Drug Levels In Brain Research Articles

ATP-binding cassette transporter inhibitor potency and substrate drug affinity are critical determinants of successful drug delivery enhancement to the brain

BackgroundPharmacotherapy for brain diseases is severely compromised by the blood-brain barrier (BBB). ABCB1 and ABCG2 are drug transporters that restrict drug entry into the brain and their inhibition can be used as a strategy to boost drug delivery and pharmacotherapy for brain diseases.MethodsWe employed elacridar and tariquidar in mice to explore the conditions for effective inhibition at the BBB. Abcg2;Abcb1a/b knockout (KO), Abcb1a/b KO, Abcg2 KO and wild-type (WT) mice received a 3 h i.p. infusion of a cocktail of 8 typical substrate drugs in combination with elacridar or tariquidar at a range of doses. Abcg2;Abcb1a/b KO mice were used as the reference for complete inhibition, while single KO mice were used to assess the potency to inhibit the remaining transporter. Brain and plasma drug levels were measured by LC-MS/MS.ResultsComplete inhibition of ABCB1 at the BBB is achieved when the elacridar plasma level reaches 1200 nM, whereas tariquidar requires at least 4000 nM. Inhibition of ABCG2 is more difficult. Elacridar inhibits ABCG2-mediated efflux of weak but not strong ABCG2 substrates. Strikingly, tariquidar does not enhance the brain uptake of any ABCG2-subtrate drug. Similarly, elacridar, but not tariquidar, was able to inhibit its own brain efflux in ABCG2-proficient mice. The plasma protein binding of elacridar and tariquidar was very high but similar in mouse and human plasma, facilitating the translation of mouse data to humans.ConclusionsThis work shows that elacridar is an effective pharmacokinetic-enhancer for the brain delivery of ABCB1 and weaker ABCG2 substrate drugs when a plasma concentration of 1200 nM is exceeded.

Evaluation of bumetanide as a potential therapeutic agent for Alzheimer's disease.

Therapeutics discovery and development for Alzheimer's disease (AD) has been an area of intense research to alleviate memory loss and the underlying pathogenic processes. Recent drug discovery approaches have utilized in silico computational strategies for drug candidate selection which has opened the door to repurposing drugs for AD. Computational analysis of gene expression signatures of patients stratified by the APOE4 risk allele of AD led to the discovery of the FDA-approved drug bumetanide as a top candidate agent that reverses APOE4 transcriptomic brain signatures and improves memory deficits in APOE4 animal models of AD. Bumetanide is a loop diuretic which inhibits the kidney Na+-K+-2Cl- cotransporter isoform, NKCC2, for the treatment of hypertension and edema in cardiovascular, liver, and renal disease. Electronic health record data revealed that patients exposed to bumetanide have lower incidences of AD by 35%-70%. In the brain, bumetanide has been proposed to antagonize the NKCC1 isoform which mediates cellular uptake of chloride ions. Blocking neuronal NKCC1 leads to a decrease in intracellular chloride and thus promotes GABAergic receptor mediated hyperpolarization, which may ameliorate disease conditions associated with GABAergic-mediated depolarization. NKCC1 is expressed in neurons and in all brain cells including glia (oligodendrocytes, microglia, and astrocytes) and the vasculature. In consideration of bumetanide as a repurposed drug for AD, this review evaluates its pharmaceutical properties with respect to its estimated brain levels across doses that can improve neurologic disease deficits of animal models to distinguish between NKCC1 and non-NKCC1 mechanisms. The available data indicate that bumetanide efficacy may occur at brain drug levels that are below those required for inhibition of the NKCC1 transporter which implicates non-NKCC1 brain mechansims for improvement of brain dysfunctions and memory deficits. Alternatively, peripheral bumetanide mechanisms may involve cells outside the central nervous system (e.g., in epithelia and the immune system). Clinical bumetanide doses for improved neurological deficits are reviewed. Regardless of mechanism, the efficacy of bumetanide to improve memory deficits in the APOE4 model of AD and its potential to reduce the incidence of AD provide support for clinical investigation of bumetanide as a repurposed AD therapeutic agent.

Abstract WP219: Repurposing The K Ca 3.1 Inhibitor Senicapoc For Treatment Of Acute Ischemic Stroke

Background: Acute ischemic stroke (AIS) is a leading cause of death and long-term disability. Both microglia (MG) and infiltrating macrophages (MP) are critical effector cell types in ischemic brain injury and recovery. K Ca 3.1 is a calcium-activated potassium channel that is upregulated in reactive MG and MP. Studies using either genetic deletion or pharmacological inhibition of K Ca 3.1 demonstrated that this channel is critical for pro-inflammatory activation of MG/MP as well as exacerbation of stroke pathophysiology. Senicapoc (ICA-17043) is a K Ca 3.1-specific inhibitor that has been used in human clinical trials for non-neurological indications (sickle cell anemia, asthma) and was proven safe. Here we evaluate the potential for repurposing senicapoc for AIS. Methods: Young adult male and female mice underwent 60 min middle cerebral artery occlusion (MCAO)/reperfusion. MCAO was monitored by laser doppler. Senicapoc’s pharmacokinetic (PK) profile was determined using high performance liquid chromatography - mass spectroscopy. Drug levels in plasma and brain were quantified at multiple time points post administration. Effects of senicapoc on post-stroke release of cytokines/chemokines was determined by multiplex ELISA. Inflammatory infiltrates were quantified with flow cytometry. Efficacy studies included: (i) infarct volume (MRI T2), white matter integrity (DTI) and longitudinal neurobehavioral outcomes (NBO). In-vitro chromogenic assay was used to assess senicapoc’s effect on proteolytic activity of tissue plasminogen activator (tPA). Results: Administration of senicapoc (40 mg/kg, i.p.) twice daily for seven days starting 12 h after MCAO resulted in ~55% reduction in infarct volume with corresponding improvements in NBO. Free senicapoc levels in brain ranged from 20 - 200 nM in stroked mice at 1, 4 & 12 hours following administration. All values exceeded senicapoc’s IC 50 (11 nM) for K Ca 3.1. Senicapoc, at up to 5 μM, had no effect on tPA’s proteolytic activity. Conclusions: We provide proof-of-concept data that senicapoc, administered in an extended temporal window, can markedly reduce infarct volume and improve NBO in a mouse model of AIS. Senicapoc demonstrated favorable PK and CNS penetration. It did not interfere with tPA.

Peptides as Pharmacological Carriers to the Brain: Promises, Shortcomings and Challenges.

Central nervous system (CNS) diseases are among the most difficult to treat, mainly because the vast majority of the drugs fail to cross the blood-brain barrier (BBB) or to reach the brain at concentrations adequate to exert a pharmacological activity. The obstacle posed by the BBB has led to the in-depth study of strategies allowing the brain delivery of CNS-active drugs. Among the most promising strategies is the use of peptides addressed to the BBB. Peptides are versatile molecules that can be used to decorate nanoparticles or can be conjugated to drugs, with either a stable link or as pro-drugs. They have been used to deliver to the brain both small molecules and proteins, with applications in diverse therapeutic areas such as brain cancers, neurodegenerative diseases and imaging. Peptides can be generally classified as receptor-targeted, recognizing membrane proteins expressed by the BBB microvessels (e.g., Angiopep2, CDX, and iRGD), "cell-penetrating peptides" (CPPs; e.g. TAT47-57, SynB1/3, and Penetratin), undergoing transcytosis through unspecific mechanisms, or those exploiting a mixed approach. The advantages of peptides have been extensively pointed out, but so far few studies have focused on the potential negative aspects. Indeed, despite having a generally good safety profile, some peptide conjugates may display toxicological characteristics distinct from those of the peptide itself, causing for instance antigenicity, cardiovascular alterations or hemolysis. Other shortcomings are the often brief lifetime in vivo, caused by the presence of peptidases, the vulnerability to endosomal/lysosomal degradation, and the frequently still insufficient attainable increase of brain drug levels, which remain below the therapeutically useful concentrations. The aim of this review is to analyze not only the successful and promising aspects of the use of peptides in brain targeting but also the problems posed by this strategy for drug delivery.

Nanocrystals as an effective strategy to improve Pomalidomide bioavailability in rodent

Pomalidomide (POM) is an FDA-approved immunomodulatory imide drug (IMiDs) an it is effectively used in the treatment of multiple myeloma. IMiDs are analogs of the drug thalidomide and they have been repurposed for the treatment of several diseases such as psoriatic arthritis and Kaposi Sarcoma. In recent years, IMiDs have been also evaluated as a new treatment for neurological disorders with an inflammatory and neuroinflammatory component. POM draws particular interest for its potent anti-TNF-α activity at significantly lower concentrations than the parent compound thalidomide. However, POM’s low water solubility underpins its low gastrointestinal permeability resulting in irregular and poor absorption. The purpose of this work was to prepare a POM nanocrystal-based formulation that could efficiently improve POM’s plasma and brain concentration after intraperitoneal injection. POM nanocrystals prepared as a nanosuspension by the media milling method showed a mean diameter of 219 nm and a polydispersity index of 0.21. POM’s nanocrystal solubility value (22.97 µg/mL) in phosphate buffer was about 1.58 folds higher than the POM raw powder. Finally, in vivo studies conducted in adult Male Sprague-Dawley rats indicated that POM nanocrystal ensured higher and longer-lasting drug levels in plasma and brain when compared with POM coarse suspension.

Abstract 6296: Preclinical evaluation of a panel of FGFR inhibitors for their normal brain and brain tumor distribution

Abstract Glioblastoma (GBM) is the most prevalent and lethal primary intrinsic brain tumor in adults with approximately 5-8% of GBM patients displaying fibroblast growth factor receptor (FGFR) gene mutations. This subset of GBM patients could potentially benefit from targeted FGFR inhibitor treatment, thus making it imperative to explore FGFR inhibitors as a therapeutic option. In this study, seven pan-FGFR inhibitors (infigratinib, AZD4547, debio1347, erdafitinib, LY2874455, pemigatinib, and TAS-120) were evaluated for their brain penetration ability employing cassette dosing strategy with an aim to identify the optimal candidates for brain tumor targeting. Cassette-dosing strategy was confirmed to be an acceptable approach for screening the panel of FGFR inhibitors by comparing pharmacokinetic parameters obtained from discrete- vs cassette-dosing studies. Subsequently, cassette-dosing strategy was utilized to investigate distributional kinetics of FGFR inhibitors in normal brain and brain tumor in wild-type and triple-knockout (Mdr1a/b-/-Bcrp1-/-) mice. The comparison of brain-to-plasma partition coefficients for total (Kp,brain) and unbound (Kp,brain) drug levels in wild-type and triple-knockout mice revealed the importance of major blood-brain barrier (BBB) efflux transporters in four out of seven FGFR inhibitors. The impact of BBB and efflux transporters were further evaluated in flank and orthotopic patient-derived xenograft mouse models of brain tumors. All FGFR inhibitors were shown to accumulate significantly in both flank and orthotopic brain tumors relative to normal brain. Furthermore, the levels in flank brain tumors were at least 10-fold higher as compared to the levels in orthotopic brain tumors with five FGFR inhibitors, indicating the critical role of efflux transporters and the impact of BBB disruption in the brain and brain tumor distribution. The predictive parameters of brain penetration calculated based on physico-chemical properties have been estimated and shown not to correlate with the experimentally obtained neuro-pharmacokinetic values. In summary, our study provides comprehensive evaluation of neuro-pharmacokinetic behavior of seven FGFR inhibitors and delivers rationale for selection of most optimal candidates for future investigation in brain tumor clinical trials. Citation Format: William Knight, Kamal Shaik, Tigran Margaryan, Shwetal Mehta, Nader Sanai, Artak Tovmasyan. Preclinical evaluation of a panel of FGFR inhibitors for their normal brain and brain tumor distribution [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6296.

Strategies to Improve Drug Strength in Nasal Preparations for Brain Delivery of Low Aqueous Solubility Drugs.

Intranasal administration is a promising route for brain drug delivery. However, it can be difficult to formulate drugs that have low water solubility into high strength intranasal solutions. Hence, the purpose of this work was to review the strategies that have been used to increase drug strength in intranasal liquid formulations. Three main groups of strategies are: the use of solubilizers (change in pH, complexation and the use cosolvents/surfactants); incorporation of the drugs into a carrier nanosystem; modifications of the molecules themselves (use of salts or hydrophilic prodrugs). The use of high amounts of cosolvents and/or surfactants and pH decrease below 4 usually lead to local adverse effects, such as nasal and upper respiratory tract irritation. Cyclodextrins and (many) different carrier nanosystems, on the other hand, could be safer for intranasal administration at reasonably high concentrations, depending on selected excipients and their dose. While added attributes such as enhanced permeation, sustained delivery, or increased direct brain transport could be achieved, a great effort of optimization will be required. On the other hand, hydrophilic prodrugs, whether co-administered with a converting enzyme or not, can be used at very high concentrations, and have resulted in a fast prodrug to parent drug conversion and led to high brain drug levels. Nevertheless, the choice of which strategy to use will always depend on the characteristics of the drug and must be a case-by-case approach.

1411. Noninvasive Assessment of Intralesional Antimicrobial Concentration-Time Profiles in Pulmonary and Central Nervous System Tuberculosis using Dynamic 18F-Pretomanid Positron Emission Tomography

1411. Noninvasive Assessment of Intralesional Antimicrobial Concentration-Time Profiles in Pulmonary and Central Nervous System Tuberculosis using Dynamic 18F-Pretomanid Positron Emission Tomography

DDRE-23. PRECLINICAL EVALUATION OF BRAIN PENETRATION PROPERTIES OF NEW FIRST-IN-CLASS LYSINE-SPECIFIC DEMETHYLASE 1 (LSD1) INHIBITOR, SP-2577

Abstract BACKGROUND Lysine specific demethylase 1 (LSD1) is a histone demethylase implicated in the maintenance of pluripotency and proliferation gene programs that give rise to a number of cancers. SP-2577 is a first-in-class selective and reversible inhibitor of LSD1. Here, we evaluated the ability of SP-2577 to cross blood-brain barrier in mouse brain. METHODS Fifteen BALB/c mice were treated with 50 mg/kg SP-2577 twice daily intraperitoneally for 4 days. At 2, 6, and 12 hours after the last treatment dose was delivered, plasma and brain samples were collected for analysis of SP-2577 bound and unbound fractions (5 mice/time point). An LC–MS/MS method was developed to measure the drug levels in mouse plasma and brain. The unbound fractions of SP-2577 in plasma and brain tissue were determined using equilibrium dialysis. RESULTS Total plasma levels of SP-2577 at the 2, 6, and 12-hour time points following the last dose were 1353 nM, 1209 nM, and 560 nM, accordingly. Total drug levels in brain measured at the same time points were 276 nM, 183 nM, and 168 nM. SP-2577 is highly bound to plasma proteins and brain tissue components with an average plasma unbound fraction value of 0.009. Unbound levels of SP-2577 were undetectable in brain tissue, potentially due to instability of the drug in brain homogenates. The total brain-to-plasma ratio (Kp) was determined as 0.032 (range, 0.027–0.046) in mice. CONCLUSION SP-2577 is well tolerated in mice and achieves reasonable total drug levels in mouse brain, yet is highly-bound to plasma proteins and brain components. Taken together, these data indicate that SP-2577 cannot reach pharmacologically-relevant drug concentrations across the mouse blood-brain barrier.

EXTH-80. PBI-200: IN VIVO EFFICACY OF A NOVEL, HIGHLY CNS-PENETRANT NEXT GENERATION TRK INHIBITOR

Abstract TRK kinases (TRK) are clinically validated targets for cancers harboring NTRK gene fusions. However, approved TRK inhibitors (TRKi) give rise to mutations reducing long-term clinical efficacy, and display limited CNS penetration that may impact their ability to address primary brain tumors (PBT), including high-grade gliomas. Second generation compounds in development address some resistance mutations, but are not sufficiently CNS penetrant. PBI-200 is a novel, orally active TRK inhibitor designed to overcome resistance, with high CNS penetration. In vitro assays demonstrate picomolar- to nanomolar potency against both wild-type TRK and the major acquired resistance mutations. A BaF3 xenograft model encoding the LMNA-NTRK1 gene fusion and G595R solvent front mutation showed superior efficacy with PBI-200 relative to first-generation compounds larotrectinib and entrectinib, and equivalent efficacy to the second-generation compound selitrectinib, in terms of tumor growth inhibition. Importantly, drug concentrations in the brains of PBI-200 treated mice showed an approximately 4-fold brain/plasma ratio (BPR). In contrast, BPR of other TRKi were < 0.17, indicating that only PBI-200 achieved sustained drug levels in brain. PBI-200 demonstrated statistically superior CNS efficacy and survival in a KM12-Luc intracranial murine model, as compared to other TRKi (chi-squared 45.6, p < 0.0001). By day 41, 50% of mice in the PBI-200 group remained alive, whereas mice in the other treatment groups had died on or before day 32. Of note, no gross or histopathological CNS adverse effects were observed in a 14d CNS Field Observation Battery study. Finally, PBI-200 was the most selective TRKi in a 125 kinase panel (ThermoFisher) with only 1 off-target kinase inhibited > 60% at 1 micromolar. Together, these data suggest that PBI-200 has potential as a best-in-class, highly CNS-penetrant next generation TRKi. A Phase 1/2 clinical trial evaluating PBI-200 in NTRK fusion-positive patients, including patients with PBT, is ongoing (NCT04901806).

EXTH-24. TUMOR PHARMACOKINETICS, PHARMACODYNAMICS AND RADIATION SENSITIZATION IN PATIENT-DERIVED XENOGRAFT MODELS OF GLIOBLASTOMA TREATED WITH THE SECOND-GENERATION HDAC INHIBITOR, QUISINOSTAT

Abstract BACKGROUND Class I Histone deacetylases (HDACs) are highly expressed in glioblastoma (GBM) and are considered promising therapeutic targets for cancer treatment. Quisinostat is a class I HDAC inhibitor with high specificity for HDAC1 and 2. In this study, we evaluated the pharmacokinetics (PK), pharmacodynamics (PD), and radiation sensitization properties of quisinostat in orthotopic and flank human glioma models of GBM. METHODS In vitro drug-uptake and PK-PD correlation experiments were conducted in patient-derived glioma stem cell (GSC) lines. Pilot PK studies were performed in mice to optimize the dosing regimen and administration route (subcutaneous (s.q.), intraperitoneal (i.p.) and oral gavage). Athymic mice with intracranial or flank tumors were treated with the most optimized treatment schedule to evaluate plasma PK profile and brain/tumor distribution of quisinostat. The drug levels in plasma, normal brain, and tumor-tissue were measured by liquid chromatography tandem mass spectrometry (LC-MS/MS). Unbound fractions were determined by equilibrium dialysis. Western blot analysis and immunocytochemistry were performed to assess changes in acetylated histone, Ki67 (proliferation) and cleaved caspase 3 (cell death) levels. Tumor-bearing mice were randomized to four cohorts – vehicle, radiotherapy, quisinostat monotherapy, and quisinostat plus radiotherapy. RESULTS Quisinostat exhibited potent growth inhibition in multiple GSC lines (IC50 ~60 nM), radiosensitization, increased histone acetylation, and decreased expression of key stem cell markers. Significant toxicity was observed at 10 mg/kg s.q. dosing while the same dosing via i.p. route was tolerated relatively well. The unbound concentration of quisinostat was 19 nmol/L, 0.03 nmol/kg and 0.03 nmol/kg in plasma, tumor, and brain tissue, respectively, at 2 hours post-dose. Pharmacodynamic changes were observed in flank but not in intracranial tumors. CONCLUSION Our data indicate low free quisinostat levels in the brain primarily due to drug instability.

Galactosylated nanoconstructs of Berberine with enhanced Biopharmaceutical and cognitive potential: A preclinical evidence in Alzheimer ‘s disease

Numerous oral treatment options have been reported for neurological diseases, particularly Alzheimer's disease (AD). Berberine (BRB), a promising phytoactive drug, has been successfully explored for effective therapeutic management of AD and other neurodegenerative disorders. However, it poses formidable challenges of exceedingly poor oral bioavailability of 0.68%, primarily owing to its poor intestinal permeation, extremely low aqueous solubility, and extensive P-glycoprotein efflux transport. The present studies, accordingly, were undertaken to develop the galactose-tethered nanostructured lipidic carriers of BRB for efficient brain uptake. NLCs were systematically developed and optimized using QbD paradigms, and thoroughly characterized for particle size, zeta potential, morphology, entrapment efficiency and in vitro drug release. U-87 cell lines studies indicated augmentation in cellular permeability and uptake of the galactosylated NLCs. Oral bioavailability was found to be highly significantly enhanced with over 8.21-folds and 3.56-folds (p < 0.001 each) improvement in the values of AUC48h and Cmax, respectively, along with substantial elevation in brain drug levels too (p < 0.05 to p < 0.001). Also, tmax was found to be notably increased by 1.41-folds (p < 0.05), indicating extended release characteristics of the galactosylated NLCs. Studies on brain homogenates exhibiting distinctly superior biochemical parameters, and behavioural studies demonstrating marked cognitive potential in β-amyloid treated rats further substantiated their promising utility in the management of AD. High degree of Level A linear correlations were established between fractions of drug dissolved (in vitro) and of drug absorbed (in vivo) BRB and its NLC formulations. Stability studies at 5 °C indicated robustness of galactosylated NLCs for 6 months. The current preclinical findings, accordingly, offer distinctive potential of the developed NLCs in enhanced drug efficacy, extended duration of action and effective therapeutic management of AD.

Nose-to-brain delivery of phenytoin and its hydrophilic prodrug fosphenytoin combined in a microemulsion - formulation development and in vivo pharmacokinetics.

Phenytoin is a low aqueous solubility antiepileptic drug, but its phosphate ester prodrug fosphenytoin is soluble, although less permeable. In a previous study, the intranasal administration of aqueous-based formulations of fosphenytoin led to high but delayed phenytoin bioavailability compared to the intravenous route. In this work, we hypothesized that formulating an association of the prodrug fosphenytoin and the drug phenytoin (the active and diffusible form), could result in a faster and/or more effective brain targeting. Hence, nano or microemulsions containing both active drug and prodrug were developed and characterized regarding viscosity, osmolality, pH, mean size and in vitro drug release. Then, in vivo pharmacokinetics of a selected microemulsion containing fosphenytoin and phenytoin was evaluated in mice following intranasal administration and compared with a similar microemulsion containing fosphenytoin only. Both microemulsions led to higher brain drug levels at short time points than previously developed simpler aqueous based fosphenytoin formulations, likely due to the microemulsion's permeation enhancing effect. In addition, having a small amount of phenytoin in the formulation led to an equivalent maximum brain drug concentration and an overall higher absolute bioavailability, with a prolonged drug exposure. Hence, it can be concluded that if there is a need for a fast and prolonged therapeutic effect, a drug/phosphate ester prodrug combination in a microemulsion is ideal, but if a fast effect is all that is needed, having the prodrug alone could be enough, while considering a formulation with permeation enhancing components.

QbD-steered development of mixed nanomicelles of galantamine: Demonstration of enhanced brain uptake, prolonged systemic retention and improved biopharmaceutical attributes

PurposeNumerous oral treatment options have been reported for neurological disorders, especially Alzheimer’s disease (AD). Galantamine (GAL) is one of such drugs duly approved for management of AD. However, it exhibits poor brain penetration, low intestinal permeation and requires frequent dosing in AD treatment. The present studies, accordingly, were undertaken to develop DSPE-PEG 2000-based micelles loaded with GAL for efficient brain uptake, improved and extended pharmacokinetics, along with reduced dosing regimen. MethodsMixed nanomicelles (MNMs) were systematically formulated using QbD approach, and characterized for morphology, entrapment efficiency andin vitrodrug release. ResultsStudies on CaCo-2 and neuronal U-87 cell lines exhibited substantial enhancement in the cellular permeability and uptake of the developed MNMs. Pharmacokinetic studies performed on rats showed significantly improved values of plasma AUC (i.e., 2.28-fold, p < 0.001), ostensibly due to bypassing of hepatic first-pass metabolism and improved intestinal permeability, together with significant rise in MRT (2.08-fold, p < 0.001) and tmax (4.80-fold; p < 0.001) values, indicating immense potential for prolonged drug residence in body.Besides, substantial elevation in brain drug levels, distinctly improved levels of biochemical parameters in brain homogenates and cognitive improvement in β-amyloid-treated rats, testify the superiority in MNMs in therapeutic management of AD. ConclusionsThe preclinical findings of the developed nanocarrier systems successfully demonstrate the notable potential of enhanced drug efficacy, extended duration of action and improved patient compliance.

Further characterization of the effect of the prototypical antidepressant imipramine on the microstructure of licking for sucrose.

We previously reported that treatment with the prototypical antidepressant imipramine induced a dose-dependent reduction of the ingestion of a 10% sucrose solution, due to reduction of the licking burst number, thus suggesting reduced motivation and/or increased satiation. Importantly, the experimental sessions were performed in an alternate order, either 1-h or 24-h after imipramine administration. The observation that imipramine effect was more pronounced in the "1-h after-treatment" sessions, i.e. at the time of the brain drug Cmax, led us to suggest that it was likely related to brain drug levels at testing time. However, such an experimental design does not allow to rule out the alternative possibility that the observed effect might be due to post-session administration, as previously observed with memantine. To determine whether imipramine-induced decrease of sucrose ingestion could be observed even in absence of post-session administration, we examined the effect of a daily 22 day treatment with imipramine (5, 10 and 20 mg/kg). In the first half of the treatment period all behavioural tests were performed 1-h after administration. In the second half of the treatment period, tests were performed alternatively either 1-h or 24-h after imipramine administration. The results confirm that imipramine reduces sucrose ingestion due to a reduction of the licking burst number. Most importantly, these results demonstrate that this effect does not require imipramine post-session administration, since it was present before the beginning of post-session administrations. This supports the interpretation of the reduction of sucrose ingestion as a consequence of reduced motivation and/or increased satiation. Thus, these findings, taken together with the results of our previous study, might be relevant in explaining the effects of imipramine in models of drug-seeking and in body weight gain reduction in rats, but not in accounting for the antidepressant therapeutic effect. At variance with the results of our previous study, an increase in burst size was present in the first half of the treatment period, which might be interpreted as a prohedonic effect and/or as a compensatory effect.

Microstructure analysis of sucrose ingestion in the course of chronic treatment with imipramine

The analysis of licking microstructure provides measures which might be interpreted in terms of psychological constructs, such as pleasure and motivation, relevant for the interpretation of the effects of antidepressant drugs. The aim of this study was to characterise the effect of the prototypical antidepressant imipramine on the microstructure of licking for a 10% sucrose solution. In particular, ten 30-min sessions were performed in the course of a daily 21 day treatment with imipramine – 5, 10 and 20 mg/kg/die administered intraperitoneally. To interpret drug effects in relation to the presumed concentration of imipramine and its active metabolite desipramine, the experimental sessions were performed in an alternate order either 1-h or 24-h after imipramine administration. In the sessions performed 1-h after drug administration, the results showed a dose-dependent reduction of sucrose ingestion, accounted for by a reduction of the licking burst number. Moreover, reduced intra-burst lick rate and increased latency to lick were observed with the highest doses. Imipramine effect in the sessions performed 24-h after drug administration was similar but less pronounced. These results are consistent with the hypothesis that the reduction of sucrose ingestion might be due to reduced motivation and/or to a potentiation of satiety signals. These effects appear to be related, at least in part, to brain drug levels at testing time, and do not seem related to the mechanisms underlying the antidepressant therapeutic effect. However, these results might be relevant in explaining the effects of imipramine in models of drug-seeking and on body weight.

Nose-to-brain delivery of lamotrigine-loaded PLGA nanoparticles.

Direct nose-to-brain delivery of drugs and faster onset of action have made intra-nasal route a much sought-after alternative to conventional routes of drug delivery to the brain. Lamotrigine is used for the treatment and management of neuropathic pain, and in the present work, lamotrigine (LTG)-PLGA nanoparticles were developed for intra-nasal delivery. The LTG-PLGA nanoparticles were prepared using modified nanoprecipitation method via high-speed homogenization and ultra-sonication techniques. Entrapment efficiency (EE%) of developed LTG-PLGA-NPs was found to be 84.87 ± 1.2% with drug loading of 10.21 ± 0.89%. The particle size of developed nanoparticles was found to be 184.6nm with PDI value of 0.082 and zeta potential of - 18.8mV. Dissolution profiles were studied in PBS (pH7.4), simulated nasal fluid, and simulated cerebrospinal fluid where almost complete release was observed within 5h in CSF. In vitro, cytotoxicity was analyzed using MTT assay where dose-dependent cytotoxicity was observed for developed LTG-PLGA-NPs. In vitro cytokine analysis showed positive effects of LTG-PLGA-NPs as pro-inflammatory cytokine suppressors. Further, in vivo studies were performed for radiolabeled formulation and drug (99mTc-LTG-PLGA-NPs and 99mTc-LTG-aqueous) using Sprague Dawley rats where with the help of gamma scintigraphy studies, various routes of administration viz. oral, intra-nasal, and intra-venous were compared. Various pharmacokinetic parameters were evaluated using biodistribution studies to estimate the drug levels in blood and brain. For 99mTc-LTG-PLGA-NPs via intra-nasal route, drug targeting efficiency (DTE%) was found to be 129.81% and drug target organ transport (DTP%) to be 22.81% in brain with Cmax of 3.82%/g within Tmax 1.5h. Thus, the developed PLGA nanoparticles for intra-nasal delivery provide a possible alternative for existing available drug formulation for neuropathic pain management.

Identification and pharmacological profile of SPP1, a potent, functionally selective and brain penetrant agonist at muscarinic M1 receptors.

We aimed to identify and develop novel, selective muscarinic M1 receptor agonists as potential therapeutic agents for the symptomatic treatment of Alzheimer's disease. We developed and utilized a novel M1 receptor occupancy assay to drive a structure activity relationship in a relevant brain region while simultaneously tracking drug levels in plasma and brain to optimize for central penetration. Functional activity was tracked in relevant native in vitro assays allowing translational (rat-human) benchmarking of structure-activity relationship molecules to clinical comparators. Using this paradigm, we identified a series of M1 receptor selective molecules displaying desirable in vitro and in vivo properties and optimized key features, such as central penetration while maintaining selectivity and a partial agonist profile. From these compounds, we selected spiropiperidine 1 (SPP1). In vitro, SPP1 is a potent, partial agonist of cortical and hippocampal M1 receptors with activity conserved across species. SPP1 displays high functional selectivity for M1 receptors over native M2 and M3 receptor anti-targets and over a panel of other targets. Assessment of central target engagement by receptor occupancy reveals SPP1 significantly and dose-dependently occupies rodent cortical M1 receptors. We report the discovery of SPP1, a novel, functionally selective, brain penetrant partial orthosteric agonist at M1 receptors, identified by a novel receptor occupancy assay. SPP1 is amenable to in vitro and in vivo study and provides a valuable research tool to further probe the role of M1 receptors in physiology and disease.

A Tailored Thermosensitive PLGA-PEG-PLGA/Emulsomes Composite for Enhanced Oxcarbazepine Brain Delivery via the Nasal Route.

The use of nanocarrier delivery systems for direct nose to brain drug delivery shows promise for achieving increased brain drug levels as compared to simple solution systems. An example of such nanocarriers is emulsomes formed from lipid cores surrounded and stabilised by a corona of phospholipids (PC) and a coating of Tween 80, which combines the properties of both liposomes and emulsions. Oxcarbazepine (OX), an antiepileptic drug, was entrapped in emulsomes and then localized in a poly(lactic acid-co-glycolic acid)-poly(ethylene glycol)-poly(lactic acid-co-glycolic acid) (PLGA-PEG-PLGA) triblock copolymer thermogel. The incorporation of OX emulsomes in thermogels retarded drug release and increased its residence time (MRT) in rats. The OX-emulsome and the OX-emulsome-thermogel formulations showed in vitro sustained drug release of 81.1 and 53.5%, respectively, over a period of 24 h. The pharmacokinetic studies in rats showed transport of OX to the systemic circulation after nasal administration with a higher uptake in the brain tissue in case of OX-emulsomes and highest MRT for OX-emulsomal-thermogels as compared to the IN OX-emulsomes, OX-solution and Trileptal® suspension. Histopathological examination of nasal tissues showed a mild vascular congestion and moderate inflammatory changes around congested vessels compared to saline control, but lower toxic effect than that reported in case of the drug solution.

Vitamin-Derived Nanolipoidal Carriers for Brain Delivery of Dimethyl Fumarate: A Novel Approach with Preclinical Evidence.

Various oral treatment options have been reported for relapsing multiple sclerosis. Recently, dimethyl fumarate (DMF) has been approved for the management of the same. Though effective, DMF is associated with concerns like multiple dosing, patient incompliance, gastrointestinal flushing, lower brain permeation, and economic hurdles. Henceforth, the objective of the present study was to develop vitamin-based solid lipid nanoparticles (SLNs) for effective brain delivery of DMF with a promise of once-a-day dosing. The developed SLNs were characterized for micromeritics, morphology, entrapment efficiency, drug loading and in vitro drug release. Caco-2 and SH-SY5Y cell lines were used to assess the intestinal permeability and neuronal uptake. Pharmacokinetic and biodistribution studies were performed on rats. The developed nanometeric lipidparticles were able to control the drug release and substantially enhance the Caco-2 as well as SH-5YSY cell permeability. The developed systems not only enhanced the oral bioavailability of the drug, but also offered substantially elevated brain drug levels to that of plain drug. The drug was protected from liver and biological residence was increased, indicating promising potential of the carriers in effective brain delivery of DMF. Enhanced bioavailability and elevated bioresidence of DMF by vitamin-based SLNs provided the evidence for once-a-day delivery potential for DMF in the management of neurological disorders.