Journal of Pharmacology and Experimental Therapeutics最新文献

Intrathecal administration of agmatine, an NMDA receptor (NMDAr) antagonist and nitric oxide synthase inhibitor, prevents neuropathic pain behavior in a dose-dependent manner by acting at the GluN2B subunit of the NMDAr. The present study investigated the pharmacological mechanism of agmatine's inhibitory effect using calcium imaging and an in vivo assay of nociceptive responses induced by NMDA. The application of NMDA-evoked calcium transients in the mouse spinal cord dorsal horn slice was inhibited by the NMDAr antagonist, 2-amino-5-phosphonovalerate. Agmatine also concentration-dependently inhibited NMDA-evoked calcium responses. To evaluate the role of the GluN2B subunit of the NMDAr in the agmatine response, we conditionally knocked-down Grin2B, the gene encoding GluN2B, in spinal cord dorsal horn neurons (GluN2B knockdown [GluN2B-KD]). In control spinal cord slices, ifenprodil inhibited NMDAr-mediated calcium transients, but it was not effective in GluN2B-KD. Surprisingly, agmatine was equally effective in reducing calcium transients in control and GluN2B-KD mouse spinal cord slices. To determine whether the effect of agmatine could be attributed to an action downstream of the NMDAr (eg, neuronal nitric oxide synthase [nNOS]), we used the PSD95-nNOS tethering inhibitor, IC87201, to disrupt the link between NMDAr and nNOS. In the presence of IC87201, agmatine's attenuation of NMDA-evoked calcium transients in ex vivo spinal cord dorsal horn was significantly reversed as was agmatine's antihyperalgesic effect in the intrathecal NMDA-evoked thermal hyperalgesia in vivo model. These results indicated that agmatine requires an intact NMDAr-PSD95-nNOS pathway to attenuate NMDAr-mediated calcium transients and thermal hyperalgesia induced by intrathecal NMDA. SIGNIFICANCE STATEMENT: Chronic pain is an urgent public health concern, and effective long-term treatments are still needed. Agmatine reduces pain in preclinical models without the side effects of motor dysfunction or addiction. Clarifying the pharmacological mechanism of agmatine's analgesic effect in spinal neurotransmission may facilitate the development of novel pain-alleviating therapeutics.

Acute respiratory distress syndrome (ARDS), often preceded by acute lung injury (ALI), is characterized by the accumulation of inflammatory fluid in the lung alveoli, leaky alveolar epithelium and endothelium, and overexpression of proinflammatory cytokines. This progression from ALI to ARDS is a major contributor to the high mortality observed in patients with COVID-19. The spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) binds to lung angiotensin-converting enzyme 2 (ACE2), and in addition to facilitating viral cell entry, it plays an important role in the development of ALI and ARDS, especially in the later phases of COVID-19 as well as long-COVID. Protein tyrosine phosphatase (PTP) 4A3 is a key mediator of ARDS pathology. This study tested the hypothesis that targeting PTP4A3 would prevent COVID-19-associated ALI. Intratracheal administration of SARS-CoV-2 spike protein subunit 1 to K18-hACE2 transgenic mice expressing human ACE2 elicited pulmonary and systemic inflammation, leaky alveoli, overexpression of cytokines, structural lung injury, and lung dysfunction; all these symptoms were ameliorated by the selective, allosteric inhibitor of PTP4A3, KVX-053. These findings provide the first evidence supporting a role for PTP4A3 in the development of SARS-CoV-2-mediated ALI. SIGNIFICANCE STATEMENT: This study tested the hypothesis that targeting PTP4A3 would prevent COVID-19-associated ALI/ARDS. Intratracheal administration of SARS-CoV-2 spike protein subunit 1 to K18-hACE2 transgenic mice expressing human ACE2 elicited pulmonary and systemic inflammation, leaky alveoli, overexpression of cytokines and chemokines, structural lung injury, and lung dysfunction; all these symptoms were ameliorated by the selective, allosteric inhibitor of PTP4A3, KVX-053. These findings suggest that this novel PTP4A3 inhibitor may be useful against COVID-19 and potentially other viral-induced ARDS.

The neuropeptide S receptor (NPSR) has been identified as a potential therapeutic target for anxiety and post-traumatic stress disorder. Central administration of neuropeptide S (NPS) in male mice produces anxiolytic-like effects, hyperlocomotion, and memory enhancement. Currently, the literature is limited in the number of studies investigating the effects of NPS in female test subjects despite females facing a higher prevalence of anxiety-related pathology, as well as greater risk for adverse effects while taking psychoactive drugs. Moreover, no previous studies have considered the influence of estrous cycle on the effects of NPS. The present study investigates whether NPS-mediated behavioral phenotypes seen in males translate to females, and whether they are affected by estrous cycle stage. Female C57BL/6NCr mice were intracerebroventricularly cannulated and underwent behavioral paradigms to test locomotion, anxiety, and memory. Estrous cycle stage was determined through examination of vaginal cytology. Our results provide evidence that NPS-mediated behaviors are influenced by the estrous cycle. Administration of NPS decreased anxiety-like behaviors more robustly when the female mice were in high estrogen stages of the estrous cycle. Therefore, the desired anxiolytic-like effects of targeting the NPSR are intact in female mice. However, these effects may to be influenced by the stage of the estrous cycle. The NPSR remains a strong potential drug target for new anxiolytic compounds and based on our initial observations further studies exploring the interaction of estrous cycle and the NPS system are warranted. SIGNIFICANCE STATEMENT: The neuropeptide S (NPS) receptor has been identified as a potential target for treating anxiety, a condition that is most prevalent in females. Therefore, the potential interaction of estrous cycle with the NPS system described in the present study is an important first step in understanding the function of the NPS system in females.

Previous studies indicated differing effects of dopamine D₁-like and D₂-like receptor (D₁R and D₂R, respectively) agonists on cocaine self-administration. Leftward shifts by D₂R agonists in the cocaine self-administration dose-effect function contrast with decreases by D₁R agonists in maximal cocaine self-administration without rightward or leftward displacement. Whether the effects of the D₁R agonists are due to actions at D₁Rs has not been determined, possibly due to the difficulty in separating the blockade by a D₁R antagonist of the effects of the D₁R agonists and those of cocaine. In the present study, pretreatment with the D₁R agonists R(+)-SKF-81297 (0.1-1.0 mg/kg) and (±)-SKF-82958 (0.032-0.32 mg/kg) dose-dependently decreased maximal cocaine self-administration at doses below those affecting food-reinforced responding. In contrast, pretreatment with the D₂R agonists R(-)-NPA (0.001-0.01 mg/kg) and (-)-quinpirole (0.01-0.1 mg/kg) dose-dependently left-shifted the cocaine self-administration dose-effect function. The decreases by D₁R agonists in maximal cocaine self-administration were dose-dependently antagonized by the D₁R antagonist SCH-39166 at doses that alone had no effects on cocaine self-administration. Doses of SCH-39166 that blocked the effects of the D₁R agonists on cocaine self-administration were like those that shifted self-administration of D₁R agonists to the right but had no effects on self-administration of D₂R agonists. Self-administration of the D₂R agonists was dose-dependently shifted to the right by the preferential D₂R antagonist L-741,626 but not by SCH-39166. These results demonstrate that the decreases by the D₁R agonists in cocaine self-administration are selectively D₁R-mediated and support findings suggesting fundamentally distinct roles of the D₁Rs and D₂Rs in cocaine reinforcement. SIGNIFICANCE STATEMENT: Dopamine D₁-like (D₁R) agonists decrease maximal cocaine self-administration, whereas D₂-like (D₂R) agonists shift the cocaine self-administration dose-effect function leftward, with mechanisms for those different effects unclear. The present study demonstrates blockade by the selective D₁R antagonist SCH-39166 of D1R-mediated decreases in maximal cocaine self-administration at doses that blocked other D₁R-mediated effects but not effects of cocaine, suggesting fundamentally distinct roles of the dopamine D₁-like and D₂-like receptors in cocaine reinforcement and development of D₁R agonists as potential treatments for cocaine use disorder.

Discoidin domain receptor 1 (DDR1) is a collagen receptor with tyrosine kinase activity, and its expression is enhanced in various disease conditions. Although previous research suggests that DDR1 contributes to renal disease progression, DDR1 inhibitors for renal fibrosis have yet to be developed. In this study, we used unilateral ureteral obstruction (UUO) mice to investigate whether CH6824025, a strong and selective DDR1 phosphorylation inhibitor, can improve renal fibrosis. Furthermore, we performed 10x Visium spatial transcriptomics (ST) analysis on the kidney. CH6824025 suppressed the phosphorylation of DDR1 in the kidney, and the amount of hydroxyproline, the Sirius red- and the F4/80-positive area, and the mRNA expression of fibrosis and inflammation-related genes in the kidney were significantly decreased. 10x Visium ST analysis suggested that DDR1 is mainly expressed in distal nephrons under normal conditions but its expression appears to increase in the injured proximal tubules in UUO mice. Comparing mRNA expression in DDR1-positive spots in the Vehicle and the CH6824025 group, oxidative phosphorylation and mitochondrial dysfunction might be improved, and pathways involved in fibrosis tended to be inhibited in the CH6824025 administration group. Downstream analysis would suggest that mRNA expression changes in the CH6824025 group contribute to the inhibition of cell movement. Taken together, our findings suggest that CH6824025 inhibited kidney fibrosis in UUO mice, which might be due to the inhibition of the migration of inflammatory cells to the injury site and the reduction of inflammation. DDR1 inhibitors are expected to be a promising treatment of renal fibrosis. SIGNIFICANCE STATEMENT: The novel discoidin domain receptor 1 inhibitor CH6824025 could ameliorate fibrosis and inflammation in unilateral ureteral obstruction (UUO) mice. CH6824025 would inhibit cell motility (e.g., migration) that prevents the progression of fibrosis and improves mitochondrial function in UUO mice. CH6824025 could provide a significant benefit to patients with kidney fibrosis.

Cisplatin is a common platinum-based chemotherapeutic that induces acute kidney injury (AKI) in about 30% of patients. Pharmacokinetic/toxicodynamic (PKTD) models of cisplatin-induced AKI have been used to understand risk factors and evaluate potential mitigation strategies. While both traditional clinical biomarkers of kidney function [e.g., serum creatinine (SCr), blood urea nitrogen (BUN), estimated glomerular filtration rate (eGFR), and creatinine clearance (CrCl)] and newer subclinical biomarkers of kidney injury [e.g., urinary kidney injury molecule 1 (KIM-1), beta-2 microglobulin (B2M), neutrophil gelatinase-associated lipocalin (NGAL), calbindin, etc.] can be used to detect cisplatin-induced AKI, published PKTD models are limited to using only traditional clinical biomarkers. Previously identified risk factors for cisplatin nephrotoxicity have included dose, age, sex, race, body surface area, genetics, concomitant medications, and comorbid conditions. However, the relationships between concentrations and the pharmacokinetics (PK) of platinum and biomarkers of kidney injury have not been well elucidated. This review discusses the evaluation of cisplatin-induced nephrotoxicity in clinical studies, mouse models, and in vitro models, and examines the available human PK and toxicodynamic (TD) data. Improved understanding of the relationships between platinum PK and TD, in the presence of identified risk factors, will enable the prediction and prevention of cisplatin kidney injury. SIGNIFICANCE STATEMENT: As cisplatin treatment continues to cause AKI in a third of patients, it is critical to improve the understanding of the relationships between platinum PK and nephrotoxicity as assessed by traditional clinical and contemporary subclinical TD markers of kidney injury. Prediction and prevention of cisplatin-induced nephrotoxicity will be advanced by the evolving development of PKTD models that incorporate kidney injury biomarkers with enhanced sensitivity and include covariates that can impact risk of developing cisplatin-induced AKI.

Single nucleotide polymorphisms (SNPs) in cytochrome P450 (CYP450) enzymes alter the metabolism of a variety of drugs. Numerous medications, including chemotherapies, are metabolized by CYP450 enzymes, making the expression of this suite of enzymes in tumor cells relevant to prescription regimens for patients with cancer. We analyzed the characteristics of mutations of the cytochrome P450 2D6 (CYP2D6) enzymes in cancer patients obtained from the Catalogue of Somatic Mutations in Cancer (COSMIC), including mutation type, age of the patient, tissue type, and histology. Mutations were analyzed through the Cancer-Related Analysis of Variants Toolkit (CRAVAT) software along with cancer-specific high-throughput annotation of somatic mutations (CHASMplus) and variant effect scoring tool (VEST4) algorithms to determine the likelihood of being a driver and/or pathogenic mutation. For mutations with significant CHASMplus and VEST4 scores, structural analysis of each corresponding mutant protein was performed. The effect of each mutation was evaluated for its impact on the overall protein stability and ligand binding using Foldit Standalone and SwissDock, respectively. Structural analysis revealed that several missense mutations in CYP2D6 resulted in altered stability after energy minimization. Three missense mutations of CYP2D6 significantly altered docking stability, and those located on alpha helices near the docking site had a more significant impact than those not found in secondary protein structures. In conclusion, we have identified a series of mutations to CYP2D6 enzymes with possible relevance to cancer pathologies. SIGNIFICANCE STATEMENT: CYP2D6 is responsible for the metabolism of many anticancer drugs. This study identified and characterized a series of mutations in the CYP2D6 enzyme that occurred in tumors. We found it likely that many of these mutations would alter enzyme function, leading to changes in drug metabolism in the tumor. We provide a basis for predicting the likelihood of a patient carrying these mutations to identify patients who may benefit from a precision medicine approach to drug selection and dosing.

Amyloid β(Aβ) deposition and neurofibrillary tangles are widely considered the primary pathological hallmarks of familial and sporadic forms of Alzheimer disease (AD). However, cerebrovascular inflammation, which is prevalent in 70% of AD patients, is emerging as another core feature of AD pathology. In our current work, we investigated the hypothesis that Aβ42 exposure drives an increase in vascular cell adhesion molecule-1 (VCAM-1) expression, a cerebrovascular inflammatory marker expressed on the blood-brain barrier (BBB) endothelium in humans and murine models. We have demonstrated that the inflammation signaling pathway is upregulated in AD patient brains, and VCAM-1 expression is increased in AD patients compared with healthy controls. Furthermore, dynamic SPEC/CT imaging in APP,PS1 transgenic mice (a mouse model that overexpresses Aβ42) demonstrated VCAM-1 upregulation at the BBB. Although there is a strong association between Aβ42 exposure and an increase in VCAM-1 expression, the underlying mechanisms remain partially understood. Molecular mechanisms driving VCAM-1 expression at the BBB were investigated in polarized human cerebral microvascular endothelial cell monolayers. Moreover, by employing reverse-phase protein array assays and immunocytochemistry we demonstrated that Aβ42 increases VCAM-1 expression via the Src/p38/MEK signaling pathway. Therefore, targeting the Src/p38/MEK pathway may help modulate VCAM-1 expression at the BBB and help mitigate cerebrovascular inflammation in Alzheimer disease. SIGNIFICANCE STATEMENT: Although considered a core pathological feature of Alzheimer disease, molecular pathways leading to cerebrovascular inflammation remain only partially understood. Moreover, clinical diagnostic methods for detecting cerebrovascular inflammation are underdeveloped. This study demonstrated the detection of VCAM-1 using radio-iodinated VCAM-1 antibody and single-photon emission computed tomography/computed tomography imaging. Additionally, exposure to Aβ42 increases VCAM-1 expression on the blood-brain barrier endothelium via the Src/p38/MEK pathway. These findings are expected to aid in the development of diagnostic and therapeutic approaches for addressing cerebrovascular inflammation in AD.