Particle formation in response to different protein formulations and containers: Insights from machine learning analysis of particle images.

Subvisible particle count is a biotherapeutics stability indicator widely used by pharmaceutical industries. A variety of stresses that biotherapeutics are exposed to during development can impact particle morphology. By classifying particle morphological differences, stresses that have been applied to monoclonal antibodies (mAbs) can be identified. This study aims to evaluate common biotherapeutic drug storage and shipment conditions that are known to impact protein aggregation. Two different studies were conducted to capture particle images using micro-flow imaging and to classify particles using a convolutional neural network. The first study evaluated particles produced in response to agitation, heat, and freeze-thaw stresses in one mAb formulated in five different formulations. The second study evaluated particles from two common drug containers, a high-density polyethylene bottle and a glass vial, in six mAbs exposed solely to agitation stress. An extension of this study was also conducted to evaluate the impact of sequential stress exposure compared to exposure to one stress alone, on particle morphology. Overall, the convolutional neural network was able to classify particles belonging to a particular formulation or container. These studies indicate that storage and shipping stresses can impact particle morphology according to formulation composition and mAb.

Automatic wild bird repellent system that is based on deep-learning-based wild bird detection and integrated with a laser rotation mechanism.

Wild bird repulsion is critical in agriculture because it helps avoid agricultural food losses and mitigates the risk of avian influenza. Wild birds transmit avian influenza in poultry farms and thus cause large economic losses. In this study, we developed an automatic wild bird repellent system that is based on deep-learning-based wild bird detection and integrated with a laser rotation mechanism. When a wild bird appears at a farm, the proposed system detects the bird's position in an image captured by its detection unit and then uses a laser beam to repel the bird. The wild bird detection model of the proposed system was optimized for detecting small pixel targets, and trained through a deep learning method by using wild bird images captured at different farms. Various wild bird repulsion experiments were conducted using the proposed system at an outdoor duck farm in Yunlin, Taiwan. The statistical test results of our experimental data indicated that the proposed automatic wild bird repellent system effectively reduced the number of wild birds in the farm. The experimental results indicated that the developed system effectively repelled wild birds, with a high repulsion rate of 40.3% each day.

Blinatumomab in Children with MRD-Positive B-Cell Precursor Acute Lymphoblastic Leukemia: A Report of 11 Cases.

Background/Objectives: Relapsed B-cell acute lymphoblastic leukemia (B-ALL) remains an unresolved matter of concern regarding adverse outcomes. This case study aimed to evaluate the effectiveness of blinatumomab, with or without door lymphocyte infusion (DLI), in treating measurable residual disease (MRD)-positive B-ALL. Methods: All patients who received blinatumomab salvage therapy were included in this study. Eleven patients were included in the study. All patients were evaluated for MRD-negativity. Results: Before starting blinatumomab therapy, seven patients tested positive for MRD, three tested negative, and one had refractory disease. Hematopoietic cell transplantation (HCT) was reserved for five patients with persistent MRD. Six patients became MRD-negative and subsequent HCT was not performed. Only two patients relapsed; one patient died of relapse, and the other one received carfilzomib-based therapy and was MRD-negative thereafter. Nine patients were MRD-negative at a median follow-up of 28 months (15-52 months). Two of three MRD-positive post-transplant patients remained in complete molecular remission after preemptive DLI at the last follow-up date. In the first salvage, blinatumomab may achieve complete remission and bridging to HCT in pediatric patients with end-of-induction MRD-positive B-cell precursor ALL. Conclusions: The decision on how to treat post-transplant relapse continues to affect survival outcomes. Blinatumomab combined with DLI may extend the armamentarium of release options for high-risk pediatric patients. This approach is encouraging for high-risk ALL patients who are MRD-positive post-transplantation.

Hydrogen-treated CoCrMo alloy: a novel approach to enhance biocompatibility and mitigate inflammation in orthopedic implants.

In recent decades, orthopedic implants have been widely used as materials to replace human bone tissue functions. Among these, metal implants play a crucial role. Metals with better chemical stability, such as stainless steel, titanium alloys, and cobalt-chromium-molybdenum (CoCrMo) alloy, are commonly used for long-term applications. However, good chemical stability can result in poor tissue integration between the tissue and the implant, leading to potential inflammation risks. This study creates hydrogenated CoCrMo (H-CoCrMo) surfaces, which have shown promise as anti-inflammatory orthopedic implants. Using the electrochemical cathodic hydrogen-charging method, the surface of the CoCrMo alloy was hydrogenated, resulting in improved biocompatibility, reduced free radicals, and an anti-inflammatory response. Hydrogen diffusion to a depth of approximately 106 ± 27 nm on the surface facilitated these effects. This hydrogen-rich surface demonstrated a reduction of 85.2% in free radicals, enhanced hydrophilicity as evidenced by a decrease in a contact angle from 83.5 ± 1.9° to 52.4 ± 2.2°, and an increase of 11.4% in hydroxyapatite deposition surface coverage. The cell study results revealed a suppression of osteosarcoma cell activity to 50.8 ± 2.9%. Finally, the in vivo test suggested the promotion of new bone formation and a reduced inflammatory response. These findings suggest that electrochemical hydrogen charging can effectively modify CoCrMo surfaces, offering a potential solution for improving orthopedic implant outcomes through anti-inflammatory mechanisms.

Neuroprotection effects of kynurenic acid-loaded micelles for the Parkinson's disease models.

Anti-glutamatergic agents may have neuroprotective effects against excitotoxicity that is known to be involved in the pathogenesis of Parkinson's disease (PD). One of these agents is kynurenic acid (KYNA), a tryptophan metabolite, which is an endogenous N-methyl-D-aspartic acid (NMDA) receptor antagonist. However, its pharmacological properties of poor water solubility and limited blood-brain barrier (BBB) permeability rules out its systemic administration in disorders affecting the central nervous system. Our aim in the present study was to investigate the neuroprotective effects of KYNA-loaded micelles (KYNA-MICs) against PD in vitro and in vivo. Lipid-based micelles (MICs) in conjunction with KYNA drug delivery have the potential to enhance the penetration of therapeutic drugs into a diseased brain without BBB obstacles. KYNA-MICs were characterized by particle size (105.8 ± 12.1 nm), loading efficiency (78.3 ± 4.23%), and in vitro drug release (approximately 30% at 24 h). The in vitro experiments showed that KYNA-MICs effectively reduced 2-fold protein aggregation. The in vivo studies revealed that KYNA was successfully delivered by 5-fold increase in neurotoxin-induced PD brains. The results showed significant enhancement of KYNA delivery into brain. We also found that the KYNA-MICs exhibited several therapeutic effects. The KYNA-MICs reduced protein aggregation of an in vitro PD model, ameliorated motor functions, and prevented loss of the striatal neurons in a PD animal model. The beneficial effects of KYNA-MICs are probably explained by the anti-excitotoxic activity of the treatment's complex. As the KYNA-MICs did not induce any appreciable side-effects at the protective dose applied to a chronic PD mouse model, our results demonstrate that KYNA provides neuroprotection and attenuates PD pathology.

Syndecan-4 is required for early-stage repair responses during zebrafish heart regeneration.

BACKGROUND: The healing process after a myocardial infarction (MI) in humans involves complex events that replace damaged tissue with a fibrotic scar. The affected cardiac tissue may lose its function permanently. In contrast, zebrafish display a remarkable capacity for scar-free heart regeneration. Previous studies have revealed that syndecan-4 (SDC4) regulates inflammatory response and fibroblast activity following cardiac injury in higher vertebrates. However, whether and how Sdc4 regulates heart regeneration in highly regenerative zebrafish remains unknown. METHODS AND RESULTS: This study showed that sdc4 expression was differentially regulated during zebrafish heart regeneration by transcriptional analysis. Specifically, sdc4 expression increased rapidly and transiently in the early regeneration phase upon ventricular cryoinjury. Moreover, the knockdown of sdc4 led to a significant reduction in extracellular matrix protein deposition, immune cell accumulation, and cell proliferation at the lesion site. The expression of tgfb1a and col1a1a, as well as the protein expression of Fibronectin, were all down-regulated under sdc4 knockdown. In addition, we verified that sdc4 expression was required for cardiac repair in zebrafish via in vivo electrocardiogram analysis. Loss of sdc4 expression caused an apparent pathological Q wave and ST elevation, which are signs of human MI patients. CONCLUSIONS: Our findings support that Sdc4 is required to mediate pleiotropic repair responses in the early stage of zebrafish heart regeneration.

The Impact of Gut Microbiota Changes on Methotrexate-Induced Neurotoxicity in Developing Young Rats.

Methotrexate (MTX) is an essential part of therapy in the treatment of acute lymphoblastic leukemia (ALL) in children, and inferior intellectual outcomes have been reported in children who are leukemia survivors. Although several studies have demonstrated that the interaction between gut microbiota changes and the brain plays a vital role in the pathogenesis of chemotherapy-induced brain injury, preexisting studies on the effect of MTX on gut microbiota changes focused on gastrointestinal toxicity only. Based on our previous studies, which revealed that MTX treatment resulted in inferior neurocognitive function in developing young rats, we built a young rat model mimicking MTX treatment in a child ALL protocol, trying to investigate the interactions between the gut and brain in response to MTX treatment. We found an association between gut microbiota changes and neurogenesis/repair processes in response to MTX treatment, which suggest that MTX treatment results in gut dysbiosis, which is considered to be related to MTX neurotoxicity through an alteration in gut-brain axis communication.

Refining risk stratification in paediatric B-acute lymphoblastic leukaemia: Combining IKZF1plus and Day 15 MRD positivity.

This study investigates the potential utility of IKZF1 deletion as an additional high-risk marker for paediatric acute lymphoblastic leukaemia (ALL). The prognostic impact of IKZF1 status, in conjunction with minimal/measurable residual disease (MRD), was evaluated within the MRD-guided TPOG-ALL-2013 protocol using 412 newly diagnosed B-ALL patients aged 1-18. IKZF1 status was determined using multiplex ligation-dependent probe amplification. IKZF1 deletions, when co-occurring with CDKN2A, CDKN2B, PAX5 or PAR1 region deletions in the absence of ERG deletions, were termed IKZF1plus. Both IKZF1 deletion (14.6%) and IKZF1plus (7.8%) independently predicted poorer outcomes in B-ALL. IKZF1plus was observed in 4.1% of Philadelphia-negative ALL, with a significantly lower 5-year event-free survival (53.9%) compared to IKZF1 deletion alone (83.8%) and wild-type IKZF1 (91.3%) (p < 0.0001). Among patients with Day 15 MRD ≥0.01%, provisional high-risk patients with IKZF1plus exhibited the worst outcomes in event-free survival (42.0%), relapse-free survival (48.0%) and overall survival (72.7%) compared to other groups (p < 0.0001). Integration of IKZF1plus and positive Day 15 MRD identified a subgroup of Philadelphia-negative B-ALL with a 50% risk of relapse. This study highlights the importance of assessing IKZF1plus alongside Day 15 MRD positivity to identify patients at increased risk of adverse outcomes, potentially minimizing overtreatment.

Impact of the implementation of the Intelligent Antimicrobial System (iAMS) on clinical outcomes among patients with bacteraemia caused by methicillin-resistant Staphylococcus aureus.

OBJECTIVES: This study aimed to investigate the clinical impact of the Intelligent Antimicrobial System (iAMS) on patients with bacteraemia due to methicillin-resistant (MRSA) and methicillin-susceptible Staphylococcus aureus (MSSA). METHODS: A total of 1008 patients with suspected SA infection were enrolled before and after the implementation of iAMS. Among them, 252 with bacteraemia caused by SA, including 118 in the iAMS and 134 in the non-iAMS groups, were evaluated. RESULTS: The iAMS group exhibited a 5.2% (from 55.2% to 50.0%; P = 0.96) increase in the 1-year survival rate. For patients with MRSA and MSSA compared to the non-iAMS group, the 1-year survival rate increased by 17.6% (from 70.9% to 53.3%; P = 0.41) and 7.0% (from 52.3% to 45.3%; P = 0.57), respectively, both surpassing the rate of the non-iAMS group. The iAMS intervention resulted in a higher long-term survival rate (from 70.9% to 52.3%; P = 0.984) for MRSA patients than for MSSA patients. MRSA patients experienced a reduced length of hospital stay (from 23.3% to 35.6%; P = 0.038), and the 45-day discharge rate increased by 20.4% (P = 0.064). Furthermore, the intervention resulted in a significant 97.3% relative decrease in near miss medication incidents reported by pharmacists (P = 0.013). CONCLUSIONS: Implementation of iAMS platform improved long-term survival rates, discharge rates, hospitalization days, and medical cost (although no significant differences were observed) among patients with MRSA bacteraemia. Additionally, it demonstrated significant benefits in ensuring drug safety.

Protocol for replacing coding intronic MiMIC and CRIMIC lines with T2A-split-GAL4 lines in Drosophila using genetic crosses.

Here, we present a protocol for generating gene-specific split-GAL4 drivers from coding intronic Minos-mediated integration cassette/CRISPR-mediated integration cassette (MiMIC/CRIMIC) lines in Drosophila. We describe steps for four rounds of in vivo genetic crosses, PCR genotyping, and fluorescence imaging to ensure correct orientation of split-GAL4 integration before establishing stable fly stocks. This protocol offers a cost-effective alternative to traditional microinjection techniques for converting coding intronic MiMIC/CRIMIC lines into gene-specific split-GAL4 lines that are adaptable for fly researchers working on different tissues. For complete details on the use and execution of this protocol, please refer to Chen et al.1.

Multipole engineering by displacement resonance: a new degree of freedom of Mie resonance.

The canonical studies on Mie scattering unravel strong electric/magnetic optical responses in nanostructures, laying foundation for emerging meta-photonic applications. Conventionally, the morphology-sensitive resonances hinge on the normalized frequency, i.e. particle size over wavelength, but non-paraxial incidence symmetry is overlooked. Here, through confocal reflection microscopy with a tight focus scanning over silicon nanostructures, the scattering point spread functions unveil distinctive spatial patterns featuring that linear scattering efficiency is maximal when the focus is misaligned. The underlying physical mechanism is the excitation of higher-order multipolar modes, not accessible by plane wave irradiation, via displacement resonance, which showcases a significant reduction of nonlinear response threshold, sign flip in all-optical switching, and spatial resolution enhancement. Our result fundamentally extends the century-old light scattering theory, and suggests new dimensions to tailor Mie resonances.

Optical Bistability in Nanosilicon with Record Low Q-Factor.

We demonstrated optical bistability in an amorphous silicon Mie resonator with a size of ∼100 nm and Q-factor as low as ∼4 by utilizing photothermal and thermo-optical effects. We not only experimentally confirmed the steep intensity transition and the hysteresis in the scattering response from silicon nanocuboids but also established a physical model to numerically explain the underlying mechanism based on temperature-dependent competition between photothermal heating and heat dissipation. The transition between the bistable states offered particularly steep superlinearity of scattering intensity, reaching an effective nonlinearity order of ∼100th power over excitation intensity, leading to the potential of advanced optical switching devices and super-resolution microscopy.

Fumarate hydratase functions as a tumor suppressor in endometrial cancer by inactivating EGFR signaling.

Fumarase hydratase (FH) is an enzyme that catalyzes the reversible hydration and dehydration of fumarate to malate in the tricarboxylic acid cycle. The present study addressed the role of FH in endometrial cancer and clinically observed that the expression of FH was significantly lower in endometrial cancer tissues compared with normal endometrial tissues and, furthermore, that the decreased FH expression in endometrial cancer tissues was significantly associated with increased tumor size and lymph node metastasis. Further analysis in in vitro study showed that cell proliferation, migration and invasion abilities were increased when the expression of FH in the endometrial cancer cells was knocked down, but, by contrast, overexpression of FH in endometrial cancer cells decreased cell proliferative, migratory and invasive abilities. Mechanistic studies showed that the expression of vimentin and twist, being two well-studied mesenchymal markers in endometrial cancer cells, were upregulated in fumarate hydratase-knockdowned cells. In addition, phosphokinase array analysis demonstrated that the expression of phospho-EGFR (Y1086), which promotes carcinogenesis in cancers, was increased in endometrial cancer cells when FH was knocked down. In conclusion, the present study suggested that FH is a tumor suppressor and inhibits endometrial cancer cell proliferation and metastasis by inactivation of EGFR. Further studies are required to clarify its role as a prognostic biomarker and therapeutic target for endometrial cancer.

Using single-cell RNA sequencing to generate predictive cell-type-specific split-GAL4 reagents throughout development.

Cell-type-specific tools facilitate the identification and functional characterization of the distinct cell types that form the complexity of neuronal circuits. A large collection of existing genetic tools in Drosophila relies on enhancer activity to label different subsets of cells and has been extremely useful in analyzing functional circuits in adults. However, these enhancer-based GAL4 lines often do not reflect the expression of nearby gene(s) as they only represent a small portion of the full gene regulatory elements. While genetic intersectional techniques such as the split-GAL4 system further improve cell-type-specificity, it requires significant time and resources to screen through combinations of enhancer expression patterns. Here, we use existing developmental single-cell RNA sequencing (scRNAseq) datasets to select gene pairs for split-GAL4 and provide a highly efficient and predictive pipeline (scMarco) to generate cell-type-specific split-GAL4 lines at any time during development, based on the native gene regulatory elements. These gene-specific split-GAL4 lines can be generated from a large collection of coding intronic MiMIC/CRIMIC lines or by CRISPR knock-in. We use the developing Drosophila visual system as a model to demonstrate the high predictive power of scRNAseq-guided gene-specific split-GAL4 lines in targeting known cell types, annotating clusters in scRNAseq datasets as well as in identifying novel cell types. Lastly, the gene-specific split-GAL4 lines are broadly applicable to any other Drosophila tissue. Our work opens new avenues for generating cell-type-specific tools for the targeted manipulation of distinct cell types throughout development and represents a valuable resource for the Drosophila community.

Sustaining the Activation of EGFR Signal by Inflammatory Cytokine IL17A Prompts Cell Proliferation and EGFR-TKI Resistance in Lung Cancer.

Non-small-cell lung cancer (NSCLC) is a typical inflammation-associated cancer, and lung adenocarcinoma (LUAD) is the most common pathological subtype. Epidermal growth factor (EGF) receptor (EGFR) mutations are the most common driver mutations of LUAD, and they have been identified as important therapeutic targets by EGFR-tyrosine kinase inhibitors (TKIs). The proinflammatory cytokine, interleukin (IL)-17A, and IL-17A-producing cells were reported to be elevated in the tumor microenvironment and peripheral blood of NSCLC patients and to be correlated with tumor progression and poor prognoses. However, the pathophysiological role of IL-17A in NSCLC remains unclear, although some studies suggested its involvement in cancer cell invasion and metastasis. Herein, we observed that expressions of IL-17A and its receptor, IL-17 receptor C (IL-17RC), were elevated in LUAD tissues and were correlated with poor survival in different lung cancer cohorts. In LUAD cells with mutant EGFR, the IL-17A/IL-17RC axis was shown to enhance phosphorylation of EGFR and Met, thereby promoting proliferation and resistance to EGFR-TKIs such as afatinib. In LUAD cells with wild-type (WT) EGFR, we found that the IL-17A/IL-17RC axis enhanced EGF-induced EGFR activation and cell proliferation through causing impairment of EGF-induced EGFR lysosomal degradation. Collectively, our results indicated diverse impacts of the IL-17A/IL-17RC axis on EGFR activation in LUAD cells with WT and mutant EGFR and suggested that developing therapeutic strategies against IL-17A/IL-17RC would be valuable for LUAD treatment.

One-Pot Double Oxidation Synthesis of N-1-Piperidonyl Amides From N-1-Piperidinyl Amides with meta-Chloroperbenzoic Acid: Rimonabant Analogue as Model Study.

A simple and efficient one-pot oxidation synthesis of N-1-piperidonyl amides was successfully developed through the double oxidation of hydrazides (involving hydrazonium formation, azodioxy-carbonyl compounds generation, and α-carbon oxidation) by using meta-chloroperbenzoic acid (mCPBA). The convenient oxidation method was also extended to Rimonabant analogue. The lactam oxidized Rimonabant analogue was first successfully synthesized for demonstrating the construction and characterized by NMR spectroscopic methods and the single-crystal X-ray diffraction study (ORTEP).

Increased Circulating ADMA in Young Male Rats Caused Cognitive Deficits and Increased Intestinal and Hippocampal NLRP3 Inflammasome Expression and Microbiota Composition Alterations: Effects of Resveratrol.

Endothelial dysfunction is characterized by disturbances in nitric oxide (NO) bioavailability and increased circulating asymmetric dimethylarginine (ADMA) due to the enormous release of free radicals. Increased circulating ADMA may cause endothelial dysfunction and a variety of clinical disorders, such as liver and kidney disease. Young male Sprague-Dawley rats at postnatal day 17 ± 1 received continuous ADMA infusion via an intraperitoneal pump to induce endothelial dysfunction. Four groups of rats (n = 10 per group) were allocated: control, control and resveratrol, ADMA infusion, and ADMA infusion and resveratrol groups. Spatial memory, NLR family pyrin-domain-containing 3 (NLRP3) inflammasome, cytokine expression, tight junction proteins in the ileum and dorsal hippocampus, and microbiota composition were examined. We found cognitive deficits; increased NLRP3 inflammasome in the plasma, ileum, and dorsal hippocampus; decreased ileum and dorsal hippocampal cytokine activation and tight junction proteins; and microbiota composition alterations in the ADMA-infusion young male rats. Resveratrol had beneficial effects in this context. In conclusion, we observed NLRP3 inflammasome activation in peripheral and central dysbiosis in young male rats with increased circulating ADMA, and found that resveratrol had beneficial effects. Our work adds to the mounting evidence that inhibiting systemic inflammation is a promising therapeutic avenue for cognition impairment, probably via the gut-brain axis.

Melatonin protects against methotrexate hepatotoxicity in young rats: Impact of PI3K/Akt/mTOR signaling.

With the improvement in children's acute lymphoblastic leukemia (ALL) care, the survival rate in children ALL has improved much. Methotrexate (MTX) plays an essential role in the success of children's ALL treatment. Since hepatotoxicity is commonly reported in individuals treated with intravenous or oral MTX, our study further examined the hepatic effect following intrathecal MTX treatment, which is an essential treatment for leukemia patients. Specifically, we examined the pathogenesis of MTX hepatotoxicity in young rats and explored the impact of melatonin treatment in protection against MTX hepatotoxicity. Successfully, we found that melatonin was able to protect against MTX hepatotoxicity.

Quality of care in the course of subcutaneous versus intravenous trastuzumab administration in patients with breast cancer: an integrated time-motion study with mixed-methods research.

OBJECTIVES: This study aimed to compare the time required and concerns raised by various perspectives of participants regarding administering subcutaneous and intravenous trastuzumab for patients with breast cancer (BC). DESIGN: This observational time-motion study design with mixed-methods research (cross-sectional surveys and semistructured interviews) was conducted. The time spent on preparing or administering trastuzumab by different healthcare professionals (HCPs) was recorded. The data were analysed by descriptive/inferential statistical analyses, followed by thematic analyses. SETTING: Outpatient and inpatient administration units of a single medical centre in Taiwan. PARTICIPANTS: The study included patients with early-stage BC who received subcutaneous or intravenous trastuzumab (n=93), and HCPs including two attending physicians, a nurse practitioner, two pharmacists and two nurses. RESULT: Based on the perspectives of patients and HCPs, the subcutaneous form of trastuzumab was more efficient, less expensive and produced less discomfort in outpatient units than inpatient units. More participants preferred the subcutaneous form over the intravenous form in both outpatient and inpatient units. Pharmacists and nurse practitioners spent threefold more time on patients when preparing and administering the intravenous form in both outpatient and inpatient units. The concerns raised by patients and HCPs varied in certain aspects, including the injection skills, speed, mental distress (eg, needle phobia) and pain associated with the subcutaneous form. Almost all patients preferred receiving the subcutaneous form in outpatient units after the initial COVID-19 outbreak. CONCLUSION: Patients with early-stage BC preferred receiving subcutaneous trastuzumab in outpatient units rather than inpatient units or the intravenous form before and after the COVID-19 outbreak. Such findings may serve as real-world evidence to facilitate better quality of care regarding administration of subcutaneous or intravenous trastuzumab in medical settings, and its feasible resolutions to balance the quality, concerns and efficiency of anticancer administration during the COVID-19 pandemic.