Journal of Pharmacology and Experimental Therapeutics最新文献

Enteroendocrine cells (EECs) in the gastrointestinal tract play a critical role in sensing dietary fat and regulating the secretion of gut hormone. However, chronic high-fat diet (HFD) intake can lead to maladaptive changes in these cells, impairing hormone regulation. Transient receptor potential ankyrin 1 (TRPA1), an ion channel endogenously expressed in EECs, is known to promote gut hormone secretion when activated. Previous studies have shown that gut TRPA1 expression is reduced in HFD-fed mice, but the underlying molecular mechanisms remained unclear. In this study, we used the secretin tumor cell-1 (STC-1) enteroendocrine cell line treated with a fatty acid (FA) mixture (oleic acid: stearic acid in a 2:3 ratio) to mimic chronic HFD exposure in vitro. Our data from label-free proteomics, flow cytometry, and western blotting revealed that FA treatment causes TRPA1 downregulation through AMP-activated protein kinase and Ca²⁺ signaling pathways. This downregulation was accompanied by altered expression of genes and proteins involved in gut hormone synthesis and secretion. We further investigated the protective effect of allicin, a natural TRPA1 agonist found in garlic. Allicin treatment prevented TRPA1 downregulation both in FA-treated STC-1 cells and in HFD-fed C57BL/6J mice. In conclusion, this study elucidates the AMP-activated protein kinase-dependent mechanisms behind FA-induced TRPA1 downregulation in EECs and highlights how this contributes to gut hormone dysregulation. Importantly, dietary TRPA1 agonists such as allicin can counteract these effects, suggesting potential for development of functional foods (eg, allicin, thiocyanates, cuminaldehyde, cinnamaldehyde) to mitigate HFD-related gut hormone disturbances. SIGNIFICANCE STATEMENT: The results of this study showed that fatty acids downregulate transient receptor potential ankyrin 1, a key ion channel involved in regulating the secretion of gut hormones. Furthermore, this study investigated the potential protective effects of allicin, a dietary transient receptor potential ankyrin 1 agonist, using both an in vitro secretin tumor cell-1 model and an in vivo high-fat diet-fed C57Bl/6 mouse model.

Pancreatic ductal adenocarcinoma (PDA) is an almost universally fatal disease. Recent advances in the understanding of PDA bioenergetic dynamic equilibrium have illuminated a potential therapeutic target in bromodomain-related protein 4 (BRD4), the most active member of the bromo- and extraterminal domain (BET) protein family of transcription factors. We previously demonstrated that BET inhibitors (BETi) decrease PDA cell proliferation and enhance chemosensitivity. We hypothesized that BETi activates mitophagy and ferroptosis in PDA. Using pharmacological and genetic BRD4 inhibition in PDA patient-derived models, we investigated the effects of BETi on mitochondrial function, mitochondrial protein complex production, ATP production, cellular respiration, autophagy/mitophagy, and murine tumor growth with BMS-986158, a BETi. We determined the role of BRD4 in PDA by evaluating mitophagy and autophagy. In PDA models, we found that BETi decreased cellular respiration (P < .01), decreased ATP production (P < .001), and increased intracellular iron uptake (P < .01) while inducing mitophagy through dysregulated mitochondria complex protein levels. Murine PDA tumors grew slower and were smaller when treated with BETi compared with the control treatment. PDA tumors from experimentally treated mice contained more lipid vacuoles than those from the vehicle control group (P < .01), consistent with ferroptosis. BETi therapy decreased isocitrate dehydrogenase-1 expression, indicating increased chemosensitivity. BETi dysregulate mitochondrial complexes inducing mitophagy. BETi is a promising therapeutic strategy for attacking oncogenic mitochondrial behavior in PDA. We demonstrated a series of mitochondrial-centered events in a temporal sequence leading to cell death. This treatment controls tumors and increases chemosensitivity, offering a novel therapeutic strategy. SIGNIFICANCE STATEMENT: Bromo- and extraterminal domain inhibition is a novel therapeutic strategy for attacking oncogenic mitochondrial behavior in pancreatic ductal adenocarcinoma. Using this strategy in patient-derived models, this study demonstrated a series of mitochondrial-centered events in a temporal sequence leading to cell death and tumor control.

Colorectal cancer remains a leading cause of cancer-related mortality, necessitating the development of novel therapeutic strategies. In this study, we investigate the pharmacological and toxicological properties of JAMC-4/108, a traditional Mongolian herbal extract, in human colonic adenocarcinoma (HT-29) and normal colonic epithelial (CCD 841 CoTr) cells. Using high-performance liquid chromatography-electrospray ionization-quadrupole time-of-flight-tandem mass spectrometry fingerprinting, we characterized its chemical composition, while a combination of cellular, biochemical and molecular assays provided mechanistic insights into its cytotoxic effects. The ethanol extract of JAMC-4/108 exhibited substantial selective cytotoxicity in HT-29 cells by inducing apoptosis and necrosis via caspase-3, cleaved caspase-7 and poly(ADP-ribose) polymerase activation, while the water extract primarily triggered caspase-9-mediated apoptosis. Both extracts modulated oxidative stress pathways, increasing Nrf2, Keap1 and LC3A/B levels, with the ethanol extract also upregulating NQO1, suggesting metabolic implications. In contrast, the ethanol extract had minimal apoptotic effects in normal colonic epithelial cells, whereas the water extract primarily influenced caspase-9 expression, indicating a distinct toxicity profile. Additionally, both extracts altered cell cycle progression-stimulated NOx release and modulated Fe³⁺ ion and DPPH radical pools in a concentration-dependent manner. Our findings highlight the selective anticancer potential of JAMC-4/108 and provide mechanistic insights into its pharmacological and toxicological effects, supporting its further evaluation for therapeutic development and safety assessment. SIGNIFICANCE STATEMENT: This study demonstrates the selective anticancer activity of JAMC-4/108, a Mongolian herbal extract, against colorectal cancer cells, with distinct apoptotic mechanisms and minimal toxicity to normal cells. These findings support the potential of JAMC-4/108 as a novel therapeutic candidate for colorectal cancer treatment, highlighting its pharmacological efficacy and safety profile for further development.

Metabolic dysfunction-associated steatohepatitis (MASH) poses a significant public health challenge, characterized by liver fat accumulation accompanied with inflammation and cell damage. Patients with MASH commonly exhibit hypercoagulability. Our previous work showed that direct oral anticoagulants exert anti-inflammatory effects in early stages of metabolic dysfunction. Mitigation of thromboinflammation ameliorated the manifestations of cardiometabolic complications. Here, we examine the protective effects of rivaroxaban in a rat model of MASH and compare it to representatives of 2 other antithrombotic drug classes, enoxaparin and clopidogrel, as well as silymarin, a bona fide hepatoprotective agent. Rats were divided into 6 groups: control, MASH (induced by an atherogenic diet), and treatment groups receiving either silymarin (50 mg/kg by mouth), rivaroxaban (20 mg/kg by mouth), enoxaparin (2 mg/kg subcutaneously), or clopidogrel (6.75 mg/kg by mouth) for 8 weeks, starting in the third week of induction. MASH rats showed elevated markers of visceral adipose thromboinflammation, along with liver injury markers (aspartate aminotransferase, alanine aminotransferase, and serum albumin), heightened hepatic levels of inflammatory cytokines (interleukin [IL]-1β and IL-6), reduced antioxidant capacity, and an imbalance in coagulation factors (elevated activated coagulation Factor X and Factor VIII/Protein C) compared with the control group. Anticoagulants and silymarin treatment led to varying degrees of amelioration of these MASH-associated abnormalities. Rivaroxaban demonstrated the most substantial improvement in thromboinflammatory markers, reaching levels comparable to the control group, with percentage improvements of approximately 52%, 49%, and 42% in activated coagulation Factor X, IL-1β, and IL-6, respectively. These findings suggest that direct oral anticoagulants hold promise as therapeutic agents for MASH by targeting the underlying thromboinflammatory state. SIGNIFICANCE STATEMENT: Metabolic dysfunction-associated steatohepatitis is a global health issue. In a rat model of the disease, rivaroxaban improves liver injury markers, outperforming bona fide hepatoprotective substances. This study emphasizes the role of thromboinflammation in metabolic dysfunction-associated steatohepatitis and highlights direct anticoagulants as a potential novel treatment approach.

The incidence of fatal drug overdoses has increased dramatically over the past decade due to the widespread availability of fentanyl and its analogs. As a complementary strategy to current overdose reversal agents, monoclonal antibodies (mAbs) are in development as therapeutics for prevention and reversal of fentanyl overdose. In the present study, the anti-fentanyl mAb HY6-F9 was tested for reversal of fentanyl-induced respiratory arrest (apnea) in a porcine model. In a first study, following fentanyl-induced apnea, chimeric HY6-F9 and naloxone control were administered as an intravenous bolus. Both chimeric HY6-F9 and naloxone restored spontaneous breathing within 90 seconds. Treatment with mAb increased the concentration of fentanyl in serum by 10-fold within the first minute after mAb bolus administration. In a second study, after induction of apnea, humanized HY6-F9 and naloxone control were administered as a slow intravenous infusion over 10 minutes to determine the ED₅₀ to restore baseline breathing. In this study, the mean ± SEM ED₅₀ of humanized HY6-F9 and naloxone to restore baseline respiratory rate were 16.0 ± 1.3 mg/kg and 6.9 ± 1.8 μg/kg, respectively. During mAb infusion, the concentration of fentanyl in serum increased proportionally to the concentration of infused mAb. The anti-fentanyl mAb ablated fentanyl-dependent opioid receptor activation in an in vitro system with concentrations of fentanyl similar to those observed in pigs after mAb treatment. These results demonstrate the efficacy of an anti-fentanyl mAb as a treatment to reverse fentanyl overdose. SIGNIFICANCE STATEMENT: Treatments for opioid use disorder and overdose are urgently needed. Here, we show that an anti-fentanyl monoclonal antibody reversed fentanyl-induced apnea in pigs, and caused rapid (<1 minute) redistribution of fentanyl into serum. Fentanyl was 99% bound by monoclonal antibodies and showed no activity at the opioid receptor.

Type I conventional dendritic cells (cDC1s) are key drivers of antitumor immunity. In human cancers, their presence correlates with better prognosis and survival benefits. In preclinical mouse tumor models, cDC1s are indispensable for successful T-cell-mediated tumor killing and therapeutic response to immune checkpoint blockade therapies. The essential role of cDC1s in antitumor immunity stems from their ability to uptake tumor-derived antigens and traffic them to the tumor-draining lymph node (tdLN) for T-cell priming. At the tdLN, cDC1s present tumor antigens to naïve CD8⁺ T cells and polarize them into tumor-specific cytotoxic T cells that eventually kill the tumor cells. Within the tumor, cDC1s secrete the chemokines CXCL9 and CXCL10 that recruit CXCR3⁺ effector natural killer and T cells and thereby sustain a local cytotoxic T-cell response. The trafficking and migration of cDC1s to the tumor and tdLNs are largely mediated by a chemokine-chemokine receptor network linking cDC1s to their interacting immune cell partners. In this review, we discuss 2 key chemokine ligand-receptor pairs, C-C chemokine ligand 5-C-C chemokine receptor 5 and X-C chemokine ligand 1-X-C chemokine receptor 1, that play essential roles in directing cDC1 migration to the tumor. Strategies that harness these cDC1-recruiting chemokine systems offer invaluable therapeutic prospects for enhancing current vaccine design and cancer immunotherapies. SIGNIFICANCE STATEMENT: The lack of type I conventional dendritic cells (cDC1s) in tumors represents a major roadblock for current cancer immunotherapies. Here, we highlight 2 chemokines, C-C chemokine ligand 5 and X-C chemokine ligand 1, that are critical for recruiting cDC1s to the tumor microenvironment, where they uptake tumor antigens and cross-present antigens to T cells following migration to the lymph nodes. We further discuss recent advances and limitations in current dendritic cell vaccine design and cancer adjuvant therapies, and propose new strategies to enhance cDC1 recruitment into tumors.

Nondrug alternative reinforcers have long been used as a component of therapeutic interventions for the management of substance use disorder; however, the conditions under which alternative reinforcers are most effective are not well characterized. This study evaluated the impact of varying the magnitude of an alternative reinforcer on oxycodone self-administration and drug-primed responding in male and female squirrel monkeys. Subjects (n = 4/sex) were trained under concurrent second-order schedules of reinforcement for intravenous oxycodone (0.001-0.1 mg/kg per injection) on one lever and sweetened condensed milk (5%, 10%, 20%, and 30% in water) on another. Oxycodone-primed responding was evaluated by administering 0.32 mg/kg of oxycodone prior to sessions in which saline was available on the drug-paired lever. During oxycodone self-administration sessions, milk availability decreased oxycodone self-administration and preference in a concentration-dependent manner; low milk concentrations were more effective at decreasing oxycodone's reinforcing potency in males. During priming tests, milk significantly attenuated oxycodone-primed responding in both males and females; low milk concentrations were more effective at decreasing the priming effects of oxycodone in females. That alternative reinforcers differentially impacted self-administration and oxycodone-primed responding in a sex-dependent manner suggests that treatment strategies that use alternative reinforcers may be more effective in males or females depending on when they are implemented. SIGNIFICANCE STATEMENT: The use of nondrug reinforcer alternatives is a key component of many therapeutic interventions for substance use disorder. However, the variables that influence response to alternative reinforcers remain incompletely understood. This study investigated how varying the magnitude of an alternative reinforcer impacts oxycodone self-administration and drug-primed responding in male and female squirrel monkeys. Overall, the findings indicate that alternative reinforcers affect self-administration and oxycodone-primed responding differently depending on sex.