Pharmacological Response Research Articles

Discovery and Optimization of Potent and Orally Available CTP Synthetase Inhibitors for Use in Treatment of Diseases Driven by Aberrant Immune Cell Proliferation.

Herein, we report the discovery of a first-in-class chemotype 2-(alkylsulfonamido)thiazol-4-yl)acetamides that act as pan-selective inhibitors of cytidine 5'-triphosphate synthetase (CTPS1/2), critical enzymes in the de novo pyrimidine synthesis pathway. Weak inhibitors identified from a high-throughput screening of 240K compounds have been optimized to a potent, orally active agent, compound 27, which has shown significant pharmacological responses at 10 mg/kg dose BID in a well-established animal model of inflammation.

Transmucosal delivery of the medical Cannabis oil via a nanoemulsion formulation

Nowadays, oral oil extracts of Cannabis represent the most relevant galenic plant-derived formulations employed in clinics. Nevertheless, the low and variable Δ9-tetrahydrocannabinol (THC) oral bioavailability due to hepatic first-pass metabolism and low solubility in biological fluids makes Cannabis pharmacological response highly unpredictable. In this scenario, new formulation strategies appear crucial to improve cannabinoids bioavailability and stability, administration comfort and patient compliance. Here, we present a nanoemulsion (NE) for the buccal administration of olive oil extract from a Cannabis sativa L. variety (Bedrocan®). A preliminary study was carried out on olive oil-based NE to identify the optimal formulation conditions to achieve a stable system. The Bedrocan®-loaded NE were then prepared and characterized for size, polydispersity index, stability upon delivery by the common buccal nebulizers.THC content and release in simulated buccal fluids and permeation studies across porcine buccal mucosa. The Bedrocan® NEs ensured THC stability and solubility in the buccal medium as compared with the Cannabis oil extract. Furthermore, the nanoemulsification process led to a THC diffusion and absorption on buccal mucosa 20-folds and 2-folds higher than olive oil extract, respectively. Overall, the results suggest that Bedrocan® NE represents a novel formulation strategy for the buccal administration of Cannabis extracts that can overcome the limits associated with conventional oily formulations.

Factors in the development of refractory primary open-angle glaucoma (part 1)

Primary open-angle glaucoma (POAG) is one of the most significant medico-social problems in the modern society. The development of its refractoriness aggravates the pathological process and inevitably leads to blindness. Despite the fact that this form accounts for up to 80% of all glaucoma cases, the problem of POAG refractoriness has not been considered in sufficient detail in either Russian or foreign sources. The article presents an overview of the main ophthalmic factors that contribute to changes in various structures of the eyeball and accelerate the formation of refractory forms of POAG. One of the reasons for that is a genetic predisposition to the development of POAG. It is worth noting that in 60% of patients with a family history of glaucoma, the risk of developing POAG increases by 10 times (among the first-degree relatives).Additional factors in the development of the refractory form of POAG are the reactive syndrome and during laser surgery. It is important to emphasize that the risk of development and progression of POAG in patients with a hereditary predisposition is much higher, while indicating a number of therapeutic measures may lead to drug resistance. Genotyping is a promising scientific and practical direction of research, allowing prediction of the pharmacological response to a particular drug and individual selection of the appropriate therapy according to the patient's genotype. This approach could help prevent a number of complications and improve the accuracy of disease prognosis.

Computational analysis of 5-fluorouracil anti-tumor activity in colon cancer using a mechanistic pharmacokinetic/pharmacodynamic model.

5-Fluorouracil (5-FU) is a standard chemotherapeutic agent to treat solid cancers such as breast, colon, head, and neck. Computational modeling plays an essential role in predicting the outcome of chemotherapy and developing optimal dosing strategies. We developed an integrated mechanistic pharmacokinetics/pharmacodynamics (PK/PD) model examining the influence of 5-FU, as an S-phase specific double-strand break (DSB)-inducing agent, on tumor proliferation. The proposed mechanistic PK/PD model simulates the dynamics of critical intermediate components and provides the accurate tumor response prediction. The integrated model is composed of PK, cellular, and tumor growth inhibition (TGI) sub-models, quantitatively capturing the essential drug-related physiological processes. In the cellular model, thymidylate synthase (TS) inhibition, resultant deoxynucleoside triphosphate (dNTP) pool imbalance, and DSB induction are considered, as well as 5-FU incorporation into RNA and DNA. The amount of 5-FU anabolites and DSBs were modeled to drive the kinetics of the pharmacological tumor response. Model parameters were estimated by fitting to literature data. Our simulation results successfully describe the kinetics of the intermediates regulating the 5-FU cytotoxic events and the pattern of tumor suppression. The comprehensive model simulated the tumor volume change under various dose regimens, and its generalizability was attested by comparing it with literature data. The potential causes of the tumor resistance to 5-FU are also investigated through Monte Carlo analysis. The simulation of various dosage regimens helps quantify the relationship between treatment protocols and chemotherapy potency, which will lead to the development of efficacy optimization.

Proposal of International Council for Harmonization (ICH) Guideline for the Approval of Biosimilars

ABSTRACT Objectives Since the initial release of biosimilars 18 years ago, regulations for their licensing have changed considerably; however, there is no global consensus on these regulations. Establishing harmonized regulatory guidelines for the approval of biosimilars with support from the ICH, an independent, non-profit association under Swiss law, will significantly enhance the affordability of biological drugs. Methods Regulatory guidelines from the Food and Drug Administration (FDA), European Medicines Agency (EMA), Medicines and Healthcare products Regulatory Agency (MHRA), and World Health Organization (WHO) were analyzed for historical changes and elements critical to the safety and efficacy of biosimilars. Results Analysis of all EMA and FDA filings show that none of the animal testing and clinical efficacy testing failed because animals do not have the required receptors to initiate pharmacologic responses, and efficacy studies cannot be statistically powered to conclude any results. New analytical technologies will enable good biosimilarity determination, avoiding both tests. Conclusion Scientifically based ICH guidelines that remove redundant studies will reduce development costs, improve safety, and allow global drug distribution based on single compliance. These guidelines are particularly necessary for emerging countries lacking the expertise and resources to evaluate biosimilar filings.

Psychiatric comorbidities among adult patients with disorders of gut-brain interaction: Prevalence and relationships to treatment outcomes.

Little is known about the impact of psychiatric comorbidity on pharmacologic treatment outcomes, including neuromodulators (medications targeting the gut-brain axis), among adult patients with disorders of gut-brain interaction (DGBI). Accordingly, we aimed to examine associations between psychiatric comorbidity and DGBI pharmacologic treatment outcomes. In a retrospective study of consecutively referred new patients (N=410; ages 18-90; 73% female) to a tertiary neurogastroenterology clinic in 2016 with follow-up through 2018, relationships between psychiatric illness (any psychiatric illness, anxiety disorders, depressive disorders) and pharmacologic treatment selection (any medication, neuromodulating medication) and treatment outcomes, respectively, were examined using multivariable logistic regression, adjusting for demographics, gastrointestinal (GI) diagnoses, and pre-existing neuromodulator use. Anxiety disorders (35%) were the most common psychiatric comorbidity, followed by depressive disorders (29%). Patients with anxiety disorders were more likely to be prescribed a neuromodulator by their gastroenterologist (OR=1.72 [95% CI 1.10-2.75]) yet less likely to respond to neuromodulators (OR=0.43 [0.21-0.90]) or any GI medication (OR=0.24 [0.12-0.50]) in fully adjusted analyses. In contrast, depressive disorders were not associated with neuromodulator prescription or response. Anxiety disorders are common among patients with DGBI and significantly reduce the likelihood of GI pharmacologic treatment response to any medication prescribed, including neuromodulators.

Solubility Enhancement of Poorly Soluble Drug by using different Techniques

Solubility is the process of a solid dissolving in a liquid phase, resulting in a homogeneous system. Solubility is a crucial characteristic for achieving the appropriate drug concentration in the systemic circulation and demonstrating pharmacological response. Drugs that are poorly water soluble require substantial dosages to achieve therapeutic plasma concentrations following oral administration. Low aqueous solubility is a major issue in the development of novel chemical entities' formulations. Any medicine that needs to be absorbed must be in the form of an aqueous solution at the absorption site. For liquid medicinal compositions, water is the preferred solvent. The majority of medications are weakly acidic and basic, with low water solubility. Micronization, chemical modification, pH adjustment, solid dispersion, complexation, cosolvency, micellar solubilization, hydrotropy, and other procedures are used to increase the solubility of weakly water-soluble pharmaceuticals. The goal of this review paper is to present solubilization approaches for achieving optimal absorption and increased bioavailability.

Physiology of chemokines in the cancer microenvironment.

Chemokines are chemotactic cytokines whose canonical functions govern movement of receptor-expressing cells along chemical gradients. Chemokines are a physiological system that is finely tuned by ligand and receptor expression, ligand or receptor oligomerization, redundancy, expression of atypical receptors, and non-GPCR binding partners that cumulatively influence discrete pharmacological signaling responses and cellular functions. In cancer, chemokines play paradoxical roles in both the directed emigration of metastatic, receptor-expressing cancer cells out of the tumor as well as immigration of tumor-infiltrating immune cells that culminate in a tumor-unique immune microenvironment. In the age of precision oncology, strategies to effectively harness the power of immunotherapy requires consideration of chemokine gradients within the unique spatial topography and temporal influences with heterogeneous tumors. In this article, we review current literature on the diversity of chemokine ligands and their cellular receptors that detect and process chemotactic gradients and illustrate how differences between ligand recognition and receptor activation influence the signaling machinery that drives cellular movement into and out of the tumor microenvironment. Facets of chemokine physiology across discrete cancer immune phenotypes are contrasted to existing chemokine-centered therapies in cancer.

Familial risk of psychosis in obsessive-compulsive disorder: Impact on clinical characteristics, comorbidity and treatment response

BackgroundFamily studies in obsessive-compulsive disorder (OCD) indicate higher rates of psychosis among their first-degree relatives (FDRs). However, the etiological and clinical relationships between the two disorders remain unclear. We compared the clinical characteristics and pharmacological treatment response in patients diagnosed with OCD with a family history of psychosis (OCD-FHP), with a family history of OCD (OCD-FHO) and those with sporadic OCD (OCD-S). MethodsA total of 226 patients who met DSM-IV criteria for OCD (OCD-FHP = 59, OCD-FHO = 112, OCD-S = 55) were included for analysis. All patients were evaluated using the Mini International Neuropsychiatric Interview (MINI 6.0.0), Yale-Brown Obsessive-Compulsive Scale (YBOCS), and the Family Interview for Genetic Studies (FIGS). Treatment response was characterized over naturalistic follow-up. ResultsThe three groups did not differ across any demographic or clinical variables other than treatment response. Patients in the OCD-FHP group were found to have received a greater number of trials with serotonin reuptake inhibitors (SRI) [F (2,223) = 7.99, p < 0.001], were more likely to have failed ≥2 trials of SRIs (χ2 = 8.45, p = 0.014), and less likely to have attained remission (χ2 = 6.57, p = 0.037) ConclusionsWe observed that having a relative with psychosis may predispose to treatment resistance in OCD. Further research on the influence of genetic liability to psychosis on treatment response in OCD may offer novel translational leads.

A novel twelve-gene signature to predict neoadjuvant chemotherapy response and prognosis in breast cancer.

Accurate evaluation of the response to neoadjuvant chemotherapy (NAC) provides important information about systemic therapies for breast cancer, which implies pharmacologicalresponse, prognosis, and guide further therapy. Gene profiles overcome the shortcomings of the relatively limited detection indicators of the classical pathological evaluation criteria and the subjectivity of observation, but are complicated and expensive. Therefore, it is essential to develop a more accurate, repeatable, and economical evaluation approach for neoadjuvant chemotherapy responses. We analyzed the transcriptional profiles of chemo-resistant breast cancer cell lines and tumors of chemo-resistant breast cancer patients in the GSE25066 dataset. We preliminarily screened out common significantly differentially expressed genes and constructed a NAC response risk model using LASSO regression and univariate and multivariate analyses. The differences in bioinformatic features of tumor cells, immune characteristics, and prognosis were compared between high and low-risk group. The potential drugs that could reverse chemotherapy resistance in breast cancer were screened by the CMap database. Thirty-six genes were commonly up/down-regulated in both NAC chemo-resistant tumors and cells compared to the sensitive tumors and wild-type cells. Through LASSO regression, we obtained a risk model composed of 12 genes. The risk model divided patients into high and low-risk groups. Univariate and multivariate Cox regression analyses suggested that the risk score is an independent prognostic factor for evaluating NAC response in breast cancer. Tumors in risk groups exhibited significant differences in molecular biological characteristics, tumor-infiltrating lymphocytes, and immunosuppressive molecule expression. Our results suggested that the risk score was also a good prognostic factor for breast cancer. Finally, we screened potential drugs that could reverse chemotherapy resistance in breast cancer. A novel 12 gene-signature could be used to predict NAC response and predict prognosis in breast cancer.

The Development of a PBPK Model for Atomoxetine Using Levels in Plasma, Saliva and Brain Extracellular Fluid in Patients with Normal and Deteriorated Kidney Function.

Atomoxetine is a treatment for attention-deficit hyperactivity disorder. It inhibits Norepinephrine Transporters (NET) in the brain. Renal impairment can reduce hepatic CYP2D6 activity and atomoxetine elimination which may increase its body exposure. Atomoxetine can be secreted in saliva. The objective of this work was to test the hypothesis that atomoxetine saliva levels (sATX) can be used to predict ATX brain Extracellular Fluid (bECF) levels and their pharmacological effects in healthy subjects and those with End-Stage Renal Disease (ESRD). The pharmacokinetics of atomoxetine after intravenous administration to rats with chemically induced acute and chronic renal impairments were investigated. A physiologically-based pharmacokinetic (PBPK) model was built and verified in rats using previously published measured atomoxetine levels in plasma and brain tissue. The rat PBPK model was then scaled to humans and verified using published measured atomoxetine levels in plasma, saliva, and bECF. The rat PBPK model predicted the observed reduced atomoxetine clearance due to renal impairment in rats. The PBPK model predicted atomoxetine exposure in human plasma, sATX and bECF. Additionally, it predicted that ATX bECF levels needed to inhibit NET are achieved at 80 mg dose. In ESRD patients, the developed PBPK model predicted that the previously reported 65% increase in plasma exposure in these patients can be associated with a 63% increase in bECF. The PBPK simulations showed that there is a significant correlation between sATX and bECF in human. Saliva levels can be used to predict atomoxetine pharmacological response.

PO252 / #911 THE IMPORTANCE OF BIOMARKERS IN THE NON-INVASIVE VAGUS NERVE STIMULATION FOR THERAPEUTIC PURPOSES: E-POSTER VIEWING

Stimulation of the vagus nerve is currently a therapeutic reality in pathologies such as epilepsy, depression, headaches, gastroesophageal disorders and tinnitus... The importance of recognizing activation biomarkers in neurostimulation therapy (electroceutics) is increasing. The National Institutes Health (NIH) established in 2000 the definition of biomarker as those “biological, biochemical, anthropometric, physiological characteristics, etc., objectively measurable, capable of identifying physiological or pathological processes, or a pharmacological response to a therapeutic intervention”(1).

Effect of Antiplatelet Activity of Aspirin on Chronic Kidney Disease

Background and aim: Individual differences in the pharmacological responsiveness to aspirin are considerable. A higher likelihood of ischemic episodes is linked to aspirin's inadequate antiplatelet actions. It has been hypothesized that chronic kidney disease (CKD) influences the pharmacologic reaction to antiplatelet drugs. The elevated chance of fatality and cardiovascular problems in CKD sufferers has already been found to be largely explained by high on-treatment platelets response (HTPR) to clopidogrel. This research aimed to examine at how aspirin affects blood clotting in CKD sufferers. Method: We performed cross-sectional research on 120 people in Medical center of Lahore taking aspirin regularly. The thromboxane production generated by AAI was used to gauge the pharmacologic reaction to aspirin. Result: Individuals with poor kidney functioning were more likely to experience HTPR to aspirin (48% vs. 23%; odds ratio, 3.17; 95% confidence interval [CI], 1.35-7.42; P = 0.007). When compared to individuals with healthy or mildly diminished kidney performance, those with mild or serious CKD had lower pharmacokinetic responsiveness to aspirin (91; interquartile ranged [IQR], 281 ng mL-1; differential in medians, 56; CI, 6-110 ng mL-1; P = 0.012). Remaining thromboxane production and glomerular filtration rate were both associated by bivariate Pearson correlation analysis (R = 0.312; R2 = 0.093; P = 0.001). Individuals with CKD were usually old age and being women. Women and aged were not significant predictors of the connection, according to multivariate linear regression analyses (R = 0.313; R2 = 0.081; P = 0.002 and P = 0.006) respectively. Conclusion: Aspirin pharmacological reaction is associated with kidney function. Individuals who have CKD are more likely to have aspirin's antiplatelet action compromised. To evaluate the therapeutic significance of this data and look into the best antithrombotic strategy for CKD individuals, additional research is required. Keywords: Chronic kidney disease, platelets, treatment, aspirin, cardiovascular

Role of Intracellular and Extracellular Annexin A1 in MIA PaCa-2 Spheroids Formation and Drug Sensitivity.

Simple SummaryIn order to improve the investigation of pancreatic cancer (PC), often supported through analyzes two-dimensional (2D) cell monolayers, we proposed to create a spheroid-based in vitro three-dimensional (3D) model using wild-type (WT) and ANXA1 knock-out (KO) MIA PaCa-2 PC cells. However, the production of spheroids still represents a technical challenge. Here, we have developed a protocol to obtain well-organized spheroids and have proved that Annexin A1 (ANXA1) affects the spheroid formation, because the WT cells have a greater ability to form this 3D model when compared to the ANXA1 KO examples. We also investigated how ANXA1 action could influence the PC pharmacological response both in basal conditions and by mimicking a tumor system through the addition of autocrine EVs. ANXA1, via EVs, significantly improves the formation, the stability and the drug resistance of this model, particularly compared to the ANXA1 KO one, which shows a structural instability and a greater drug sensitivity.Among solid tumors, pancreatic cancer (PC) remains a leading cause of death. In PC, the protein ANXA1 has been identified as an oncogenic factor acting in an autocrine/paracrine way, and also as a component of tumor-deriving extracellular vesicles. Here, we proposed the experimental protocol to obtain spheroids from the two cell lines, wild-type (WT) and Annexin A1 (ANXA1) knock-out (KO) MIA PaCa-2, this last previously obtained through CRISPR/Cas9 genome editing system. The use of three-dimensional (3D) models, like spheroids, can be useful to mimic tumor characteristics and for preclinical chemo-sensitivity studies. By using PC spheroids, we have assessed the activity of intracellular and extracellular ANXA1. Indeed, we have proved that the intracellular protein influences in vitro tumor development and growth by spheroids analysis, in addition to defining the modification about cell protein pattern in ANXA1 KO model compared to the WT one. Moreover, we have tested the response to FOLFIRINOX chemotherapy regimen whose cytostatic effect appeared notably increased in ANXA1 KO spheroids. Additionally, this study has highlighted that the extracellular ANXA1 action is strengthened through the EVs supporting spheroids growth and resistance to drug treatment, mainly affecting tumor progression. Thus, our data interestingly suggest the relevance of ANXA1 as a potential therapeutic PC marker.

Human-induced pluripotent stem cell-atrial-specific cardiomyocytes and atrial fibrillation.

Patient-specific human-induced pluripotent stem cell-derived atrial cardiomyocytes (hiPSC-aCMs) may be produced, genome-edited, and differentiated into multiple cell types for regenerative medicine, disease modeling, drug testing, toxicity screening, and three-dimensional tissue fabrication. There is presently no complete model of atrial fibrillation (AF) available for studying human pharmacological responses and evaluating the toxicity of potential medication candidates. It has been demonstrated that hiPSC-aCMs can replicate the electrophysiological disease phenotype and genotype of AF. The hiPSC-aCMs, however, are immature and do not reflect the maturity of aCMs in the native myocardium. Numerous laboratories utilize a variety of methodologies and procedures to improve and promote aCM maturation, including electrical stimulation, culture duration, biophysical signals, and changes in metabolic variables. This review covers the current methods being explored for use in the maturation of patient-specific hiPSC-aCMs and their application towards a personalized approach to the pharmacologic therapy of AF.

Generation of a TPH2-EGFP reporter cell line for purification and monitoring of human serotonin neurons in vitro and in vivo

SummaryGeneration of serotonin neurons (SNs) from human pluripotent stem cells (hPSCs) provides a promising platform to explore the mechanisms of serotonin-associated neuropsychiatric disorders. However, neural differentiation always yields heterogeneous cell populations, making it difficult to identify and purify SNs in vitro or track them in vivo following transplantation. Herein, we generated a TPH2-EGFP reporter hPSC line with insertion of EGFP into the endogenous tryptophan hydroxylase 2 (TPH2) locus using CRISPR-Cas9-mediated gene editing technology. This TPH2-reporter, which faithfully indicated TPH2 expression during differentiation, enabled us to obtain purified SNs for subsequent transcriptional analysis and study of pharmacological responses to antidepressants. In addition, the reporter system showed strong EGFP expression to indicate SNs, which enabled us to explore in vitro and ex vivo electrophysiological properties of SNs. In conclusion, this TPH2-EGFP reporter cell line might be of great significance for studies on human SN-related development and differentiation, drug screening, disease modeling, and cell replacement therapies.

Pharmacognostical, Phytochemical and Pharmacological Evaluation of Mallotus philippensis

The present study was aimed at Pharmacognostical, Phytochemical and Pharmacological evaluation of the plant Mallotus Philippinensis family- Euphorbiaceae. Pharmacognostical investigation were carried out by performing organoleptic, microscopical and physicochemical evaluation i.e. ash values, extractive values, moisture content, swelling index, foaming index and foreign matter. The obtained results showed that the moisture content was found to be 1.74%. Similarly swelling index was recorded to be (0.8cm). Foaming index (less than 100). Phytochemical investigation included successive soxhlet extraction, the obtained extractive values were 4.9%, 5.45%, 9.77%, 8.75%, and 4.4% for petroleum ether, chloroform, ethyl acetate, ethanol and distilled water respectively. The preliminary qualitative phytochemical screening revealed the presence of carbohydrates, phenolic compounds, alkaloids, glycosides, flavonoids, steroids, fixed oils and fats. Pharmacological investigation included the Anthelmentic activity by using the Mallotus Philippinensis plant ethyl acetate extract. The 200 mg/kg dose to be found positively in Pharmacological responses.&#x0D; Keywords: Mallotus Philippinensis, Anthelmentic activity, Phytochemical investigation, Pharmacological investigation, Pharmacognostical investigation

Circulating MicroRNAs Expression Profile in Lung Inflammation: A Preliminary Study.

Background: Bronchial asthma is an inflammatory airway disease with an ever-increasing incidence. Therefore, innovative management strategies are urgently needed. MicroRNAs are small molecules that play a key role in lungs cellular functions and are involved in chronic inflammatory diseases, such as bronchial asthma. This study aims to compare microRNA serum expression between subjects with asthma, obesity, the most common co-morbidity in asthma, and healthy controls to obtain a specific expression profile specifically related to lung inflammation. Methods: We collected serum samples from a prospective cohort of 25 sex-matched subjects to determine circulating miRNAs through a quantitative RT-PCR. Moreover, we performed an in silico prediction of microRNA target genes linked to lung inflammation. Results: Asthmatic patients had a significant lower expression of hsa-miR-34a-5p, 181a-5p and 146a-5p compared to both obese and healthy ones suggesting microRNAs’ specific involvement in the regulation of lungs inflammatory response. Indeed, using in silico analysis, we identified microRNAs novel target genes as GATA family, linked to the inflammatory-related pathway. Conclusions: This study identifies a novel circulating miRNAs expression profile with promising potentials for asthma clinical evaluations and management. Further and larger investigations will be needed to confirm the potential role of microRNA as a clinical marker of bronchial asthma and eventually of pharmacological treatment response.

Taste-immune associative learning amplifies immunopharmacological effects and attenuates disease progression in a rat glioblastoma model

Mechanistic target of rapamycin (mTOR)-signaling is one key driver of glioblastoma (GBM), facilitating tumor growth by promoting the shift to an anti-inflammatory, pro-cancerogenic microenvironment. Even though mTOR inhibitors such as rapamycin (RAPA) have been shown to interfere with GBM disease progression, frequently chaperoned toxic drug side effects urge the need for developing alternative or supportive treatment strategies. Importantly, previous work document that taste-immune associative learning with RAPA may be utilized to induce learned pharmacological placebo responses in the immune system. Against this background, the current study aimed at investigating the potential efficacy of a taste-immune associative learning protocol with RAPA in a syngeneic GBM rat model. Following repeated pairings of a novel gustatory stimulus with injections of RAPA, learned immune-pharmacological effects could be retrieved in GBM-bearing animals when re-exposed to the gustatory stimulus together with administering 10 % amount of the initial drug dose (0.5 mg/kg). These inhibitory effects on tumor growth were accompanied by an up-regulation of central and peripheral pro-inflammatory markers, suggesting that taste-immune associative learning with RAPA promoted the development of a pro-inflammatory anti-tumor microenvironment that attenuated GBM tumor growth to an almost identical outcome as obtained after 100 % (5 mg/kg) RAPA treatment. Together, our results confirm the applicability of taste-immune associative learning with RAPA in animal disease models where mTOR overactivation is one key driver. This proof-of-concept study may also be taken as a role model for implementing learning protocols as alternative or supportive treatment strategy in clinical settings, allowing the reduction of required drug doses and side effects without losing treatment efficacy.

Utility of constraints reflecting system stability on analyses for biological models.

Simulating complex biological models consisting of multiple ordinary differential equations can aid in the prediction of the pharmacological/biological responses; however, they are often hampered by the availability of reliable kinetic parameters. In the present study, we aimed to discover the properties of behaviors without determining an optimal combination of kinetic parameter values (parameter set). The key idea was to collect as many parameter sets as possible. Given that many systems are biologically stable and resilient (BSR), we focused on the dynamics around the steady state and formulated objective functions for BSR by partial linear approximation of the focused region. Using the objective functions and modified global cluster Newton method, we developed an algorithm for a thorough exploration of the allowable parameter space for biological systems (TEAPS). We first applied TEAPS to the NF-κB signaling model. This system shows a damped oscillation after stimulation and seems to fit the BSR constraint. By applying TEAPS, we found several directions in parameter space which stringently determines the BSR property. In such directions, the experimentally fitted parameter values were included in the range of the obtained parameter sets. The arachidonic acid metabolic pathway model was used as a model related to pharmacological responses. The pharmacological effects of nonsteroidal anti-inflammatory drugs were simulated using the parameter sets obtained by TEAPS. The structural properties of the system were partly extracted by analyzing the distribution of the obtained parameter sets. In addition, the simulations showed inter-drug differences in prostacyclin to thromboxane A2 ratio such that aspirin treatment tends to increase the ratio, while rofecoxib treatment tends to decrease it. These trends are comparable to the clinical observations. These results on real biological models suggest that the parameter sets satisfying the BSR condition can help in finding biologically plausible parameter sets and understanding the properties of biological systems.