Brianna Dalesandro

Methods Mol Biol. 2024;2727:205-214. doi: 10.1007/978-1-0716-3491-2_16.

ABSTRACT

The development of advanced microscopy techniques has ushered in a new era of research as it helps understand biological processes on a deeper, mechanistic, and molecular level like never before. Live-cell fluorescence microscopy has importantly allowed us to visualize subcellular protein localization and incorporation of various fluorophores compatible with living cells in real time. As such, this technique offers valuable insights at the single-cell level and enables us to monitor phenotypic differences that were easily overlooked at a population level. One area of research that has benefited greatly from these advances is the study of the bacterial cell envelope biogenesis and cell division process. In this report, we provide detailed protocols, optimized in our lab, for imaging these processes in the Gram-positive organisms Bacillus subtilis and Staphylococcus aureus.

PMID:37815719 | DOI:10.1007/978-1-0716-3491-2_16

Eur J Pharm Sci. 2023 Oct 6:106602. doi: 10.1016/j.ejps.2023.106602. Online ahead of print.

ABSTRACT

Staphylococcus aureus is an important pathogenic bacterium responsible for various organ infections. The serious side effects and the development of antibiotic resistance have rendered the antibiotic therapy against S. aureus increasingly challenging, emphasizing the pressing need for the exploration of novel therapeutic agents. Our research has uncovered the promising antimicrobial properties of 8-octyl berberine (OBBR), a novel compound derived from berberine (BBR), against S. aureus. OBBR exhibited a minimum inhibitory concentration (MIC) of 1.0 μg/mL, which closely approximated that of levofloxacin. Intriguingly, a multipassage resistance assay demonstrated that the MIC of OBBR against S. aureus remained relatively stable, while levofloxacin exhibited a 4-fold increase over 20 days, suggesting that OBBR was less prone to inducing resistance. Mechanistically, our investigation, employing Zeta potential measurements, flow cytometry, scanning electron microscopy, and transmission electron microscopy, unveiled that OBBR induced morphological alterations in the bacteria. Furthermore, it disrupted the bacterial cell wall and membrane by altering membrane potential and compromising membrane integrity. These actions culminated in bacterial disintegration and apoptosis. Transcriptomic analysis shed light on significant downregulation of gene ontology terms, predominantly associated with membranes. The Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis implicated OBBR in disturbing peptidoglycan biosynthesis, with the membrane protein MraY emerging as a potential target for OBBR's action against S. aureus. Notably, experiments involving the overexpression of MraY confirmed OBBR's inhibitory effect on peptidoglycan synthesis. Furthermore, molecular docking and cellular thermal shift assay revealed OBBR's direct interaction with MraY, potentially leading to the inhibition of the enzymatic activity of MraY and, consequently, impeding peptidoglycan synthesis. In summary, OBBR, by targeting MraY and inhibiting peptidoglycan synthesis, emerges as a promising alternative antibiotic against S. aureus, offering potential advantages in terms of limited drug resistance development.

PMID:37806408 | DOI:10.1016/j.ejps.2023.106602

Lancet Gastroenterol Hepatol. 2023 Oct 5:S2468-1253(23)00241-8. doi: 10.1016/S2468-1253(23)00241-8. Online ahead of print.

NO ABSTRACT

PMID:37806315 | DOI:10.1016/S2468-1253(23)00241-8

Lancet Gastroenterol Hepatol. 2023 Oct 5:S2468-1253(23)00198-X. doi: 10.1016/S2468-1253(23)00198-X. Online ahead of print.

ABSTRACT

BACKGROUND: HU6 is a controlled metabolic accelerator that is metabolised in the liver to the mitochondrial uncoupler 2,4-dinitrophenol and increases substrate utilisation so that fat and other carbon sources are oxidised in the body rather than accumulated. We aimed to assess the safety and efficacy of HU6 compared with placebo in people with non-alcoholic fatty liver disease (NAFLD) and high BMI.

METHODS: This randomised, double-blind, placebo-controlled, phase 2a trial was done at a single community site in the USA. Adults (aged 28-65 years) with a BMI of 28-45 kg/m², a FibroScan controlled attenuation parameter score of more than 270 decibels per metre, and at least 8% liver fat by MRI-proton density fat fraction (MRI-PDFF) were randomly assigned (1:1:1:1) to receive, under fasting conditions, either once-daily HU6 100 mg, HU6 300 mg, HU6 450 mg, or matching placebo by oral administration for 61 days. Randomisation was blocked (groups of four) and stratified by baseline glycated haemoglobin (<5·7% vs ≥5·7%; 39 mmol/mol). All participants and study personnel involved with outcome assessments were masked to treatment assignment. The primary endpoint was the relative change in liver fat content from baseline to day 61, as assessed by MRI-PDFF, and was analysed in the full analysis set (FAS), which comprised all participants who were randomly assigned, received at least one dose of treatment, and had less than 4·5 kg of weight gain or weight loss from the time of screening to day 1 of treatment. The safety population included all participants who were randomly assigned and received at least one dose of study drug. This study was registered at ClinicalTrials.gov, NCT04874233, and is complete.

FINDINGS: Between April 28, 2021, and Nov 29, 2021, 506 participants were assessed for eligibility and 80 adults (39 [49%] women and 41 [51%] men) were enrolled and randomly assigned to placebo (n=20), HU6 150 mg (n=20), HU6 300 mg (n=21), or HU6 450 mg (n=19). One participant in the HU6 450 mg group was excluded from the FAS due to weight gain. Relative mean change in liver fat content from baseline to day 61 was -26·8% (SD 17·4) for the HU6 150 mg group, -35·6% (13·8) for the HU6 300 mg group, -33·0% (18·4) for the HU6 450 mg group, and 5·4% (19·8) for the placebo group. Three people treated with HU6 (two treated with 150 mg and one treated with 300 mg) and two people treated with placebo discontinued treatment due to treatment-emergent adverse events (TEAEs). No serious TEAEs were reported. In those treated with HU6, flushing (19 [32%] participants), diarrhoea (15 [25%] participants), and palpitations (seven [12%] participants) were the most frequently reported TEAEs (in the placebo group, two [10%] participants had flushing, none had diarrhoea, and one [5%] had palpitations). There were no deaths.

INTERPRETATION: HU6 could be a promising pharmacological agent for treating patients with obesity and NAFLD and its metabolic complications.

FUNDING: Rivus Pharmaceuticals.

PMID:37806314 | DOI:10.1016/S2468-1253(23)00198-X

JAMA. 2023 Oct 5:e2319574. doi: 10.1001/jama.2023.19574. Online ahead of print.

NO ABSTRACT

PMID:37796527 | PMC:PMC10557026 | DOI:10.1001/jama.2023.19574

Chem Commun (Camb). 2023 Oct 6. doi: 10.1039/d3cc03583b. Online ahead of print.

ABSTRACT

The production of β-lactamases by bacterial pathogens endangers antimicrobial therapy, and new inhibitors for β-lactamases are urgently needed. We report the development of a luminescent-based biosensor that quantifies β-lactamase inhibition in a cellular context, based on the activation of transcriptional factor AmpR following the exposure of bacterial cells to β-lactams. This rapid method can account for factors like membrane permeability and can be employed to identify new β-lactamase inhibitors.

PMID:37801331 | DOI:10.1039/d3cc03583b

mBio. 2023 Oct 5:e0134923. doi: 10.1128/mbio.01349-23. Online ahead of print.

ABSTRACT

Staphylococcus aureus (S. aureus) represents a major clinical challenge due to its explicit capacity to select mutations that increase antibiotic resistance and immune evasion. However, the molecular mechanisms are poorly defined, especially for adaptive immunity. Cancer immunotherapy targeting programmed cell death protein 1 (PD-1) enhances T-cell activity and is emerging for the treatment of certain viral infections, while its potential against bacterial infections remains elusive. We show that an S. aureus clpP mutant, selected during clinical antibiotic therapy, inhibits T-cell activity by directly interacting with PD-1 on human T cells. Specificity of the interaction was confirmed using recombinant PD-1, as well as PD-1 overexpressing and knock out cells. Moreover, the PD-1-binding S. aureus inhibited intracellular calcium mobilization, T-cell proliferation, CD25 expression, and IL-2 secretion, while the key effects were alleviated by antibody-mediated PD-1 blockade using an engineered IgG1-based anti-PD-1 antibody. Our results suggest that clpP mutant S. aureus directly targets PD-1 to evade immune activation and that therapeutic targeting of PD-1 may be used against certain staphylococcal infections. IMPORTANCE Therapies that target and aid the host immune defense to repel cancer cells or invading pathogens are rapidly emerging. Antibiotic resistance is among the largest threats to human health globally. Staphylococcus aureus (S. aureus) is the most common bacterial infection, and it poses a challenge to the healthcare system due to its significant ability to develop resistance toward current available therapies. In long-term infections, S. aureus further adapt to avoid clearance by the host immune defense. In this study, we discover a new interaction that allows S. aureus to avoid elimination by the immune system, which likely supports its persistence in the host. Moreover, we find that blocking the specific receptor (PD-1) using antibodies significantly relieves the S. aureus-imposed inhibition. Our findings suggest that therapeutically targeting PD-1 is a possible future strategy for treating certain antibiotic-resistant staphylococcal infections.

PMID:37796131 | DOI:10.1128/mbio.01349-23

Med Microbiol Immunol. 2023 Oct 5. doi: 10.1007/s00430-023-00782-9. Online ahead of print.

ABSTRACT

Methicillin-resistant Staphylococcus aureus (MRSA) is a pathogenic bacterium responsible for a range of severe infections, such as skin infections, bacteremia, and pneumonia. Due to its antibiotic-resistant nature, current research focuses on targeting its virulence factors. Sortase A (SrtA) is a transpeptidase that anchors surface proteins to the bacterial cell wall and is involved in adhesion and invasion to host cells. Through fluorescence resonance energy transfer (FRET), we identified echinacoside (ECH), a natural polyphenol, as a potential SrtA inhibitor with an IC₅₀ of 38.42 μM in vitro. It was demonstrated that ECH inhibited SrtA-mediated S. aureus fibrinogen binding, surface protein A anchoring, and biofilm formation. The fluorescence quenching assay determined the binding mode of ECH to SrtA and calculated the K_A-binding constant of 3.09 × 10⁵ L/mol, demonstrating the direct interaction between the two molecules. Molecular dynamics simulations revealed that ECH-SrtA interactions occurred primarily at the binding sites of A92G, A104G, V168A, G192A, and R197A. Importantly, the combination of ECH and vancomycin offered protection against murine models of MRSA-induced pneumonia. Therefore, ECH may serve as a potential antivirulence agent against S. aureus infections, either alone or in combination with vancomycin.

PMID:37796314 | DOI:10.1007/s00430-023-00782-9

Free Radic Biol Med. 2023 Oct 2:S0891-5849(23)00664-0. doi: 10.1016/j.freeradbiomed.2023.09.039. Online ahead of print.

ABSTRACT

Podocyte injury is a hallmark of glomerular disease and one of the leading causes of chronic kidney disease (CKD). Peroxisome proliferator-activated receptor α (PPARα) plays a key role in podocyte fatty acid oxidation (FAO). However, the underlying regulatory mechanisms remain unresolved. Trim63 is an E3 ubiquitin ligase that has been shown to inhibit PPARα activity; however, its role in fatty acid metabolism in the kidney has not been elucidated to date. In this study, we investigated the effects of overexpression and knockdown of Trim63 in Adriamycin (ADR)-induced nephropathy and diabetic nephropathy models and a podocyte cell line. In both rodents and human patients with proteinuric CKD, Trim63 was upregulated, particularly in the podocytes of injured glomeruli. In the ADR-induced nephropathy model, ectopic Trim63 application aggravated FAO deficiency and mitochondrial dysfunction and triggered intense lipid deposition, podocyte injury, and proteinuria. Notably, Trim63 inhibition alleviated FAO deficiency and mitochondrial dysfunction, and markedly restored podocyte injury and renal fibrosis in ADR-induced and diabetic nephropathy (DN) models. Additionally, Trim63 was observed to mediate PPARα ubiquitination and degradation, leading to podocyte injury. We demonstrate the pathological role of Trim63, which was previously unrecognized in kidney tissue, in FAO deficiency and podocyte injury. Targeting Trim63 may represent a viable therapeutic strategy for podocyte injury and proteinuria.

PMID:37793501 | DOI:10.1016/j.freeradbiomed.2023.09.039

J Med Virol. 2023 Oct;95(10):e29135. doi: 10.1002/jmv.29135.

ABSTRACT

Personalized cancer immunotherapies, combined with nanotechnology (nano-vaccines), are revolutionizing cancer treatment strategies, explicitly targeting Human papilloma virus (HPV)-related cancers. Despite the availability of preventive vaccines, HPV-related cancers remain a global concern. Personalized cancer nano-vaccines, tailored to an individual's tumor genetic mutations, offer a unique and promising solution. Nanotechnology plays a critical role in these vaccines by efficiently delivering tumor-specific antigens, enhancing immune responses, and paving the way for precise and targeted therapies. Recent advancements in preclinical models have demonstrated the potential of polymeric nanoparticles and high-density lipoprotein-mimicking nano-discs in augmenting the efficacy of personalized cancer vaccines. However, challenges related to optimizing the nano-carrier system and ensuring safety in human trials persist. Excitingly, the integration of nanotechnology with Proteolysis-Targeting Chimeras (PROTACs) provides an additional avenue to enhance the effectiveness of personalized cancer treatment. PROTACs selectively degrade disease-causing proteins, amplifying the impact of nanotechnology-based therapies. Overcoming these challenges and leveraging the synergistic potential of nanotechnology, PROTACs, and Proteolysis-Targeting Antibodies hold great promise in pursuing novel and effective therapeutic solutions for individuals affected by HPV-related cancers.

PMID:37792364 | DOI:10.1002/jmv.29135

Front Microbiol. 2023 Sep 19;14:1232413. doi: 10.3389/fmicb.2023.1232413. eCollection 2023.

ABSTRACT

The microbiota of the gut has continued to co-evolve alongside their human hosts conferring considerable health benefits including the production of nutrients, drug metabolism, modulation of the immune system, and playing an antagonistic role against pathogen invasion of the gastrointestinal tract (GIT). The gut is said to provide a habitat for diverse groups of microorganisms where they all co-habit and interact with one another and with the immune system of humans. Phages are bacterial parasites that require the host metabolic system to replicate via the lytic or lysogenic cycle. The phage and bacterial populations are regarded as the most dominant in the gut ecosystem. As such, among the various microbial interactions, the phage-bacteria interactions, although complex, have been demonstrated to co-evolve over time using different mechanisms such as predation, lysogenic conversion, and phage induction, alongside counterdefense by the bacterial population. With the help of models and dynamics of phage-bacteria interactions, the complexity behind their survival in the gut ecosystem was demystified, and their roles in maintaining gut homeostasis and promoting the overall health of humans were elucidated. Although the treatment of various gastrointestinal infections has been demonstrated to be successful against multidrug-resistant causative agents, concerns about this technique are still very much alive among researchers owing to the potential for phages to evolve. Since a dearth of knowledge exists regarding the use of phages for therapeutic purposes, more studies involving experimental models and clinical trials are needed to widen the understanding of bacteria-phage interactions and their association with immunological responses in the gut of humans.

PMID:37795308 | PMC:PMC10546012 | DOI:10.3389/fmicb.2023.1232413

Life Sci. 2023 Nov 15;333:122132. doi: 10.1016/j.lfs.2023.122132. Epub 2023 Oct 2.

ABSTRACT

A diverse and stable microbiota promotes a healthy state, nevertheless, an imbalance in gut or oral bacterial composition, called dysbiosis, can cause gastrointestinal disorders, systemic inflammatory states and oxidative stress, among others. Recently, gut and oral dysbiosis has been linked to Alzheimer's disease (AD), which is considered the most common form of dementia and a public health priority due to its high prevalence and incidence. The aim of this review is to highlight the implications of gut and oral microbiota in the neuroinflammation characteristic of AD pathology and the subsequent cognitive impairment. It is a systematic review of the current literature obtained by searching the PubMed, Web of Science and Scopus databases. The characteristic intestinal dysbiosis in AD patients leads to increased permeability of the intestinal barrier and activates immune cells in the central nervous system due to translocation of microbiota-derived metabolites and/or bacteria into the circulation leading to increased neuroinflammation and neuronal loss, thus generating the cognitive impairment characteristic of AD. The presence in the central nervous system of Porphyromonas gingivalis can cause an increased neuroinflammation and beta-amyloid peptide accumulation.

PMID:37793482 | DOI:10.1016/j.lfs.2023.122132

Biochim Biophys Acta Mol Basis Dis. 2023 Oct 2;1870(1):166908. doi: 10.1016/j.bbadis.2023.166908. Online ahead of print.

ABSTRACT

Metabolic disorders such as type 2 diabetes, fatty liver disease, hyperlipidemia, and obesity commonly co-occur but clinical treatment options do not effectively target all disorders. Calorie restriction, semaglutide, rosiglitazone, and mitochondrial uncouplers have all demonstrated efficacy against one or more obesity-related metabolic disorders, but it currently remains unclear which therapeutic strategy best targets the combination of hyperglycaemia, liver fat, hypertriglyceridemia, and adiposity. Herein we performed a head-to-head comparison of 5 treatment interventions in the female db/db mouse model of severe metabolic disease. Treatments included ∼60 % calorie restriction (CR), semaglutide, rosiglitazone, BAM15, and niclosamide ethanolamine (NEN). Results showed that BAM15 and CR improved body weight and liver steatosis to levels superior to semaglutide, NEN, and rosiglitazone, while BAM15, semaglutide, and rosiglitazone improved glucose tolerance better than CR and NEN. BAM15, CR, semaglutide, and rosiglitazone all had efficacy against hypertriglyceridaemia. These data provide a comprehensive head-to-head comparison of several key treatment strategies for metabolic disease and highlight the efficacy of mitochondrial uncoupling to correct multiple facets of the metabolic disease milieu in female db/db mice.

PMID:37793464 | DOI:10.1016/j.bbadis.2023.166908

Front Microbiol. 2023 Sep 8;14:1170418. doi: 10.3389/fmicb.2023.1170418. eCollection 2023.

ABSTRACT

Antibiotics remain the frontline agents for treating deadly bacterial pathogens. However, the indiscriminate use of these valuable agents has led to an alarming rise in AMR. The antibiotic pipeline is insufficient to tackle the AMR threat, especially with respect to the WHO critical category of priority Gram-negative pathogens, which have become a serious problem as nosocomial and community infections and pose a threat globally. The AMR pandemic requires solutions that provide novel antibacterial agents that are not only effective but against which bacteria are less likely to gain resistance. In this regard, natural or engineered phage-encoded lysins (enzybiotics) armed with numerous features represent an attractive alternative to the currently available antibiotics. Several lysins have exhibited promising efficacy and safety against Gram-positive pathogens, with some in late stages of clinical development and some commercially available. However, in the case of Gram-negative bacteria, the outer membrane acts as a formidable barrier; hence, lysins are often used in combination with OMPs or engineered to overcome the outer membrane barrier. In this review, we have briefly explained AMR and the initiatives taken by different organizations globally to tackle the AMR threat at different levels. We bring forth the promising potential and challenges of lysins, focusing on the WHO critical category of priority Gram-negative bacteria and lysins under investigation for these pathogens, along with the challenges associated with developing them as therapeutics within the existing regulatory framework.

PMID:37789862 | PMC:PMC10542408 | DOI:10.3389/fmicb.2023.1170418

J Infect Dis. 2023 Oct 3;228(Suppl 4):S291-S296. doi: 10.1093/infdis/jiad077.

ABSTRACT

BACKGROUND: Microbial-based cancer treatments are an emerging field, with multiple bacterial species evaluated in animal models and some advancing to clinical trials. Noninvasive bacteria-specific imaging approaches can potentially support the development and clinical translation of bacteria-based cancer treatments by assessing the tumor and off-target bacterial colonization.

METHODS: 18F-Fluorodeoxysorbitol (18F-FDS) positron emission tomography (PET), a bacteria-specific imaging approach, was used to visualize an attenuated strain of Yersinia enterocolitica, currently in clinical trials as a microbial-based cancer treatment, in murine models of breast cancer.

RESULTS: Y. enterocolitica demonstrated excellent 18F-FDS uptake in in vitro assays. Whole-body 18F-FDS PET demonstrated a significantly higher PET signal in tumors with Y. enterocolitica colonization compared to those not colonized, in murine models utilizing direct intratumor or intravenous administration of bacteria, which were confirmed using ex vivo gamma counting. Conversely, 18F-fluorodeoxyglucose (18F-FDG) PET signal was not different in Y. enterocolitica colonized versus uncolonized tumors.

CONCLUSIONS: Given that PET is widely used for the management of cancer patients, 18F-FDS PET could be utilized as a complementary approach supporting the development and clinical translation of Y. enterocolitica-based tumor-targeting bacterial therapeutics.

PMID:37788499 | DOI:10.1093/infdis/jiad077

Cancer Cell Int. 2023 Sep 30;23(1):225. doi: 10.1186/s12935-023-03062-x.

ABSTRACT

Post-translational modifications (PTMs) of proteins contribute to the occurrence and development of tumors. Previous studies have suggested that canonical PTMs such as ubiquitination, glycosylation, and phosphorylation are closely implicated in different aspects of gastrointestinal tumors. Recently, emerging evidence showed that non-canonical PTMs play an essential role in the carcinogenesis, metastasis and treatment of gastrointestinal tumors. Therefore, we summarized recent advances in sumoylation, neddylation, isoprenylation, succinylation and other non-canonical PTMs in gastrointestinal tumors, which comprehensively describe the mechanisms and functions of non-classical PTMs in gastrointestinal tumors. It is anticipated that targeting specific PTMs could benefit the treatment as well as improve the prognosis of gastrointestinal tumors.

PMID:37777749 | PMC:PMC10544213 | DOI:10.1186/s12935-023-03062-x

Exp Mol Med. 2023 Oct 2. doi: 10.1038/s12276-023-01087-w. Online ahead of print.

ABSTRACT

Posttranslational modification of proteins via ubiquitination determines their activation, translocation, dysregulation, or degradation. This process targets a large number of cellular proteins, affecting all biological pathways involved in the cell cycle, development, growth, and differentiation. Thus, aberrant regulation of ubiquitination is likely associated with several diseases, including various types of metabolic diseases. Among the ubiquitin enzymes, E3 ubiquitin ligases are regarded as the most influential ubiquitin enzymes due to their ability to selectively bind and recruit target substrates for ubiquitination. Continued research on the regulatory mechanisms of E3 ligases and their adaptors in metabolic diseases will further stimulate the discovery of new targets and accelerate the development of therapeutic options for metabolic diseases. In this review, based on recent discoveries, we summarize new insights into the roles of E3 ubiquitin ligases and their adaptors in the pathogenesis of metabolic diseases by highlighting recent evidence obtained in both human and animal model studies.

PMID:37779139 | DOI:10.1038/s12276-023-01087-w

Genome Biol Evol. 2023 Sep 30:evad175. doi: 10.1093/gbe/evad175. Online ahead of print.

ABSTRACT

The detection of invasive pathogens is critical for host immune defense. Cell surface receptors play a key role in the recognition of diverse microbe-associated molecules, triggering leukocyte recruitment, phagocytosis, release of antimicrobial compounds, and cytokine production. The intense evolutionary forces acting on innate immune receptor genes have contributed to their rapid diversification across plants and animals. However, the functional consequences of immune receptor divergence are often unclear. Formyl peptide receptors (FPRs) comprise a family of animal G-protein coupled receptors which are activated in response to a variety of ligands including formylated bacterial peptides, pathogen virulence factors, and host-derived antimicrobial peptides. FPR activation in turn promotes inflammatory signaling and leukocyte migration to sites of infection. Here we investigate patterns of gene loss, diversification, and ligand recognition among FPRs in primates and carnivores. We find that FPR1, which plays a critical role in innate immune defense in humans, has been lost in New World primates. Amino acid variation in FPR1 and FPR2 among primates and carnivores is consistent with a history of repeated positive selection acting on extracellular domains involved in ligand recognition. To assess the consequences of FPR divergence on bacterial ligand interactions, we measured binding between primate FPRs and the FPR agonist Staphylococcus aureus enterotoxin B, as well as S. aureus FLIPr-like, an FPR inhibitor. We found that few rapidly evolving sites in primate FPRs are sufficient to modulate recognition of bacterial proteins, demonstrating how natural selection may serve to tune FPR activation in response to diverse microbial ligands.

PMID:37776517 | DOI:10.1093/gbe/evad175

Sci Rep. 2023 Sep 29;13(1):16403. doi: 10.1038/s41598-023-43762-y.

ABSTRACT

Cancer immunotherapy employing checkpoint inhibitors holds great promise across diverse cancers; nonetheless, a substantial proportion of patients (ranging from 55 to 87%) remain unresponsive to this treatment. To amplify therapeutic efficiency, we propose a synergistic therapeutic strategy that entails the deployment of targeted nano-sized particles carrying Toll-like receptor (TLR) agonists to the tumor site. This innovative approach seeks to activate intratumoral antigen-presenting cells using bioengineered outer membrane vesicles (OMVs) derived from gram-negative bacteria. These OMVs possess inherent attributes of surface-exposed immune stimulators and TLR-activating components, rendering them intriguing candidates for investigation. These OMVs were meticulously designed to selectively target cancer cells exhibiting an overexpression of epidermal growth factor receptor (EGFR). To gauge the precision of this targeting, the conducted affinity-based assays aimed at determining the equilibrium dissociation constant of the single-chain variable fragment employed for this purpose. In vitro experiments confirmed the OMVs' proficiency in adhering to EGFR-overexpressed cancer cells. Moreover, the evaluation extended to an in vivo context, where the therapeutic effect of nanovesicles was appraised within the tumor microenvironment of the triple-negative breast cancer mouse model. Notably, both intraperitoneal and intratumoral administrations of nanovesicles exhibited the ability to activate natural killer cells and skew M2 macrophage towards an M1 phenotype. The combined scrutiny of in vitro and in vivo findings underscores the potential efficiency of OMVs as a promising strategy for future anti-tumor endeavors.

PMID:37775519 | PMC:PMC10541432 | DOI:10.1038/s41598-023-43762-y

Toxicol Res. 2023 May 29;39(4):611-623. doi: 10.1007/s43188-023-00189-x. eCollection 2023 Oct.

ABSTRACT

FCCP (carbonyl cyanide-4-(trifluoromethoxy)phenylhydrazone) is known to inhibit oxidative phosphorylation as a protonophore, dissipating the proton gradient across the inner mitochondrial membrane. To understand the toxicity of FCCP, 3-day, 2- and 4-week repeated oral dose studies were performed in male rats. In the 3-day and 2-week repeated dose toxicity studies, observations included salivation, increased body temperature, and dead and moribund animals. Increased liver weight was observed in conjunction with hydropic degeneration and centrilobular necrosis of hepatocytes. In addition, pathological changes were observed in the pancreas, testis, epididymal duct, stomach and parotid gland. Electron microscopic examination revealed mitochondrial pleomorphism in the hepatocytes. Swelling of mitochondria was observed in the alpha cells and beta cells of the pancreas. Dilatation of rough endoplasmic reticulum, Golgi bodies and loss of secretory granules were also noted in the beta cells of the pancreas. FCCP was also compared with three other mUncouplers (DNP, OPC-163493 and tolcapone) with regard to in vitro mitochondrial uncoupling (mUncoupling) activities. FCCP produced the peak ΔOCR (oxygen consumption rate) at the lowest concentration (0.4 μM), followed by OPC-163493, tolcapone, and DNP, based on peak values in ascending order of concentration (2.5, 10, and 50 μM, respectively). Considering the relationship between the mUncoupling activity and toxicity profile of the four mUncouplers, there is no parallel relationship between the in vitro mUncoupling activity and the degree of in vivo toxicity. These findings may contribute to the efficient development of new mitochondrial uncoupler candidates.

SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s43188-023-00189-x.

PMID:37779591 | PMC:PMC10541353 | DOI:10.1007/s43188-023-00189-x