Molecular Pharmacology最新文献

The pregnane X receptor (PXR) is a ligand-activated regulator of cytochrome P450 (CYP)3A enzymes. Among the ligands of human PXR is hyperforin, a constituent of St John's wort (SJW) extracts and potent inducer of human CYP3A4. It was the aim of this study to compare the effect of hyperforin and SJW formulations controlled for its content on CYP3A23-3A1 in rats. Hyperiplant was used as it contains a high hyperforin content and Rebalance because it is controlled for a low hyperforin content. In silico analysis revealed a weak hyperforin-rPXR binding affinity, which was further supported in cell-based reporter gene assays showing no hyperforin-mediated reporter activation in presence of rPXR. However, cellular exposure to Hyperiplant and Rebalance transactivated the CYP3A reporter 3.8-fold and 2.8-fold, respectively, and they induced Cyp3a23-3a1 mRNA expression in rat hepatoma cells compared with control 48-fold and 18-fold, respectively. In Wistar rats treated for 10 days with 400 mg/kg of Hyperiplant, we observed 1.8 times the Cyp3a23-3a1 mRNA expression, a 2.6-fold higher CYP3A23-3A1 protein amount, and a 1.6-fold increase in activity compared with controls. For Rebalance we only observed a 1.8-fold hepatic increase of CYP3A23-3A1 protein compared with control animals. Even though there are differing effects on rCyp3a23-3a1/CYP3A23-3A1 in rat liver reflecting the hyperforin content of the SJW extracts, the modulation is most likely not linked to an interaction of hyperforin with rPXR. SIGNIFICANCE STATEMENT: Treatment with St John's wort (SJW) has been reported to affect CYP3A expression and activity in rats. Our comparative study further supports this finding but shows that the pregnane X receptor-ligand hyperforin is not the driving force for changes in rat CYP3A23-3A1 expression and function in vivo and in vitro. Importantly, CYP3A induction mimics findings in humans, but our results suggest that another so far unknown constituent of SJW is responsible for the expression- and function-modifying effects in rat liver.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by amyloid-β (Aβ) protein accumulation in the brain. Passive immunotherapies using monoclonal antibodies for targeting Aβ have shown promise for AD treatment. Indeed, recent US Food and Drug Administration approval of aducanumab and lecanemab, alongside positive donanemab Phase III results demonstrated clinical efficacy after decades of failed clinical trials for AD. However, the pharmacological basis distinguishing clinically effective from ineffective therapies remains unclear, impeding development of potent therapeutics. This study aimed to provide a quantitative perspective for effectively targeting Aβ with antibodies. We first reviewed the contradicting results associated with the amyloid hypothesis and the pharmacological basis of Aβ immunotherapy. Subsequently, we developed a quantitative systems pharmacology (QSP) model that describes the non-linear progression of Aβ pathology and the pharmacologic actions of the Aβ-targeting antibodies. Using the QSP model, we analyzed various scenarios for effective passive immunotherapy for AD. The model revealed that binding exclusively to the Aβ monomer has minimal effect on Aβ aggregation and plaque reduction, making the antibody affinity toward Aβ monomer unwanted, as it could become a distractive mechanism for plaque reduction. Neither early intervention, high brain penetration, nor increased dose could yield significant improvement of clinical efficacy for antibodies targeting solely monomers. Antibodies that bind all Aβ species but lack effector function exhibited moderate effects in plaque reduction. Our model highlights the importance of binding aggregate Aβ species and incorporating effector functions for efficient and early plaque reduction, guiding the development of more effective therapies for this devastating disease. SIGNIFICANCE STATEMENT: Despite previous unsuccessful attempts spanning several decades, passive immunotherapies utilizing monoclonal antibodies for targeting amyloid-beta (Aβ) have demonstrated promise with two recent FDA approvals. However, the pharmacological basis that differentiates clinically effective therapies from ineffective ones remains elusive. Our study offers a quantitative systems pharmacology perspective, emphasizing the significance of selectively targeting specific Aβ species and importance of antibody effector functions. This perspective sheds light on the development of more effective therapies for this devastating disease.

Glucocorticoids act via the glucocorticoid receptor (GR; NR3C1) to downregulate inflammatory gene expression and are effective treatments for mild to moderate asthma. However, in severe asthma and virus-induced exacerbations, glucocorticoid therapies are less efficacious, possibly due to reduced repressive ability and/or the increased expression of proinflammatory genes. In human A549 epithelial and primary human bronchial epithelial cells, toll-like receptor (TLR)-2 mRNA and protein were supra-additively induced by interleukin-1β (IL-1β) plus dexamethasone (IL-1β+Dex), interferon-γ (IFN-γ) plus dexamethasone (IFN-γ+Dex), and IL-1β plus IFN-γ plus dexamethasone (IL-1β+IFN-γ+Dex). Indeed, ∼34- to 2100-fold increases were apparent at 24 hours for IL-1β+IFN-γ+Dex, and this was greater than for any single or dual treatment. Using the A549 cell model, TLR2 induction by IL-1β+IFN-γ+Dex was antagonized by Org34517, a competitive GR antagonist. Further, when combined with IL-1β, IFN-γ, or IL-1β+IFN-γ, the enhancements by dexamethasone on TLR2 expression required GR. Likewise, inhibitor of κB kinase 2 inhibitors reduced IL-1β+IFN-γ+Dex-induced TLR2 expression, and TLR2 expression induced by IL-1β+Dex, with or without IFN-γ, required the nuclear factor (NF)-κB subunit, p65. Similarly, signal transducer and activator of transcription (STAT)-1 phosphorylation and γ-interferon-activated sequence-dependent transcription were induced by IFN-γ These, along with IL-1β+IFN-γ+Dex-induced TLR2 expression, were inhibited by Janus kinase (JAK) inhibitors. As IL-1β+IFN-γ+Dex-induced TLR2 expression also required STAT1, this study reveals cooperation between JAK-STAT1, NF-κB, and GR to upregulate TLR2 expression. Since TLR2 agonism elicits inflammatory responses, we propose that synergies involving TLR2 may occur within the cytokine milieu present in the immunopathology of glucocorticoid-resistant disease, and this could promote glucocorticoid resistance. SIGNIFICANCE STATEMENT: This study highlights that in human pulmonary epithelial cells, glucocorticoids, when combined with the inflammatory cytokines interleukin-1β (IL-1β) and interferon-γ (IFN-γ), can synergistically induce the expression of inflammatory genes, such as TLR2. This effect involved positive combinatorial interactions between NF-κB/p65, glucocorticoid receptor, and JAK-STAT1 signaling to synergistically upregulate TLR2 expression. Thus, synergies involving glucocorticoid enhancement of TLR2 expression may occur in the immunopathology of glucocorticoid-resistant inflammatory diseases, including severe asthma.

Hematopoietic cell transplantation (HCT) is often considered a last resort leukemia treatment, fraught with limited success due to microbial infections, a leading cause of mortality in leukemia patients. To address this critical issue, we explored a novel approach by synthesizing antileukemic agents containing antibacterial substances. This innovative strategy involves conjugating fluoroquinolone antibiotics, such as ciprofloxacin (CIP) or levofloxacin (LVX), with the cell-penetrating peptide transportan 10 (TP10). Here, we demonstrate that the resultant compounds display promising biologic activities in preclinical studies. These novel conjugates not only exhibit potent antimicrobial effects but are also selective against leukemia cells. The cytotoxic mechanism involves rapid disruption of cell membrane asymmetry leading to membrane damage. Importantly, these conjugates penetrated mammalian cells, accumulating within the nuclear membrane without significant effect on cellular architecture or mitochondrial function. Molecular simulations elucidated the aggregation tendencies of TP10 conjugates within lipid bilayers, resulting in membrane disruption and permeabilization. Moreover, mass spectrometry analysis confirmed efficient reduction of disulfide bonds within TP10 conjugates, facilitating release and activation of the fluoroquinolone derivatives. Intriguingly, these compounds inhibited human topoisomerases, setting them apart from traditional fluoroquinolones. Remarkably, TP10 conjugates generated lower intracellular levels of reactive oxygen species compared with CIP and LVX. The combination of antibacterial and antileukemic properties, coupled with selective cytostatic effects and minimal toxicity toward healthy cells, positions TP10 derivatives as promising candidates for innovative therapeutic approaches in the context of antileukemic HCT. This study highlights their potential in search of more effective leukemia treatments. SIGNIFICANCE STATEMENT: Fluoroquinolones are commonly used antibiotics, while transportan 10 (TP10) is a cell-penetrating peptide (CPP) with anticancer properties. In HCT, microbial infections are the primary cause of illness and death. Combining TP10 with fluoroquinolones enhanced their effects on different cell types. The dual pharmacological action of these conjugates offers a promising proof-of-concept solution for leukemic patients undergoing HCT. Strategically designed therapeutics, incorporating CPPs with antibacterial properties, have the potential to reduce microbial infections in the treatment of malignancies.

G protein-coupled receptors (GPCRs) exhibit a wide range of pharmacological efficacies, yet the molecular mechanisms responsible for the differential efficacies in response to various ligands remain poorly understood. This lack of understanding has hindered the development of a solid foundation for establishing a mathematical model for signaling efficacy. However, recent progress has been made in delineating and quantifying receptor conformational states and associating function with these conformations. This progress has allowed us to construct a mathematical model for GPCR signaling efficacy that goes beyond the traditional ON/OFF binary switch model. In this study, we present a quantitative conformation-based mathematical model for GPCR signaling efficacy using the adenosine A_2A receptor (A_2AR) as a model system, under the guide of ¹⁹F quantitative nuclear magnetic resonance experiments. This model encompasses two signaling states, a fully activated state and a partially activated state, defined as being able to regulate the cognate Gα _s nucleotide exchange with respective G protein recognition capacity. By quantifying the population distribution of each state, we can now in turn examine GPCR signaling efficacy. This advance provides a foundation for assessing GPCR signaling efficacy using a conformation-based mathematical model in response to ligand binding. SIGNIFICANCE STATEMENT: Mathematical models to describe signaling efficacy of GPCRs mostly suffer from considering only two states (ON/OFF). However, research indicates that a GPCR possesses multiple active-(like) states that can interact with Gαβγ independently, regulating varied nucleotide exchanges. With the guide of ¹⁹F-qNMR, the transitions among these states are quantified as a function of ligand and Gαβγ, serving as a foundation for a novel conformation-based mathematical signaling model.