole membrane anchor, tryptophan made a contribution of 0.4 kcal/mol to TM complex stability in bicelles. In membranes, it proved more difficult to replace tryptophan even by tyrosine, indicating a superior capacity to interact with heterogeneous lipids of biological membranes. Interestingly, at intracellular TM helix ends, where integrin activation is initiated, sequence motifs that interact with lipids via opposing polarity patterns were found to restrict TM helix orientations beyond tryptophan anchoring. In contrast to bicelles, phenylalanine became the least accepted substitute in membranes, demonstrating an increased role of the hydrophobic effect. Altogether, our study implicates a wide amphiphilic range of tryptophan, membrane complexity, and the hydrophobic effect to be important factors in tryptophan membrane anchoring.

Despite similarities relevant age- and gender-specific issues exist in the care of patients after allogeneic hematopoietic SCT (HSCT). Female genital chronic GVHD (cGVHD) has been markedly underreported in the past but has a significant impact on the patients’ health and quality of life. Data on prevention and treatment of this complication are still limited. Here we present a comprehensive review summarizing the current knowledge, which was discussed during several meetings of the German, Austrian and Swiss Consensus Project on clinical practice in cGVHD. In this report, we provide recommendations for post-transplant gynecological care of cGVHD manifestations agreed upon by all participants. This includes guidelines for diagnosis, prevention, and therapeutic options and topical treatments in female patients with genital cGVHD and hormonal replacement treatment of premature ovarian failure for adult and pediatric patients and underlines the necessity for regular gynecological care and screening programs for women after HSCT.

Doped and stabilized amorphous selenium (a-Se) alloys in a multilayer form are currently used as a photoconductor in direct conversion flat panel x-ray imagers in mammography and tomosynthesis. While much progress has been made on the physics of such detectors, there are still unresolved questions on such issues as the principles of operation of the so-called p-i-n detector structure in extinguishing the dark current. The present paper examines dark current transients after the application of a voltage in seven types of a-Se alloy based devices: i-layer, i-n, n-i, i-p, p-i, p-i-n, and n-i-p structures. The substrate was ITO coated glass, the top (radiation receiving) electrode was chromium, and the films were fabricated by vacuum deposition. The nominal device thickness was 200 μm, similar to commercial mammographic detectors. It is shown that n-i, i-n, n-i-p, and p-i-n devices have dark currents less than 1 pA mm−2 at an applied field of 10 V/μm. The dark current in the p-i-n device is the lowest at approximately 0.01 pA mm−2 at an applied field of 10 V μm−1. Experiments have been carried out by subjecting the detector to a staircase voltage-time profile during the voltage application (turn-on) and a staircase voltage ramp-down during the turning-off of the bias voltage. Step-voltage ramp-up and step-voltage ramp-down current transient are typical of expected dark current transient behavior in a semiconductor with traps in which carriers are captured and released from various trap centers in the bandgap. The dark current transients are qualitatively similar to those expected from a capacitor in parallel with a large resistor and both in series with a much smaller resistor. Current transients during charging and discharging experiments were integrated to find how much of the injected charge is released during discharge experiments. It is shown that the majority of this trapped charge is stored in the n-type and p-type blocking layers, near the contacts. The trapped carriers in the i-layer represent a very small portion. The evolution of the dark current upon single and repeated x-ray exposure has also been examined in n-i and p-i-n devices. Right after the cessation of irradiation, there is an excess or residual dark current whose magnitude is roughly ∼20 times higher in p-i-n and about ∼2 times higher in the case of n-i under an exposure of 3.36 R and a mean photon energy of 34.2 keV. The absorbed dose is 1.53 Gy. The excess dark current has a fast decay component with a time constant ∼10 s and a slow component with a time constant ∼100 s. It is shown that the decay in the irradiation induced excess dark current is very similar to the initial dark current. There is no permanent change in the dark current, and within a few hundred seconds, the dark current reaches the same level as that in the unexposed detector. The experimental results in this work highlight the distinct advantages of p-i-n and n-i-type a-Se multilayer structures in x-ray detection applications.Doped and stabilized amorphous selenium (a-Se) alloys in a multilayer form are currently used as a photoconductor in direct conversion flat panel x-ray imagers in mammography and tomosynthesis. While much progress has been made on the physics of such detectors, there are still unresolved questions on such issues as the principles of operation of the so-called p-i-n detector structure in extinguishing the dark current. The present paper examines dark current transients after the application of a voltage in seven types of a-Se alloy based devices: i-layer, i-n, n-i, i-p, p-i, p-i-n, and n-i-p structures. The substrate was ITO coated glass, the top (radiation receiving) electrode was chromium, and the films were fabricated by vacuum deposition. The nominal device thickness was 200 μm, similar to commercial mammographic detectors. It is shown that n-i, i-n, n-i-p, and p-i-n devices have dark currents less than 1 pA mm−2 at an applied field of 10 V/μm. The dark current in the p-i-n device is the lowest at appr...

In July 2012 a revised terminology on colposcopic examinations of the cervix uteri was adopted by IFCPC. Central aspect is a description of characteristics that should aid in the definition of the disease entity cervix uteri. The nomenclature is built up in such a way that the examiner can evaluate colposcopic criteria according to a specific scheme. Firstly it is assessed whether the colposcopy is representative. Then it should be clarified whether or not the findings are normal. If the findings are not normal but rather abnormal the severity of the lesion is classified according to the so-called grade 1 (minor change) and grade 2 findings (major change). Specific abnormal findings such as leukoplakia, erosion and Lugol's finding are also not defined at this point. Characteristics suspicious for an invasion are described separately, in particular, atypical vessel patterns are mentioned here. As already held in the previous revised nomenclature (Barcelona), various findings are described in a last group: congenital transformation zones (CTZ), congenital anomalies, condylomas (papillomas), endometriosis, polyps (ectocervical, endocervical), inflammation, stenosis, postoperative changes (scarred portio, vaginal stump).Im Juli 2012 wurde eine überarbeitete Nomenklatur zur kolposkopischen Untersuchung der Cervix uteri durch die IFCPC verabschiedet. Im Mittelpunkt steht die Beschreibung von Merkmalen, die helfen sollen, Krankheitsbilder an der Cervix uteri zu definieren. Die Nomenklatur ist so aufgebaut, dass der Untersucher nach einem bestimmten Schema kolposkopische Kriterien beurteilt. Zunächst wird geprüft, ob die Kolposkopie repräsentativ ist. Dann soll geklärt werden, ob ein Normalbefund vorliegt. Liegen keine Normalbefunde, sondern abnorme Befunde vor, wird die Schwere der Läsion in sog. Grad-1- (minor change) und Grad-2-Befunde (major change) festgelegt. Es werden auch nicht spezifische abnorme Befunde – die Leukoplakie, die Erosion und die Lugol-Probe – definiert. Gesondert werden Merkmale beschrieben, die verdächtig auf eine Invasion sind, insbesondere wird hier auf die atypischen Gefäßmuster hingewiesen. Wie schon in der letzten überarbeiteten Nomenklatur (Barcelona) werden in einer letzten Gruppe verschiedene Befunde beschrieben: kongenitale Transformationszone (KTZ), kongenitale Anomalie, Kondylome (Papillome), Endometriose, Polypen (ektozervikal, endozervikal) Entzündung, Stenose, postoperative Veränderung (vernarbte Portio, Scheidenblindsack).

Type 1 diabetes is an autoimmune disease in which insulin-producing pancreatic isletβcells are the target of self-reactive B and T cells. T cells reactive with epitopes derived from insulin and/or IGRP are critical for the initiation and maintenance of disease, but T cells reactive with other islet antigens likely have an essential role in disease progression. We sought to identify candidate CD8+T cell epitopes that are pathogenic in type 1 diabetes. Proteins that elicit autoantibodies in human type 1 diabetes were analyzed by predictive algorithms for candidate epitopes. Using several different tolerizing regimes using synthetic peptides, two new predicted tolerogenic CD8+T cell epitopes were identified in the murine homolog of the major human islet autoantigen zinc transporter ZnT8 (aa 158–166 and 282–290) and one in a non-βcell protein, dopamineβ-hydroxylase (aa 233–241). Tolerizing vaccination of NOD mice with a cDNA plasmid expressing full-length proinsulin prevented diabetes, whereas plasmids encoding ZnT8 and DβH did not. However, tolerizing vaccination of NOD mice with the proinsulin plasmid in combination with plasmids expressing ZnT8 and DβH decreased insulitis and enhanced prevention of disease compared to vaccination with the plasmid encoding proinsulin alone.

Myeloid-derived suppressor cells (MDSC) are a heterogeneous population expanded in cancer and other chronic inflammatory conditions. Here the authors identify the challenges and propose a set of minimal reporting guidelines for mouse and human MDSC., Myeloid-derived suppressor cells (MDSCs) have emerged as major regulators of immune responses in cancer and other pathological conditions. In recent years, ample evidence supports key contributions of MDSC to tumour progression through both immune-mediated mechanisms and those not directly associated with immune suppression. MDSC are the subject of intensive research with >500 papers published in 2015 alone. However, the phenotypic, morphological and functional heterogeneity of these cells generates confusion in investigation and analysis of their roles in inflammatory responses. The purpose of this communication is to suggest characterization standards in the burgeoning field of MDSC research.

A split plot experiment with 72 male pigs weighing 52.9 ± 0.39 kg (mean ± SEM) was conducted to examine AA partitioning and body protein deposition (PD) in response to increasing dietary sulfur amino acids (SAA) with or without immune system (IS) activation. The main plot was with and without IS activation, and 4 diets containing different amounts of standardized ileal digestible (SID) SAA (SAA to Lys ratios of 0.45, 0.55, 0.65 and 0.75) were the subplots. Activation of IS was achieved by intramuscular injection of Escherichia coli lipopolysaccharides (LPS; serotype 055:B5, Sigma; 30 μg/kg BW) every Monday and Thursday, with control pigs injected with sterile saline. Maximum body PD, measured by dual-energy X-ray absorptiometry (DXA), and minimum plasma urea content were achieved at SID SAA:Lys ratio of 0.55 in saline-injected pigs but were achieved at a SID SAA:Lys ratio of 0.75 in IS-activated pigs. Immune system activation increased rectal temperature (P < 0.05), plasma haptoglobin (1.1 vs. 2.0 mg/mL; P < 0.001), and the proportion of neutrophils (0.39 vs. 0.42; P < 0.05) and decreased serum albumin content (38.4 vs. 36.8 g/L; P < 0.01). Increasing dietary SAA had no effects on these variables. Immune system-activated pigs had lower levels of homocysteine (Hcy; P < 0.001) and a lower Ser content (P < 0.05). Results showed that increasing dietary SAA as DL-methionine in growing and/or finishing pigs altered plasma AA contents, and that use efficiency of the AA was improved when greater levels of SAA were supplemented in IS-activated pigs.