Reverse Translational Study Research Articles

Methodological advances in gastrointestinal tract physiology measurements: relevance to nutritional studies.

The gastrointestinal (GI) tract plays a critical role in nutrition and the pathophysiology of disease, and there is an increasing variety of methodologies available for the assessment of various aspects of GI physiology. Advancements in assessment methods, including techniques to study gut motility, fermentation, permeability, and microbiota composition, have provided researchers with powerful tools to investigate the impact of diet on GI tract physiology and the microbiota-gut-brain axis. Mechanistic evidence from reverse translational studies, which apply findings from human studies to preclinical models in a "bedside-to-bench" approach, have also enhanced our understanding of the bidirectional interactions and candidate signalling molecules among the diet-gut-brain relationship. Interpreting data from these advanced techniques and study designs requires a thorough understanding of their principles, applications, and limitations. This review aims to summarise the methodological advances in GI tract physiology measurements and their application in nutritional studies, focusing on gut motility, fermentation, and permeability. We will present examples of how these techniques have been utilised in recent research, discuss their advantages and limitations, and provide insights on their use and interpretation in research. Understanding the capabilities and limitations of these tools is crucial for designing robust studies and elucidating the complex interplay between diet and the GI tract. The scope of this review encompasses recent advancements in GI tract assessment methodologies and their implications for nutritional research, providing a comprehensive overview for researchers in the field.

Progress towards understanding risk factor mechanisms in the development of autism spectrum disorders.

Autism spectrum disorders (ASD) are a heterogenous set of syndromes characterised by social impairment and cognitive symptoms. Currently, there are limited treatment options available to help people with ASD manage their symptoms. Understanding the biological mechanisms that result in ASD diagnosis and symptomatology is an essential step in developing new interventional strategies. Human genetic studies have identified common gene variants of small effect and rare risk genes and copy number variants (CNVs) that substantially increase the risk of developing ASD. Reverse translational studies using rodent models based on these genetic variants provide new insight into the biological basis of ASD. Here we review recent findings from three ASD associated CNV mouse models (16p11.2, 2p16.3 and 22q11.2 deletion) that show behavioural and cognitive phenotypes relevant to ASD. These models have identified disturbed excitation-inhibition neurotransmitter balance, evidenced by dysfunctional glutamate and GABA signalling, as a key aetiological mechanism. These models also provide emerging evidence for serotoninergic neurotransmitter system dysfunction, although more work is needed to clarify the nature of this. At the brain network level, prefrontal cortex (PFC) dysfunctional connectivity is also evident across these models, supporting disturbed PFC function as a key nexus in ASD aetiology. Overall, published data highlight the utility and valuable insight gained into ASD aetiology from preclinical CNV mouse models. These have identified key aetiological mechanisms that represent putative novel therapeutic targets for the treatment of ASD symptoms, making them useful translational models for future drug discovery, development and validation.

Plasma Venetoclax Concentrations in Patients with Acute Myeloid Leukemia Treated with CYP3A4 Inhibitors.

Venetoclax (VEN) is used in patients with acute myeloid leukemia (AML) and is primarily metabolized by CYP3A4, a major drug-metabolizing enzyme. Patients with AML simultaneously administered VEN and CYP3A4 inhibitors require a more appropriate management of drug-drug interactions (DDIs). Here, we report two cases of patients with AML (54-year-old man and 22-year-old woman) administrated VEN and CYP3A4 inhibitors, such as posaconazole, cyclosporine, or danazol. In the first case, we evaluated the appropriateness of timing for adjusting VEN dosage subsequent to the cessation of posaconazole. Consequently, modifying the VEN dosage in conjunction with the cessation of Posaconazole simultaneously may result in elevated plasma VEN levels. In the second case, plasma VEN concentrations were markedly elevated when co-administered with several CYP3A4 inhibitors. Additionally, in vitro assays were conducted for reverse translational studies to analyze CYP3A4 inhibition. CYP3A4 inhibition by combinatorial administration of cyclosporine A and danazol was demonstrated in vitro, which potentially explains the increasing plasma VEN concentrations observed in clinical settings. Although the acquisition of therapeutic effects is a major priority for patients, frequent therapeutic drug monitoring and dosage adjustments considering DDIs would be important factors in chemotherapy.

Integration of individual preclinical and clinical anti-infective PKPD data to predict clinical study outcomes.

The AIDA randomized clinical trial found no significant difference in clinical failure or survival between colistin monotherapy and colistin-meropenem combination therapy in carbapenem-resistant Gram-negative infections. The aim of this reverse translational study was to integrate all individual preclinical and clinical pharmacokinetic-pharmacodynamic (PKPD) data from the AIDA trial in a pharmacometric framework to explore whether individualized predictions of bacterial burden were associated with the trial outcomes. The compiled dataset included for each of the 207 patients was (i) information on the infecting Acinetobacter baumannii isolate (minimum inhibitory concentration, checkerboard assay data, and fitness in a murine model), (ii) colistin plasma concentrations and colistin and meropenem dosing history, and (iii) disease scores and demographics. The individual information was integrated into PKPD models, and the predicted change in bacterial count at 24 h for each patient, as well as patient characteristics, was correlated with clinical outcomes using logistic regression. The invivo fitness was the most important factor for change in bacterial count. A model-predicted growth at 24 h of ≥2-log10 (164/207) correlated positively with clinical failure (adjusted odds ratio, aOR = 2.01). The aOR for one unit increase of other significant predictors were 1.24 for SOFA score, 1.19 for Charlson comorbidity index, and 1.01 for age. This study exemplifies how preclinical and clinical anti-infective PKPD data can be integrated through pharmacodynamic modeling and identify patient- and pathogen-specific factors related to clinical outcomes - an approach that may improve understanding of study outcomes.

Characterization of cardiovascular profile of anti-influenza drug peramivir: A reverse-translational study using the isoflurane-anesthetized dog

An injectable anti-influenza drug peramivir has been reported to induce QT-interval prolongation in some phase III studies, although its thorough QT/QTc study was negative. We investigated the discrepancy among those clinical studies using isoflurane-anesthetized beagle dogs (n = 4). Peramivir in doses of 1 mg/kg/10 min (sub-therapeutic dose) followed by 10 mg/kg/10 min (clinically-relevant dose) was intravenously administered. Peramivir prolonged QT interval/QTcV and Tpeak-Tend, and tended to delay ventricular repolarization in a reverse-frequency dependent manner, indicating IKr inhibition in vivo. Meanwhile, peramivir did not alter P-wave duration, PR interval or QRS width, indicating a lack of impact on cardiac conduction via Na+ or Ca2+ channel inhibition in vivo. Peramivir prolonged Tpeak-Tend and tended to prolong terminal repolarization period, which would develop substrates for initiating and maintaining spiral reentry, respectively. Meanwhile, peramivir did not prolong J-Tpeakc, which could not induce early afterdepolarization, a trigger inducing torsade de pointes. Thus, our results support that clinical dose exposure of peramivir can delay the ventricular repolarization in influenza patients. Peramivir has only a small potential to induce torsade de pointes in patients with the intact hearts, but caution should be paid on its use for patients formerly having the trigger for torsade de pointes.

Inactivation of phosphodiesterase-4B gene in rat nucleus accumbens shell by CRISPR/Cas9 or positive allosteric modulation of the protein affects the motivation to chronically self-administer nicotine

Large scale human genome wide association studies (GWAS) have identified a growing pool of genes associated with cigarette smoking. One of the most prominent, phosphodiesterase-4B (PDE4B), has been associated with multiple smoking phenotypes. Although PDE4B modulates the half-life of neuronal cAMP, its precise role in smoking behaviors is unknown. To address this knowledge gap, we used a reverse translational approach. We inactivated PDE4B in bilateral medial nucleus accumbens shell (NAcs) neurons by injecting AAV containing a specific gRNA in female transgenic Cas9+ Long Evans rats. These rats then were given 23-h chronic access to nicotine intravenous self-administration (IVSA) under a schedule of increasing fixed ratios (FR). With the increased effort required at FR7, nicotine SA (i.e. active presses and drug infusions) declined significantly in controls, whereas it was maintained in the mutagenized group. A progressive ratio (PR) study also showed significantly greater cumulative nicotine infusions in the PDE4B-edited group. Hence, we hypothesized that enhanced PDE4B protein activity would reduce nicotine IVSA. A positive allosteric modulator, 2-(3-(4-chloro-3-fluorophenyl)-5-ethyl-1H-1,2,4-triazol-1-yl)-N-(3,5-dichlorobenzyl)acetamide (MR-L2), was microinfused into NAcs bilaterally at FR3 or FR5; in both cohorts, MR-L2 acutely reduced nicotine IVSA. In summary, these studies show that the activity of PDE4B regulates the capacity of NAcs to maintain nicotine IVSA in face of the cost of increasing work. This finding and the results of the PR study indicate that PDE4B affects the motivation to obtain nicotine. These reverse translational studies in rats provide insight into the motivational effects of NAcs PDE4B that advance our understanding of the smoking behaviors mapped in human GWAS.

A reverse translational study of PPAR-α agonist efficacy in human and rodent models relevant to alcohol use disorder

Alcohol Use Disorder (AUD) is a chronic relapsing disorder affecting an estimated 283 million individuals worldwide, with substantial health and economic consequences. Peroxisome proliferator-activated receptors (PPARs), particularly PPAR-α and PPAR-γ, have shown promise in preclinical studies as potential therapeutic targets for AUD. In this human laboratory study, we aimed to translate preclinical findings on the PPAR-α agonist fenofibrate to a human population with current AUD. We hypothesized that, relative to placebo, fenofibrate at the highest FDA-approved dose of 145 mg/d would attenuate responsiveness to in vivo alcohol cues in the lab and reduce drinking under natural conditions. However, the results did not show significant differences in craving and alcohol consumption between the fenofibrate and placebo groups. Reverse translational studies in rodent models confirmed the lack of fenofibrate effect at human-equivalent doses. These findings suggest that inadequate translation of drug dose from rodents to humans may account for the lack of fenofibrate effects on alcohol craving and consumption in humans with AUD. The results highlight the need for new brain-penetrant PPAR-α agonists to adequately test the therapeutic potential of PPAR-α agonists for AUD, and the importance of reverse translational approaches and selection of human-equivalent doses in drug development.

Balancing the beautiful and the good in pursuit of biomarkers for depression.

Balancing the beautiful and the good in pursuit of biomarkers for depression.

Intracellular Adhesion Molecule-1 Improves Responsiveness to Immune Checkpoint Inhibitor by Activating CD8+ T Cells.

Immune checkpoint inhibitor (ICI) clinically benefits cancer treatment. However, the ICI responses are only achieved in a subset of patients, and the underlying mechanisms of the limited response remain unclear. 160 patients with non-small cell lung cancer treated with anti-programmed cell death protein-1(anti-PD-1) or anti-programmed death ligand-1 (anti-PD-L1)are analyzed to understand the early determinants of response to ICI. It is observed that high levels of intracellular adhesion molecule-1 (ICAM-1) in tumors and plasma of patients are associated with prolonged survival. Further reverse translational studies using murine syngeneic tumor models reveal that soluble ICAM-1 (sICAM-1) is a key molecule that increases the efficacy of anti-PD-1 via activation of cytotoxic T cells. Moreover, chemokine (CXC motif) ligand 13 (CXCL13) in tumors and plasma is correlated with the level of ICAM-1 and ICI efficacy, suggesting that CXCL13 might be involved in the ICAM-1-mediated anti-tumor pathway. Using sICAM-1 alone and in combination with anti-PD-1 enhances anti-tumor efficacy in anti-PD-1-responsive tumors in murine models. Notably, combinatorial therapy with sICAM-1 and anti-PD-1 converts anti-PD-1-resistant tumors to responsive ones in a preclinical study. These findings provide a new immunotherapeutic strategy for treating cancers using ICAM-1.

Outgrowth Endothelial Cell Conditioned Medium Negates TNF-α-Evoked Cerebral Barrier Damage: A Reverse Translational Research to Explore Mechanisms

Improved understanding of the key mechanisms underlying cerebral ischemic injury is essential for the discovery of efficacious novel therapeutics for stroke. Through detailed analysis of plasma samples obtained from a large number of healthy volunteers (n = 90) and ischemic stroke patients (n = 81), the current study found significant elevations in the levels of TNF-α at baseline (within the first 48 h of stroke) and on days 7, 30, 90 after ischaemic stroke. It then assessed the impact of this inflammatory cytokine on an invitro model of human blood–brain barrier (BBB) and revealed dramatic impairments in both barrier integrity and function, the main cause of early death after an ischemic stroke. Co-treatment of BBB models in similar experiments with outgrowth endothelial cell-derived conditioned media (OEC-CM) negated the deleterious effects of TNF-α on BBB. Effective suppression of anti-angiogenic factor endostatin, stress fiber formation, oxidative stress, and apoptosis along with concomitant improvements in extracellular matrix adhesive and tubulogenic properties of brain microvascular endothelial cells and OECs played an important role in OEC-CM-mediated benefits. Significant increases in pro-angiogenic endothelin-1 and monocyte chemoattractant protein-1 in OEC-CM compared to the secretomes of OEC and HBMEC, detected by proteome profiling assay, accentuate the beneficial effects of OEC-CM. In conclusion, this reverse translational study identifies TNF-α as an important mediator of post-ischemic cerebral barrier damage and proposes OEC-CM as a potential vasculoprotective therapeutic strategy by demonstrating its ability to regulate a wide range of mechanisms associated with BBB function. Clinical trial registration NCT02980354.Graphical

Assessment of disease outcome measures in systemic sclerosis.

The assessment of disease activity in systemic sclerosis (SSc) is challenging owing to its heterogeneous manifestations across multiple organ systems, the variable rate of disease progression and regression, and the relative paucity of patients in early-phase therapeutic trials. Despite some recent successes, most clinical trials have failed to show efficacy, underscoring the need for improved outcome measures linked directly to disease pathogenesis, particularly applicable for biomarker studies focused on skin disease. Current outcome measures in SSc-associated interstitial lung disease and SSc skin disease are largely adequate, although advancing imaging technology and the incorporation of skin mRNA biomarkers might provide opportunities for earlier detection of the therapeutic effect. Biomarkers can further inform pathogenesis, enabling early phase trials to act as reverse translational studies through the incorporation of routine high-throughput sequencing.

Decreased plasma acetaminophen glucuronide/acetaminophen concentration ratio warns the onset of acetaminophen-induced liver injury.

Acetaminophen (APAP)-induced liver injury (AILI) is the most common cause of acute liver failure. Although the mechanisms that trigger AILI are well known, it is less understood how to halt AILI progression and facilitate liver recovery. Therefore, it is necessary to understand the pathophysiology of APAP hepatotoxicity in patients and to examine predictive/preventive markers. In a clinical study, we had a case in which aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels increased in a patient with a low ratio of APAP glucuronide concentration (AP-G)/APAP plasma concentration. Then a reverse translational study was conducted for clarifying this clinical question. The relationship between plasma AP-G/APAP concentration ratio and the levels of AST and ALT was examined by in vivo and in vitro experiments. In in vivo experiments, 10-week-old rats showed lower UGT activity, lower AP-G/APAP concentration ratios, and higher AST and ALT levels than 5-week-old rats. This suggests an inverse correlation between the AP-G/APAP concentration ratio and the AST, ALT levels in APAP-treated rats. Furthermore, as a result of the in vitro experiment, it was confirmed that the cell viability decreased when the AP-G/APAP concentration ratio in the culture medium decreased. Since the decrease in the plasma AP-G/APAP concentration ratio appears earlier than the increase of AST and ALT levels, the ratio might be a presymptomatic marker of AILI. When APAP is used for a long time, it is recommended to perform therapeutic drug monitoring of the AP-G/APAP concentration ratio, which is a predictive/preventive marker of AILI.

Elevated Serum Purine Levels in Schizophrenia: A Reverse Translational Study to Identify Novel Inflammatory Biomarkers.

BackgroundImmunological markers and related signaling molecules in the blood are altered in schizophrenia mouse models, in acutely relapsed patients with schizophrenia, and in persons at a clinically high risk for subsequently developing psychosis, highlighting their potential as prognostic and theranostic biomarkers. Therefore, we herein aimed to identify novel potential biomarkers in the serum that are associated with purinergic signaling.MethodsTo our knowledge, this is the first study to assess the correlations among the levels of human serum adenine nucleotides (ATP, ADP), adenosine, P2X7 receptor, and disease activity in patients hospitalized due to an acute relapse of schizophrenia (n = 53) and healthy controls (n = 47). In addition, to validate these findings using a reverse translational approach, we examined the same parameters in an acute phencyclidine-induced schizophrenia mouse model.ResultsWe found consistently elevated levels of ATP, ADP, interleukin (IL)-6, and IL-10 in both schizophrenia groups compared with the controls. The levels of adenosine, IL-1β, IL-12, and C-reactive protein were also increased in the human patient samples. Moreover, ATP and ADP were significantly positively correlated with the Positive and Negative Symptom Scale item “lack of judgment and insight”; IL-1β, IL-12, and tumour necrosis factor alpha were significantly positively correlated with “tension” and “depression”; and “disorientation” and “poor attention” were correlated significantly with IL-6 and IL-8.ConclusionsOur study suggests the promising potential of blood purines and inflammatory markers as future prognostic tools.

Hypoglycemia Impairs the Heat Shock Protein Response: A Risk for Heat Shock in Cattle?

BackgroundHeat stress (HS) in cattle is a major debilitating problem, affecting health and milk yield. Physiologically, HS has been shown to lower blood glucose levels to 2.5 mmol/l (45 mg/dl) and results in upregulation of heat shock proteins (HSPs), eliciting the heat shock response (HSR) of which HSP90, 70 and 27 have been shown to be protective. However, it is unclear if the HSP response is blunted by decreased glucose, thereby preventing adaptive mechanisms. To address this question, this exploratory reverse translational study on the effects of hypoglycemia on the HSP pathway was undertaken.MethodsA human prospective, study in healthy control individuals (n = 23) was undertaken. Subjects underwent hyperinsulinemic-induced hypoglycemia [≤2.0 mmol/L (36 mg/dl)] with blood sampling at baseline, at hypoglycemia and for a 24-h post-hypoglycemia follow-up period. Proteomic analysis of the heat shock-related protein pathway, the pathway associated with HS in cattle, was performed.ResultsIn response to hypoglycemia, HS pathway proteins were significantly decreased (p < 0.05): HSP70 and HSP27 (at hypoglycemia); DnaJ homolog subfamily B member 1 (DNAJB1), Stress-induced-phosphoprotein 1 (STIP1) and the ubiquitin pathway proteins, Ubiquitin-conjugating enzyme (UBE2L3) and Ubiquitin-conjugating enzyme E2 N (UBE2N) (at 30-min post-hypoglycemia); HSP90 (at 2-h post-hypoglycemia). STIP1, UBE2L3, and UBE2N remained suppressed at 24-h.ConclusionHeat stress in cattle reduces blood glucose that, in turn, may blunt the HS pathway protective response, including HSP 90, 70, 27 and the ubiquitin proteins, leading to adverse outcomes. Monitoring of blood glucose in susceptible cattle may allow for earlier intervention and may also identify those animals at greatest risk to ensure that milk yield is not compromised.

Analysis of clinically-reported, memantine-induced cardiovascular adverse responses using the halothane-anesthetized dogs: Reverse translational study

Although NMDA receptor antagonist memantine is considered to be better tolerated than cholinesterase inhibitors on treating Alzheimer's disease, several types of cardiovascular adverse events have been associated with memantine treatment, including hypertension, myocardial infarction, severe bradycardia and QT-interval prolongation. In order to clarify how memantine induces these cardiovascular adverse events, we assessed its electropharmacological effects using the halothane-anesthetized dogs (n = 4). Memantine hydrochloride was intravenously administered in doses of 0.01, 0.1 and 1 mg/kg over 10 min, providing subtherapeutic, clinically-relevant and supratherapeutic concentrations, respectively. The low to high doses increased the mean blood pressure and left ventricular contraction and enhanced the atrioventricular nodal conduction, suggesting an increase of sympathicotonic output from the central nervous system similarly to donepezil, which might induce myocardial ischemia in patients with coronary artery disease. Meanwhile, the high dose suppressed the intra-atrial conduction and the low to high doses inhibited the intra-ventricular conduction, indicating potential to induce severe bradycardic adverse event by advanced cardiac conduction block in susceptible patients. Memantine alone did not induce repolarization delay, indicating lack of risk for inducing torsade de pointes. Thus, these in vivo experimental findings may provide basic information to better understand the clinically observed adverse events of memantine.

High-sensitivity reverse-translational studies in cell-specific genes: modified disease model animal using multidimensional cell-specific analysis of disease iPS cell differentiation-induced cells

High-sensitivity reverse-translational studies in cell-specific genes: modified disease model animal using multidimensional cell-specific analysis of disease iPS cell differentiation-induced cells

Is platelet responsiveness to clopidogrel attenuated in overweight or obese patients and why? A reverse translational study in mice

Overweight or obese patients exhibit poorer platelet responses to clopidogrel. However, the mechanisms behind this phenotype remain to be elucidated. Here, we sought to discover whether and why obesity could affect the metabolic activation of and/or platelet response to clopidogrel in obese patients and high-fat diet-induced obese mice. A post hoc stratified analysis of an observational clinical study was performed to investigate changes in residual platelet reactivity with increasing body weight in patients taking clopidogrel. Furthermore, high-fat diet-induced obese mice were used to reveal alterations in systemic exposure of clopidogrel thiol active metabolite H4, ADP-induced platelet activation and aggregation, the expression of genes involved in the metabolic activation of clopidogrel, count of circulating reticulated and mature platelets, and proliferation profiles of megakaryocytes in bone marrow. The relevant genes and potential signalling pathways were predicted and enriched according to the GEO datasets available from obese patients. Obese patients exhibited significantly attenuated antiplatelet effects of clopidogrel. In diet-induced obese mice, systemic exposure of clopidogrel active metabolite H4 was reduced but that of its hydrolytic metabolite was increased due to down-regulation of certain P450s but up-regulation of carboxylesterase-1 in the liver. Moreover, enhanced proliferation of megakaryocytes and elevated platelet count also contributed. Obesity attenuated metabolic activation of clopidogrel and increased counts of circulating reticulated and mature platelets, leading to impaired platelet responsiveness to the drug in mice, suggesting that clopidogrel dosage may need to be adjusted adequately in overweight or obese patients.

338-OR: Reversal of Insulin Resistance by Resveratrol and Hesperetin Combination in Overweight and Obese Subjects Correlates with Decrease in Expression of Thioredoxin Interacting Protein

Trans-Resveratrol and hesperetin combination (tRES-HESP) in randomized, double-blind, placebo-controlled crossover study in overweight and obese subjects corrected insulin resistance; healthy aging through functional food study (HATFF, Clinicaltrials.gov identifier NCT02095873). tRES-HESP is an optimized supplement for induction of expression of glyoxalase 1 (Glo1) via activation of transcription factor Nrf2. Increased Glo1 expression decreases reactive metabolite, methylglyoxal (MG), contributing to insulin resistance through glycation-linked unfolded protein formation and activation of the unfolded protein response (UPR). Herein, we report further analysis of study variables and follow-up reverse translational studies. With tRES-HESP treatment, plasma MG correlated negatively with peripheral blood mononuclear cell (PBMC) quinone reductase activity - a marker of Nrf2 activation (r = - 0.68, P&amp;lt;0.01). For change from baseline of PBMC gene expression with tRES-HESP treatment (assessed by the Nanostring method), change in area-under-the-curve plasma glucose (ΔAUGg) during oral glucose tolerance tests correlated negatively with change in Glo1 expression (r = - 0.56, P&amp;lt;0.05) and correlated positively with change in expression of thioredoxin interacting protein (TXNIP) (r = 0.59, P&amp;lt;0.05). In reverse translational studies, we found tRES-HESP decreased TXNIP expression by 39 ± 1% (P&amp;lt;0.001) in human hepatocyte-like HepG2 cells and 14 ± 3% in BJ fibroblasts in vitro (P&amp;lt;0.01). tRES-HESP may decrease TXNIP expression by decreasing the MG-driven UPR and by direct Nrf2-driven down regulation. TXNIP is a mediator of insulin resistance in liver and skeletal muscle and impaired pancreatic beta-cell insulin secretion. tRES-HESP is suitable for further evaluation for prevention and treatment of type 2 diabetes where decreased TXNIP may contribute to the therapeutic response. Disclosure M. Xue: None. M. Weickert: None. N. Rabbani: None. P. Thornalley: None. Funding Qatar Foundation; Qatar University

Repeated Dosing of Ketamine in the Forced Swim Test: Are Multiple Shots Better Than One?

The anesthetic drug ketamine has been successfully repurposed as an antidepressant in human subjects. This represents a breakthrough for clinical psychopharmacology, because unlike monoaminergic antidepressants, ketamine has rapid onset, including in Major Depressive Disorder (MDD) that is resistant to conventional pharmacotherapy. This rapid therapeutic onset suggests a unique mechanism of action, which continues to be investigated in reverse translational studies in rodents. A large fraction of rodent and human studies of ketamine have focused on the effects of only a single administration of ketamine, which presents a problem because MDD is typically a persistent illness that may require ongoing treatment with this drug to prevent relapse. Here we review behavioral studies in rodents that used repeated dosing of ketamine in the forced swim test (FST), with an eye toward eventual mechanistic studies. A subset of these studies carried out additional experiments with only a single injection of ketamine for comparison, and several studies used chronic psychosocial stress, where stress is a known causative factor in some cases of MDD. We find that repeated ketamine can in some cases paradoxically produce increases in immobility in the FST, especially at high doses such as 50 or 100 mg/kg. Several studies however provide evidence that repeated dosing is more effective than a single dose at decreasing immobility, including behavioral effects that last longer. Collectively, this growing literature suggests that repeated dosing of ketamine has prominent depression-related effects in rodents, and further investigation may help optimize the use of this drug in humans experiencing MDD.

White Button Mushroom Disrupts AR Signaling in Prostate Cancer: The Scientific Basis for a Diet-Based Chemoprevention

Abstract White button mushroom (WBM) (Agaricus bisporus) is a potential prostate cancer (PCa) chemo-preventive and therapeutic agent. Our clinical phase trial of WBM powder in patients with biochemically recurrent PCa indicated that WBM intake reduced the circulating levels of prostate-specific antigen (PSA), with minimal side effects [1]. We hypothesized that WBM exerts its effects on PCa through the androgen receptor (AR) signaling axis. We thus conducted the reverse translational study. Androgen-sensitive PCa cell lines (LNCaP and VCaP) and patient-derived-xenografts (PDX), of a prostate tumor (TM00298) were used. In both LNCaP and VCaP cells, western blots and qRT-PCR assays indicated that WBM extract (6~30 mg/mL) suppressed DHT-induced PSA expression and cell proliferation in a dose-dependent manner. Immunofluorescence on AR revealed that the nuclear localization of AR was reduced upon WBM extract treatment, which agreed with the results of a PSA promotor-luciferase assay, suggesting that WBM extract inhibited DHT-induced luciferase activity. RNA-Seq on WBM-treated LNCaP cells confirmed that WBM treatment suppressed androgen response pathways and cell-cycle control pathways. Our prostate cancer PDX showed that oral intake of WBM extract (200 mg/kg/day) significantly suppressed tumor growth, as well as decreased PSA levels in both tumors and serum. Both in vitro and in vivo studies suggested that chemical(s) in WBM extract behave as AR antagonist(s). We previously identified a conjugated linoleic acid isomer (CLA-9Z11E) as an active component in WBM extract. In the present study, we extended these findings by performing LanthaScreen™ TR-FRET AR Coactivator Interaction Assays for a direct interaction of CLA-9Z11E with AR. We report here that CLA-9Z11E exerts a strong antagonist potency against the recruitment of an AR coactivator peptide towards AR. The inhibitory effect of CLA-9Z11E (IC50: 350 nM) was nearly two times stronger than the known AR antagonist, cyproterone acetate (IC50: 672 nM). The information gained from this study improves the overall understanding of how WBM may contribute to the prevention and treatment of PCa. It also serves as an important, scientific basis for developing diet-based chemoprevention and integrative therapeutic strategies for prostate cancer (supported by NIH R01 CA227230). Reference: [1] Twardowski P, et al. A phase I trial of mushroom powder in patients with biochemically recurrent prostate cancer: Roles of cytokines and myeloid-derived suppressor cells for Agaricus bisporus-induced prostate-specific antigen responses. Cancer. 2015.121(17):2942-50.