Browsing by Author "Chan, Cliburn"
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Item Open Access A randomized phase 2 trial of pembrolizumab versus pembrolizumab and acalabrutinib in patients with platinum-resistant metastatic urothelial cancer.(Cancer, 2020-08-05) Zhang, Tian; Harrison, Michael R; O'Donnell, Peter H; Alva, Ajjai S; Hahn, Noah M; Appleman, Leonard J; Cetnar, Jeremy; Burke, John M; Fleming, Mark T; Milowsky, Matthew I; Mortazavi, Amir; Shore, Neal; Sonpavde, Guru P; Schmidt, Emmett V; Bitman, Bojena; Munugalavadla, Veerendra; Izumi, Raquel; Patel, Priti; Staats, Janet; Chan, Cliburn; Weinhold, Kent J; George, Daniel JBACKGROUND:Inhibition of the programmed cell death protein 1 (PD-1) pathway has demonstrated clinical benefit in metastatic urothelial cancer (mUC); however, response rates of 15% to 26% highlight the need for more effective therapies. Bruton tyrosine kinase (BTK) inhibition may suppress myeloid-derived suppressor cells (MDSCs) and improve T-cell activation. METHODS:The Randomized Phase 2 Trial of Acalabrutinib and Pembrolizumab Immunotherapy Dual Checkpoint Inhibition in Platinum-Resistant Metastatic Urothelial Carcinoma (RAPID CHECK; also known as ACE-ST-005) was a randomized phase 2 trial evaluating the PD-1 inhibitor pembrolizumab with or without the BTK inhibitor acalabrutinib for patients with platinum-refractory mUC. The primary objectives were safety and objective response rates (ORRs) according to the Response Evaluation Criteria in Solid Tumors, version 1.1. Secondary endpoints included progression-free survival (PFS) and overall survival (OS). Immune profiling was performed to analyze circulating monocytic MDSCs and T cells. RESULTS:Seventy-five patients were treated with pembrolizumab (n = 35) or pembrolizumab plus acalabrutinib (n = 40). The ORR was 26% with pembrolizumab (9% with a complete response [CR]) and 20% with pembrolizumab plus acalabrutinib (10% with a CR). The grade 3/4 adverse events (AEs) that occurred in ≥15% of the patients were anemia (20%) with pembrolizumab and fatigue (23%), increased alanine aminotransferase (23%), urinary tract infections (18%), and anemia (18%) with pembrolizumab plus acalabrutinib. One patient treated with pembrolizumab plus acalabrutinib had high MDSCs at the baseline, which significantly decreased at week 7. Overall, MDSCs were not correlated with a clinical response, but some subsets of CD8+ T cells did increase during the combination treatment. CONCLUSIONS:Both treatments were generally well tolerated, although serious AE rates were higher with the combination. Acalabrutinib plus pembrolizumab did not improve the ORR, PFS, or OS in comparison with pembrolizumab alone in mUC. Baseline and on-treatment peripheral monocytic MDSCs were not different in the treatment cohorts. Proliferating CD8+ T-cell subsets increased during treatment, particularly in the combination cohort. Ongoing studies are correlating these peripheral immunome findings with tissue-based immune cell infiltration.Item Open Access Computational Methods for Comparative Analysis of Rare Cell Subsets in Flow Cytometry(2013) Frelinger, Jacob JeffreyAutomated analysis techniques for flow cytometry data can address many of the limitations of manual analysis by providing an objective approach for the identification of cellular subsets. While automated analysis has the potential to significantly improve automated analysis, challenges remain for automated methods in cross sample analysis for large scale studies. This thesis presents new methods for data normalization, sample enrichment for rare events of interest, and cell subset relabeling. These methods build upon and extend the use of Gaussian mixture models in automated flow cytometry analysis to enable practical large scale cell subset identification.
Item Open Access Establishment of normative ranges of the healthy human immune system with comprehensive polychromatic flow cytometry profiling.(PloS one, 2019-01) Yi, John S; Rosa-Bray, Marilyn; Staats, Janet; Zakroysky, Pearl; Chan, Cliburn; Russo, Melissa A; Dumbauld, Chelsae; White, Scott; Gierman, Todd; Weinhold, Kent J; Guptill, Jeffrey TExisting normative flow cytometry data have several limitations including small sample sizes, incompletely described study populations, variable flow cytometry methodology, and limited depth for defining lymphocyte subpopulations. To overcome these issues, we defined high-dimensional flow cytometry reference ranges for the healthy human immune system using Human Immunology Project Consortium methodologies after carefully screening 127 subjects deemed healthy through clinical and laboratory testing. We enrolled subjects in the following age cohorts: 18-29 years, 30-39, 40-49, and 50-66 and enrolled cohorts to ensure an even gender distribution and at least 30% non-Caucasians. From peripheral blood mononuclear cells, flow cytometry reference ranges were defined for >50 immune subsets including T-cell (activation, maturation, T follicular helper and regulatory T cell), B-cell, and innate cells. We also developed a web tool for visualization of the dataset and download of raw data. This dataset provides the immunology community with a resource to compare and extract data from rigorously characterized healthy subjects across age groups, gender and race.Item Open Access Evidence2Practice (E2P): Leveraging Implementation Science to Promote Careers in HIV Research Among Students From Historically Black Colleges and Universities.(Journal of acquired immune deficiency syndromes (1999), 2023-10) Okeke, Nwora Lance; Ware, Kenric B; Campbell, Russell; Taylor, Jamilah; Hung, Frances; Questell, Caroline; Brickler, Mildred P; Smith, Ukamaka D; Nawas, George T; Hanlen-Rosado, Emily; Chan, Cliburn; Bosworth, Hayden B; Aifah, Angela; Corneli, AmyBackground
The HIV research workforce is not representative of populations most affected by the epidemic. Innovative educational programs are needed to motivate diverse student populations to pursue careers in HIV research.Methods
The Duke University Center for AIDS Research Evidence2Practice (E2P) program is a 3-day interactive workshop that introduces students from Historically Black Colleges and Universities (HBCU) to HIV pre-exposure prophylaxis, implementation science, and human-centered design. Participants develop 1-page action plans to increase awareness and uptake of pre-exposure prophylaxis on their campus. The program was evaluated using a partially mixed-method concurrent equal status study design with pre-program and post-program surveys and in-depth interviews.Results
Among the 52 participating students, 44 completed the preworkshop survey, 45 completed the postworkshop survey, and 10 participated in an in-depth interview. Most participants identified as Black or African American and cisgender female. Participating in the E2P program was associated with: (1) an increase in median interest in pursuing a career in HIV research (P < 0.01) and (2) a decrease in median perceived difficulty in starting a career in HIV research (P < 0.01). Several students described that a lack of knowledge about initiating an HIV research career, a perceived lack of qualifications and knowledge about HIV science, and limited experience were major barriers to considering careers in HIV research.Conclusions
The E2P program enhanced HBCU students' interest in careers related to HIV research and improved their self-efficacy to pursue such careers. On-campus educational enrichment initiatives, led by active HIV researchers and clinicians, should be a critical part of diversifying the HIV workforce.Item Open Access Flow Cytometry Characterization of Cerebrospinal Fluid Monocytes in Patients With Postoperative Cognitive Dysfunction: A Pilot Study.(Anesthesia and analgesia, 2019-05-03) Berger, Miles; Murdoch, David M; Staats, Janet S; Chan, Cliburn; Thomas, Jake P; Garrigues, Grant E; Browndyke, Jeffrey N; Cooter, Mary; Quinones, Quintin J; Mathew, Joseph P; Weinhold, Kent J; MADCO-PC Study TeamAnimal models suggest postoperative cognitive dysfunction may be caused by brain monocyte influx. To study this in humans, we developed a flow cytometry panel to profile cerebrospinal fluid (CSF) samples collected before and after major noncardiac surgery in 5 patients ≥60 years of age who developed postoperative cognitive dysfunction and 5 matched controls who did not. We detected 12,654 ± 4895 cells/10 mL of CSF sample (mean ± SD). Patients who developed postoperative cognitive dysfunction showed an increased CSF monocyte/lymphocyte ratio and monocyte chemoattractant protein 1 receptor downregulation on CSF monocytes 24 hours after surgery. These pilot data demonstrate that CSF flow cytometry can be used to study mechanisms of postoperative neurocognitive dysfunction.Item Open Access FlowKit: A Python Toolkit for Integrated Manual and Automated Cytometry Analysis Workflows.(Frontiers in immunology, 2021-01) White, Scott; Quinn, John; Enzor, Jennifer; Staats, Janet; Mosier, Sarah M; Almarode, James; Denny, Thomas N; Weinhold, Kent J; Ferrari, Guido; Chan, CliburnAn important challenge for primary or secondary analysis of cytometry data is how to facilitate productive collaboration between domain and quantitative experts. Domain experts in cytometry laboratories and core facilities increasingly recognize the need for automated workflows in the face of increasing data complexity, but by and large, still conduct all analysis using traditional applications, predominantly FlowJo. To a large extent, this cuts domain experts off from the rapidly growing library of Single Cell Data Science algorithms available, curtailing the potential contributions of these experts to the validation and interpretation of results. To address this challenge, we developed FlowKit, a Gating-ML 2.0-compliant Python package that can read and write FCS files and FlowJo workspaces. We present examples of the use of FlowKit for constructing reporting and analysis workflows, including round-tripping results to and from FlowJo for joint analysis by both domain and quantitative experts.Item Open Access HLA Loci and Recurrence of Focal Segmental Glomerulosclerosis in Pediatric Kidney Transplantation.(Transplantation direct, 2021-10) Shaw, Brian I; Ochoa, Alejandro; Chan, Cliburn; Nobuhara, Chloe; Gbadegesin, Rasheed; Jackson, Annette M; Chambers, Eileen TRecurrent focal segmental glomerulosclerosis (FSGS) after kidney transplantation accounts for the majority of allograft failures in children with primary FSGS. Although current research focuses on FSGS pathophysiology, a common etiology and mechanisms of disease recurrence remain elusive.Methods
We performed a retrospective review of the Scientific Registry of Transplant Recipients to determine the association of specific HLA recurrence of FSGS. Kidney transplants recipients under the age of 19 who were diagnosed with FSGS, who were transplanted after January 1, 2000, and who had complete HLA data were included in the study. We performed simple logistic regression on all HLA A, B, C, DR, and DQ represented in the dataset and FSGS recurrence and then determined those associated with recurrence using the Benjamini-Hochberg method for multiple comparisons. For those HLAs that were associated with recurrence, we further determined the effect of matching recipient and donor HLA with recurrence.Results
HLA DR7, DR53, DQ2, DR52, and DQ7 were associated with increased or decreased risk of recurrent disease after transplantation. We identified a risk haplotype consisting of HLA-DR7, DR53, and DQ2 that was consistently associated with an increased risk of recurrence (odds ratio 1.91; 95% confidence interval, 1.44-2.54, P < 0.001). We also found that donor/recipient concordance for HLA-DQ7 was associated with a decreased risk of recurrence (odds ratio 0.42; 95% confidence interval, 0.37-0.53, P = 0.009).Conclusions
HLA profiles may be used for risk stratification of recurrence of FSGS in pediatric kidney transplant recipients and deserves further study.Item Open Access Immunomodulatory lipid mediator profiling of cerebrospinal fluid following surgery in older adults.(Scientific reports, 2021-02-04) Terrando, Niccolò; Park, John J; Devinney, Michael; Chan, Cliburn; Cooter, Mary; Avasarala, Pallavi; Mathew, Joseph P; Quinones, Quintin J; Maddipati, Krishna Rao; Berger, Miles; MADCO-PC Study TeamArachidonic acid (AA), docosahexaenoic acid (DHA), and eicosapentaenoic acid (EPA) derived lipids play key roles in initiating and resolving inflammation. Neuro-inflammation is thought to play a causal role in perioperative neurocognitive disorders, yet the role of these lipids in the human central nervous system in such disorders is unclear. Here we used liquid chromatography-mass spectrometry to quantify AA, DHA, and EPA derived lipid levels in non-centrifuged cerebrospinal fluid (CSF), centrifuged CSF pellets, and centrifuged CSF supernatants of older adults obtained before, 24 h and 6 weeks after surgery. GAGE analysis was used to determine AA, DHA and EPA metabolite pathway changes over time. Lipid mediators derived from AA, DHA and EPA were detected in all sample types. Postoperative lipid mediator changes were not significant in non-centrifuged CSF (p > 0.05 for all three pathways). The AA metabolite pathway showed significant changes in centrifuged CSF pellets and supernatants from before to 24 h after surgery (p = 0.0000247, p = 0.0155 respectively), from before to 6 weeks after surgery (p = 0.0000497, p = 0.0155, respectively), and from 24 h to 6 weeks after surgery (p = 0.0000499, p = 0.00363, respectively). These findings indicate that AA, DHA, and EPA derived lipids are detectable in human CSF, and the AA metabolite pathway shows postoperative changes in centrifuged CSF pellets and supernatants.Item Open Access Multivariate analysis of FcR-mediated NK cell functions identifies unique clustering among humans and rhesus macaques.(Frontiers in immunology, 2023-01) Tuyishime, Marina; Spreng, Rachel L; Hueber, Brady; Nohara, Junsuke; Goodman, Derrick; Chan, Cliburn; Barfield, Richard; Beck, Whitney E; Jha, Shalini; Asdell, Stephanie; Wiehe, Kevin; He, Max M; Easterhoff, David; Conley, Haleigh E; Hoxie, Taylor; Gurley, Thaddeus; Jones, Caroline; Adhikary, Nihar Deb; Villinger, Francois; Thomas, Rasmi; Denny, Thomas N; Moody, Michael Anthony; Tomaras, Georgia D; Pollara, Justin; Reeves, R Keith; Ferrari, GuidoRhesus macaques (RMs) are a common pre-clinical model used to test HIV vaccine efficacy and passive immunization strategies. Yet, it remains unclear to what extent the Fc-Fc receptor (FcR) interactions impacting antiviral activities of antibodies in RMs recapitulate those in humans. Here, we evaluated the FcR-related functionality of natural killer cells (NKs) from peripheral blood of uninfected humans and RMs to identify intra- and inter-species variation. NKs were screened for FcγRIIIa (human) and FcγRIII (RM) genotypes (FcγRIII(a)), receptor signaling, and antibody-dependent cellular cytotoxicity (ADCC), the latter mediated by a cocktail of monoclonal IgG1 antibodies with human or RM Fc. FcγRIII(a) genetic polymorphisms alone did not explain differences in NK effector functionality in either species cohort. Using the same parameters, hierarchical clustering separated each species into two clusters. Importantly, in principal components analyses, ADCC magnitude, NK contribution to ADCC, FcγRIII(a) cell-surface expression, and frequency of phosphorylated CD3ζ NK cells all contributed similarly to the first principal component within each species, demonstrating the importance of measuring multiple facets of NK cell function. Although ADCC potency was similar between species, we detected significant differences in frequencies of NK cells and pCD3ζ+ cells, level of cell-surface FcγRIII(a) expression, and NK-mediated ADCC (P<0.001), indicating that a combination of Fc-FcR parameters contribute to overall inter-species functional differences. These data strongly support the importance of multi-parameter analyses of Fc-FcR NK-mediated functions when evaluating efficacy of passive and active immunizations in pre- and clinical trials and identifying correlates of protection. The results also suggest that pre-screening animals for multiple FcR-mediated NK function would ensure even distribution of animals among treatment groups in future preclinical trials.Item Open Access Prognosis and treatment effects of HIV-associated talaromycosis in a real-world patient cohort.(Medical mycology, 2021-04) Klus, Jonathan; Ly, Vo Trieu; Chan, Cliburn; Le, ThuyTalaromycosis is a leading cause of AIDS-associated opportunistic infections and death in Southeast Asia. We have recently shown in the Itraconazole versus Amphotericin for Talaromycosis (IVAP) trial that induction therapy with amphotericin B reduced mortality over 24 weeks, but not during the first 2 weeks. Antifungal treatment effects in real-world settings have not been rigorously evaluated. Using data obtained from patient records at the Hospital for Tropical Diseases, Ho Chi Minh City, Vietnam from 2004 to 2009, we first developed a prognostic model using Bayesian logistic regression to identify predictors of death. Second, we developed a causal model using propensity score matching to assess the treatment effects of amphotericin B and itraconazole. Our prognostic model identified intravenous drug use (odds ratio [OR] = 2.01), higher respiratory rate (OR = 1.12), higher absolute lymphocyte count (OR = 1.62), a concurrent respiratory infection (OR = 1.67) or central nervous system infection (OR = 2.66) as independent predictors of death. Fever (OR = 0.56) was a protective factor. Our prognostic model exhibits good in-sample performance and out-of-sample validation, with a discrimination power of 0.85 and 0.91, respectively. Our causal model showed no significant difference in treatment outcomes between amphotericin B and itraconazole over the first 2 weeks (95% credible interval: 0.62, 2.50). Our prognostic model provides a simple tool based on routinely collected clinical data to predict individual patient outcome. Our causal model shows similar results to the IVAP trial at 2 weeks, demonstrating an agreement between real-world data and clinical trial data.Item Open Access Setting objective thresholds for rare event detection in flow cytometry.(J Immunol Methods, 2014-07) Richards, Adam J; Staats, Janet; Enzor, Jennifer; McKinnon, Katherine; Frelinger, Jacob; Denny, Thomas N; Weinhold, Kent J; Chan, CliburnThe accurate identification of rare antigen-specific cytokine positive cells from peripheral blood mononuclear cells (PBMC) after antigenic stimulation in an intracellular staining (ICS) flow cytometry assay is challenging, as cytokine positive events may be fairly diffusely distributed and lack an obvious separation from the negative population. Traditionally, the approach by flow operators has been to manually set a positivity threshold to partition events into cytokine-positive and cytokine-negative. This approach suffers from subjectivity and inconsistency across different flow operators. The use of statistical clustering methods does not remove the need to find an objective threshold between between positive and negative events since consistent identification of rare event subsets is highly challenging for automated algorithms, especially when there is distributional overlap between the positive and negative events ("smear"). We present a new approach, based on the Fβ measure, that is similar to manual thresholding in providing a hard cutoff, but has the advantage of being determined objectively. The performance of this algorithm is compared with results obtained by expert visual gating. Several ICS data sets from the External Quality Assurance Program Oversight Laboratory (EQAPOL) proficiency program were used to make the comparisons. We first show that visually determined thresholds are difficult to reproduce and pose a problem when comparing results across operators or laboratories, as well as problems that occur with the use of commonly employed clustering algorithms. In contrast, a single parameterization for the Fβ method performs consistently across different centers, samples, and instruments because it optimizes the precision/recall tradeoff by using both negative and positive controls.Item Open Access The INTUIT Study: Investigating Neuroinflammation Underlying Postoperative Cognitive Dysfunction.(Journal of the American Geriatrics Society, 2019-01-23) Berger, Miles; Oyeyemi, Deborah; Olurinde, Mobolaji O; Whitson, Heather E; Weinhold, Kent J; Woldorff, Marty G; Lipsitz, Lewis A; Moretti, Eugene; Giattino, Charles M; Roberts, Kenneth C; Zhou, Junhong; Bunning, Thomas; Ferrandino, Michael; Scheri, Randall P; Cooter, Mary; Chan, Cliburn; Cabeza, Roberto; Browndyke, Jeffrey N; Murdoch, David M; Devinney, Michael J; Shaw, Leslie M; Cohen, Harvey Jay; Mathew, Joseph P; INTUIT InvestigatorsBACKGROUND/OBJECTIVES:Every year, up to 40% of the more than 16 million older Americans who undergo anesthesia/surgery develop postoperative cognitive dysfunction (POCD) or delirium. Each of these distinct syndromes is associated with decreased quality of life, increased mortality, and a possible increased risk of Alzheimer's disease. One pathologic process hypothesized to underlie both delirium and POCD is neuroinflammation. The INTUIT study described here will determine the extent to which postoperative increases in cerebrospinal fluid (CSF) monocyte chemoattractant protein 1 (MCP-1) levels and monocyte numbers are associated with delirium and/or POCD and their underlying brain connectivity changes. DESIGN:Observational prospective cohort. SETTING:Duke University Medical Center, Duke Regional Hospital, and Duke Raleigh Hospital. PARTICIPANTS:Patients 60 years of age or older (N = 200) undergoing noncardiac/nonneurologic surgery. MEASUREMENTS:Participants will undergo cognitive testing before, 6 weeks, and 1 year after surgery. Delirium screening will be performed on postoperative days 1 to 5. Blood and CSF samples are obtained before surgery, and 24 hours, 6 weeks, and 1 year after surgery. CSF MCP-1 levels are measured by enzyme-linked immunosorbent assay, and CSF monocytes are assessed by flow cytometry. Half the patients will also undergo pre- and postoperative functional magnetic resonance imaging scans. 32-channel intraoperative electroencephalogram (EEG) recordings will be performed to identify intraoperative EEG correlates of neuroinflammation and/or postoperative cognitive resilience. Eighty patients will also undergo home sleep apnea testing to determine the relationships between sleep apnea severity, neuroinflammation, and impaired postoperative cognition. Additional assessments will help evaluate relationships between delirium, POCD, and other geriatric syndromes. CONCLUSION:INTUIT will use a transdisciplinary approach to study the role of neuroinflammation in postoperative delirium and cognitive dysfunction and their associated functional brain connectivity changes, and it may identify novel targets for treating and/or preventing delirium and POCD and their sequelae.Item Open Access The Multiscale Systems Immunology project: software for cell-based immunological simulation.(Source Code Biol Med, 2008-04-28) Mitha, Faheem; Lucas, Timothy A; Feng, Feng; Kepler, Thomas B; Chan, CliburnBACKGROUND: Computer simulations are of increasing importance in modeling biological phenomena. Their purpose is to predict behavior and guide future experiments. The aim of this project is to model the early immune response to vaccination by an agent based immune response simulation that incorporates realistic biophysics and intracellular dynamics, and which is sufficiently flexible to accurately model the multi-scale nature and complexity of the immune system, while maintaining the high performance critical to scientific computing. RESULTS: The Multiscale Systems Immunology (MSI) simulation framework is an object-oriented, modular simulation framework written in C++ and Python. The software implements a modular design that allows for flexible configuration of components and initialization of parameters, thus allowing simulations to be run that model processes occurring over different temporal and spatial scales. CONCLUSION: MSI addresses the need for a flexible and high-performing agent based model of the immune system.Item Open Access Toward development of a comprehensive external quality assurance program for polyfunctional intracellular cytokine staining assays(Journal of Immunological Methods, 2014-01-01) Staats, Janet S; Enzor, Jennifer H; Sanchez, Ana M; Rountree, Wes; Chan, Cliburn; Jaimes, Maria; Chan, Ray Chun-Fai; Gaur, Amitabh; Denny, Thomas N; Weinhold, Kent JThe External Quality Assurance Program Oversight Laboratory (EQAPOL) Flow Cytometry Program assesses the proficiency of NIH/NIAID/DAIDS-supported and potentially other interested research laboratories in performing Intracellular Cytokine Staining (ICS) assays. The goal of the EQAPOL Flow Cytometry External Quality Assurance Program (EQAP) is to provide proficiency testing and remediation for participating sites. The program is not punitive; rather, EQAPOL aims to help sites identify areas for improvement. EQAPOL utilizes a highly standardized ICS assay to minimize variability and readily identify those sites experiencing technical difficulties with their assays. Here, we report the results of External Proficiency 3 (EP3) where participating sites performed a 7-color ICS assay. On average, sites perform well in the Flow Cytometry EQAP (median score is "Good"). The most common technical issues identified by the program involve protocol adherence and data analysis; these areas have been the focus of site remediation. The EQAPOL Flow Cytometry team is now in the process of expanding the program to 8-color ICS assays. Evaluating polyfunctional ICS responses would align the program with assays currently being performed in support of HIV immune monitoring assays. © 2014 Elsevier B.V.