Centella asiatica Water Extract Shows Low Potential for Cytochrome P450-Mediated Drug Interactions.

Centella asiatica (CA) shows considerable promise for development as a botanical drug for cognitive decline. Its primary bioactive components include triterpene glycosides asiaticoside and madecassoside and their corresponding aglycones asiatic acid and madecassic acid. Exploration of the bioactivity of CA's caffeoylquinic acids is ongoing. In this study, an aqueous extract of CA (CAW-R61J) was evaluated for drug interaction potential through inhibition or induction of P450 enzymes, as required by the US Food and Drug Administration. CAW-R61J was assessed for induction potential of CYP1A2, CYP2B6, and CYP3A4 using transporter-certified cryopreserved human hepatocytes in sandwich culture. Gene expression of these target P450s was quantified, and enzyme activities were determined to confirm gene expression results. No induction was observed up to 16.7 µg/ml CAW-R61J (equivalent to 1.1 µM asiaticoside, 0.8 µM madecassoside, 0.09 µM asiatic acid, and 0.12 µM madecassic acid). Reversible and time-dependent inhibitory effects of CAW-R61J on CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP3A4/5 were evaluated using human liver microsomes. CAW-R61J showed weak reversible inhibition of most of the P450 forms tested, with the strongest being CYP2C9 (IC50 of 330 µg/ml). CAW-R61J (≤1000 µg/ml) was not a time-dependent inhibitor of any of these P450 enzymes. In summary, CAW-R61J had no, or only a weak impact, on P450 induction and inhibition in vitro. The clinical relevance of these results will depend on the in vivo concentration of CAW-R61J components achieved in humans. Plasma triterpene concentrations measured in our recent clinical studies suggest minimal risk of P450-mediated drug interactions by these components. SIGNIFICANCE STATEMENT: A preparation of Centella asiatica is currently under clinical development for the prevention or treatment of cognitive decline. The US Food and Drug Administration required an evaluation of its potential for drug interactions mediated through drug-metabolizing enzymes. This in vitro study revealed minimal induction or inhibition of a range of P450 enzymes, including CYP3A4, by the C. asiatica extract, suggesting a low potential for drug interactions modulated by P450 metabolism.

Integration of mass spectral fingerprinting analysis with precursor ion (MS1) quantification for the characterisation of botanical extracts: application to extracts of Centella asiatica (L.) Urban.

INTRODUCTION: The phytochemical composition of plant material governs the bioactivity and potential health benefits as well as the outcomes and reproducibility of laboratory studies and clinical trials. OBJECTIVE: The objective of this work was to develop an efficient method for the in-depth characterisation of plant extracts and quantification of marker compounds that can be potentially used for subsequent product integrity studies. Centella asiatica (L.) Urb., an Ayurvedic herb with potential applications in enhancing mental health and cognitive function, was used as a case study. METHODS: A quadrupole time-of-flight analyser in conjunction with an optimised high-performance liquid chromatography (HPLC) separation was used for in-depth untargeted fingerprinting and post-acquisition precursor ion quantification to determine levels of distinct phytochemicals in various C. asiatica extracts. RESULTS: We demonstrate the utility of this workflow for the characterisation of extracts of C. asiatica. This integrated workflow allowed the identification or tentative identification of 117 compounds, chemically interconnected based on Tanimoto chemical similarity, and the accurate quantification of 24 phytochemicals commonly found in C. asiatica extracts. CONCLUSION: We report a phytochemical analysis method combining liquid chromatography, high resolution mass spectral data acquisition, and post-acquisition interrogation that allows chemical fingerprints of botanicals to be obtained in conjunction with accurate quantification of distinct phytochemicals. The variability in the composition of specialised metabolites across different C. asiatica accessions was substantial, demonstrating that detailed characterisation of plant extracts is a prerequisite for reproducible use in laboratory studies, clinical trials and safe consumption. The methodological approach is generally applicable to other botanical products.

Centella asiatica attenuates hippocampal mitochondrial dysfunction and improves memory and executive function in ß-amyloid overexpressing mice.

Centella asiatica is a medicinal plant used to enhance memory. We have previously shown that a water extract of Centella asiatica (CAW) attenuates ß-amyloid (Aß)-induced spatial memory deficits in mice and improves neuronal health. Yet the effect of CAW on other cognitive domains remains unexplored as does its in vivo mechanism of improving Aß-related cognitive impairment. This study investigates the effects of CAW on learning, memory and executive function as well as mitochondrial function and antioxidant response in the 5xFAD model of Aß accumulation. Seven month old 5xFAD female mice were treated with CAW (2 mg/mL) in their drinking water for two weeks prior to behavioral testing. Learning, memory and executive function were assessed using the object location memory task (OLM), conditioned fear response (CFR) and odor discrimination reversal learning (ODRL) test. Mitochondrial function was profiled using the Seahorse XF platform in hippocampal mitochondria isolated from these animals and tissue was harvested for assessment of mitochondrial, antioxidant and synaptic proteins. CAW improved performance in all behavioral tests in the 5xFAD but had no effect on WT animals. Hippocampal mitochondrial function was improved and hippocampal and cortical expression of mitochondrial genes was increased in CAW-treated 5xFAD mice. Gene expression of the transcription factor NRF2, as well as its antioxidant target enzymes, was also increased with CAW treatment in both WT and 5xFAD mice. CAW treatment also decreased Aß-plaque burden in the hippocampus of treated 5xFAD mice but had no effect on plaques in the cortex. These data show that CAW can improve many facets of Aß-related cognitive impairment in 5xFAD mice. Oral treatment with CAW also attenuates hippocampal mitochondrial dysfunction in these animals. Because mitochondrial dysfunction and oxidative stress accompany cognitive impairment in many pathological conditions beyond Alzheimer's disease, this suggests potentially broad therapeutic utility of CAW.

Centella asiatica - Phytochemistry and mechanisms of neuroprotection and cognitive enhancement.

This review describes in detail the phytochemistry and neurological effects of the medicinal herb Centella asiatica (L.) Urban. C. asiatica is a small perennial plant that grows in moist, tropical and sub-tropical regions throughout the world. Phytochemicals identified from C. asiatica to date include isoprenoids (sesquiterpenes, plant sterols, pentacyclic triterpenoids and saponins) and phenylpropanoid derivatives (eugenol derivatives, caffeoylquinic acids, and flavonoids). Contemporary methods for fingerprinting and characterization of compounds in C. asiatica extracts include liquid chromatography and/or ion mobility spectrometry in conjunction with high-resolution mass spectrometry. Multiple studies in rodent models, and a limited number of human studies support C. asiatica's traditional reputation as a cognitive enhancer, as well as its anxiolytic and anticonvulsant effects. Neuroprotective effects of C.asiatica are seen in several in vitro models, for example against beta amyloid toxicity, and appear to be associated with increased mitochondrial activity, improved antioxidant status, and/or inhibition of the pro-inflammatory enzyme, phospholipase A2. Neurotropic effects of C. asiatica include increased dendritic arborization and synaptogenesis, and may be due to modulations of signal transduction pathways such as ERK1/2 and Akt. Many of these neurotropic and neuroprotective properties of C.asiatica have been associated with the triterpene compounds asiatic acid, asiaticoside and madecassoside. More recently, caffeoylquinic acids are emerging as a second important group of active compounds in C. asiatica, with the potential of enhancing the Nrf2-antioxidant response pathway. The absorption, distribution, metabolism and excretion of the triterpenes, caffeoylquinic acids and flavonoids found in C. asiatica have been studied in humans and animal models, and the compounds or their metabolites found in the brain. This review highlights the remarkable potential for C. asiatica extracts and derivatives to be used in the treatment of neurological conditions, and considers the further research needed to actualize this possibility.

Mode of administration influences plasma levels of active Centella asiatica compounds in 5xFAD mice while markers of neuroinflammation remain unaltered.

Introduction: A water extract of Centella asiatica (L.) Urban [Apiaceae] (CAW) has demonstrated cognitive-enhancing effects in mouse models of Alzheimer's disease and aging, the magnitude of which is influenced by whether CAW is delivered in the drinking water or the diet. These cognitive benefits are accompanied by improvements in oxidative stress and mitochondrial function in the brain, two pathways related to the neuroinflammatory response. The effect of CAW on neuroinflammation, however, has not been directly studied. Here, we investigated the effect of CAW on neuroinflammation in 5xFAD mice and compared plasma levels of CAW's active compounds following two modes of CAW administration. Methods: Eight-to-nine-month-old male and female 5xFAD mice and their wild-type littermates were administered CAW in their diet or drinking water (0 or 1,000 mg/kg/day) for five weeks. Immunohistochemistry was performed for ß-amyloid (Aß), glial fibrillary acidic protein (GFAP), and Griffonia simplicifolia lectin I (GSL I) in the cortex and hippocampus. Gene expression of inflammatory mediators (IL-6, TNFα, IL-1ß, TREM2, AIF1, CX3CR1, CX3CL1, CD36, C3AR1, RAGE, CCR6, CD3E) was measured in the deep grey matter. Results: CAW decreased cortical Aß plaque burden in female 5xFAD mice administered CAW in the drinking water but had no effect on Aß plaques in other treatment groups. CAW did not impact elevated levels of GFAP or GSL I in 5xFAD mice, regardless of sex, brain region, or mode of CAW administration. In the deep grey matter, CAW increased C3AR1 expression in 5xFAD females administered CAW in the drinking water and decreased IL-1ß expression in 5xFAD males administered CAW in the diet. CAW had no effect, however, on gene expression levels of any other inflammatory mediator in the deep grey, for either sex or mode of CAW administration. Mice administered CAW in the drinking water versus the diet had significantly higher plasma levels of CAW compounds. Discussion: CAW had little impact on the neuroinflammatory markers selected for evaluation in the present study, suggesting that the cognitive benefits of CAW may not be mediated by an anti-inflammatory effect or that additional molecular markers are needed to fully characterize the effect of CAW on neuroinflammation.

Quantification of Caffeoylquinic Acids and Triterpenes as Targeted Bioactive Compounds of Centella asiatica in Extracts and Formulations by Liquid Chromatography Mass Spectrometry.

Centella asiatica (CA) is a culinary vegetable and well-known functional food that is widely used as a medicinal herb and dietary supplement. CA is rich in pentacyclic triterpenes (TTs), including asiaticoside (AS), madecassoside (MS) and the related aglycones asiatic acid (AA), madecassic acid (MA). Traditionally, TTs have been associated with the bioactivity and health promoting effect of CA. Recently, mono-caffeoylquinic acids (MonoCQAs) and di-caffeoylquinic acids (DiCQAs) have been found to contribute to the bioactivity of CA as well. This work reports an analytical strategy based on liquid chromatography coupled to multiple reaction monitoring mass spectrometry (LC-MRM-MS) for the simultaneous rapid and accurate quantification of 12 bioactive compounds in CA, namely AS, MS, AA, MA, 5-CQA, 4-CQA, 3-CQA, 1,3-DiCQA, 3,4-DiCQA, 1,5-DiCQA, 3,5-DiCQA, 4,5-DiCQA. Method selectivity, accuracy, precision, repeatability, robustness, linearity range, limit of detection (LOD), and limit of quantitation (LOQ) were validated. The validated LC-MRM-MS method has been successfully applied to quantify the 12 bioactive compounds in CA aqueous extracts and two related formulations: a standardized CA product (CAP) used in a phase I clinical trial and formulated CA rodent diets used in preclinical studies. The validated method allows us to support the standardization of CA products used for clinical trials and conduct routine LC-MRM-MS analyses of formulated preclinical diets to confirm correct levels of CA phytochemical markers.

Bioanalytical method validation and application to a phase 1, double-blind, randomized pharmacokinetic trial of a standardized Centella asiatica (L.) Urban water extract product in healthy older adults.

Introduction: Centella asiatica is an herbaceous plant reputed in Eastern medicine to improve memory. Preclinical studies have shown that C. asiatica aqueous extract (CAW) improves neuronal health, reduces oxidative stress, and positively impacts learning and cognition. This study aimed to develop and validate bioanalytical methods for detecting known bioactive compounds from C. asiatica in human biological matrices and apply them to a human pharmacokinetic trial in healthy older adults. Methods: High performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was used for detecting triterpenes and caffeoylquinic acids from C. asiatica, or their metabolites, in human plasma and urine. Validation parameters including linearity, precision, accuracy, recovery and thermal stability were evaluated. The method was applied to a Phase I, randomized, double-blind, crossover trial of two doses (2 or 4 g) of a standardized C. asiatica water extract product (CAP) in eight healthy older adults. Pharmacokinetic parameters were measured over a 12-h post administration period and acute safety was assessed. Results: The method satisfied US Food & Drug Administration criteria for linearity and recovery of the analytes of interest in human plasma and urine. The method also satisfied criteria for precision and accuracy at medium and high concentrations. Single administration of 2 and 4 g of CAP was well tolerated and safe in healthy older adults. The parent triterpene glycosides, asiaticoside and madecassoside, were not detected in plasma and in minimal amounts in urinary excretion analyses, while the aglycones, asiatic acid and madecassic acid, showed readily detectable pharmacokinetic profiles. Similarly, the di-caffeoylquinic acids and mono-caffeoylquinic acids were detected in low quantities, while their putative metabolites showed readily detectable pharmacokinetic profiles and urinary excretion. Discussion: This method was able to identify and calculate the concentration of triterpenes and caffeoylquinic acids from C. asiatica, or their metabolites, in human plasma and urine. The oral absorption of these key compounds from CAP, and its acute safety in healthy older adults, support the use of this C. asiatica product in future clinical trials.

Pharmacokinetics and Pharmacodynamics of Key Components of a Standardized Centella asiatica Product in Cognitively Impaired Older Adults: A Phase 1, Double-Blind, Randomized Clinical Trial.

Centella asiatica is reputed in Eastern medicine to improve cognitive function in humans. Preclinical studies have demonstrated that aqueous extracts of C. asiatica improve cognition in mouse models of aging and Alzheimer's disease (AD) through the modulation of mitochondrial biogenesis and nuclear factor-erythroid-2-related factor 2 (Nrf2)-dependent antioxidant response genes. This randomized, double-blind, crossover Phase I trial explored the oral bioavailability and pharmacokinetics of key compounds from two doses (2 g and 4 g) of a standardized C. asiatica aqueous extract product (CAP), over 10 h, in four mildly demented older adults on cholinesterase inhibitor therapy. The analysis focused on triterpenes (TTs) and caffeoylquinic acids (CQAs), which are known to contribute to C. asiatica's neurological activity. The acute safety of CAP and the effects on NRF2 gene expression in peripheral blood mononuclear cells were evaluated. Single administration of 2 g or 4 g of CAP was safe and well-tolerated. The TT aglycones, asiatic acid and madecassic acid, were identified in plasma and urine, while the parent glycosides, asiaticoside and madecassoside, although abundant in CAP, were absent in plasma and had limited renal excretion. Similarly, mono- and di-CQAs showed delayed absorption and limited presence in plasma or urine, while the putative metabolites of these compounds showed detectable plasma pharmacokinetic profiles and urinary excretion. CAP elicited a temporal change in NRF2 gene expression, mirroring the TT aglycone's pharmacokinetic curve in a paradoxical dose-dependent manner. The oral bioavailability of active compounds or their metabolites, NRF2 target engagement, and the acute safety and tolerability of CAP support the validity of using CAP in future clinical studies.

Withania somnifera and Centella asiatica Extracts Ameliorate Behavioral Deficits in an In Vivo Drosophila melanogaster Model of Oxidative Stress.

Due to an increase in the aging population, age-related diseases and age-related changes, such as diminished cognition and sleep disturbances, are an increasing health threat. It has been suggested that an increase in oxidative stress underlies many of these changes. Current treatments for these diseases and changes either have low efficacy or have deleterious side effects preventing long-time use. Therefore, alternative treatments that promote healthy aging and provide resilience against these health threats are needed. The herbs Withania somnifera and Centella asiatica may be two such alternatives because both have been connected with reducing oxidative stress and could therefore ameliorate age-related impairments. To test the effects of these herbs on behavioral phenotypes induced by oxidative stress, we used the Drosophila melanogaster sniffer mutant which has high levels of oxidative stress due to reduced carbonyl reductase activity. Effects on cognition and mobility were assessed using phototaxis assays and both, W. somnifera and C. asiatica water extracts improved phototaxis in sniffer mutants. In addition, W. somnifera improved nighttime sleep in male and female sniffer flies and promoted a less fragmented sleep pattern in male sniffer flies. This suggests that W. somnifera and C. asiatica can ameliorate oxidative stress-related changes in behavior and that by doing so they might promote healthy aging in humans.

Unobtrusive Sensing Technology Detects Ecologically Valid Spatiotemporal Patterns of Daily Routines Distinctive to Persons With Mild Cognitive Impairment.

BACKGROUND: The ability to capture people's movement throughout their home is a powerful approach to inform spatiotemporal patterns of routines associated with cognitive impairment. The study estimated indoor room activities over 24 hours and investigated relationships between diurnal activity patterns and mild cognitive impairment (MCI). METHODS: One hundred and sixty-one older adults (26 with MCI) living alone (age = 78.9 ± 9.2) were included from 2 study cohorts-the Oregon Center for Aging & Technology and the Minority Aging Research Study. Indoor room activities were measured by the number of trips made to rooms (bathroom, bedroom, kitchen, living room). Trips made to rooms (transitions) were detected using passive infrared motion sensors fixed on the walls for a month. Latent trajectory models were used to identify distinct diurnal patterns of room activities and characteristics associated with each trajectory. RESULTS: Latent trajectory models identified 2 diurnal patterns of bathroom usage (high and low usage). Participants with MCI were more likely to be in the high bathroom usage group that exhibited more trips to the bathroom than the low-usage group (odds ratio [OR] = 4.1, 95% CI [1.3-13.5], p = .02). For kitchen activity, 2 diurnal patterns were identified (high and low activity). Participants with MCI were more likely to be in the high kitchen activity group that exhibited more transitions to the kitchen throughout the day and night than the low kitchen activity group (OR = 3.2, 95% CI [1.1-9.1], p = .03). CONCLUSIONS: The linkage between bathroom and kitchen activities with MCI may be the result of biological, health, and environmental factors in play. In-home, real-time unobtrusive-sensing offers a novel way of delineating cognitive health with chronologically-ordered movement across indoor locations.

Effects of Withania somnifera (Ashwagandha) on Stress and the Stress- Related Neuropsychiatric Disorders Anxiety, Depression, and Insomnia.

BACKGROUND: Withania somnifera (WS), also known as Ashwagandha, is commonly used in Ayurveda and other traditional medicine systems. WS has seen an increase in worldwide usage due to its reputation as an adaptogen. This popularity has elicited increased scientific study of its biological effects, including a potential application for neuropsychiatric and neurodegenerative disorders. OBJECTIVE: This review aims to provide a comprehensive summary of preclinical and clinical studies examining the neuropsychiatric effects of WS, specifically its application in stress, anxiety, depression, and insomnia. METHODS: Reports of human trials and animal studies of WS were collected primarily from the PubMed, Scopus, and Google Scholar databases. RESULTS: WS root and leaf extracts exhibited noteworthy anti-stress and anti-anxiety activity in animal and human studies. WS also improved symptoms of depression and insomnia, though fewer studies investigated these applications. WS may alleviate these conditions predominantly through modulation of the hypothalamic-pituitary-adrenal and sympathetic-adrenal-medullary axes, as well as through GABAergic and serotonergic pathways. While some studies link specific withanolide components to its neuropsychiatric benefits, there is evidence for the presence of additional, as yet unidentified, active compounds in WS. CONCLUSION: While benefits were seen in the reviewed studies, significant variability in the WS extracts examined prevents a consensus on the optimum WS preparation or dosage for treating neuropsychiatric conditions. WS generally appears safe for human use; however, it will be important to investigate potential herb-drug interactions involving WS if used alongside pharmaceutical interventions. Further elucidation of active compounds of WS is also needed.

Loss of NRF2 accelerates cognitive decline, exacerbates mitochondrial dysfunction, and is required for the cognitive enhancing effects of Centella asiatica during aging.

The water extract of Centella asiatica (CAW) improves cognitive and mitochondrial function and activates the nuclear factor erythroid 2-related factor 2 (NRF2) regulated antioxidant response pathway in aged mice. Here we investigate whether NRF2 activation is required for the cognitive and mitochondrial effects of prolonged CAW exposure during aging. Five-month-old NRF2 knockout (NRF2KO) and wild-type mice were treated with CAW for 1, 7, or 13 months. Each cohort underwent cognitive testing and hippocampal mitochondrial analyses. Age-related cognitive decline was accelerated in NRF2KO mice and while CAW treatment improved cognitive performance in wild-type mice, it had no effect on NRF2KO animals. Hippocampal mitochondrial function also declined further with age in NRF2KO mice and greater hippocampal mitochondrial dysfunction was associated with poorer cognitive performance in both genotypes. Long-term CAW treatment did not affect mitochondrial endpoints in animals of either genotype. These data indicate that loss of NRF2 results in accelerated age-related cognitive decline and worsened mitochondrial deficits. NRF2 also appears to be required for the cognitive enhancing effects of CAW during aging.

Prolonged Treatment with Centella asiatica Improves Memory, Reduces Amyloid-ß Pathology, and Activates NRF2-Regulated Antioxidant Response Pathway in 5xFAD Mice.

BACKGROUND: The medicinal herb Centella asiatica has been long been used for its neuroprotective and cognitive enhancing effects. We have previously shown that two weeks of treatment with a water extract of Centella asiatica (CAW) improves cognition and activates the endogenous antioxidant response pathway without altering amyloid-ß (Aß) plaque burden. OBJECTIVE: Here, we assess the effect of long-term treatment of CAW in the 5xFAD mouse model of Aß accumulation. METHODS: Four-month-old 5xFAD mice were treated with CAW in their drinking water (2 g/L) for three months at which point they underwent cognitive testing as well as analysis of Aß plaque levels and antioxidant and synaptic gene expression. In order to confirm the involvement of the antioxidant regulatory transcription factor NRF2 on the effects of CAW on synaptic plasticity, neurons isolated from 5xFAD mice were also treated with CAW and the targeted inhibitor ML385. RESULTS: Three months of treatment with CAW improved spatial and contextual memory as well as executive function in 5xFAD mice. This improvement was accompanied by increased antioxidant gene expression and a decrease in Aß plaque burden relative to untreated 5xFAD animals. In isolated neurons, treatment with ML385 blocked the effects of CAW on dendritic arborization and synaptic gene expression. CONCLUSION: These results suggest that prolonged CAW exposure could be beneficial in Alzheimer's disease and that these effects likely involve NRF2 activation. Moreover, these findings suggest that targeting NRF2 itself may be a relevant therapeutic strategy for improving synaptic plasticity and cognitive function in Alzheimer's disease.

Centella asiatica Alters Metabolic Pathways Associated With Alzheimer's Disease in the 5xFAD Mouse Model of ß-Amyloid Accumulation.

Centella asiatica is an herb used in Ayurvedic and traditional Chinese medicine for its beneficial effects on brain health and cognition. Our group has previously shown that a water extract of Centella asiatica (CAW) elicits cognitive-enhancing effects in animal models of aging and Alzheimer's disease, including a dose-related effect of CAW on memory in the 5xFAD mouse model of ß-amyloid accumulation. Here, we endeavor to elucidate the mechanisms underlying the effects of CAW in the brain by conducting a metabolomic analysis of cortical tissue from 5xFAD mice treated with increasing concentrations of CAW. Tissue was collected from 8-month-old male and female 5xFAD mice and their wild-type littermates treated with CAW (0, 200, 500, or 1,000 mg/kg/d) dissolved in their drinking water for 5 weeks. High-performance liquid chromatography coupled to high-resolution mass spectrometry analysis was performed and relative levels of 120 annotated metabolites were assessed in the treatment groups. Metabolomic analysis revealed sex differences in the effect of the 5xFAD genotype on metabolite levels compared to wild-type mice, and variations in the metabolomic response to CAW depending on sex, genotype, and CAW dose. In at least three of the four treated groups (5xFAD or wild-type, male or female), CAW (500 mg/kg/d) significantly altered metabolic pathways related to purine metabolism, nicotinate and nicotinamide metabolism, and glycerophospholipid metabolism. The results are in line with some of our previous findings regarding specific mechanisms of action of CAW (e.g., improving mitochondrial function, reducing oxidative stress, and increasing synaptic density). Furthermore, these findings provide new information about additional, potential mechanisms for the cognitive-enhancing effect of CAW, including upregulation of nicotinamide adenine dinucleotide in the brain and modulation of brain-derived neurotrophic factor. These metabolic pathways have been implicated in the pathophysiology of Alzheimer's disease, highlighting the therapeutic potential of CAW in this neurodegenerative disease.

Developing a Rational, Optimized Product of Centella asiatica for Examination in Clinical Trials: Real World Challenges.

Botanical products are frequently sold as dietary supplements and their use by the public is increasing in popularity. However, scientific evaluation of their medicinal benefits presents unique challenges due to their chemical complexity, inherent variability, and the involvement of multiple active components and biological targets. Translation away from preclinical models, and developing an optimized, reproducible botanical product for use in clinical trials, presents particular challenges for phytotherapeutic agents compared to single chemical entities. Common deficiencies noted in clinical trials of botanical products include limited characterization of the product tested, inadequate placebo control, and lack of rationale for the type of product tested, dose used, outcome measures or even the study population. Our group has focused on the botanical Centella asiatica due to its reputation for enhancing cognition in Eastern traditional medicine systems. Our preclinical studies on a Centella asiatica water extract (CAW) and its bioactive components strongly support its potential as a phytotherapeutic agent for cognitive decline in aging and Alzheimer's disease through influences on antioxidant response, mitochondrial activity, and synaptic density. Here we describe our robust, scientific approach toward developing a rational phytotherapeutic product based on Centella asiatica for human investigation, addressing multiple factors to optimize its valid clinical evaluation. Specific aspects covered include approaches to identifying an optimal dose range for clinical assessment, design and composition of a dosage form and matching placebo, sourcing appropriate botanical raw material for product manufacture (including the evaluation of active compounds and contaminants), and up-scaling of laboratory extraction methods to available current Good Manufacturing Practice (cGMP) certified industrial facilities. We also address the process of obtaining regulatory approvals to proceed with clinical trials. Our study highlights the complexity of translational research on botanicals and the importance of identifying active compounds and developing sound analytical and bioanalytical methods for their determination in botanical materials and biological samples. Recent Phase I pharmacokinetic studies of our Centella asiatica product in humans (NCT03929250, NCT03937908) have highlighted additional challenges associated with designing botanical bioavailability studies, including specific dietary considerations that need to be considered.

Photoperiod-dependent floral reversion in the natural allopolyploid Arabidopsis suecica.

Flower reversion is the result of genetic or environmental effects that reverse developmental steps in the transition from the vegetative to the reproductive phase in plants. Here, we describe peculiar floral abnormalities, homeotic conversions, and flower reversion in several wild-type accessions of the natural allopolyploid Arabidopsis suecica. Microscopy was used to illustrate the phenotype in detail and we experimented with varying photoperiod lengths to establish whether or not the phenotype was responsive to the environment. We also profiled the transcriptional activity of several floral regulator genes during flower reversion using real-time PCR. We showed that the frequency of floral reversion was affected by day length and the position of the flower along the inflorescence axis. In reverting flowers we found unusual gene expression patterns of floral promoters and inflorescence maintenance genes, including lower mRNA levels of AGAMOUS-LIKE-24 (AGL-24), APETALA1 (AP1), and SHORT VEGETATIVE PHASE (SVP), and higher mRNA levels of SUPRESSOR OF CONSTANS1 (SOC1) compared with normal flowers. We conclude that the floral reversion frequency in A. suecica is susceptible to photoperiod changes, and that the floral abnormalities coincide with the competing expression of floral promoters and floral repressors in reverting floral tissue.

An Ecologically Valid, Longitudinal, and Unbiased Assessment of Treatment Efficacy in Alzheimer Disease (the EVALUATE-AD Trial): Proof-of-Concept Study.

BACKGROUND: The current clinical trial assessment methodology relies on a combination of self-report measures, cognitive and physical function tests, and biomarkers. This methodology is limited by recall bias and recency effects in self-reporting and by assessments that are brief, episodic, and clinic based. Continuous monitoring of ecologically valid measures of cognition and daily functioning in the community may provide a more sensitive method to detect subtle, progressive changes in patients with cognitive impairment and dementia. OBJECTIVE: This study aimed to present an alternative trial approach using a home-based sensing and computing system to detect changes related to common treatments employed in Alzheimer disease (AD). This paper introduces an ongoing study that aims to determine the feasibility of capturing sensor-based data at home and to compare the sensor-based outcomes with conventional outcomes. We describe the methodology used in the assessment protocol and present preliminary results of feasibility measures and examples of data related to medication-taking behavior, activity levels, and sleep. METHODS: The EVALUATE-AD (Ecologically Valid, Ambient, Longitudinal and Unbiased Assessment of Treatment Efficacy in Alzheimer's Disease) trial is a longitudinal naturalistic observational cohort study recruiting 30 patients and 30 spouse coresident care partners. Participants are monitored continuously using a home-based sensing and computing system for up to 24 months. Outcome measures of the automated system are compared with conventional clinical outcome measures in AD. Acceptance of the home system and protocol are assessed by rates of dropout and protocol adherence. After completion of the study monitoring period, a composite model using multiple functional outcome measures will be created that represents a behavioral-activity signature of initiating or discontinuing AD-related medications, such as cholinesterase inhibitors, memantine, or antidepressants. RESULTS: The home-based sensing and computing system has been well accepted by individuals with cognitive impairment and their care partners. Participants showed good adherence to the completion of a weekly web-based health survey. Daily activity, medication adherence, and total time in bed could be derived from algorithms using data from the sensing and computing system. The mean monitoring time for current participants was 14.6 months. Medication adherence, as measured with an electronic pillbox, was 77% for participants taking AD-related medications. CONCLUSIONS: Continuous, home-based assessment provides a novel approach to test the impact of new or existing dementia treatments generating objective, clinically meaningful measures related to cognition and everyday functioning. Combining this approach with the current clinical trial methodology may ultimately reduce trial durations, sample size needs, and reliance on a clinic-based assessment. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/17603.

Caffeoylquinic Acids in Centella asiatica Reverse Cognitive Deficits in Male 5XFAD Alzheimer's Disease Model Mice.

Centella asiatica (CA) is an edible plant and a popular botanical dietary supplement. It is reputed, in Ayurveda, to mitigate age-related cognitive decline. There is a considerable body of preclinical literature supporting CA's ability to improve learning and memory. This study evaluated the contribution of CA's triterpenes (TT), widely considered its active compounds, and caffeoylquinic acids (CQA) to the cognitive effects of CA water extract (CAW) in 5XFAD mice, a model of Alzheimer's disease. 5XFAD mice were fed a control diet alone, or one containing 1% CAW or compound groups (TT, CQA, or TT + CQA) equivalent to their content in 1% CAW. Wild-type (WT) littermates received the control diet. Conditioned fear response (CFR) was evaluated after 4.5 weeks. Female 5XFAD controls showed no deficit in CFR compared to WT females, nor any effects from treatment. In males, CFR of 5XFAD controls was attenuated compared to WT littermates (p = 0.005). 5XFAD males receiving CQA or TT + CQA had significantly improved CFR (p < 0.05) compared to 5XFAD male controls. CFR did not differ between 5XFAD males receiving treatment diets and WT males. These data confirm a role for CQA in CAW's cognitive effects.

Lipoic Acid and Other Antioxidants as Therapies for Multiple Sclerosis.

Oxidative stress (OS), when oxidative forces outweigh endogenous and nutritional antioxidant defenses, contributes to the pathophysiology of multiple sclerosis (MS). Evidence of OS is found during acute relapses, in active inflammatory lesions, and in chronic, longstanding plaques. OS results in both ongoing inflammation and neurodegeneration. Antioxidant therapies are a rational strategy for people with MS with all phenotypes and disease durations. PURPOSE OF REVIEW: To understand the function of OS in health and disease, to examine the contributions of OS to MS pathophysiology, and to review current evidence for the effects of selected antioxidant therapies in people with MS (PwMS) with a focus on lipoic acid (LA). RECENT FINDINGS: Studies of antioxidant interventions in both animal and in vivo models result in reductions in serum markers of OS and increases in levels and activity of antioxidant enzymes. Antioxidant trials in PwMS, while generally underpowered, detect short-term improvements in markers of OS and antioxidant defenses, and to a lesser extent, in clinical symptoms (fatigue, depression). The best evidence to date is a 2-year trial of LA in secondary progressive MS which demonstrated a significant reduction of whole-brain atrophy and trend toward improvement in walking speed. Antioxidant therapy is a promising approach to treat MS across the spectrum and duration of disease. Rigorous and well-powered trials are needed to determine their therapeutic benefits.

Centella Asiatica Improves Memory and Promotes Antioxidative Signaling in 5XFAD Mice.

Centella asiatica (CA) herb is a traditional medicine, long reputed to provide cognitive benefits. We have reported that CA water extract (CAW) treatment improves cognitive function of aged Alzheimer's disease (AD) model Tg2576 and wild-type (WT) mice, and induces an NRF2-regulated antioxidant response in aged WT mice. Here, CAW was administered to AD model 5XFAD female and male mice and WT littermates (age: 7.6 +/ - 0.6 months), and object recall and contextual fear memory were tested after three weeks treatment. CAW's impact on amyloid-ß plaque burden, and markers of neuronal oxidative stress and synaptic density, was assessed after five weeks treatment. CAW antioxidant activity was evaluated via nuclear transcription factor (erythroid-derived 2)-like 2 (NRF2) and NRF2-regulated antioxidant response element gene expression. Memory improvement in both genders and genotypes was associated with dose-dependent CAW treatment without affecting plaque burden, and marginally increased synaptic density markers in the hippocampus and prefrontal cortex. CAW treatment increased Nrf2 in hippocampus and other NRF2 targets (heme oxygenase-1, NAD(P)H quinone dehydrogenase 1, glutamate-cysteine ligase catalytic subunit). Reduced plaque-associated SOD1, an indicator of oxidative stress, was observed in the hippocampi and cortices of CAW-treated 5XFAD mice. We postulate that CAW treatment leads to reduced oxidative stress, contributing to improved neuronal health and cognition.