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Intrinsic breast cancer molecular subtyping (IBCMS) provides significant prognostic information for patients with breast cancer and helps determine treatment. This study compared IBCMS methods on various gene-expression platforms in PALOMA-2 and PALLET trials. PALOMA-2 tumor samples were profiled using EdgeSeq and nanostring and subtyped with AIMS, PAM50, and research-use-only (ruo)Prosigna. PALLET tumor biopsies were profiled using mRNA sequencing and subtyped with AIMS and PAM50. In PALOMA-2 (n = 222), a 54% agreement was observed between results from AIMS and gold-standard ruoProsigna, with AIMS assigning 67% basal-like to HER2-enriched. In PALLET (n = 224), a 69% agreement was observed between results from PAM50 and AIMS. Different IBCMS methods may lead to different results and could misguide treatment selection; hence, a standardized clinical PAM50 assay and computational approach should be used.Trial number: NCT01740427.

Abstract Intrinsic breast cancer molecular subtyping (IBCMS) provides significant prognostic information for patients with breast cancer and helps determine treatment. This study compared IBCMS methods on various gene-expression platforms in PALOMA-2 and PALLET trials. PALOMA-2 tumor samples were profiled using EdgeSeq and nanostring and subtyped with AIMS, PAM50, and research-use-only (ruo)Prosigna. PALLET tumor biopsies were profiled using mRNA sequencing and subtyped with AIMS and PAM50. In PALOMA-2 (n = 222), a 54% agreement was observed between results from AIMS and gold-standard ruoProsigna, with AIMS assigning 67% basal-like to HER2-enriched. In PALLET (n = 224), a 69% agreement was observed between results from PAM50 and AIMS. Different IBCMS methods may lead to different results and could misguide treatment selection; hence, a standardized clinical PAM50 assay and computational approach should be used. Trial number: NCT01740427

AbstractHyperthyroidism is considered the most common endocrinopathy in middle-aged and old cats. The increased level of thyroid hormones influences many organs, including the heart. Cardiac functional and structural abnormalities in cats with hyperthyroidism have indeed been previously described. Nonetheless, myocardial vasculature has not been subjected to analysis. Also, no comparison with hypertrophic cardiomyopathy has been previously described. Although it has been shown that clinical alterations resolve after the treatment of hyperthyroidism, no detailed data have been published on the cardiac pathological or histopathological image of field cases of hyperthyroid cats that received pharmacological treatment. The aim of this study was to evaluate the cardiac pathological changes in feline hyperthyroidism and to compare them to alterations present in cardiac hypertrophy due to hypertrophic cardiomyopathy in cats. The study was conducted on 40 feline hearts divided into three groups: 17 hearts from cats suffering from hyperthyroidism, 13 hearts from cats suffering from idiopathic hypertrophic cardiomyopathy and 10 hearts from cats without cardiac or thyroid disease. A detailed pathological and histopathological examination was performed. Cats with hyperthyroidism showed no ventricular wall hypertrophy in contrast to cats with hypertrophic cardiomyopathy. Nonetheless, histological alterations were similarly advanced in both diseases. Moreover, in hyperthyroid cats more prominent vascular alterations were noted. In contrast to hypertrophic cardiomyopathy, the histological changes in hyperthyroid cats involved all ventricular walls and not mainly the left ventricle. Our study showed that despite normal cardiac wall thickness, cats with hyperthyroidism show severe structural changes in the myocardium.

Mild-to-moderate ulcerative colitis (UC) management is centred on 5-aminosalicylic acid (5-ASA) derivatives. Whether supplementing 5-ASA with nutraceuticals can provide real advantages in UC-relevant outcomes is unclear. This retrospective multicentre study compared clinical remission, response rates, and faecal calprotectin levels in a two-arm design, including patients treated with 5-ASA alone and those with additional H. erinaceus-based multi-compound supplementation. In the 5-ASA alone group, clinical response rates were 41% at three months (T1) and 60.2% at six months (T2), while corresponding clinical remission rates were 16.9% and 36.1%. In the nutraceutical supplementation group, clinical response rates were 49.6% (T1) and 70.4% (T2), with clinical remission rates of 30.4% (T1) and 50.9% (T2). No significant differences in clinical response rates between the groups at T1 (p = 0.231) and T2 (p = 0.143) emerged. Clinical remission rates differed significantly at both time points (p = 0.029 and p = 0.042, respectively). Faecal calprotectin levels decreased significantly in both groups during the retrospective follow-up (p < 0.05), and this was more pronounced in nutraceutical supplementation patients at both T1 (p = 0.005) and T2 (p = 0.01). No adverse events were reported. This multi-component nutraceutical supplementation offers real-world potential in controlling disease activity in patients with mild-to-moderate UC.

The bacterial amyloid curli, produced by Enterobacteriales including Salmonella species and Escherichia coli, is implicated in the pathogenesis of several complex autoimmune diseases. Curli binds to extracellular DNA, and these complexes drive autoimmunity via production of anti-double-stranded DNA autoantibodies. Here, we investigated immune activation by phenol-soluble modulins (PSMs), the amyloid proteins expressed by Staphylococcus species. We confirmed the amyloid nature of PSMs expressed by S. aureus using a novel specific amyloid stain, (E,E)-1-fluoro-2,5-bis(3-hydroxycarbonyl-4-hydroxy) styrylbenzene (FSB). Direct interaction of one of the S. aureus PSMs, PSMα3, with oligonucleotides promotes fibrillization of PSM amyloids and complex formation with bacterial DNA. Finally, utilizing a mouse model with an implanted mesh-associated S. aureus biofilm, we demonstrated that exposure to S. aureus biofilms for six weeks caused anti-double-stranded DNA autoantibody production in a PSM-dependent manner. Taken together, these results highlight how the presence of PSM-DNA complexes in S. aureus biofilms can induce autoimmune responses, and suggest an explanation for how bacterial infections trigger autoimmunity.

Dietary interventions represent an interesting alternative to pharmacological treatments for improving the quality of life (QoL) of subjects suffering from gastroesophageal reflux disease (GERD). This randomized, double-blind, placebo-controlled study aimed to evaluate the efficacy of a food supplement (FS) containing a probiotic strain, bioactive peptides, and vitamins in relieving heartburn/dyspeptic symptoms in subjects with mild-to-moderate GERD. Fifty-six adult participants were randomly assigned to receive the placebo or the active FS for 28 days. Subjects were asked to record daily the frequency and intensity of heartburn episodes and the intake of over- the-counter (OTC) medications. GERD-QoL and self-assessment questionnaires were also completed every two weeks and at the end of the treatment, respectively. FS was effective in achieving a progressive and significant reduction of heartburn frequency and severity, with an intergroup significant difference at the end of the treatment period. FS group also reported a reduction in the OTC medication intake, whereas placebo administration did not modify the OTC intake. Results from the QoL and self-assessment questionnaires showed that FS administration achieved a progressive and statistically significant intragroup and intergroup improvement in the QoL score and a higher positive response with respect to the placebo treatment.

Technology companies across the country receive requests from law enforcement agencies for cell phone location information near the scenes of crimes. These requests rely on the traditional warrant process and are known as geofence warrants, or reverse location search warrants. By obtaining location information, law enforcement can identify potential suspects or persons of interest who were near the scene of a crime when they have no leads. But the use of this investigative technique is controversial, as it threatens to intrude upon the privacy of innocent bystanders who had the misfortune of being nearby when the crime took place. Innocent bystanders are swept up in a geofence warrant because the warrant seeks information about all devices within a certain area at a certain time, instead of a more traditional search warrant, which focuses on a specific individual. The Washington Constitution provides heightened individual privacy protections compared to what the Constitution of the United States affords. As a result, some law enforcement techniques may be allowable under the federal constitution but forbidden within Washington under the Washington Constitution. This Comment considers whether geofence warrants are compatible with the Washington Constitution and recommends a framework for courts in Washington to adopt when reviewing a geofence warrant request. Alternatively, this Comment calls on the Washington State Legislature to regulate the use of geofence warrants., INTRODUCTION The familiar riddle asks, 'If a tree falls in a forest and no one is around to hear it, does it make a sound?' (1) Some have suggested the [...]

ObjectiveBiosimilars represent a new opportunity for inflammatory bowel disease (IBD) treatment and economic sustainability of therapies. This study aimed to evaluate the efficacy and long-term safety of the adalimumab biosimilar ABP 501 in biologic-naïve vs. biologic-switched IBD patients.MethodsA retrospective observational study was conducted using a database of patients with IBD treated with ABP 501, biologic-naïve or switched from the original, at eight IBD centers. We included adult patients with at least one year of follow-up. The primary objective of this study was to assess the efficacy (persistence) and safety (adverse event rate) of ABP 501 therapy.ResultsA total of 118 patients with IBD were included in the analysis: 84 patients with Crohn’s disease (CD) (39 women, 45 men, mean age 40.4 ± 14.3 years; 33% biologic-naïve) and 34 patients with ulcerative Colitis (UC) (16 women, 18 men, mean age 38.9 ± 14.9 years; 61.8% biologic-naïve). Regarding the primary endpoint, no difference was observed in the efficacy between biologic-naïve patients and patients with Adalimumab (ADA) originator replacement for non-medical reasons in terms of long-term persistence. However, ABP 501 showed a higher percentage of sustained clinical remission at 2 years in patients with CD (64 patients, 77%) than in those with UC (15 patients, 45.5%; p=0.00091). Nine patients (six with CD and three with UC) experienced adverse events that led to drug discontinuation in three.ConclusionsAPB 501 showed a good safety and efficacy profile in maintaining clinical response at 2 years in patients with IBD, both as a treatment-naïve and as a replacement for ADA originator for non-medical reasons.

Measurement(s) expression profiling Technology Type(s) cytometry time of flight assay Sample Characteristic - Organism Homo sapiens Sample Characteristic - Location Australia

Here, Parzych et al describe the development of a DNA-delivered SARS-CoV-2 mAb cocktail displaying synergistic RBD binding that confers broad neutralizing activity against variants of concern and prophylactic efficacy in multiple small animal models.

Glioblastoma is an aggressive tumor with poor survival rates. Bispecific T cell engagers (BTEs) against different cancers are in various stages of clinical development. Toxicity resulting from cytokine release syndrome and the short half-life of BTEs, which necessitates continuous infusion, complicating delivery and increasing costs, are major challenges in the field. Here we describe the development of in vivo DNA-launched BTEs (dBTEs) with highly focused targeting of interleukin-13 receptor α2 (IL-13Rα2), a glioblastoma cell-surface target. We developed 4 BTEs targeting 2 epitopes of IL-13Rα2 and studied how heavy-light chain orientation affects BTE function. The dBTEs induced T cell activation, cytokine production, and tumor cytolysis in the presence of IL-13Rα2+ tumor cells, but we observed unique patterns of immune activation. We found a strong correlation between granzyme B secretion and dBTE-induced cytolysis of specific and nonspecific tumors. We down-selected dBTE PB01-forward based on lower cytokine induction profile and highest activation specificity. In vivo, dBTE PB01-forward demonstrated an improved half-life versus intravenous recombinant BTE delivery. In an orthotopic glioblastoma model, dBTE PB01-forward controlled tumor growth, improving animal survival, supporting the hypothesis that the blood-brain barrier does not affect the function of systemically delivered dBTE. Further study of PB01-forward for targeting glioblastoma and other IL-13Rα2+ cancers is warranted.

Abstract Novel approaches for malaria prophylaxis remain important. Synthetic DNA-encoded monoclonal antibodies (DMAbs) are a promising approach to generate rapid, direct in vivo host-generated mAbs with potential benefits in production simplicity and distribution coupled with genetic engineering. Here, we explore this approach in a malaria challenge model. We engineered germline-reverted DMAbs based on human mAb clones CIS43, 317, and L9 which target a junctional epitope, major repeat, and minor repeat of the Plasmodium falciparum circumsporozoite protein (CSP) respectively. DMAb variants were encoded into a plasmid vector backbone and their expression and binding profiles were characterized. We demonstrate long-term serological expression of DMAb constructs resulting in in vivo efficacy of CIS43 GL and 317 GL in a rigorous mosquito bite mouse challenge model. Additionally, we engineered an Fc modified variant of CIS43 and L9-based DMAbs to ablate binding to C1q to test the impact of complement-dependent Fc function on challenge outcomes. Complement knockout variant DMAbs demonstrated similar protection to that of WT Fc DMAbs supporting the notion that direct binding to the parasite is sufficient for the protection observed. Further investigation of DMAbs for malaria prophylaxis appears of importance.

The essay reproduces the speech given at the Conference on "Burden of Proof and Fact Finding in the New Civil Trial," organized at Università Cattolica del Sacro Cuore in Milan on April 20, 2023. The analysis focuses on the topic of so-called defensive employer’s controls: a notion of jurisprudential elaboration aimed at subtracting - at least in part - covert controls carried out to protect 4company's assets, from the limiting discipline of "remote" controls in Article 4 of the "Workers' Statute." The brief essay is divided in two parts: the first devoted to focusing on some critical profiles of the theory of defensive controls; the second devoted to some essential evidentiary and procedural aspects.

The mechanisms by which the gut luminal environment is disturbed by the immune system to foster pathogenic bacterial growth and survival remain incompletely understood. Here, we show that STAT2 dependent type I IFN signaling contributes to the inflammatory environment by disrupting hypoxia enabling the pathogenic S. Typhimurium to outgrow the microbiota. Stat2-/- mice infected with S. Typhimurium exhibited impaired type I IFN induced transcriptional responses in cecal tissue and reduced bacterial burden in the intestinal lumen compared to infected wild-type mice. Although inflammatory pathology was similar between wild-type and Stat2-/- mice, we observed decreased hypoxia in the gut tissue of Stat2-/- mice. Neutrophil numbers were similar in wild-type and Stat2-/- mice, yet Stat2-/- mice showed reduced levels of myeloperoxidase activity. In vitro, the neutrophils from Stat2-/- mice produced lower levels of superoxide anion upon stimulation with the bacterial ligand N-formylmethionyl-leucyl-phenylalanine (fMLP) in the presence of IFNα compared to neutrophils from wild-type mice, indicating that the neutrophils were less functional in Stat2-/- mice. Cytochrome bd-II oxidase-mediated respiration enhances S. Typhimurium fitness in wild-type mice, while in Stat2-/- deficiency, this respiratory pathway did not provide a fitness advantage. Furthermore, luminal expansion of S. Typhimurium in wild-type mice was blunted in Stat2-/- mice. Compared to wild-type mice which exhibited a significant perturbation in Bacteroidetes abundance, Stat2-/- mice exhibited significantly less perturbation and higher levels of Bacteroidetes upon S. Typhimurium infection. Our results highlight STAT2 dependent type I IFN mediated inflammation in the gut as a novel mechanism promoting luminal expansion of S. Typhimurium.

Despite numerous clinically available vaccines and therapeutics, aged patients remain at increased risk for COVID-19 morbidity. Furthermore, various patient populations, including the aged can have suboptimal responses to SARS-CoV-2 vaccine antigens. Here, we characterized vaccine-induced responses to SARS-CoV-2 synthetic DNA vaccine antigens in aged mice. Aged mice exhibited altered cellular responses, including decreased IFNγ secretion and increased TNFα and IL-4 secretion suggestive of TH2-skewed responses. Aged mice exhibited decreased total binding and neutralizing antibodies in their serum but significantly increased TH2-type antigen-specific IgG1 antibody compared to their young counterparts. Strategies to enhance vaccine-induced immune responses are important, especially in aged patient populations. We observed that co-immunization with plasmid-encoded adenosine deaminase (pADA)enhanced immune responses in young animals. Ageing is associated with decreases in ADA function and expression. Here, we report that co-immunization with pADA enhanced IFNγ secretion while decreasing TNFα and IL-4 secretion. pADA expanded the breadth and affinity SARS-CoV-2 spike-specific antibodies while supporting TH1-type humoral responses in aged mice. scRNAseq analysis of aged lymph nodes revealed that pADA co-immunization supported a TH1 gene profile and decreased FoxP3 gene expression. Upon challenge, pADA co-immunization decreased viral loads in aged mice. These data support the use of mice as a model for age-associated decreased vaccine immunogenicity and infection-mediated morbidity and mortality in the context of SARS-CoV-2 vaccines and provide support for the use of adenosine deaminase as a molecular adjuvant in immune-challenged populations.

Cancer immunotherapy has demonstrated great promise with several checkpoint inhibitors being approved as the first-line therapy for some types of cancer, and new engineered cytokines such as Neo2/15 now being evaluated in many studies. In this work, we designed antibody-cytokine chimera (ACC) scaffolding cytokine mimetics on a full-length tumor-specific antibody. We characterized the pharmacokinetic (PK) and pharmacodynamic (PD) properties of first-generation ACC TA99-Neo2/15, which synergized with DLnano-vaccines to suppress in vivo melanoma proliferation and induced significant systemic cytokine activation. A novel second-generation ACC TA99-HL2-KOA1, with retained IL-2Rβ/γ binding and attenuated but preserved IL-2Rα binding, induced lower systemic cytokine activation with non-inferior protection in murine tumor studies. Transcriptomic analyses demonstrated an upregulation of Type I interferon responsive genes, particularly ISG15, in dendritic cells, macrophages and monocytes following TA99-HL2-KOA1 treatment. Characterization of additional ACCs in combination with cancer vaccines will likely be an important area of research for treating melanoma and other types of cancer.