Analysis Of Subpopulations Research Articles

Change in breast cancer risk perception related to perceived COVID-19 risk: A WISDOM sub-population analysis.

e24135 Background: Obstacles to access to care due to the perceptions of the risk posed by COVID-19 have led to unprecedented disruptions in cancer care. Yet, little is understood about whether perceived COVID-19 risk influences perceptions of cancer risk. We examined how COVID-19 risk perception was associated with perceptions of breast cancer risk over one year of the pandemic among women enrolled in the WISDOM study, a PCORI-funded pragmatic trial testing risk-based cancer screening that began before the pandemic. Methods: We conducted four longitudinal surveys among the 13,002 women enrolled in the WISDOM study from May - December 2020. Responses from 8,285 women are eligible for inclusion in this analysis leading to a total sample size of 16,859 survey responses. Surveys were conducted online and asked women’s perceived lifetime chance of developing breast cancer (0-100%). COVID-19 risk perception was reported on a 5-point scale from Very Low to Very High. We computed the difference between breast cancer risk perception at each COVID-19 survey to pre-COVID breast cancer risk perception, measured as a secondary aim of the WISDOM study, and compared that to COVID-19 risk perception at each time point. Results: Across the four survey waves, most women perceived low COVID risk: 29% very low, 42% moderately low, 23% neither high nor low and 6% high or very high. Overall, breast cancer risk perception declined for those with very low COVID-19 risk perception and rose for women in the highest levels of COVID-19 risk perception. However, changes in breast cancer risk perception associated with COVID risk perception were small. For example, in survey wave 4, breast cancer risk change was -2.4% very low, -1.4% low, 2.5% not high or low and 3.1% high or very high. (Table). Conclusions: Among women participating in a pragmatic trial testing risk-based cancer screening, COVID risk perception had a small relationship with change in breast cancer risk perception. Change in breast cancer risk perception paralleled COVID-19 risk perception. This calls for exploration of the underpinnings of these risk changes and may have implications for changes in cancer screening behavior related to COVID-19.[Table: see text]

Immune Response in Vitamin D Deficient Metastatic Colorectal Cancer Patients: A Player That Should Be Considered for Targeted Vitamin D Supplementation.

Simple SummaryIn this study, we investigated the role of vitamin D levels and the impact on immune response in patients with mCRC and propose a vitamin D cut-off level. Among several hematological, biochemical and immunological variables, we identified the neutrophils-to-lymphocytes ratio, CD4+ T lymphocytes and B lymphocytes as being closely related to vitamin D status with an impact on survival.Background: Vitamin D deficiency is a poor prognostic factor in metastatic colorectal cancer (mCRC); however, targeted supplementation trials have so far yielded limited results. We investigated clinical-laboratory parameters influencing vitamin D deficiency, with a particular focus on immune response, and the effect on survival. These parameters could help optimize targeted supplementation therapy. Methods: Association of plasma 25-hydroxyvitamin D (25(OH])D) with overall survival (OS) was assessed with the Hazard Ratio Smoothed Curve with Restricted Cubic Splines (HRSC-RCS) and maximally selected rank statistics (MSRS) in mCRC patients who underwent first-line chemotherapy. Several hematobiochemical variables were evaluated as predictors of vitamin D deficiency by means of Least Absolute Shrinkage and Selection Operator (LASSO) analysis. In a patient subset, peripheral lymphocyte subpopulations were also analyzed. Results: One hundred thirty-three mCRC patients were included. The median(m) baseline 25(OH)D was 10.8 ng/mL (range 3–53.4). HRSC-RCS revealed a linear association between 25(OH)D and OS. MSRS found 10 ng/mL as the optimal 25(OH)D cut-off. The median OS for 25(OH)D < 10 (n = 60) vs. > 10 ng/mL (n = 73) was 12.3 and 24.5 months, respectively (p = 0.002). The LASSO analysis identified high neutrophil-to-lymphocyte ratio (NLR > 3.5) as the strongest predictor of vitamin D deficiency (Odds Ratio 3.35, p 0.0009). Moreover, patients with low 25(OH)D levels (< 10 ng/mL) and high NLR (>3.5) had the shortest survival and patients with 25(OH)D >10 ng/mL and NLR <3.5 had the longest: mOS 8.1 and 28.1 months, respectively, HR 3.40 (1.76–6.59), p 0.0004. Besides the significant difference in NLR between 25(OH)D < and > 10 ng/mL patients (mNLR 3.6 vs. 2.9, p 0.03), the lymphocyte subpopulation analysis revealed that vitamin D deficiency was associated with high T- CD4+ (p = 0.04) and low B (p = 0.03) lymphocyte frequency. Conclusions: NLR is a powerful predictor of Vitamin D deficiency and can further help in stratifying prognosis. Vitamin D deficiency was associated with significant variations in peripheral immune cells. We hypothesize that integrated targeted interventions to both vitamin D and immune system would improve the prognosis of mCRC patients.

Sociodemographic Factors and Health Insurance Coverage Are Associated with Invasive Breast Cancer in Tennessee: Appalachian and Non-Appalachian County Comparison.

Background:Tennessean women experience the 12th highest breast cancer (BC) mortality in the United States. Yet, few studies have examined BC outcomes among Tennessean women in and outside of Appalachia. We examined whether sociodemographic factors and health insurance status were associated with invasive BC in Tennessee by Appalachian and non-Appalachian county designation.Materials and Methods:Using the Tennessee Cancer Registry, we identified 52,187 women, aged ≥18, diagnosed with BC between 2005 and 2015. Multivariable logistic regression was performed to examine associations between invasive BC and sociodemographic characteristics, health insurance coverage, and county designation (Appalachian/non-Appalachian). Regression analyses stratified by county designation were subsequently performed.Results:In Tennessee, younger women had lower odds of invasive BC diagnosis (<45: odds ratio [OR] = 0.74, 95% confidence interval [CI] = 0.67–0.81; 55–64: OR = 0.91, 95% CI = 0.84–0.97) compared to women ≥65. Married Tennessean women had 12% (95% CI = 1.04–1.21) higher odds of invasive BC than single women. Further, both public (OR = 1.81, 95% CI = 1.41–2.33) and private (OR = 1.36, 95% CI = 1.06–1.76) health insurance were found to increase odds of invasive BC compared to no insurance/self-pay. Results from the subpopulation analyses were largely consistent with overall findings. In Appalachian counties, women on public health insurance had increased odds (OR = 1.42, 95% CI = 1.00–2.03) of invasive BC compared to uninsured/self-pay women, while in non-Appalachian counties, women insured both publicly (OR = 2.25, 95% CI = 1.57–3.24) and privately (OR = 1.68, 95% CI = 1.16–2.24) had increased odds of invasive BC.Conclusions:The results identify risk factors for Tennessean women in Appalachian and non-Appalachian counties whose malignancies evaded early detection, increasing risk of mortality.

Analysis of cerebrospinal fluid cells by flow cytometry: Comparison to conventional cytology.

This study compared the results obtained by basic immunophenotyping of cerebrospinal fluid (CSF) cells by flow cytometry (FC) to the results of conventional cytology and evaluated the possibility of detailed analyses of CSF B-cell subpopulations. Samples from 42 patients were examined by conventional cytology (native and/or pre-centrifuged CSF) and FC. The results from 15 patients without evidence of organic neurological disease were used to estimate reference ranges. Pre-centrifugated CSF had significantly higher cell yield on the cytologic slide, but cell subpopulation percentages were altered; the percentage of lymphocytes was significantly higher and monocytes significantly lower compared to both native CSF slides and FC. The percentage of granulocytes was higher in FC compared to cytology. For leukocyte count, the following reference ranges were estimated for Fuchs-Rosenthal chamber (FR) counting and FC, respectively: leukocytes ≤4.7/μL and ≤2.5/μL, lymphocytes ≤4.1/μL and ≤1.8/μL, monocytes ≤1.2/μL and ≤0.9/μL, and granulocytes 0/μL and ≤0.2/μL. The following reference ranges were estimated for basic subpopulations: T-lymphocytes 84.1-100%, B lymphocytes 0.0-1.5%, NK cells 0.0-6.3%, NKT cells 0-9.5%, and CD3+CD4+/CD3+CD8+ 0.8-4.9. Using a volume of 1.2-2.4 mL, the number of B lymphocytes was too low (<20) in samples with ≤2.7 cells/μL in the FR. Even normal CSF samples are amenable to basic mononuclear cell subpopulation analysis by FC. However, analysis of the B-cell subpopulations requires either a larger sample volume or selection of samples with ≥ 3 cells/μL.

Role of Axonal Translation in the Upregulation of Nav1.1 Following Nerve Injury

Recent data indicates that an increase in axonal NaV1.1 enables action potential (AP) propagation in a subpopulation of axons in the trigeminal nerve following injury, and that selective block of this channel can attenuate chronic constriction injury (CCI)-induced mechanical hypersensitivity. The increase in axonal NaV1.1 is not associated with an increase in protein or mRNA in the trigeminal ganglion (TG), suggesting this upregulation is due to stabilization, local translation, or increased trafficking. The goal of present study to begin to assess contribution of local translation as well as identify the afferent subpopulation(s) in which the increase occurs. To address this, quantitative RT-PCR and Western blot (WB) of the infra-orbital nerve (ION) with and without pharmacological manipulations translation and trafficking were initiated. To enable analysis of afferent subpopulations in which NaV1.1 was upregulated, GCaMP6s expression in trigeminal ganglia was induced by AAV injection in neonatal rats. Results from this series of experiments confirms that mRNA is detectable in the ION, consistent with previous data reported by Korczeniewska et al (Eur. J Pain 2020, PMID: 32100907), though no apparent differences were observed with CCI-ION. Additionally, neonatal AAV9-GCaMP6s injection provided robust expression throughout the TG for in-vivo imaging. The absence of a detectable decrease in axonal NaV1.1 mRNA is consistent with an increase in mRNA trafficking to compensate for an increase in local translation. Ongoing experiments will be used to confirm these initial observations. Grant support from 1F31NS125993-01.

Dynamic evolution of imipenem/relebactam resistance in a KPC-producing Klebsiella pneumoniae from a single patient during ceftazidime/avibactam-based treatments.

The novel carbapenem/β-lactamase inhibitor combination imipenem/cilastatin/relebactam has been developed for the treatment of infections due to carbapenemase-producing Enterobacteriaceae (CPE). Herein, we describe the in vivo evolution of imipenem/cilastatin/relebactam resistance in longitudinal intra-patient Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae (KPC-Kp) strains isolated from a patient following ceftazidime/avibactam-based treatments. WGS analysis was performed on KPC-Kp strains isolated at different times and during antimicrobial treatments from the same patient. Genome assemblies were performed using a hybrid approach using Illumina iSeq 100 and Minion Oxford Nanopore platforms. Subpopulation analysis and allele frequency determination was performed by mapping Illumina reads to blaKPC. During antimicrobial treatment, resistance to ceftazidime/avibactam was observed following 16 days of antimicrobial therapy. WGS results showed that all KPC-Kp exhibited a low SNP rate of divergence, belonged to ST512 and shared similar antimicrobial resistance and porin gene patterns. Genetic analysis demonstrated that the first ceftazidime/avibactam-resistant KPC-Kp strain harboured a blaKPC-53 gene in a Tn4401 transposon moved from IncFII(K) to a 43 kb IncX3 plasmid, while a imipenem/cilastatin/relebactam-resistant strain exhibited two copies of the Tn4401 transposon in IncFII(K) and IncX3 plasmids, resulting in an increased blaKPC copy number. Of note, frequency analysis demonstrated that imipenem/cilastatin/relebactam-resistant KPC-Kp consisted of mixed subpopulations harbouring blaKPC-40 and blaKPC-53 alleles. Our results show the in vivo evolution of genetic rearrangement conferring resistance to imipenem/relebactam in a patient with KPC-Kp infection and treated with different ceftazidime/avibactam-based treatments. The rapid development of mutations and the high adaptability of its genome highlight the potential threat of KPC-Kp.

Safety and efficacy of apalutamide in Japanese patients with metastatic castration-sensitive prostate cancer receiving androgen deprivation therapy: Final report for the Japanese subpopulation analysis of the randomized, placebo-controlled, phase III TITAN study.

ObjectivesThe TITAN study is a randomized, double‐blind, placebo‐controlled, multinational trial that evaluated apalutamide with androgen deprivation therapy in patients with metastatic castration‐sensitive prostate cancer. At the first interim analysis in the Japanese subpopulation (median follow‐up 25.7 months), there was an improvement in overall survival and radiological progression‐free survival with apalutamide versus placebo. Here, we report the final analysis results for the Japanese subpopulation.MethodsPatients were randomized 1:1 to receive apalutamide 240 mg or placebo. After the first interim analysis, protocol treatment was unblinded, and crossover was allowed. Efficacy and safety were evaluated in the preplanned, event‐driven final analysis.ResultsFifty‐one patients were Japanese (apalutamide n = 28; placebo n = 23). After a median follow‐up of 46.0 months, the median overall survival was not reached neither in the apalutamide nor the placebo group; the hazard ratio was 0.45, favoring apalutamide, which was consistent with the overall population. Hazard ratios for time to cytotoxic chemotherapy (0.39), time to pain progression (0.87), and time to chronic opioid use (0.82) also favored apalutamide and were comparable with those of the overall population. Time to prostate‐specific antigen progression and progression‐free survival 2, respectively, was favored in the apalutamide group (0.21 and 0.44). Apalutamide was associated with higher incidences of rash and fracture in the Japanese subpopulation compared with the overall population.ConclusionsThe efficacy of apalutamide with androgen deprivation therapy in Japanese patients was consistent with efficacy demonstrated in the overall population. No new safety concerns emerged with long‐term follow‐up.

Diagnostic accuracy of urinary antigen tests for legionellosis: A systematic review and meta-analysis.

Urinary antigen tests (UATs) have been used for the early detection of legionellosis and have demonstrated moderate sensitivity and high specificity. However, the most recent systematic review and meta-analysis published in 2009 evaluated the accuracy of UATs; since then, UAT accuracy may have changed owing to advances and developments in UAT technology and epidemiological changes in the frequency of Legionella species that cause legionellosis. Therefore, this systematic review and meta-analysis aimed to update the accuracy of UATs for legionellosis among patients with suspected pneumonia. Overall, 1326 studies were screened, 21 of which fulfilled the eligibility criteria for quality assessment and meta-analysis. Data from 5772 patients, including 1368 (23.7%) with the target condition (i.e., suspected legionellosis), were included in the analysis. The overall quality of the included studies, which was assessed using the Quality Assessment of Diagnostic Accuracy Studies-2 tool, was unclear. The calculated pooled sensitivity and specificity were 0.79 (95% confidence interval [CI], 0.71-0.85) and 1.00 (95% CI, 0.99-1.00), respectively. Subpopulation analysis revealed that the accuracy of UATs for sensitivity and specificity for Legionella pneumophilia serogroup 1 was 0.86 (95% CI, 0.78-0.91) and 1.00 (95% CI, 0.99-1.00), respectively. This study demonstrated that the sensitivity and specificity of UATs were moderate and high, respectively, which is comparable to the results reported in 2009. Therefore, UATs may be a useful method for the early detection of legionellosis caused by Legionella pneumophila serogroup 1. The review protocol was prospectively registered with the University Hospital Medical Information Network Clinical Trials Registry (UMIN000041080).

Pathological response and predictive role of tumour-infiltrating lymphocytes in HER2-positive early breast cancer treated with neoadjuvant pyrotinib plus trastuzumab and chemotherapy (Panphila): a multicentre phase 2 trial

PurposePanphila evaluated pyrotinib plus trastuzumab, docetaxel and carboplatin as neoadjuvant therapy for early breast cancer (BC), and investigated the predictive role of immune cell subpopulations. Patients and methodsIn this multicentre phase 2 study, patients with human epidermal growth factor receptor 2-positive, stage T2-3N0-3M0 BC received pyrotinib 400 mg once daily plus docetaxel (75 mg/m2, day 1), carboplatin (6 mg/mL/min, day 1) and trastuzumab (8 mg/kg loading dose and 6 mg/kg maintenance dose, day 1) for 6 cycles of 21 days each. Simon's 2-stage design was adopted. The primary end-point was pathological complete response (pCR, ypT0/is ypN0) rate. Tumour-infiltrating lymphocytes (TILs) were assessed by haematoxylin and eosin staining and multiplex immunohistochemistry. ResultsIn the modified intention-to-treat population (n = 69), 38 patients (55.1%) achieved pCR. In the safety population (n = 74), the most common grade ≥3 adverse events were diarrhoea (43.2%), anaemia (37.8%), vomiting (16.2%) and platelet count decrease (10.8%). No treatment-related deaths occurred. Analysis of single immune subpopulations revealed a significant association of pCR with higher baseline infiltration by stromal (s)-CD20+, s-CD8+ and s-CD4+ TILs. Unsupervised hierarchical clustering of stromal immune markers identified a group of patients characterised by high s-CD20+, s-CD8+, s-CD4+ and s-FOXP3+ immune cells infiltration, which was independently associated with pCR. ConclusionNeoadjuvant pyrotinib plus trastuzumab-based chemotherapy exhibits promising efficacy and manageable toxicity in patients with human epidermal growth factor receptor 2-positive early BC, and thus phase 3 trials are warranted. Our findings also contribute to understanding the potential role of the immune microenvironment in response to neoadjuvant pyrotinib-based therapy.

A Randomized Placebo-Controlled Trial of Combination Therapy With Post-triple-antibiotic-therapy Fecal Microbiota Transplantation and Alginate for Ulcerative Colitis: Protocol.

BackgroundFecal microbiota transplantation (FMT) has been widely performed for ulcerative colitis (UC) treatment at the clinical trial stage. Previous reports have used multiple FMT methods to enhance the colonization of healthy donor microbiota in the recipient's intestines. FMT following triple antibiotic therapy with amoxicillin, fosfomycin, and metronidazole (A-FMT) is not only effective but also requires only one FMT, which improves dysbiosis caused by reduced Bacteroidetes diversity in patients with UC. Alginate and its derivatives have the potential to induce the growth of intestinal bacteria including Bacteroides members and produce short-chain fatty acids (SCFAs), which are beneficial in regulating overactive autoimmunity. Our trial aims to investigate whether post-intervention with alginate, which can improve the intestinal environment, will enhance the therapeutic effect of A-FMT in UC and increase the long-term remission rate.Methods and AnalysisThis trial is a double-blinded, randomized, placebo-controlled, parallel assignment trial. Patients with UC and fecal donation candidates will undergo strict screening before being involved in the trial. Eligible patients are randomly divided into two groups: one group will drink one bottle of alginate twice a day for 8 consecutive weeks after A-FMT, while the other group will take a placebo instead of the alginate drink. The primary endpoints are the changes in the Total Mayo Score at 8 weeks after study initiation and A-FMT from baseline. The secondary endpoint is the comparison of clinical features, microbiota, and metabolomic analysis before and after 8 weeks of study food intake. Changes at 6, 12, 18, and 24 months after A-FMT will be assessed. Finally, a subpopulation analysis of the relationship between patients and donors is an exploratory endpoint.DiscussionThe FMT post-treatment used in this study is an oral alginate drink that is easily accepted by patients. If the regimen achieves the desired results, it can further improve the A-FMT regimen and provide evidence for clinical practice guidelines for UC.Clinical Trial Registrationhttps://jrct.niph.go.jp/latest-detail/jRCTs031200103, identifier: jRCTs031200103.

Allergic rhinitis, allergic contact dermatitis and disease comorbidity belong to separate entities with distinct composition of T-cell subsets, cytokines, immunoglobulins and autoantibodies

BackgroundAllergic rhinitis (AR) and allergic contact dermatitis (ACD) are prevalent allergic diseases and have significant impacts on patients’ daily life. Despite many studies on AR or ACD have been conducted separately, little is known about the immune responses in patients of AR combined with ACD and the interplay between AR and ACD. Our study compared various aspects of immune elements in patients with AR or/and ACD, aiming to characterize the immune responses in AR, ACD, and AR combined with ACD.MethodsA total of 57 patients diagnosed with AR or/and ACD and 28 healthy volunteers were included. AR patients were further divided into seasonal AR (SAR) and perennial AR (PAR). All subjects’ blood samples were taken to assess the concentration of immunoglobulins, complement C3, C4, autoantibodies and cytokines in serum by immunoturbidimetry, ELISA or Luminex200 platform. Peripheral blood mononuclear cells (PBMCs) were subjected to the analysis of lymphocyte subpopulations by flow cytometry.ResultsIt indicated that AR disease caused elevated levels of IgE, IgA, IgG, IgG4, as well as IL-4, IL-15, IL-8 and IL-6 in serum. AR patients possessed a decreased CD4/CD8 ratio and an increased proportion of memory CD4 + T-cell subset, with a skewed Th2 response and an enhanced CD8 + T-cell activation. Compared with patients with sole AR or ACD condition, AR + ACD patients presented with a significantly increased proportion of memory CD8 + T-cell subset and were prone to autoimmune disorders as indicated by the increased autoantibodies. The immune elements in patients with ACD only were least affected compared with those in other conditions. Additionally, seasonal or perennial AR patients exhibited different cytokine profiles and proportions of memory T-cell subsets.ConclusionsIn this study, we illuminated the respective characteristics of immune responses in AR, ACD, and AR combined with ACD. Meanwhile, we discovered that the PAR and SAR patients possessed different cytokine profiles and T-cell compartments. It suggested that these allergic conditions belong to different disease entities. Characterizing the detailed immune changes in these allergic diseases would help to develop proper treatments targeting particular immune elements in different allergic diseases.

Acceptability and further potential of an app for maternal notes, the eMutterPass: A patient-centered online survey at the University Hospital Zurich

ObjectivesWe wanted to characterize the acceptability of and women’s satisfaction with the eMutterPass application. Particular attention was placed on concerns about data confidentiality and on willingness to use the app in an interactive way to share information about medication consumption. Study designThe present analysis is based on self-reported data from obstetric patients participating in an anonymous online survey between April 6th 2021 and April 20th 2021. ResultsDuring the 2-week timeframe, 1123 questionnaires were completed. The eMutterPass application was widely appreciated by our survey population and almost all participants would recommend the application to other pregnant women. A subpopulation analysis indicates that concerns about data confidentiality were higher among younger, multigravid and non-German-speaking pregnant women. The majority of women would be willing to report their medication use by taking pictures, filling in medication dosages or submitting assessments of perceived drug effectiveness. ConclusionThe development of our eMutterPass application meets the spirit of the times and gives pregnant women uncomplicated access to their own data. Concerns about data confidentiality can be adequately countered with additional information about the system structure. The largely positive adherence to the idea of reporting medication use on the app lays the groundwork for potential use of the eMutterPass for documentation of non-prescribed drugs.

A method for rapid flow-cytometric isolation of endothelial nuclei and RNA from archived frozen brain tissue

AbstractEndothelial cells are important contributors to brain development, physiology, and disease. Although RNA sequencing has contributed to the understanding of brain endothelial cell diversity, bulk analysis and single-cell approaches have relied on fresh tissue digestion protocols for the isolation of single endothelial cells and flow cytometry-based sorting on surface markers or transgene expression. These approaches are limited in the analysis of the endothelium in human brain tissues, where fresh samples are difficult to obtain. Here, we developed an approach to examine endothelial RNA expression by using an endothelial-specific marker to isolate nuclei from abundant archived frozen brain tissues. We show that this approach rapidly and reliably extracts endothelial nuclei from frozen mouse brain samples, and importantly, from archived frozen human brain tissues. Furthermore, isolated RNA transcript levels are closely correlated with expression in whole cells from tissue digestion protocols and are enriched in endothelial markers and depleted of markers of other brain cell types. As high-quality RNA transcripts could be obtained from as few as 100 nuclei in archived frozen human brain tissues, we predict that this approach should be useful for both bulk analysis of endothelial RNA transcripts in human brain tissues as well as single-cell analysis of endothelial sub-populations.Understanding of endothelial functions would be accelerated by methods for the specific isolation of these cells from archived human specimens. Here, the authors use the endothelial transcription factor Erg to isolate nuclei from mouse and human tissues, paving the way for high-throughput characterization of the function of endothelium in homeostasis and disease.

First-line nivolumab + ipilimumab in advanced NSCLC: CheckMate 227 subpopulation analyses in Asian patients

BackgroundNivolumab plus ipilimumab demonstrated clinically meaningful improvement in efficacy versus chemotherapy with a manageable safety profile in patients with advanced non-small cell lung cancer (NSCLC) and tumor programmed death-ligand 1 (PD-L1) expression ≥1% or <1% in Part 1 of CheckMate 227. Here we report efficacy and safety results for the Asian subpopulation.MethodsPatients with stage IV/recurrent NSCLC were randomized 1 : 1 : 1 to nivolumab plus ipilimumab, nivolumab monotherapy, or chemotherapy (PD-L1 ≥1%) or nivolumab plus ipilimumab, nivolumab plus chemotherapy, or chemotherapy (PD-L1 <1%). Overall survival (OS), progression-free survival, objective response rate, duration of response, and safety were evaluated among patients in Japan, South Korea, and Taiwan.ResultsIn the Asian subpopulation with PD-L1 ≥1%, 81 patients received nivolumab plus ipilimumab and 81 received chemotherapy. Median OS was not reached with nivolumab plus ipilimumab versus 24.8 months with chemotherapy; 3-year OS rate was 53% versus 37% [hazard ratio (HR), 0.72; 95% confidence interval (CI) 0.47-1.11]. The 3-year progression-free survival rate was 26% versus 7% (HR, 0.65; 95% CI 0.45-0.96), objective response rate was 56% versus 37%, and median duration of response was 29.0 months (95% CI 15.0 months-not reached) versus 6.9 months (95% CI 3.9-11.1 months). Similar results were observed regardless of tumor PD-L1 expression and in Japanese patients. Grade 3-4 treatment-related adverse events occurred in 40% of patients receiving nivolumab plus ipilimumab and 36% receiving chemotherapy, in the overall Asian subpopulation (tumor PD-L1 expression ≥1% and <1%); no new safety signals were identified.ConclusionsAt 3-year follow-up, nivolumab plus ipilimumab provided durable long-term efficacy benefits versus chemotherapy regardless of tumor PD-L1 expression in the Asian subpopulation, including Japanese patients. Consistent with findings for all randomized patients, these data support the use of nivolumab plus ipilimumab as first-line treatment of Asian patients with advanced NSCLC.

Bridging atomistic simulations and thermodynamic hydration models of aqueous electrolyte solutions.

Chemical thermodynamic models of solvent and solute activities predict the equilibrium behavior of aqueous solutions. However, these models are semi-empirical. They represent micro-scale ion and solvent behaviors controlling the macroscopic properties using small numbers of parameters whose values are obtained by fitting to activities and other partial derivatives of the Gibbs energy measured for the bulk solutions. We have conducted atomistic simulations of aqueous electrolyte solutions (MgCl2 and CaCl2) to determine the parameters of thermodynamic hydration models. We have implemented a cooperative hydration model to categorize the water molecules in electrolyte solutions into different subpopulations. The value of the electrolyte-specific parameter, k, was determined from the ion-affected subpopulation with the lowest absolute value of the free energy of removing the water molecule. The other equilibrium constant parameter, K1, associated with the first degree of hydration, was computed from the free energy of hydration of hydrated clusters. The hydration number, h, was determined from a reorientation dynamic analysis of the water subpopulations compared to bulk-like behavior. The reparameterized models [R. H. Stokes and R. H. Robinson, J. Solution Chem. 2, 173 (1973) and Balomenos et al., Fluid Phase Equilib. 243, 29 (2006)] using the computed values of the parameters lead to the osmotic coefficients of MgCl2 solutions that are consistent with measurements. Such an approach removes the dependence on the availability of experimental data and could lead to aqueous thermodynamic models capable of estimating the values of solute and solvent activities as well as thermal and volumetric properties for a wide range of compositions and concentrations.

Enriched CD45RA-CD62L+ central memory T and decreased CD3+CD56+ natural killer T lymphocyte subsets in the rectum of ulcerative colitis patients.

ObjectivesTo investigate the distinctive features of lymphocytes promoting inflammation in ulcerative colitis.MethodsWe performed flow cytometric analysis of peripheral blood mononuclear cells (PBMCs) and colorectal mucosa lymphocytes in ulcerative colitis patients (n = 13) and control patients (n = 5).ResultsCD62L+/CD3+CD4+ (35.7 ± 14.0% vs. 19.9 ± 6.4%) and CD62L+/CD3+CD4− cells (17.1 ± 17.4% vs. 2.4 ± 3.9%) were higher in the rectum of ulcerative colitis patients than in control patients. Subpopulation analysis revealed that CD45RA−CD62L+/CD3+CD4+, that is, central memory T cell fraction in CD4+ T cells, was significantly increased in the rectum of ulcerative colitis, compared to that in control patients (23.3 ± 10.5% vs. 8.2 ± 4.0%). Comparison of rectum and colon samples in ulcerative colitis patients indicated that CD56+/CD3+ was decreased in the rectum compared to that in the colon (11.3 ± 12.5% vs. 21.3 ± 16.5%). The ratio of CD56+/CD3+ was also decreased in the rectum of active ulcerative colitis patients compared to that in ulcerative colitis patients at the endoscopic remission stages (2.8 ± 1.7% vs. 18.5 ± 13.3%).ConclusionWe demonstrated that CD62L+ T lymphocytes, particularly the CD45RA−CD62L+ T cell subset that represents central memory T cells, were increased in the rectum of patients with ulcerative colitis. In addition, the CD56+/CD3+ subset (natural killer T cells) was decreased in the rectum compared to that of less inflamed colonic mucosa. These results suggest that the enrichment of central memory T lymphocytes and the reduction of natural killer T cells in the gut mucosa are involved in the pathogenesis of ulcerative colitis.

Changes in glycemic variability, gastric emptying and vascular endothelial function after switching from twice-daily to once-weekly exenatide in patients with type 2 diabetes: a subpopulation analysis of the twin-exenatide study

BackgroundWe investigated the changes in blood glucose fluctuation, gastric emptying, and vascular endothelial function by switching from an exenatide twice-daily formulation (BID) to a once-weekly formulation (QW) since the evaluation of postprandial glucose excursion and glycemic variability (GV) by continuous glucose monitoring (CGM) after switching was lacking.MethodsTwenty-nine patients with type 2 diabetes treated with exenatide BID were included in this study and switched to exenatide QW for 24 weeks. GV assessed by CGM, gastric emptying (by 13 C-acetate breath test) and vascular endothelial function (by reactive hyperemia - peripheral arterial tonometry) were evaluated at baseline and 24 weeks after switching.ResultsHbA1c decreased significantly from the baseline to week 24, while postprandial glucose levels after breakfast and dinner significantly increased (both P <0.05). However, the increases in GV indices were modest and not statistically significant at week 24. Vascular endothelial function was also not significantly changed after switching (P >0.05). Gastric emptying was significantly accelerated at week 24 (Tmax 83.4 ± 12.1 min vs. 58.2 ± 16.4 min) (P <0.001) and correlated with increased postprandial glucose levels after breakfast and dinner (both P <0.05).ConclusionsDespite the increase in postprandial glucose associated with accelerated gastric emptying after switching from exenatide BID to QW, change in GV was modest and no significant deterioration in vascular endothelial function was observed after switching. These results support the superiority of treatment with exenatide QW over exenatide BID in clinical practice; however, attention should be paid to the monitoring and management of postprandial glucose levels when selecting exenatide QW.Trial registrationClinical trial registry number; UMIN000016390 and jRCTs031180320.Approval date of Registry and the Registration: December 12, 2014.

3211 – MULTIOMIC ANALYSIS OF BONE MARROW HEMATOPOIETIC STEM/PROGENITOR CELLS REVEALS A POTENTIAL EARLY STAGE OF MALIGNANCY OF CLONAL HEMATOPOIESIS OF INDETERMINATE POTENTIAL

Clonal hematopoiesis of indeterminate potential (CHIP) increases the risk to develop hematological cancers such as acute myeloid leukemia (AML) tenfold. Since AML has been shown to originate from a hematopoietic stem/progenitor cell (HSPC) we compared the transcriptional landscape of bone marrow HSPCs from a healthy donor and a CHIP carrier (mutations in DNMT3A, 1.1% VAF and TET2, 5.3% VAF) by performing single cell RNA and ATAC sequencing. Cell frequency analysis of HSCP subpopulations, defined by analysis of conserved genes in clusters after dimensional reduction, showed that the proportion of HSC was twofold higher in the CHIP donor whereas the lymphocyte progenitor and pre-B cells compartment was fourfold lower, indicating an overall myeloid bias in the CHIP donor. Gene Ontology analysis of differentially expressed genes in the HSC compartment showed an upregulation of genes involved in metabolic processes in CHIP HSCs, among which genes that are associated with the onset of AML such as MEIS1 and PBX1. Furthermore, the expression levels of genes that are involved in niche retention, such as CD44 and VLA-4 were significantly higher in CHIP HSCs, suggesting alterations in the architecture of CHIP bone marrow. In addition to differences in gene expression, chromatin accessibility analysis showed unique genomic peaks in the CHIP HSC. Analysis of these sequences for binding motifs of transcription factors showed an overabundance of GATA, CEBP and RUNX1 motifs, all believed to play major roles in the onset of AML. The transcriptional landscapes of HSPC from healthy and CHIP donors show distinct differences, indicating that even the absence of signs of hematological disease, changes associated with a possible onset of malignancy can already be observed and provide possible targets for early intervention. Clonal hematopoiesis of indeterminate potential (CHIP) increases the risk to develop hematological cancers such as acute myeloid leukemia (AML) tenfold. Since AML has been shown to originate from a hematopoietic stem/progenitor cell (HSPC) we compared the transcriptional landscape of bone marrow HSPCs from a healthy donor and a CHIP carrier (mutations in DNMT3A, 1.1% VAF and TET2, 5.3% VAF) by performing single cell RNA and ATAC sequencing. Cell frequency analysis of HSCP subpopulations, defined by analysis of conserved genes in clusters after dimensional reduction, showed that the proportion of HSC was twofold higher in the CHIP donor whereas the lymphocyte progenitor and pre-B cells compartment was fourfold lower, indicating an overall myeloid bias in the CHIP donor. Gene Ontology analysis of differentially expressed genes in the HSC compartment showed an upregulation of genes involved in metabolic processes in CHIP HSCs, among which genes that are associated with the onset of AML such as MEIS1 and PBX1. Furthermore, the expression levels of genes that are involved in niche retention, such as CD44 and VLA-4 were significantly higher in CHIP HSCs, suggesting alterations in the architecture of CHIP bone marrow. In addition to differences in gene expression, chromatin accessibility analysis showed unique genomic peaks in the CHIP HSC. Analysis of these sequences for binding motifs of transcription factors showed an overabundance of GATA, CEBP and RUNX1 motifs, all believed to play major roles in the onset of AML. The transcriptional landscapes of HSPC from healthy and CHIP donors show distinct differences, indicating that even the absence of signs of hematological disease, changes associated with a possible onset of malignancy can already be observed and provide possible targets for early intervention.

Efficient exosome subpopulation isolation and proteomic profiling using a Sub-ExoProfile chip towards cancer diagnosis and treatment.

Exosomes have been extensively studied as liquid biopsy biomarkers in the past decade. However, the origin and molecular heterogeneity of exosomes hinder the research development moving from proof-of-concept to clinical applications. Herein, we report an integrated microfluidic platform termed Sub-ExoProfile chip, to achieve the selective isolation and subsequent proteomic profiling of multiplex exosome subpopulations simultaneously. The Sub-ExoProfile chip comprises three cylindrical self-assembled nanopillars, on which specific exosome capture antibodies (CD81, EpCAM, HER2) were immobilized to capture and sort different exosome subpopulations. It is worth noting that the 3D porous nanopillars afford enhanced interface binding efficiency; thus, a tumor-specific exosome subpopulation with lowly-expressed surface marker was still isolated with satisfactory capture efficiency. Moreover, the amphiphilic mesoporous silica nanoparticle self-assembled nanopillars also served as a nanoreactor for the enrichment and in situ digestion of exosomal proteins, providing improved performance for the mass-spectrometry based molecular characterization of exosome subpopulations. The Sub-ExoProfile chip was investigated on standard exosome samples from different breast cancer cell lines. The isolation and quantitative detection of exosome subpopulations were in line with the molecular subtype of breast cancer cell lines, and the molecular makeup was confirmed using classic microplate ELISA. Clinical samples from HER2-positive and triple-negative breast cancer patients were also examined using the Sub-ExoProfile chip. The quantitative results of three exosome subpopulations distinguished the three subtypes of breast cancer significantly. Most importantly, the molecular characterization of three exosome subpopulations revealed that the distinct exosome subpopulation participated in a different signaling pathway and performed distinct biological functions. It is envisioned that the analysis of the exosome subpopulation on the Sub-ExoProfile chip will facilitate the screening of tumor-specific exosomal biomarkers and open a new avenue for exosome-based liquid biopsy.

Coordinate control of basal epithelial cell fate and stem cell maintenance by core EMT transcription factor Zeb1.

Maintenance of undifferentiated, long-lived, and often quiescent stem cells in the basal compartment is important for homeostasis and regeneration of multiple epithelial tissues, but the molecular mechanisms that coordinately control basal cell fate and stem cell quiescence are elusive. Here, we report an epithelium-intrinsic requirement for Zeb1, a core transcriptional inducer of epithelial-to-mesenchymal transition, for mammary epithelial ductal side branching and for basal cell regenerative capacity. Our findings uncover an evolutionarily conserved role of Zeb1 in promoting basal cell fate over luminal differentiation. We show that Zeb1 loss results in increased basal cell proliferation at the expense of quiescence and self-renewal. Moreover, Zeb1 cooperates with YAP to activate Axin2 expression, and inhibition of Wnt signaling partially restores stem cell function to Zeb1-deficient basal cells. Thus, Zeb1 is a transcriptional regulator that maintains both basal cell fate and stem cell quiescence, and it functions in part through suppressing Wnt signaling.