Chlorogenic Acid: A Systematic Review on the Biological Functions, Mechanistic Actions, and Therapeutic Potentials

Chlorogenic acid (CGA) is a type of polyphenol compound found in rich concentrations in many plants such as green coffee beans. As an active natural substance, CGA exerts diverse therapeutic effects in response to a variety of pathological challenges, particularly conditions associated with chronic metabolic diseases and age-related disorders. It shows multidimensional functions, including neuroprotection for neurodegenerative disorders and diabetic peripheral neuropathy, anti-inflammation, anti-oxidation, anti-pathogens, mitigation of cardiovascular disorders, skin diseases, diabetes mellitus, liver and kidney injuries, and anti-tumor activities. Mechanistically, its integrative functions act through the modulation of anti-inflammation/oxidation and metabolic homeostasis. It can thwart inflammatory constituents at multiple levels such as curtailing NF-kB pathways to neutralize primitive inflammatory factors, hindering inflammatory propagation, and alleviating inflammation-related tissue injury. It concurrently raises pivotal antioxidants by activating the Nrf2 pathway, thus scavenging excessive cellular free radicals. It elevates AMPK pathways for the maintenance and restoration of metabolic homeostasis of glucose and lipids. Additionally, CGA shows functions of neuromodulation by targeting neuroreceptors and ion channels. In this review, we systematically recapitulate CGA’s pharmacological activities, medicinal properties, and mechanistic actions as a potential therapeutic agent. Further studies for defining its specific targeting molecules, improving its bioavailability, and validating its clinical efficacy are required to corroborate the therapeutic effects of CGA.

CGA has limited bioavailability in plant foods due to the esterification with cell wall components such as proteins, lignin, and cellulose [5]; thus, appropriate food processing is needed to facilitate release [6].CGAs are enriched in green coffee bean extract (GCE), which may comprise 54% of its contents [7].Particularly, 5-CQA and 3-CQA present about 35−40% and 10−15% among CGA components, respectively [8].The roasting process leads to a dramatic decrease in the total amount of CGAs and changes in CGA compositions with main contents of 3,4-di-CQA, 5-CQA, 4-CQA, and 3-CQA [9].One-third of CGA is metabolized quickly after direct absorption in the upper gastrointestinal tract upon oral administration [3,10].The esterase secreted by the intestine microbes (such as Lactobacillus gasseri, Bifidobacterium lactis, and Escherichia coli) can hydrolyze the remaining CGAs and release CGA and quinic acid to be absorbed in the intestines [11,12].
CGA exhibits a good safety profile, which has not shown any obvious adverse effect and toxicity to normal cells or tissues, and is well-tolerated by humans [13,14].In an acute toxicity experiment, no side effects are observed in mice for two weeks upon an intake of CGA-enriched GCE (1 g/kg) [15].A single dose of GCE (2 g/kg) (containing 50% CGA) in rats does not cause any type of toxicity.Rats with an intake of CGA (250, 500, and 1000 mg/kg) show no adverse effects in three months [16].Cautiously, a high-dosage consumption of CGA (2 g/day) or black tea (4 g/4 L/day) four times in 7 days can moderately increase plasma homocysteine levels by 12% or 11% in humans, respectively [17].
In this review, the literature search was conducted between 2005 and 2024 in the database of PubMed for articles related to the subjects using the specific keywords of "chlorogenic acid" and ("inflammation" or "oxidation").Inclusion criteria included fulltext publications in English.Exclusion criteria included preprints and extracts without mention of CGA as the active ingredient in the abstract.The publications in this search result served as core literature for this review.

Glucose and Lipid Metabolic Homeostasis Modulation (Figure 1B)
CGAs can facilitate the maintenance of metabolic homeostasis of glucose and lipids [38].One mechanism involves the modulation of activities of AMP-activated protein kinase (AMPK) and ERK1/2 [4] (Figure 2C,D).AMPK is a master energy sensor that regulates cellular glucose and lipid metabolism.These functions directly underlie the CGA effects on the mitigation of chronic metabolic-associated syndromes such as obesity, diabetes mellitus (DM), and their complications.CGA or CGA-containing extracts can inhibit pancreatic lipase activity [39].CGA exhibits inhibitory effects on the function of many lipid metabolic enzymes including fatty acid synthase, HMG-CoA reductase, and cholesterol acyltransferase in mice fed on a high-fat diet (HFD) [40] (Figure 2C).CGA can upregulate AMPK and carnitine palmitoyltransferase I (CPT-1) and inhibit acetyl-CoA carboxylase (ACC), thus reducing hepatic and blood levels of triglyceride (TG) and free fatty acids (FFA) in HFD rats [41] (Figure 2C).CGA can facilitate cholesterol elimination by modulating homeostasis of bilirubin and bile acids via farnesoid X receptor (FXR) and peroxisome proliferator-activated receptor (PPAR) gamma coactivator 1-alpha (PGC-1α) or fibroblast growth factor (FGF) 15 pathways [42,43].
CGA can reduce glucose release.CGA can inhibit glucose-6-phosphatase (G6Pase), the enzyme converting glycogen to glucose [44].CGA inhibits the expression and activity of hepatic α-glucosidase and G6Pase, reduces the hydrolysis of hepatic glycogen, and activates AMPK pathways, resulting in attenuation of hepatic steatosis and improvements of metabolic indexes including fasting serum glucose (FSG) level, glucose tolerance, glucose uptake, insulin sensitivity, and lipid profiles [38,48,49] (Figure 2D).
CGA can modulate plasma levels of glucose and lipids.In HFD golden hamsters or rats, CGA upregulates hepatic PPAR-α levels, increases the activity of hepatic lipase (HL), decreases hepatic levels of TG and FFA and fasting serum levels of TG, FFA, total cholesterol (TC), low-density lipoprotein cholesterol (LDL-c), high-density lipoprotein cholesterol (HDL-c), FSG, and insulin (FSI), as well as attenuates the activity of lipoprotein lipase (LPL) in skeletal muscle [50,51].CGA-containing GCEs (100 mg/kg, 6 weeks) can decrease blood glucose levels, body weight, and fat mass in mice fed on an HFD [52].CGA (oral gavage 80 mg/kg/day, 12 weeks) in db/db mice can lower FSG, adiponectin, and TG, and increase muscle glycogen via up-regulating hepatic PPAR-α and inhibiting G6Pase expression [53].Post-meal CGA treatment (60 min) decreases the level of blood sugar compared to the placebo in rats [54].

Human Subject Studies
In a randomized crossover study, healthy postmenopausal women (BMI 25-40, n = 16) with consumption of the bioactive yogurt containing curcumin and CGA showed significantly lower plasma levels of TNFα compared to the placebo group and the baseline [55].In an acute pilot study, healthy subjects (n = 31) were given a single dose of a polyphenol-rich beverage (PRB) or placebo.The plasma levels of 8-iso-PGF2-alpha and advanced oxidation protein products were decreased, and hydroxyl radical antioxidant capacity at one-hour post intake of PRB was increased compared to the baseline [56].
In a cohort of 15 patients with impaired glucose tolerance (IGT), CGA (400 mg three times per day for 3 months) decreased FSG, insulinogenic index, body weight, body mass, waist circumference, TG, TC, LDL-c, and very low-density lipoprotein levels, with an upregulated Matsuda index [57].In a randomized, double-blind controlled trial, participants (n = 65) were given an 8-week cooked ham enriched with a pool of antioxidants (including 22.5 mg CGA/100 g cooked ham) or received a placebo.Subjects with intervention showed significantly lower levels of ox-LDL, malondialdehyde (MDA), TC, high-sensitive C-reactive protein (hs-CRP), and IL-6 [58].In a cohort of overweight dyslipidemic subjects (n = 90), a nutraceutical (containing bergamot, phytosterols, vitamin C, and CGA) or placebo was administered for 8 weeks.The subjects with the treatment showed improved lipid and glucose metabolism, which were associated with reduced levels of TG, LDL-c, non-HDL-c, the ratio of leptin/adiponectin, hs-CRP, and TNFα [59].In a randomized, cross-over, controlled study, hypercholesterolemic subjects (n = 27) were administered soluble green/roasted (35:65) coffee or placebo for 8 weeks.The subjects showed lower lipid parameters (TC, TG, LDL-c, VLDL-c), MDA, and protein carbonyl group oxidation, systolic and diastolic blood pressures (SBP, DBP), heart rate, and body weight compared with the baselines [60].Habitual coffee intake decreased serum levels of IL-18 and 8-isoprostane but increased adiponectin and HDL-c in healthy subjects (n = 47) [61].In a study, healthy, overweight subjects (n = 142, BMI ≥25 to <30 kg/m 2 ) were given a high-CGA (369 mg CGA/serving) or control (35 mg CGA/serving) coffee for 12 weeks.subjects with an intake of high-CGA coffee showed significant improvements in lowering the visceral fat area (VFA), total abdominal fat area (TFA), BMI, and waist circumference compared to those in the control group [62].In a cohort of 21 patients with metabolic syndrome, CGA-containing GCE (400 mg twice per day for 2 months) showed a decrease in levels of FSG, insulin resistance, weight, and BMI in patients [8].In a study of healthy Japanese women (n = 57), plasma CGA showed a negative association with FSG, glycated hemoglobin, and CRP [63].
CGAs showed diverse effects on neuroprotection for neurodegenerative disorders and diabetic peripheral neuropathy, mitigation of cardiovascular disorders, skin diseases, diabetic mellitus, liver and kidney injuries, and anti-tumor activities (Figure 1).Mechanistically, their anti-inflammation and anti-oxidation properties and metabolic modulations underlie these pharmacological activities for protection against cell injuries, restoration of cellular function, and maintenance of physiological and metabolic homeostasis (Figure 2), which is discussed across various tissues and disorders in this review.

Cardiovascular Protective Effect
CGA can exert protective roles at multiple levels in various cardiovascular complications, including mitigating hypotension, improving endothelial cell function, alleviating atherosclerosis, and ameliorating cardiomyopathy.

Human Subject Studies for Cardiovascular Protection
CGA could lower SBP and DBP in patients with mild hypertension [7,14,103,104].For example, Kozuma et al. showed that daily oral ingestion of GCE (93 or 185 mg for 4 weeks) could lead to a reduction of 4.7 and 5.6 mmHg in levels of systolic blood pressure (SBP) and a decrease of 3.3 and 3.9 mmHg in levels of diastolic blood pressure (DBP), respectively, in patients with hypertension [7].In a randomized trial in Japanese patients with mild essential hypertension (HPT), CGA (140 mg/day) for 12 weeks could lower 10 mmHg of SBP and 6 mmHg of DBP [14].Mild HPT patients taking CGA (228 mg/day for 1 month) showed a reduction of 3.3 and 2.8 mmHg in levels of SBP and DBP, respectively [104].Ferulic acid is considered one of the active substances of CGA for producing a strong hypotensive effect via muscarinic acetylcholine receptors after short-and long-term ingestions [64,65].In a clinical trial of patients with borderline or stage 1 hypertension (n = 37), a single intake of coffee with a high content of CGAs and low content of hydroxyhydroquinone (HHQ) significantly improved postprandial flowmediated vasodilation and decreased circulating 8-isoprostane levels, which was effective for improving postprandial endothelial dysfunction [105].A separate study showed that healthy male adults with ingestion of CGA without HHQ for four weeks could significantly increase postprandial fat oxidation and the ratio of postprandial biological antioxidant potential (BAP) to the derivatives of reactive oxygen metabolites (d-ROMs) compared to those with an intake of CGA with HHQ [106].CGAs can incorporate specific phenolic acids into LDL particles to lower the risk of their oxidations in human subjects [70,71,107].
In a randomized controlled trial, healthy adults with an 8-week consumption of CGA-enriched coffee beverages showed a significant decrease in levels of twelve urine oxylipins compared to the baseline.Oxylipins are generated during foam cell formation in atherogenesis and thus are biomarkers for CVDs [108].In a cohort of healthy subjects (n = 25), the impact of consumption of coffee containing 787 mg or 407 mg CGAs on CVD risk markers such as oxysterols and FFAs was assessed.Subjects with an intake of coffee showed a decrease in oxysterols and FFAs and an increase in cholesteryl esters.While subjects in the placebo group showed an elevation of oxysterols and FFAs and a reduction in cholesteryl esters [109].Healthy subjects with consumption of decaffeinated GCE (CGAs accounting for about 51.2% constituents) showed an acute improvement in flow-mediated dilation (%FMD) of the brachial artery [110].A study from two randomized trials with healthy male subjects (n = 15) showed that coffee intake could acutely improve human vascular function, likely through 5-CQA and its physiological metabolites [111].A higher response of FMD induced by CGA-rich coffee was also reported in a study with 12 healthy subjects [112].Healthy adults with an intake of a coffee berry beverage (containing 440 mg chlorogenic acid) could increase subjective energic arousal and hemodynamic responses from cerebral blood flow compared with the baseline [113].In a placebo-controlled doubleblind pilot study with healthy Japanese men (n = 16), subjects with the intake of GCE showed significantly greater changes in cardio-ankle vascular index (CAVI) (e.g., increasing FMD and decreasing sympathetic nervous activity) than those in the placebo group [114].In a randomized, double-blind, placebo-controlled study, subjects (n = 50, BMI ≥ 25 to <30 kg/m 2 ) were given a nutraceutical containing CGA and luteolin extracts for 6 months.Participants in the treatment group showed significantly decreased body weight, glycemic and lipid parameters (TC, TG, LDL-c) as well as improved hepatic functionality, carotidmedia thickness (CIMT), and endothelial function compared to the subjects in the placebo group [115].In a separate study of subjects with metabolic syndrome (n = 50), a 6-month intake of the same nutraceutical significantly improved hepatic and cardio-metabolic parameters in the patients [116].

Mitigative Effects on Diabetes Mellitus (DM)
CGA has shown its functions in protecting β cells from apoptosis, improving β cell function, facilitating glycemic control, and mitigating DM complications.

Protective Effects on β Cells (Figure 1B)
CGA can competitively reduce α-amylase activity [81,117].CGA shows inhibition on porcine pancreatic α-amylase (PPA), PPA-I, and PPA-II [118].CGA can enhance insulin secretion in β cells and Langerhans from rat islets [119,120].CGA can reduce obesity-related insulin resistance in mice fed on HFD or high-fat milk, spontaneously obese mice, or rats fed on HFD [121][122][123].One mechanism underlying CGA's effects on decreasing insulin resistance and increasing insulin sensitivity is related to antioxidative stress.CGA reduces levels of lipid hydrogen peroxide and increases plasma antioxidants such as glutathione (GSH), vitamin C, vitamin E, and ceruloplasmin in DM model rats [124].CGA scavenges thiobarbituric acid reactive substances and hydroperoxide through upregulation of SOD, catalase (CAT), glutathione peroxidase (GSH-Px), and glutathione S-transferase (GST) in the liver and kidney [125].CGA suppresses inflammatory response by downregulation of F4/80+ and CD68+ macrophages in the liver and white adipose tissues [121].CGA increases GSH and GSH-Px and reduces ROS, thus protecting β cells from exposure to streptozotocin (STZ) [120].In STZ-induced DM rats, CGA (5 mg/kg/day, 45 days) in combination with tetrahydrocurcumin (80 mg/kg/day, 45 days) can mitigate the STZ-induced aberrances of enzymes related to gluconeogenesis (G6Pase and fructose-1,6-bisphosphatase) and glycolysis (glucokinase and hexokinase), thus lowering the levels of blood glucose and glycosylated hemoglobin (HbA(1C)) and elevating the levels of insulin, C-peptide, hemoglobin, and glycogen [126].1D) CGA reduces glomerular hypertrophy and proliferation and mesangial cell expansions, decreases kidney malondialdehyde (MDA) levels, increases antioxidants (such as SOD, CAT, and GSH-Px), and reduces factors associated with oxidation and inflammation (such as IL-6, TNF-α, COX-2, and IL-1β) in the kidney of a diabetic nephropathy rat model [127,128].CGAcontaining extracts suppress vascular proliferation in kidneys induced by STZ and decrease serum VEGF levels induced by HIF-1α in DM mice [129,130].In a diabetic retinopathy rat model, CGA shows restoration of the impaired tight junction protein occludin, mitigation of aberrant retinal vascular permeability, and protection of the integrity of the blood-retinal barrier [131].In DM mice, CGA alleviates diabetic peripheral neuropathy (DPN)-induced auditory dysfunction by functional restoration of cochlear hair cells and protection of the external auditory canal [132].CGA can relieve DM-induced neuropathic pain [133].

Human Subject Studies for Glycemic Control
CGA can attenuate FSG and insulin production in patients [57].In a randomized, double-blind, placebo-controlled crossover study to evaluate acute response, a one-time intake of green tea catechins (GTC) together with coffee CGA significantly increased GLP-1 and decreased blood sugar levels and GIP secretion in healthy subjects compared with the placebo group after consumption of a 75 g glucose load [134].This data was echoed by a related study showing that a three-week intake of GTC + CGA-enriched beverages exhibited similar beneficial effects in postprandial glycemic control and diabetic prevention [135].In a cohort of subjects with prediabetic impaired fasting glucose (IFG), CGA-rich Cynarascolymus (Cs) extracts (n = 27) or placebo (n = 27) were administered.The subjects in the treatment group showed significant improvements in glycemic control, insulin sensitivity, and many metabolic parameters (TC, LDL-c, HDL-c, TG, ApoA, ApoB, and glycated hemoglobin) [136].In a randomized clinical trial in patients with metabolic syndrome, participants with an intake of GCE (400 mg, twice per day, 8 weeks) significantly decreased SBP, FBS, homoeostatic model of assessment of insulin resistance, waist circumference, and appetite scores in comparison to those in the placebo group [8].

Hepatoprotection
CGA can mediate hepatoprotective roles in various pathological conditions of the liver via antioxidant and anti-inflammatory features [4].(1) It can inhibit TLR4-mediated activation of NF-κB, thus suppressing pro-inflammatory responses; (2) it can activate the AMPK pathway to modulate metabolic homeostasis; (3) it can increase the activity of the Nrf2 pathway, thus exerting antioxidant effects; and (4) it can inhibit caspases' activation to suppress hepatic apoptosis induced by chemicals or toxins.

Mitigative Effects on Metabolic-Associated Fatty Liver Disease (MAFLD) (Figure 1E)
CGA inhibits HMG-CoA reductase, thus reducing the quantity of palmitic acid, oleic acid, or linoleic acid-induced large lipid droplets in the hepatic cell line HepG2 [162,163].CGA attenuates MAFLD in HFD mice by increasing the production of glucagon-like peptide-1, reducing ER stress, suppressing mucosa barrier injury in the intestine, and inhibiting JNK signaling, as a result of autophagic suppression and insulin-resistant mitigation [123,164,165].
In an α-naphthylisothiocyanate-induced mouse model with cholestatic liver injury, CGA suppresses cell death and neutrophilic and monocytic infiltration and reverses dysregulated hepatocyte transporters and enzymes related to synthesis, uptake, metabolism, and efflux of bile acids [43,174].In a rat model of hepatic ischemia/reperfusion injury, CGA attenuated liver damage by suppressing HMGB1/TLR-4/NF-κB signaling and mitochondriamediated apoptosis [175].

Human Subject Studies for Hepatic Protection
In a clinical study with subjects with NDFLD in type 2 DM, neither CGA nor caffeine showed significant effects on improving stiffness of the liver and other hepatic outcomes.The TC was lower in the caffeine group and insulin was higher in the CGA plus caffeine group than in the placebo group, respectively [185].In a randomized controlled clinical trial with HCC patients (n = 291) transcatheter arterial chemoembolization (TACE) therapy was administered, with or without FZJDXJ, a Chinese medicine formulation, for 48 weeks.The active ingredients of FZJDXJ included formononetin, CGA, caffeic acid, luteolin, gallic acid, diosgenin, ergosterol endoperoxide, and lupeol, which might potentially target AKT/CyclinD1/p21/p27 pathways.In addition, molecular docking showed that CGA and gallic acid could effectively interact with the phosphorylation site Thr308 of AKT1.FZJDXJ and TACE treatment significantly prolonged one-year overall survival (OS) and progression-free survival (PFS) of patients compared with TACE treatment alone [186].

Neuroprotection
CGA has shown diverse neuroprotective effects on various neuropathological conditions which may be exerted through inhibition of neuroinflammation, reduction in ROS production, prevention of oxidation, and suppression of neuronal apoptosis [187][188][189][190].

Mitigative Effects on Alzheimer's Disease (AD) (Figure 1F)
CGA or extracts containing CGA can inhibit Aβ aggregation-caused cellular injury in SH-SY5Y cells, a neuroblastoma cell line, and PC12 cells [210][211][212][213].It suppresses the Aβ1-42 self-induced aggregation in PC12 cells [213].In Aβ-treated hippocampal neurons, CGA increases survival and decreases apoptosis via decreasing activities of lactate dehydrogenase (LDH) and the levels of MDA and raising the levels of SOD and GSH-Px [214].CGA facilitates Aβ clearance and cognitive improvement by enhancing the expression of hippocampal LDL receptor-related protein 1 and restoring perivascular deposition of aquaporin 4 [215].
CGA prevents Aβ deposition and neuronal loss and ameliorates learning and memory deterioration in APP/PS2 mice [216].CGA restores spatial learning and memory in SAMP8 mice, a mouse model showing plaques with Aβ depositions and age-related cognitive defects [217].CGA inhibits acetylcholinesterase (AChE) activity in rat brains, suggesting its beneficial effect against cognitive impairment [218,219].Molecular docking simulations suggest that CGA can bind towards AChE [220].CGA inhibits AChE, decreases the hippocampal and frontal cortical levels of MDA, and improves the deteriorated short-term or working memory and defective cognition induced by scopolamine, a muscarinic receptor antagonist [203].

Mitigative Effects on Parkinson's Disease (PD) (Figure 1F)
CGA has demonstrated preventative effects against PD.CGA improves the decrease in α-synuclein-induced cell viability and blocks the interplay between oxidized dopamine and α-synuclein [221].CGA attenuates the 6-OHDA-caused apoptosis of SH-SY5Y cells [222,223].CGA combined with caffeic acid prevents rotenone-caused Parkinsonian pathology in nigral dopaminergic and intestinal enteric neurons [224].CGA enhances the expression of tyrosine hydroxylase and anti-inflammatory cytokine IL-10 and reduces the drug-induced neuroinflammatory factors such as IL-1β, TNF-α, and NF-κB in substantia nigra [192,225].CGA inhibits the activation of pro-apoptotic proteins including Bax and caspase-3 and elevates the levels of anti-apoptotic factors such as Bcl-2 [226].

Effects on Cognitive Function (Figure 1F)
CGA protects against anxiolytic and depressive processes in a mouse model of anxiety [234].CGA improves cognitive impairments in sleep-deprived mice via immunomodulatory effects and gut microbial metabolic modulation.The potential contributive mechanism was Nrf2/PPAR activation [235].CGA attenuates the polarization of macrophages and alleviates cognitive impairments in an LPS-induced neuroinflammation mouse model by targeting the TNFα signaling pathway [236].CGA improves memory dysfunction and attenuates frontal cortex inflammation in diabetic rats [237].Dried loquat fruit extract containing CGA improves corticosterone-induced depression-like behaviors in mice [238].

Modulation of Neuropathic Pain (Figure 1F)
Neuropathic pain is related to immunomodulation and inflammatory response [18].CGA shows antinociceptive efficacies in pains related to tonic and inflammations and chronic neuropathy [133,[239][240][241], which may be a result of CGA's anti-inflammatory activities on suppression of peripheral release of many pro-inflammatory factors, including TNF-α, NO, and ILs [239,242,243].Oxidative stress involves all stages of neuropathy and its related pain since free radicals are key mediators causing peripheral nerve injury [244,245].Data have demonstrated that ROS is a crucial contributor to the development of neuropathic and inflammatory pain [246][247][248][249][250], which can be attenuated by various phenolic antioxidants [251][252][253].CGA has strong antioxidant activities for scavenging free radicals such as ROS [254].It is reasonable to posit that CGA can reduce neuropathic pain by scavenging ROS.
CGA-enriched herb extracts execute antinociceptive actions in various animal models [255,256].Acidosis-induced and trigeminal nociceptive pain can be reduced by CGA [257,258].CGA can suppress the inflammatory cascade and decrease mechanical and cold hyperalgesia in the rat model of chronic constrictive nerve injury (CCI) [240,241].The underlying mechanism is probably realized through facilitating the activation of gammaaminobutyric acid A (GABA A ) receptors in the spinal cord, a major inhibitory neuronal transmission for pain modulation [259,260] (Figure 2E).However, CGA seems ineffective in mitigating acute pain [133].
CGA may directly act on ion channels related to neuropathic pain for its mitigative effects.For example, voltage-gated potassium channel subfamily A member 4 (Kv1.4),which is specifically expressed in nociceptive sensory neurons in small diameters (A δ and C fibers), is involved in neuropathic pain when its function is suppressed [261,262].Kv activities are upregulated by CGA in trigeminal ganglions on the basal level and PGE 2 -induced inflammations [263,264], leading to an attenuation of neuronal excitability-related pain induction [264][265][266][267] (Figure 2E).Furthermore, CGA can suppress acid-sensing ion channels in sensory ganglions [257,268], presenting another potential peripheral antinociceptive pathway.

Human Subject Studies for Neuroprotection
Several studies show that regularly prolonged intake of CGA has positive effects on cognitive function in humans [269][270][271].In a cohort of healthy subjects with self-description of memory decline (n = 38, 50-69 years old), individuals were given a CGA-enriched beverage or placebo for 4 months.The data showed that CGA improved some categories of cognitions (such as attention shifting, function of execution, and motor and psychomotor speed) and increased plasma levels of early cognitive impairment biomarkers such as apolipoprotein A1 and transthyretin [270].In another cohort of the elderly with subjective memory complaints (n = 8), subjects were administered CGA (330 mg) for 6 months, and similar improvements were observed including memory for composition and verb use, cognition of flexibility, function of execution and attention, and motor speed.Furthermore, there were reductions in the plasma levels of Aβ42 and Aβ42/Aβ40 and an increase in the plasma level of dehydroepiandrosterone sulfate [271].In a recent randomized controlled trial on 34 individuals with mild cognitive impairment who were administered two periods of CGA (554 mg of CGA or placebo, twice/day) for 3 months with a monthly interval, data showed improvements in cognitive functions, especially attention and executive function [269].In a randomized, double-blind, placebo-controlled crossover study, healthy humans with consumption of CGA-enriched coffee berry extracts increased arousal, but limited cognitive effects were observed [272].Ingestion of CGA (600 mg) over 5 days in healthy subjects (n = 9) shortened sleep latency without effects on sleep architecture, enhanced parasympathetic activity, and increased fat oxidation during sleep [273].

Anticancer Effect
CGA has the role of an anticancer agent in various types of cancer cells by arresting cell proliferation, promoting apoptosis, and facilitating intracellular DNA impairment [13] (Figure 1G).

Breast Cancer
CGA exhibits cytotoxicity in breast cancer cell lines such as MCF-7 with an IC 50 of 127 µM, resulting in DNA injury, cell cycle stall, and apoptosis [274,275].A possible mechanism is that CGA can bind to PKC in the cytosol and translocate it to the plasma membrane, thus disturbing the cell cycle, arresting cells at the G1, and reducing cells in the S phase [274].CGA shows cytotoxicity on breast cancer cell lines such as MDA-MB-231, MDA-MB-453, and 4T1 in dose-and time-dependent manners through downregulation of NF-κB pathway [276].It also modulates the epithelial-mesenchymal transition (EMT) process of breast cancer cells by downregulation of N-cadherin and upregulation of Ecadherin [276].In a breast cancer cell-bearing BALB/c mouse model, CGA suppresses tumor growth by increasing the expression of p53, Bax, and the ratio of Bax/Bcl-2 [276,277].

Colorectal Cancer
CGA can stall the cells in the S phase and cause DNA injury in human colon cancer cell lines such as HCT116 and HT29 by increasing ROS production, upregulation of phosphorylated p53, HO-1, and Nrf2 [278].CGA activates the mitochondrial apoptotic pathway in cancer cells by showing DNA breakdown, cleavage of pro-caspase-9 and PARP-1, and upregulation of Bax and the Bax/Bcl-2 ratio [279].CGA and its metabolites can increase the levels of pro-caspase-3 and activated caspase-3 in human colon cancer cell lines such as Caco-2 [280].CGA combined with lactoferrin arrests SW480 cells at the G 0 /G 1 phase and decreases cell viability [281].

Esophageal Cancer
Evidence reveals that CGA can suppress proliferation and colony formation on many esophageal cancer cell lines such as KYSE30/70/140/150/180/510 [282].In esophageal cancer cell line-bearing non-obese diabetic (NOD)/severe combined immunodeficiency disease (SCID) mouse models, CGA (50 mg/kg) inhibits the propagation and size of the tumor and reduces esophageal hyperplasia, thus extending mouse lifespan.CGA decreases expressions of survivin and SOX2 in esophageal squamous carcinoma [282].

Leukemia
CGA (10-25 µg/mL) causes the apoptosis of Bcr-Abl + leukemia cell lines by an increase in intracellular H 2 O 2, O 2 − and levels of caspases, as well as PARP degradation and suppression of p-STAT-5 and p-CrkL [283].Similar results have been reported in U937 and HL-60 leukemia cells.CGA (50-200 µM) facilitates cancer cell death through the induction of ROS and activation of caspase-dependent signaling, leading to reduction in membrane potentials of mitochondria, DNA damage, and apoptosis [284,285].

Lung Cancer
CGA (2-50 µM) can suppress the progression of human lung cancer cell line A549 by increasing the levels of annexin-V, Bax, and CASP3, activating p38 and Jun, and decreasing Bcl-2 and tumor stem cell markers including NANOG, POU5F1, and SOX2, indicating multiple kinase pathways and ROS signaling underlying CGA-mediated anti-lung cancer activity [286].This finding has been echoed by in vivo experiments using an A549-bearing nude mouse (BALB/c) model, in which CGA (120 mg/kg) reduces their tumor mass and size by binding with annexin A2 and inhibiting the expression of NF-κB downstream antiapoptotic genes, thus suppressing cancer cell growth and migration [287].

Melanoma
CGA (1-1.5 mM) reduces the growth of melanoma C32 cells by increasing the expression of antioxidant molecules such as SOD and GSH-Px, thus decreasing oxidation [288].CGA prevents B16F10 melanoma cell proliferation by facilitating the tumor-associated macrophage (TAM) polarization from M2 to M1. CGA with an anti-PD1 antibody can decrease the CD4 + Foxp3 + T cell ratio and increase the CD8 + T cell ratio, leading to an enhancement of immunotherapeutic activity in vivo [289].

Osteosarcoma
CGA reduces the proliferation of osteosarcoma cell lines such as U2OS, MG-63, and Saos-2 by increasing the activity of caspase-3, caspase-7, and PARP, and inducing apoptosis through the blockage of the STAT3/Snail pathway [294,295].CGA in combination with doxorubicin suppresses cellular metabolic activity, colony formation, and cell growth of U2OS and MG-63 cells by upregulating caspase-3 and PARP and suppressing the p44/42 MAPK pathway, thus inducing apoptosis [296].

Pancreatic Cancer
CGA (100-300 µM) can stall cells at the G 2 /M phase and suppress cell proliferation and colony formation of pancreatic carcinoma cells (PANC-1), which can be synergically enhanced in combination with thermal cycling hyperthermia (TC-HT) (10 cycles) with or without a low-intensity pulsed electric field (LIPEF) [297,298].The underlined mechanism involves CGA-mediated excessive ROS production, causing mitochondrial dysfunction, leading to increases in cleaved levels of caspase-3, caspase-9, PARP, and Bax/Bcl-2 ratio [297,298].These data are further validated by in vivo experiments showing that CGA can reduce tumor growth and volume in pancreatic cancer cell-bearing nude mice by modifying cancer cell metabolism through decreasing levels of cyclin D1, c-Myc, and cyclin-dependent kinase-2 (CDK-2), interrupting mitochondrial respiration, and suppressing aerobic glycolysis [299].

Renal Cell Carcinoma (RCC)
CGA (IC 50 40 µM) selectively suppresses cell proliferation and colony formation of human RCC A498 cells but without effects on human embryonic kidney (HEK293) cells through upregulation of cleaved levels of caspase-3, caspase-9, and PARP and the ratio of Bax/Bcl-2 and inhibition of the PI3K/Akt/mTOR pathway [301].

Human Subject Studies for Cancer Management
In an open-label, dose-escalation phase I trial on patients with recurrent high-grade glioma after standard-of-care treatments (n = 26), CGA was intramuscularly injected into patients once daily for 28 days.The median OS after CGA treatment was 11.3 months, which showed a prolonged trend as compared with the median OS (5.7 to 7.5 months) for patients in similar stages under standard-of-care therapeutics [302].

Skin Protection
CGA has shown diverse dermal protective roles in various skin conditions such as anti-UV-induced photoaging, promoting skin slap survival, improving skin barrier function, mitigating systemic lupus erythematosus (SLE)-like symptoms, and suppressing melanogenesis.
8.1.Dermal Protection against Skin Pathologies (Figure 1H) (1) CGA shows anti-inflammatory and antiaging effects by inhibiting UVA-activated TGF/Smad2/3 signaling, decreasing ROS, pro-inflammatory factors IL-1β and TNF-a, reducing apoptosis and necrosis, attenuating DNA damage, promoting cell repair, and increasing synthesis of collagens in dermal fibroblasts [303,304]; CGA ameliorates deoxynivalenol-induced dermal injury by activating Nrf2 and inhibiting MAPK/NFkb/NLRP3 pathways [305]; (2) CGA promotes skin flap survival in rats by downregulating MDA and NO, upregulating GSH and SOD, and elevating VEGF expression and capillary density, leading to blood perfusion [306]; (3) CGA restores the epidermal skin barrier by upregulation of filaggrin, involucrin, and envoplakin and induction of diverse responses of cytokines in epidermal keratinocytes [307]; (4) CGA has anti-acne vulgaris effects.CGA rescues P. acnes-induced skin lesions in ears including redness, swelling, and erythema, downregulates the levels of pro-inflammatory factors by suppressing NF-κB signaling, and inhibits lipogenesis by attenuating AKT/mTOR/SREBP signaling [308]; and (5) CGA relieves SLE-like skin lesions.CGA down-regulates IL-17 levels, mitigates SLE-caused injuries in the skin and mucous membranes, and improves arthritis-like syndromes in MRL/lpr mice [309]; (6) CGA-containing hydrogel promotes the formation of microvessels from HUVEC cells and proliferation of HaCAT cells.In a skin-wound rat model, CGA hydrogel facilitates the wound-healing process by modulating macrophage polarization, alleviating the production of pro-inflammatory cytokines, enhancing collagen deposition, and increasing the expression of CD31 and VEGF [310].

Anti-Melanogenesis Effects (Figure 1H)
In melanoma B16 cells, CGA likely acts on melanin as a substrate, but its metabolites may inhibit melanogenesis by suppressing tyrosinase activity [311].CGA and caffeic acid derivatives inhibit melanocyte-stimulating hormone (α-MSH)-induced melanogenesis [312][313][314].CGA binds to tyrosinase.The molecular docking simulation of CGA on tyrosinase shows the binding energy of −4.59 kcal/mol through interactions with ARG 321 and ARG 374 residues of tyrosinase.Therefore, CGA has the potential as an anti-hyperpigmentation agent through the inhibition of tyrosinase [315].

Human Subject Studies for Skin Protection
In a randomized, double-blind, controlled clinical study, subjects (n = 46) were administered jujube syrup containing gallic acid (1140 ± 17.65 µg/mL) and CGA (1520 ± 25.77 µg/mL) or placebo (23 in each group) twice a day for 8 weeks.The number of facial pigment spots and pigmented areas and percentages were significantly lower in the participants taking jujube syrup than in those in the placebo group [316].In a double-blind, placebo-controlled study, female subjects with mildly xerotic skin (n = 49) were given a beverage containing coffee polyphenols (CPPs) (270 mg/100 mL/day) or placebo for 8 weeks.The intake of CPPs improved skin barrier and microcirculatory functions by lowering skin dryness, transepidermal water loss, and skin surface pH, increasing free fatty acids and lactic acid in the stratum corneum, and promoting skin blood flow [317].

Anti-Allergic Effect
CGA reduces the allergic response induced by shrimp food in mice, likely by suppressing Acetyl-CoA carboxylase (ACC) and increasing carnitine palmitoyltransferase-1 (CPT-1) and AMPK and ACC phosphorylation [330].

Extending Lifespan in Worms
CGA reduces the generation of ROS in worms and increases their lifespan through the DAF-16/FOXO and Nrf2/SKN-1 signaling axis under normal conditions or in challenge to oxidation [331].CGA can prolong about 20.1% of C. elegans' lifespan by attenuating the ageassociated decrease in body mobility and enhancing stress challenge via DAF-16-regulated insulin/IGF-1 signaling [332].CGA prolongs about 24% and 9% of the lifespans of DAF-16a-and DAF-16f-rescued worms, respectively, through the activation of Nrf2/SKN-1 [333] (Figure 1J).

Other Protective Roles of CGA 11.1. Lung Protective Effects
CGA counteracts paraquat-induced oxidative, fibrotic, and inflammatory injuries to the lungs in rats [334].KAT2A is the crucial regulatory gene for the expression of pro-inflammatory factors.CGA acts as a KAT2A inhibitor, attenuating the acute lung inflammation and improving the impaired respiratory function in a mouse model of LPS-induced acute lung injury [335].In LPS and polyinosinic:polycytidylic acid (POLY I:C)-induced acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) models, CGA counteracts the inflammatory and oxidative stress in human airway epithelial cells and in BALB/c mice through targeting the TLR4/TLR3/NLRP3 inflammasome axis [336].

Intestinal Protective Effects
CGA alleviates intestinal inflammation and injury in broilers induced by necrotic enteritis challenge through suppressing the mtDNA-cGAS-STING signaling pathway [337].In a rat model of post-infectious irritable bowel syndrome (PI-IBS), rectal application of CGA ameliorated PI-IBS-related pathologies, probably by increasing glycine levels and modulating gut microbial-released extracellular vesicles [338].

Ovarian Protective Effects
CGA significantly counteracts oxidative stress, pro-inflammatory, and pro-apoptotic markers in cisplatin (CDDP)-induced ovarian damage in rats [339].CGA mitigates symptoms in patients with polycystic ovarian syndrome (PCOS) and improves follicular development, hormone status, and oxidative stress in PCOS rats, likely through modulating HIF-1alpha signaling [340].

Human Subject Studies for Menopausal Symptom Management
In a randomized, placebo-controlled, double-blind, parallel-group trial with healthy women (n = 82), the effects of CGAs on menopausal symptoms were examined.The subjects were administered CGAs (270 mg) or the placebo for 4 weeks.CGAs significantly decreased the modified Kupperman index of menopausal symptoms and reduced the number of hot flushes, the severity of hot flushes during sleep, and the severity of daytime sweats compared to the placebo group.No adverse effects were observed in the CGAs group [341].
GTC together with coffee CGA; CGA-rich Cs extracts; GCE
Though our current understanding of CGA has been substantially expanded in contemporary years, there are many important scientific gaps yet to be addressed in future studies.(1) The specific molecular targets of CGA on NF-κB, MAPK, and Nrf2 pathways remain elusive.Molecular docking modeling can provide an insightful lead to the direct interaction of CGA and targeting molecules, followed by functional characterizations.Furthermore, an in-depth analysis including next-generation sequencing and multiome approaches at tissue and single-cell levels should be applied to reveal a systemic and comprehensive picture of CGA's biological effects at transcriptional, translational, epigenetic, and intermolecular levels.(2) The pharmacokinetic data of CGA is inadequate due to its limited bioavailability.Upon oral ingestion, a significant portion of CGA remains in the colon and becomes metabolized and absorbed into circulation.Therefore, the observed pharmacological effects are likely to result from CGA and its bioactive metabolites, which further impedes the mechanistic interpretation of the data.Furthermore, efforts to modify the structure of CGA or develop novel and effective drug delivery systems such as liposomes, micelles, and nanoparticles for CGA are ongoing and need further validation for bioavailability, tissue distribution, and efficacy.(3) Clinical studies are required to translate CGA efficacy from bench to bedside for patients.Most current studies are performed on in vitro or in vivo models, which may not truthfully recapitulate the pathological conditions in real patients.Moreover, supraphysiological concentrations of CGA are used in many studies, which may lead to a misinterpretation of the value of CGA effects.(4) There has been a rising popularity of green coffee bean powder recently.The consumption of CGA-enriched natural products such as GCE, fruits, and vegetables at a dose equivalent to daily intake may empower a versatile way to extend its health benefits to the general population without any safety concerns.The development of novel approaches for raw material processing to preserve the CGA and other bioactive substances and improve their release and absorption upon ingestion is a task for the dietary supplementary industry.
In summary, recent advances in our understanding of CGAs have supported its therapeutic potential in many disorders.It is necessary to propel properly designed clinical trials and prospective studies to further elucidate and validate its efficacy in clinics.

Figure 1 .
Figure 1.A summary of therapeutic effects of CGG on multiorgan.CGA shows various beneficial roles in many pathological conditions.It can (A) mitigate inflammatory response and oxidative stress; (B) modulate glucose and lipid homeostasis and alleviate DMs; (C-E) protect cardiovascular system, kidneys, and liver; (F) facilitate the recovery from neurological impairments such as neurodegenerative disorders and diabetic peripheral neuropathy; (G) inhibit tumor cell proliferation and migration; (H) ameliorate skin pathologies; (I) execute anti-pathogen effects, and (J) exert antiaging effects.↑, increasing; ↓, decreasing.The graph was created with Biorender.com.

Figure 2 .
Figure 2. The potential mechanistic actions of CGG on multi-targets.CGA can potentially (A) target NF-kB, MPAKs, and JAK pathways to mitigate inflammation; (B) activate Nrf2-dependent and independent pathways to execute antioxidation function; (C) regulate lipid metabolism through increasing lipolysis and fatty acid oxidation and suppressing synthesis of cholesterol and fatty acids; (D) modulate glucose metabolism through increasing glycolysis and suppressing glucose uptake and glucose synthesis; and (E) exhibit neuromodulation through targeting multiple neuroreceptors and ion channels.The graph was created with Biorender.com.

Figure 1 .
Figure 1.A summary of therapeutic effects of CGA on multiorgan.CGA shows various beneficial roles in many pathological conditions.It can (A) mitigate inflammatory response and oxidative stress; (B) modulate glucose and lipid homeostasis and alleviate DMs; (C-E) protect cardiovascular system, kidneys, and liver; (F) facilitate the recovery from neurological impairments such as neurodegenerative disorders and diabetic peripheral neuropathy; (G) inhibit tumor cell proliferation and migration; (H) ameliorate skin pathologies; (I) execute anti-pathogen effects, and (J) exert antiaging effects.↑, increasing; ↓, decreasing.The graph was created with Biorender.com.

Figure 1 .
Figure 1.A summary of therapeutic effects of CGG on multiorgan.CGA shows various beneficial roles in many pathological conditions.It can (A) mitigate inflammatory response and oxidative stress; (B) modulate glucose and lipid homeostasis and alleviate DMs; (C-E) protect cardiovascular system, kidneys, and liver; (F) facilitate the recovery from neurological impairments such as neurodegenerative disorders and diabetic peripheral neuropathy; (G) inhibit tumor cell proliferation and migration; (H) ameliorate skin pathologies; (I) execute anti-pathogen effects, and (J) exert antiaging effects.↑, increasing; ↓, decreasing.The graph was created with Biorender.com.

Figure 2 .
Figure 2. The potential mechanistic actions of CGG on multi-targets.CGA can potentially (A) target NF-kB, MPAKs, and JAK pathways to mitigate inflammation; (B) activate Nrf2-dependent and independent pathways to execute antioxidation function; (C) regulate lipid metabolism through increasing lipolysis and fatty acid oxidation and suppressing synthesis of cholesterol and fatty acids; (D) modulate glucose metabolism through increasing glycolysis and suppressing glucose uptake and glucose synthesis; and (E) exhibit neuromodulation through targeting multiple neuroreceptors and ion channels.The graph was created with Biorender.com.

Figure 2 .
Figure 2. The potential mechanistic actions of CGA on multi-targets.CGA can potentially (A) target NF-kB, MPAKs, and JAK pathways to mitigate inflammation; (B) activate Nrf2-dependent and independent pathways to execute antioxidation function; (C) regulate lipid metabolism through increasing lipolysis and fatty acid oxidation and suppressing synthesis of cholesterol and fatty acids; (D) modulate glucose metabolism through increasing glycolysis and suppressing glucose uptake and glucose synthesis; and (E) exhibit neuromodulation through targeting multiple neuroreceptors and ion channels.The graph was created with Biorender.com.

Author Contributions:
Conceptualization, W.T.; writing-original draft preparation, V.N., E.G.T. and W.T.; writing-review and editing, T.C. and W.T.; visualization, D.M.; supervision, W.T.; project administration, V.N.; funding acquisition, T.C. and W.T. All authors have read and agreed to the published version of the manuscript.Funding: This work was supported by grants from the National Institute of Arthritis and Musculoskeletal and Skin Diseases of the National Institutes of Health (R01AR073172 and 1R21AR083066 to W.T.) and Department of Defense (HT9425-23-10008 to W.T.), and National Heart, Lung, and Blood Institute (R01HL160541 to T.C.) and National Center for Advancing Translational Science (R03TR004450 to T.C.).

Table 1 .
A summary of the potential mechanisms underlying CGA's pharmacological activities and related experimental models.