Influence of the Circadian Clock Gene DBP on Pharmacokinetic Parameters: A Comprehensive Review

DBP, a representative PAR leucine zipper transcriptional activator intricately involved in circadian rhythm regulation, has garnered considerable attention for its potential therapeutic role in ameliorating metabolic disorders and modulating pharmacokinetic processes. In this review, we systematically elucidate the mechanisms underlying DBP’s involvement within the circadian rhythm regulatory network and its subsequent influence on pharmacokinetics. DBP, functioning as a D-box binding protein, is transcriptionally regulated by the CLOCK/BMAL1 signaling pathway and interacts synergistically with ROR/REV-ERB-mediated transcriptional regulation of NFIL3, ultimately controlling the expression of core circadian genes, such as PER. Furthermore, DBP significantly affect drug absorption and metabolism in peripheral tissues by modulating various drug-metabolizing enzymes, transporter proteins, and hormonal pathways. These findings underscore the potential of DBP as a key regulator of circadian rhythm disorders and a pivotal factor in optimizing drug bioavailability. The comprehensive summary presented herein highlights DBP’s clinical significance in addressing circadian-related pathologies, managing associated chronic diseases, and improving pharmacotherapeutic efficacy. Additionally, this review provides novel insights and therapeutic targets that could inform future advancements in drug delivery systems and pharmacological research. Influence of the Circadian Clock Gene DBP on Pharmacokinetic Parameters: A Comprehensive Review Hao Jiang a,b,c#, Xiaoqing Zheng a#, Ying Chen a, Jinyun Luo a, Xiantao Zheng a,Quanlei Qiu d, Lvbo Wu e, Xiaoyan Liu a,b,c * , Affiliations a Guangdong Provincial Key Laboratory of Lingnan Specialty Food Science and Technology, college of Light Industry and Food, Zhongkai University of Agriculture and Engineering, 510080 d Academy of Contemporary Agricultural Engineering Innovations, Zhongkai University of Agriculture and Engineering, Guangzhou 510080, China; c Key Laboratory of Lingnan Specialty Food Green Processing and Intelligent Manufacturing, Ministry of Rural Agriculture, Guangzhou 510080, China d Tibet Bomi Plateau Tibetan Gastrodia Elata Industry Development Co., LTD. Nyingchi 860311, China e Guangdong Xiaoxiaoxie Yizhi Technology Co., LTD. Chaozhou 515638 # Hao Jiang and Xiaoqing Zheng，these authors share first authorship. * Corresponding author: Xiaoyan Liu College of Food Science and Technology, Zhongkai University of Agriculture and Engineering, Guangzhou, 510225, China Tel: +86 20 89003827 Fax: +86 20 89003227 E-mail: [email protected]

DBP, a representative PAR leucine zipper transcriptional activator intricately involved in circadian rhythm regulation, has garnered considerable attention for its potential therapeutic role in ameliorating metabolic disorders and modulating pharmacokinetic processes. In this review, we systematically elucidate the mechanisms underlying DBP’s involvement within the circadian rhythm regulatory network and its subsequent influence on pharmacokinetics. DBP, functioning as a D-box binding protein, is transcriptionally regulated by the CLOCK/BMAL1 signaling pathway and interacts synergistically with ROR/REV-ERB-mediated transcriptional regulation of NFIL3, ultimately controlling the expression of core circadian genes, such as PER. Furthermore, DBP significantly affect drug absorption and metabolism in peripheral tissues by modulating various drug-metabolizing enzymes, transporter proteins, and hormonal pathways. These findings underscore the potential of DBP as a key regulator of circadian rhythm disorders and a pivotal factor in optimizing drug bioavailability. The comprehensive summary presented herein highlights DBP’s clinical significance in addressing circadian-related pathologies, managing associated chronic diseases, and improving pharmacotherapeutic efficacy. Additionally, this review provides novel insights and therapeutic targets that could inform future advancements in drug delivery systems and pharmacological research. Key Words: Circadian Rhythm; DBP; Circadian Clock Gene; Pharmacokinetics 1 Introduction Circadian rhythms, commonly referred to as biological clocks, represent intrinsic oscillations in physiological and behavioral processes that follow approximately 24-hour cycles. These rhythms are primarily regulated and sustained by an endogenous biological clock system consisting of autonomous oscillations persisting even in the absence of external entrainment stimuli (Walker, Walton, DeVries, & Nelson, 2020). Nevertheless, several extrinsic factors, including genetic mutations, substance abuse, jet lag, and shift work, can disrupt circadian rhythm synchronization, leading to various circadian rhythm disorders and metabolic dysfunction (Manoogian & Panda, 2017). The Circadian Rhythm Factor (CRF) is particularly sensitive to rhythmic environmental changes, such as day-night cycles, ambient temperature variations, and sunlight exposure. CRF orchestrates a multitude of endogenous physiological responses that collectively facilitate the adaptation, synchronization, and regulation of cyclic hormonal secretions, thermoregulatory processes, sleep-wake cycles, and overall behavioral patterns (Crislip et al., 2022). When the expression of circadian rhythm factors is disrupted or desynchronized from environmental signals due to circumstances such as jet lag, sleep deprivation, or irregular dietary habits, it results in altered amplitude, period, and overall rhythmicity of circadian gene expression. Such perturbations profoundly influence appetite and sleep patterns, ultimately contributing to the development of metabolic and psychological disorders, including diabetes, obesity and depression (Neves, Albuquerque, Quintela, & Costa, 2022). The intrinsic clock system is central to these mechanisms and is responsible for generating and maintaining the circadian rhythms. This system influences various physiological functions by modulating intracellular hormone secretion and complex behavioral patterns. Notably, circadian factors significantly affect the bioavailability of nutrients and pharmacokinetic profiles of therapeutic agents, thereby altering their absorption, distribution, metabolism, and elimination. Indeed, variations in drug efficacy influenced by circadian factors can exhibit more than ten-fold differences (Dallmann, Okyar, & Lévi, 2016). Consequently, nutrient utilization efficiency and drug metabolic kinetics are intricately linked to the circadian timing systems. Recent studies have highlighted the critical role of the circadian protein D-site-binding protein (DBP) in modulating rhythmic gene expression related to metabolic enzymes and transporters. DBP regulates the circadian expression patterns of drug-metabolizing enzymes, particularly the cytochrome P450 (CYP450) family, through specific interactions with its target genes at the D-site. This regulatory pathway profoundly affects drug efficacy and toxicity, underscoring DBP’s significance in mediating pharmacokinetic variability and nutrient metabolism efficiency (Yanke Lin et al., 2019). Therefore, elucidating the mechanistic interplay between circadian rhythm factors, particularly DBP, and pharmacokinetics is essential. Optimizing drug administration timing and frequency according to individual circadian profiles promises substantial improvements in therapeutic efficacy and a reduction in adverse effects. In this review, we specifically focus on DBP to examine their regulatory influence on metabolic enzyme systems and transporters, aiming to clarify the underlying mechanisms linking circadian biology to pharmacokinetic responses. 2 Molecular mechanisms of the circadian system Circadian rhythms are orchestrated and maintained by an intricate biological clock system fundamentally composed of three key components: input pathways responding to temporal cues (e.g., light, nutrition, and temperature), a self-sustained central oscillator (the pacemaker), and output pathways responsible for regulating behavioral, physiological, and metabolic processes (Takahashi, 2018). Peripheral biological clocks, such as those in the kidney, liver, and muscle, synchronize their rhythmic activity according to environmental factors, such as variations in light intensity and feeding schedules. At the molecular scale, the core biological clock operates through a sophisticated transcriptional network involving three major regulatory elements: E-box, D-box, and RORE motifs (Lin, Feng, DeOliveira, & Crane, 2023; Patke, Young, & Axelrod, 2020). Central to this regulatory network is CLOCK, a heterodimer formed by the circadian locomotor output cycles kaput (CLOCK) and brain and muscle ARNT-like protein 1 (BMAL1). This heterodimer functions as a primary circadian transcriptional activator that drives the expression of core clock genes (Period [Per1, Per2, Per3], Cryptochrome [Cry1, Cry2], Rev-erb, Ror, Dbp, Tef, Hlf, and E4bp4) and various clock-controlled genes (CCGs), subsequently activating negative feedback loops crucial to the rhythmic oscillations of the circadian system (Figure 1). As rhythmic proteins such as PER and CRY accumulate in the cytoplasm, they dimerize and associate with casein kinase 1 isoforms δ（CK1δ）or ε (CK1ε) (Fan, Song, Qin, & Lin, 2023), facilitating their recognition by E3 ubiquitin ligases and subsequent proteasomal degradation (X. Cao, Yang, Selby, Liu, & Sancar, 2021). The resulting trimeric complex (PER: CRY:CK1δ/ε) translocates to the nucleus, where it interacts directly with CLOCK/BMAL1, suppressing their transcriptional activity. This cyclical transcription-translation feedback loop (TTFL) constitutes the central molecular mechanism governing the circadian rhythm, maintaining the transcriptional homeostasis of core clock components within the organism (Chiou et al., 2016; Janoski, Aiello, Lundberg, & Finkielstein, 2024; Ye et al., 2014). In parallel, circadian regulation involves a complementary feedback mechanism mediated by retinoic acid-related orphan receptors (RORα, β, and γ) and BMAL1. Specifically, ROR proteins bind to the RORE element located in the promoter region of the BMAL1 gene, activating BMAL1 transcription, and subsequently facilitating the expression of nuclear factor interleukin 3-regulated protein (NFIL3or E4BP4), which promotes the transcription of REV-ERB proteins. Conversely, REV-ERB proteins negatively regulate BMAL1 transcription by competitively binding to RORE elements, serving as critical repressors within the circadian feedback loop (Amador et al., 2018; Ikeda et al., 2019). Accumulating evidence highlights that ROR (α, β, γ) and REV-ERB proteins selectively bind to DNA response elements across various tissues, establishing a dynamic regulatory loop that regulates the expression levels of nuclear receptors REV-ERBα/β and CLOCK/BMAL1. Additionally, CLOCK/BMAL1-mediated expression of D-box binding protein (DBP) and ROR/REV-ERB-mediated transcription of NFIL3 form a tertiary feedback loop within the circadian system (Costello, Johnston, Juffre, Crislip, & Gumz, 2022) (Figure 2). DBP and NFIL3, which act as transcription factors that interact with D-box elements, modulate a broad spectrum of circadian clock gene expressions. For example, the DBP-mediated pathway predominantly drives gene activation during the day, whereas the NFIL3-mediated inhibitory pathway primarily functions at night (Takahashi, 2017). The synergistic and temporally regulated interactions between DBP and NFIL3 ensure robust transcriptional control of the circadian genes. Consequently, the interplay among these three central regulatory elements, the E-box, D-box, and RORE, underpins the intricate auto-regulatory feedback loop essential for the precise maintenance of circadian homeostasis and temporal coordination of biological clock gene expression. 3 Role of DBP in the circadian system DBP (D Site Albumin Promoter Binding Protein) is a transcriptional activator that belongs to the PAR leucine zipper transcription factor family. It dimerizes through its carboxy-terminal leucine zipper and binds to DNA via an adjacent alkaline region. This structural feature enables DBP to regulate circadian rhythms (Yuan et al., 2015). Its regulatory capacity is closely associated with variations in its DNA-binding sites and sequence specificity. Previous studies have demonstrated that the availability and number of DBP-binding sites fluctuate throughout the circadian cycle. For instance, comparisons of nuclear extracts from mouse liver tissues at different circadian time points revealed 6,066 overlapping binding sites for DBP and E4BP4 at zeitgeber time 12 (ZT12), whereas only 1,490 common binding sites were identified at ZT24, primarily clustered around the transcription start site (TSS) (Fang et al., 2014). This temporal variation underscores the circadian modulation of DBP expression levels, binding site specificity and frequency. The TTATGCAA sequence exhibited the highest binding affinity at sites commonly recognized by both DBP and its antagonist E4BP4, highlighting DBP’s essential role within the circadian system. DBP performs this function by binding to specific DNA sequences, particularly D-box elements, thereby coordinating downstream transcriptional input and output within the circadian regulatory network (Yoshitane et al., 2019)(Figure 2). Interestingly, the D-box sequence recognized by both DBP and E4BP4 also encompasses the promoter region of the Per2 gene (TTATGTAA). This finding elucidates a potential direct regulatory relationship between DBP and the circadian marker Per2, suggesting that DBP binds to and activates the transcription of the mPer2 gene promoter. In peripheral tissues, DBP is implicated in the synchronization of central circadian rhythms, influencing physiological metabolism, and behavioral responses. Yoshikazu Morishita demonstrated that phosphatidylinositol 3-kinase (PI3K) directly modulates the transcript levels of key circadian proteins and regulates DBP via the heterodimerization of BMAL1 and CLOCK, essential components in rhythmic DBP transcription (Sopasakis et al., 2010). PI3K, a crucial mediator of intracellular signaling pathways, participates in diverse biological functions, including cell proliferation, motility, and glucose metabolism. Emerging evidence indicates an intricate relationship between peripheral circadian transcriptional mechanisms and intracellular signaling pathways, highlighting DBP’s pivotal role in modulating gene expression in peripheral tissues (Morishita, Miura, & Kida, 2016; Sopasakis et al., 2010). Notably, DBP expression exhibits distinct oscillatory cycles and amplitudes in the suprachiasmatic nucleus (SCN) and peripheral tissues. Unlike typical circadian peaks during the daytime (ZT4) and troughs early at night (ZT16), DBP mRNA expression in the cerebral cortex demonstrates a delayed rhythm with oscillation periods of 4-8 h. This finding suggests that DBP can autonomously regulate divergent circadian cycles within peripheral biological clocks, independent of the SCN, and may even influence rhythms within the central biological clock. Peripheral circadian rhythms are significantly modulated by environmental cues such as light exposure and feeding schedules. For instance, food intake enhances DBP gene expression in cardiac tissue, thereby advancing the circadian oscillatory phases; however, the same feeding regimen does not affect DBP expression in renal tissues (Wu et al., 2012). Consequently, external factors, including sleep disorders, shift work, and irregular dietary habits, can partially disrupt peripheral circadian rhythms, causing temporal misalignment relative to the central clock. These disruptions further exacerbate physiological dysfunction in organisms. Laboratory and epidemiological studies have indicated that at least 15% of circadian genes within the SCN are directly regulated by peripheral circadian clocks. Chronic dysregulation of the central and peripheral biological clocks is associated with numerous chronic conditions, such as obesity and metabolic syndrome, ultimately impairing immune and metabolic functions (Bozek et al., 2009). In particular, research involving mouse models of type 2 diabetes mellitus (T2DM) revealed a significantly reduced amplitude of DBP daily oscillations compared to healthy controls (Su et al., 2012). This reduction impairs translational efficiency, affecting protein accumulation and degradation pathways, thus contributing substantially to glucose metabolism disorders and T2DM pathogenesis. Collectively, these findings underscore that DBP critical function in maintaining glucose homeostasis by regulating circadian rhythms. 4 Pharmacokinetics of DBP in the circadian system DBP plays a crucial role in regulating circadian rhythm disorders, ameliorating associated metabolic diseases, and influencing the pharmacokinetics, bioavailability, and therapeutic efficacy of drugs. Pharmacokinetic processes are conventionally divided into absorption, distribution, metabolism, and excretion (collectively known as ‘ADME’), each of which significantly influences drug and metabolite concentrations within target tissues and organs (Figure 3). Consequently, these processes are pivotal determinants of drug efficacy and toxicity (Dong, Yang, Lin, Wang, & Wu, 2020). Investigating the mechanisms linking the circadian rhythm factor DBP with pharmacokinetic processes from pharmacokinetic and pharmacotoxicological perspectives can profoundly impact drug bioavailability. Such insights are invaluable for optimizing drug delivery systems and developing precise dosing regimens. 4.1 Effect of the circadian factor DBP on frug absorption Drug absorption critically dictates the bioavailability and therapeutic efficacy of pharmaceuticals within the body and is influenced by several physiological parameters, including gastric acid secretion, gastric emptying time, gastrointestinal motility, and blood flow (Smolensky, Siegel, Haus, Hermida, & Portaluppi, 2011)(Figure 3). Once orally administered drugs enter the stomach, digestion and enzymatic interactions are initiated and mediated by gastric acids and digestive enzymes. Thus, variations in gastric acid secretion and metabolic enzyme activity, which are known to be modulated by circadian rhythms, significantly affect drug absorption. Empirical evidence indicates that pharmacokinetics under circadian control directly regulate drug and metabolite concentrations in plasma and tissues. For instance, studies have demonstrated that the area under the plasma concentration-time curve (AUC) for nifedipine was higher following administration at 8:00 a.m. than following doses administered at 4:00 p.m. or midnight, with the shortest time to peak plasma concentration (C max ) observed following 8:00 a.m. dosing. This underscores the direct influence of circadian rhythms on drug absorption and implies that absorption capacity is closely associated with the expression patterns of circadian rhythm-related proteins (Q. R. Cao, Kim, Choi, & Lee, 2005). Furthermore, studies have reported increased gastric acid secretion at night than during the day, with disturbances such as irregular eating habits, shift work, or extensive travel correlated with gastrointestinal symptoms, including abdominal distension, pain, altered bowel habits, constipation, and diarrhea. These findings suggest that nutrient absorption and drug transporter proteins are under circadian regulation, with certain processes directly controlled by clock genes (Ohdo, 2010; Voigt, Forsyth, & Keshavarzian, 2019). Saito et al. (Saito, Terada, Shimakura, Katsura, & Inui, 2008) observed that the circadian rhythmic fluctuations of intestinal PEPT1 coincided with the expression pattern of the circadian gene DBP under two distinct feeding schedules. Conversely, intestinal transcription factors, such as Sp1, Cdx2, and PPARα, were not associated with the circadian regulation of DBP mRNA expression. The intestinal H/peptide cotransporter protein 1 (PEPT1) is a proton-dependent oligopeptide transporter predominantly localized at the brush border membrane of epithelial cells in the small intestine, facilitating the absorption of protein degradation products, specifically di-and tripeptides (Stelzl, 2018). Subsequent analyses demonstrated that DBP transactivates the expression of rPEPT1 by interacting with a binding site located between base pairs -6,314 and -6,305 in the D-box, suggesting DBP’s regulatory role through distal promoter interactions. This binding consequently influences PEPT1 transcriptional activity, modulating the cellular oligopeptide uptake via the inward proton gradient across the brush border membrane. These findings underline the critical role of DBP as a circadian factor in the rhythmic oscillation of PEPT1 expression (Daniel, 2004). Moreover, a growing body of evidence indicates that the absorption of various therapeutic drugs is significantly affected by the expression and activity of specific intestinal transport proteins. Studies have demonstrated that PEPT1 transporter activity is notably inhibited in rats with disrupted circadian rhythms due to altered fasting patterns, resulting in substantial differences in the intestinal absorption and bioavailability of the antibiotic cefbutene when compared to normal controls (Pan, Terada, Okuda, & Inui, 2003). This observation prompted the hypothesis that targeting intestinal PEPT1 may enhance the drug absorption efficiency. Furthermore, the circadian clock gene DBP, a critical transcriptional regulator of intestinal PEPT1, has the potential to modulate drug absorption by influencing the expression of intestinal transport-related proteins. This hypothesis was supported experimentally by Hayashi et al. (Hayashi et al., 2010), who reported that ABCB1A gene expression is directly regulated by circadian clock genes. Specifically, they demonstrated that time-restricted feeding triggered oscillatory alterations in DBP expression, subsequently affecting ABCB1A mRNA levels and the activity of P-glycoprotein (P-gp). Their study highlighted that not only was DBP mRNA expression rhythmically altered by advancing feeding schedules by 12 h compared to ad libitum feeding, but similar circadian oscillations were also observed for ABCB1A, a critical transcriptional factor for orally administered drug absorption. P-glycoprotein (P-gp), encoded by the ABCB1A gene (also referred to as MDR-1), belongs to subfamily B of the ATP-binding cassette (ABC) superfamily of transport proteins. It plays an indispensable role in drug absorption processes, facilitating lipid transport from the gastrointestinal tract into systemic circulation and mediating the cellular efflux of drugs, such as digoxin. Notably, the absorption of lipophilic drugs and lipid uptake display pronounced circadian variability, with higher absorption during the daytime than at night, a rhythmic pattern that is absent in CLOCK mutant mice (Cascorbi, 2013; Sukumaran, Almon, DuBois, & Jusko, 2010). Further research has indicated significant suppression of cholecystokinin A (CCK-A) receptor and lipase mRNA expression in the pancreas of CLOCK mutant mice, adversely affecting lipid absorption and suggesting that disrupted circadian rhythms significantly influence lipid metabolism (Ge et al., 2023). These alterations in lipid absorption may be associated with the transcriptional activities of downstream circadian rhythm genes, such as thyrotropic embryonic factor (TEF), hepatic leukemia factor (HLF), and E4BP4, which are regulated by DBP. Further investigations revealed competitive binding interactions between HLF and E4BP4 at the same DNA loci of the MDR1A gene, indicating a regulatory mechanism involving these transcription factors. Specifically, DBP, TEF, and HLF enhanced MDR1A transcriptional activity, whereas E4BP4 acted as a transcriptional repressor, modulating the amplitude of MDR1A gene expression (Leliavski, 2014). Additionally, the expression of HLF and E4BP4 is governed by CLOCK, a core component of the circadian clock machinery (Murakami, Higashi, Matsunaga, Koyanagi, & Ohdo, 2008). Thus, dysfunction in CLOCK substantially disrupts circadian-driven rhythmicity in lipid absorption, highlighting the complex regulatory interplay between circadian genes and intestinal nutrient and drug absorption mechanisms. Other studies have shown that the absence of the DBP upstream factor Per1, a pivotal circadian regulator implicated in intestinal lipid absorption and energy metabolism, significantly disrupts daily fat absorption and its accumulation. Specifically, Per1 loss impairs intestinal fatty acid absorption and perturbs the circadian rhythmicity of bile acid synthesis. This phenomenon may arise due to the regulatory role of Per1, which modulates bile acid synthase activity through a phosphorylation-dependent mechanism mediated by the PER-PKA signaling pathway. Consequently, this modulation affects the enzymatic activities of cholesterol 7α-hydroxylase (Cyp7a1) and sterol 12α-hydroxylase (Cyp8b1), which are critical enzymes involved in lipid metabolism regulation (Ge et al., 2023). Notably, the modulatory function of Per1 in fat absorption is markedly pronounced under fasting and high-fat diet conditions. The underlying molecular mechanism may involve the transcriptional activation of co-regulated genes, such as mPers and mCrys, by CLOCK/BMAL1 heterodimer binding to the E-box promoter elements. Upon reaching a critical threshold, the Per-Cry heterodimer subsequently provides negative feedback, inhibiting CLOCK/BMAL1-mediated transcription. Importantly, DBP acts as a positive regulator of Per and Cry expression, whereas CLOCK/BMAL1 ensures stability within the transcription-translation feedback loop by controlling the rhythmic transcription of Rev-erbα and DBP (Mekbib, 2020). Therefore, the simultaneous deletion of CLOCK and Per1 significantly disrupts DBP expression, leading to dysregulated circadian transcription of downstream target genes, impaired lipid uptake, and diminished synthesis of associated lipases. In addition to lipid uptake, glucose uptake exhibits circadian oscillations closely linked to cyclic changes in the expression of the circadian gene DBP. Dyar et al. demonstrated a marked reduction in muscle glucose uptake capacity in Bmal1 knockout mice in response to glucose stimulation. This diminished glucose can be attributed to the disrupted circadian expression of glucose transporter genes and the rhythmicity of key circadian regulatory genes, such as Per1, Rev-erbα, and DBP. Perturbation of rhythmic gene expression consequently impairs glucose uptake and glycolytic pathways, compromising glucose homeostasis and ultimately resulting in hyperglycemia and glucose intolerance (Dyar et al., 2014). In summary, the circadian clock gene DBP is a promising therapeutic target for individuals predisposed to obesity and those with circadian rhythm disorders. DBP is instrumental in regulating circadian fluctuations that influence the absorption of proteins, carbohydrates, lipids, and drugs. Collectively, these findings provide a robust theoretical framework that enables clinicians to enhance therapeutic strategies and optimize drug efficacy, safety, and cost-effectiveness for disease prevention and treatment. 4.2 Effect of circadian factor DBP on drug distribution The distribution of drugs within various tissues and organs in the human body is governed by multiple factors, including the physicochemical properties of the drug, regional blood flow, functionality of drug transporters and efflux pumps, and the drug affinity to plasma proteins (Currie, 2018)(Figure 3). Additionally, circadian rhythms significantly influence drug distribution due to the rhythmic expression of biological clock factors. Among these, the clock gene D-site Binding Protein (DBP) has emerged as a crucial regulator that modulates drug-targeting efficiency and optimizes the timing of drug release at specific lesion sites. Blood flow profoundly impacts drug distribution in tissues and organs and is subject to considerable regulation by circadian clock mechanisms (Sukumaran et al., 2010). For instance, gastrointestinal blood flow demonstrates substantial diurnal variation, being significantly higher during the daytime than at night, resulting in time-dependent fluctuations in drug availability and efficacy within the body (Bicker, Alves, Falcão, & Fortuna, 2020). Generally, blood flow distribution is primarily directed towards the gastrointestinal tract, skeletal muscles, liver, and adipose tissue, with notable variations depending on physiological states. During physical activity, blood flow is substantially enhanced in the muscles, liver, and other metabolically active organs. In particular, intense exercise can elevate muscular blood flow by approximately 10 – 20-fold compared to that under resting conditions (Joyner & Casey, 2015; Saltin & Mortensen, 2012). This increased perfusion accelerates drug distribution to the target tissue and decreases retention time in the bloodstream, facilitating a more rapid achievement of peak pharmacological effects (Beesley, Noguchi, & Welsh, 2016). Conversely, during the resting state, blood flow to the muscle tissue markedly diminishes, whereas perfusion predominantly shifts towards visceral organs, such as the gastrointestinal tract and adipose tissue. Notably, gastrointestinal blood flow during rest approximates 50% -70% of the levels observed during exercise (Saltin & Mortensen, 2012). Reduced regional blood flow prolongs drug retention within the systemic circulation and slows the rate of achieving peak tissue concentration, thereby extending the duration of the therapeutic effects (Rowland & Tozer, 2010). Hence, circadian fluctuations in biological clock factors, including changes in blood flow, significantly affect drug distribution and therapeutic outcomes. Moreover, DBP influences vascular dynamics by regulating the expression of genes associated with vasodilation, thereby modulating blood flow to various tissues and organs. DBP mediates transcriptional activation pathways involving the CLOCK/BMAL1/PER complex interacting with D-box elements, which subsequently affects the secretion and systemic circulation of vasoactive hormones such as adrenaline and prostaglandins. These hormones critically regulate vasoconstriction and vasodilation, thereby influencing blood flow (Oishi, Koyanagi, & Ohkura, 2011). Animal studies involving Per2 mutant mice have demonstrated endothelial dysfunction characterized by diminished synthesis of vasodilatory mediators, such as nitric oxide and prostaglandins, and enhanced production of vasoconstrictive substances (Beesley et al., 2016). Consequently, DBP emerges as a pivotal regulator of both systemic and localized tissue blood flow through its modulatory effects on gene expression and hormonal balance, thereby significantly impacting drug distribution pharmacokinetics. The ability of drugs and nutrients to penetrate tissue and organ barriers significantly influences their distribution within organisms. Recent studies have shown that lipid-soluble drugs primarily depend on membrane-bound proteins, such as transporters, and their facilitated translocation through specific channels for cross-tissue distribution (Vanni, NP, Di Maggio, & Cooney, 2024). Importantly, the functionality of these transporters is modulated by circadian regulatory systems (Pácha, Balounová, & Soták, 2021). One such transporter, P-glycoprotein (P-gp), an ATP-dependent efflux pump, exhibits pronounced circadian fluctuations in various tissues, including the liver, intestine, and kidney. The selective expression and activity of P-gp substantially limit drug distribution. Research has indicated that the active metabolite of the antihistamine ebastine undergoes P-gp-mediated efflux, leading to marked circadian variations in its intestinal concentrations (Lu, Zhao, Chen, & Wu, 2020). Similarly, the permeability of the Drosophila blood-brain barrier is influenced by magnesium ion concentrations, displaying increased permeability at night compared to during the day. Structurally, the blood-brain barrier comprises endothelial cells lining the capillaries, astrocytic end-feet encapsulating the capillaries, and pericytes embedded in the capillary basement membrane. Circadian rhythms critically regulate permeability, as biological clock proteins modulate intracellular magnesium ion concentrations by controlling gap junction formation. This modulation affects tight junction protein levels within the blood-brain barrier, thereby influencing its permeability (S. L. Zhang, Yue, Arnold, Artiushin, & Sehgal, 2018). Collectively, these findings suggest that drug distribution within tissues and organs is intricately linked to circadian gene expression patterns. The circadian regulation of drug distribution can be further explained by the neuronal activity-driven modulation of PAR bZip transcription factors in brain endothelial cells. Neuronal activity regulates PAR bZip factors through the expression of Nr1d2 (Rev-erb), which participates in Bmal1-positive and Cry1-negative feedback loops, ultimately influencing the expression of circadian proteins, such as DBP, Tef, and Hlf. Elevated neuronal activity at night results in circadian gene expression variations that affect transporter function and drug distribution (Pulido et al., 2020). Additionally, critical components of the biological clock modulate drug distribution by influencing drug binding to plasma proteins. Circadian variations in drug-plasma protein interactions have been demonstrated in multiple studies. For instance, cisplatin, a platinum-based chemotherapeutic extensively employed in cancer treatment, exhibits pronounced circadian fluctuations in plasma protein binding (ranging from 65% to 98%). Peak binding levels correspond closely to circadian rhythmic changes in plasma albumin concentration (Messori & Merlino, 2016). The transcription factor DBP influences plasma protein activity through D-box element on the DNA, thereby altering the binding capacity of cisplatin and affecting its distribution and therapeutic efficacy. Once bound to serum albumin in plasma proteins, cisplatin enters cells through sulfur donor groups and sulfur-containing residues such as Cys34 and forms cross-linking with DNA, disrupting the double helix structure of DNA and thereby inducing apoptosis of cancer cells (Merlino, 2023). Understanding the circadian variations in cisplatin binding can contribute to optimizing chemotherapeutic strategies. Likewise, nonsteroidal anti-inflammatory drugs (NSAIDs) exhibit significant circadian rhythmicity in their binding capacity to plasma protein-binding capacities, peaking notably at zeitgeber times ZT 6 and ZT 18 (Al-Waeli et al., 2020; Bicker et al., 2020). Al-Waeli et al. (Al-Waeli et al., 2020) reported diurnal variations in plasma protein binding, where administering carprofen (20 mg/kg) during the active phase in mice enhanced plasma protein binding rates, stimulated the release of anti-inflammatory cytokines (e.g., IL-13 and IL-4), and increased serum vascular endothelial growth factor levels. These effects were mediated through the enhanced expression of circadian genes, such as Per2 and DBP, ultimately alleviating postoperative pain and facilitating wound healing. Moreover, biological clock genes may optimize drug efficacy by modulating enzymatic activity, thereby improving the therapeutic index. NSAIDs achieve their therapeutic effects by inhibiting the activity of cyclooxygenase (COX) enzymes, consequently suppressing the secretion of the pro-inflammatory factor interleukin-1 beta (IL-1β) and promoting the release of the anti-inflammatory cytokines IL-13 and IL-4 (Al-Waeli et al., 2020; Y. K. Zhang, Yeager, & Klaassen, 2009). Further studies have shown that manipulating the biological clock gene Period 2 (Per2) and its downstream signaling pathways, such as the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), epidermal growth factor receptor (EGFR), and beta-catenin pathways, can enhance drug efficacy and minimize adverse effects (Guo et al., 2015; Tao et al., 2015; Yang et al., 2009). Additionally, DBP, acting as a positive regulator within the circadian feedback loop, modulates the expression of Per genes and other rhythmic factors, thereby influencing drug binding and delivery efficiency. In conclusion, the components of the biological clock significantly affect drug distribution through mechanisms such as blood flow modulation, transporter functionality, and plasma protein binding. Investigating circadian genes, such as DBP and Per, in the context of drug distribution not only facilitates therapeutic optimization, efficacy enhancement, and adverse effect minimization but also lays a robust scientific foundation for precise clinical drug administration. 4.3 Regulation of metabolism by circadian factor DBP DBP is a pivotal regulator of an intricate network of drug-metabolizing enzymes. Drug metabolism involves a sophisticated interplay between phase I enzymes, such as cytochrome P450 family members, quinone oxidoreductase, paraoxonase, and aldehyde dehydrogenase, and phase II enzymes, including glutathione S-transferase, sulfotransferase (SULT), cysteine dioxygenase, and other sulfotransferases (Fukami, Yokoi, & Nakajima, 2022). Collectively, these enzymes constitute an extensive metabolic system for drug biotransformation. The liver is particularly critical for drug metabolism because of its abundance of enzymatic machinery localized within subcellular compartments, including the microsomes, cytosol, and mitochondria (Almazroo, Miah, & Venkataramanan, 2017) (Figure 3). The circadian clock significantly influences hepatic drug metabolism by precisely modulating the metabolic kinetic profiles of various hepatic enzymes, such as cytochrome P450 cholesterol 7α-hydroxylase (Cyp7a1), Cyp3a11, steroid 15α-hydroxylase (Cyp2a4), coumarin 7-hydroxylase (Cyp2a5), and flavin-containing monooxygenases (Fmo), thereby affecting their enzymatic activities. Lipid metabolism requires bile acids to be emulsified before absorption in the intestinal lumen. Interestingly, the secretion of bile acids in organisms exhibits pronounced circadian oscillations, reflecting the integration between circadian rhythm and negative feedback loops associated with bile acid homeostasis (Segers & Depoortere, 2021). Recent studies have elucidated the role of D-binding protein (DBP) in modulating the circadian expression of hepatic Cyp7a1, a pivotal enzyme in bile acid biosynthesis. Remarkably, the expression level of Cyp7a1 displayed a robust circadian variation, with a significantly elevated expression at night compared to daytime, with a night-to-day ratio of approximately 100:1. DNAzyme I footprinting analyses further demonstrated that DBP, through circadian fluctuations in its own activity, can activate Cyp7a1 gene transcription. This activation is achieved by binding to multiple distinct regulatory sites within the Cyp7a1 promoter region, specifically sites B (nucleotides -115/-125), C (nucleotides -172/-195), and D (nucleotides -214/-230), thereby enhancing the enzymatic activity of 7α-hydroxylase (Lee, Alberta, Gonzalez, & Waxman, 1994). Similarly, Lin et al. (Y. Lin et al., 2019)reported that both the circadian rhythmic cycle and oscillation amplitude of the cytochrome P450 enzyme Cyp3a11 were suppressed in mice with the BMAL1 gene. This suppression was accompanied by a trend toward decreased metabolism of drugs such as ouabain and triprolactone, suggesting a potential link between the circadian factor BMAL1 and the metabolic processes of these drugs. Subsequent investigations have indicated that BMAL1 regulates drug metabolism through the transcriptional activation of DBP and Hnf4α, which, in turn, induce Cyp3a11 expression. This regulatory mechanism primarily involves the binding of DBP and Hnf4α to the D-box (located at 234/224 bp) and DR1 elements in the Cyp3a11 promoter, leading to enhanced Cyp3a11 transcription. Notably, DBP overexpression significantly increased the mRNA expression of human Cyp3a4 homolog of mouse Cyp3a11, while leaving other circadian genes, including PER2, CRY1, and REV-ERBα, unaffected (Tong et al., 2019). This observation underscores the pivotal role of the circadian factor DBP in modulating Cyp3a11 and Cyp3a4 expression. Moreover, a strong affinity exists between the Cyp3a4 promoter and DBP-binding sites, which activate transcription through DNA sequences upstream of the transcriptional start site. This interaction subsequently affects detoxification pathways involving xenobiotics and pharmaceuticals (Takiguchi et al., 2007). Similarly, consensus sequences for DBP binding have been identified in the promoter regions (sites A, B, and C) of steroid 15α-hydroxylase (Cyp2a4) and coumarin 7-hydroxylase (Cyp2a5) genes. Cyp2a4 and Cyp2a5 possess characteristic structures of the cytochrome P450 enzyme family and are actively involved in the metabolism of drugs with a steroidal structure, as well as in the metabolism of steroidal compounds, steroid analogs, and coumarin derivatives (Lavery et al., 1999). Studies have demonstrated that hepatic detoxification and drug metabolism pathways are severely compromised in DBP mutant and DBP/Tef/HIf triple knockout mice. Specifically, administration of a low-dose pentobarbital (25 mg/kg), a central nervous system depressant, significantly attenuated the amplitude of Cyp2a4 and Cyp2a5 mRNA expression levels. Furthermore, pentobarbital administration resulted in a substantial prolongation of sleep duration in DBP/Tef/Hlf knockout mice compared to that in wild-type controls (Gachon, Olela, Schaad, Descombes, & Schibler, 2006) (Figure1). These findings collectively suggest that the circadian factor DBP plays a critical role, either directly or indirectly, in regulating the expression of drug-metabolizing enzymes Cyp2a4 and Cyp2a5. In addition to the cytochrome P450 enzyme family, flavin-containing monooxygenase (Fmo) is a significant intrahepatic phase I detoxification enzyme that is highly expressed in the liver of both humans and rodents. The expression of Fmo demonstrates a dose- and time-dependent correlation with drug efficacy, toxicity, and tolerability, and exhibits circadian rhythmic fluctuations during drug metabolism. Pharmacokinetic data indicate a logarithmic relationship between Fmo5 expression and the concentration of hexoketone cacodylate, a metabolite specifically dependent on Fmo5 (Uney, Tras, Corum, Yildiz, & Maden, 2019). Notably, the timing of hexoketone cacodylate administration significantly influenced Fmo5expression; administration at Zeitgeber Time 14 (ZT14) resulted in markedly higher metabolite levels in both hepatic tissue and plasma than administration at Zeitgeber Time 2 (ZT2). This strongly suggests a critical relationship between circadian rhythm and fmo5-mediated metabolism of hexoketone cacodylate. The underlying mechanism of this phenomenon involves the presence of E-box and D-box regulatory elements located in the promoter region of the Fmo5 gene, which serve as binding sites for circadian-regulatory proteins. Specifically, albumin D-site binding protein (DBP) activates Fmo5 expression via its interaction with D-box elements, whereas BMAL1/CLOCK influences DBP and Rev-erbα/E4BP4, which are key factors within this regulatory axis, by interacting with E-box elements. This interplay subsequently modulated Fmo5 transcription (M. Chen et al., 2019. Furthermore, E4BP4 represses Fmo5 expression via a negative feedback loop mechanism (Mitsui, Yamaguchi, Matsuo, Ishida, & Okamura, 2001). Collectively, these regulatory interactions elucidate the observed circadian fluctuations in Fmo5 mRNA levels, protein abundance, and enzymatic activity, which peak at ZT10/14 and decline to trough levels at ZT2–ZT22, respectively. During the active phase, Fmo5 expression predominates because of DBP-driven activation, whereas during the inactive phase, Fmo5 expression is attenuated by the antagonistic activity of E4BP4. Consequently, this regulatory circuit generates a circadian rhythm in drug metabolism. Moreover, recent evidence has demonstrated that carboxylesterase (CES), another critical enzyme within the esterase family, undergoes similar regulatory control mediated by DBP, Tef, and Hlf (X. Chen et al., 2021). These findings highlight dibutyl phthalate (DBP) as a potential modulator of drug metabolism and detoxification through its regulatory effects on the expression of drug-metabolizing enzymes. Furthermore, DBP is a significant nexus linking circadian regulatory pathways to drug metabolism, underscoring its relevance in pharmacokinetic and chronotherapeutic studies. 4.4 Regulation of elimination by the circadian rhythm factor DBP In recent years, drug efflux has been increasingly acknowledged as an essential strategy for mitigating drug-induced toxicity and its adverse effects. This physiological process involves multiple organs, notably the liver, kidneys, and intestines, and entails complex regulatory mechanisms. Accumulating evidence highlights the role of biological clock factors in the regulation of genes involved in drug efflux. This regulation leads to rhythmic fluctuations in renal excretory processes, such as glomerular filtration, tubular secretion, and reabsorption, which are closely aligned with circadian rhythms (Figure 3). For example, the glomerular filtration rate (GFR) and urine output are characteristically higher during the active period (daytime) than during the rest period (nighttime), indicating a strong coupling between excretory function circadian regulatory mechanisms (Firsov & Bonny, 2018; Johnston & Pollock, 2018). Huang et al. (Huang et al., 2013) investigated the localization and circadian rhythmicity of biological clock proteins (BMAL1andPER2) and the clock-controlled output protein, DBP, in the residual kidneys of 5/6 nephrectomized rats (STNx). In sham-operated control rats, the biological clock proteins exhibited normal circadian rhythmicity. However, in the residual kidneys of STNx rats, the rhythmic patterns were notably disrupted, with the peak expression times of BMAL1, DBP, and PER2 advancing by approximately four hours compared to the controls. Moreover, alterations in the localization of BMAL1 and PER2 were observed in the STNx group. BMAL1 expression, typically present in glomerular endothelial cells and tubular interstitial vessels, was also detected in the nuclei of cortical and medullary renal tubular cells. PER2 was primarily expressed in proximal renal tubular cells at the corticomedullary junction, whereas DBP was widely distributed in the nuclei and cytoplasm of glomerular and tubular interstitial microvascular cells. Further studies focusing on the specific localization and regulatory mechanisms of DBP in glomerular filtration and tubular reabsorption revealed that these processes heavily rely on transporters, such as P-glycoprotein (P-gp), organic anion/cation transporters, and members of the solute carrier protein (SLC) family. These transporters exhibit circadian rhythmicity in renal expression, significantly influencing the processing of drugs and metabolic waste products (Yonezawa, Masuda, Yokoo, Katsura, & Inui, 2006). For instance, cisplatin is transported into renal proximal tubular cells by organic cation transporter 2 (SLC22A2/OCT2) and subsequently secreted into the tubular lumen via multidrug and toxin extrusion protein 1 (MATE1), encoded by Slc47a1 (Nakamura, Yonezawa, Hashimoto, Katsura, & Inui, 2010; Oda et al., 2014). Concurrent investigations have established a robust association between the circadian proteins CLOCK and DBP and the organic cation transporters. Specifically, CLOCK and DBP modulate the expression of Slc22a2 mRNA by interacting with peroxisome proliferator-activated receptor-α (PPARα), thereby regulating OCT2’s rhythmic expression and its ability to transport cisplatin (Oda et al., 2014). Furthermore, emerging evidence suggests a biphasic regulatory relationship between cisplatin exposure and DBP Expression, in which cisplatin-induced nephrotoxicity may suppress DBP expression in renal tissue (B.-B. Cao et al., 2018). Collectively, these findings underscore the pivotal role of DBP in renal drug excretion, emphasizing its function in modulating transporter expression and, consequently, renal drug clearance. Biliary excretion is an essential pathway for eliminating drugs and their metabolites. A multispecific transporter protein, known as multidrug resistance-associated protein 2 (MRP2/ABCC2), significantly contributes to the excretion of endogenous substances and xenobiotics, particularly lipophilic compounds conjugated with glutathione, glucuronide, sulfate, and unmetabolized drugs. This process predominantly occurs within the tubular membranes of intestinal cells, proximal tubular epithelial cells in the kidneys, and hepatocytes (Jemnitz et al., 2010). The biliary excretion rate constant of spiramycin is substantially reduced by approximately 10-fold in MRP2 knockout mice, strongly indicating that MRP2 is a critical transporter mediating spiramycin excretion and serves as a central conduit for the elimination of various pharmaceutical agents (Tian et al., 2007). Furthermore, phenolphthalein, commonly employed to assess renal function and evaluate renal tubular secretory-excretory capabilities, exhibits distinct circadian rhythmic fluctuations. Specifically, phenolphthalein concentrations are lower during the daytime and higher at night, indicating enhanced hepatic excretion during the dark phase. Notably, biliary and biliary excretion rates at night are elevated 2.37-fold and 1.74-fold, respectively, compared to those during the daytime (Oh, Lee, Han, Cho, & Lee, 2017). Yu et al. (F. Yu et al., 2019)further demonstrated in that BMAL1/CLOCK activates Rev-erbα through E-box, indirectly inhibiting the expression of E4BP4. And DBP activates its transcription by directly binding to the D-box of the MRP2 promoter. Within this regulatory network, DBP acts as an MRP2 activator, whereas E4BP4 serves as a negative regulator. In addition to MRP2, the Na+/taurine bile salt cotransporter polypeptide (NTCP) is another pivotal transporter responsible for the hepatic uptake of bile acids from portal circulation. NTCP expression is also subject to circadian regulation, exhibiting lower levels during the day and higher levels at night (Z. Yu et al., 2020). Studies have indicated that DBP directly influences bile acid uptake and homeostasis by binding to specific promoter sequences of NTCP, thereby regulating its transcriptional activity (Ma et al., 2009). These findings underscore the critical role of DBP in coordinating bile acid metabolism and facilitating its excretion. The biological clock gene DBP exhibits pronounced circadian rhythmicity and significantly influences drug and metabolite excretion by modulating the expression of essential transport proteins, including OCT2, MRP2, and NTCP, in renal and hepatic tissues. Consequently, DBP plays a central role in synchronizing the excretory processes. However, despite evidence indicating that knockdown of PARbzip proteins (DBP, TEF, and HLF) markedly reduces the expression levels of intrarenal transporters, such as multidrug resistance-associated protein 4 (Mrp4) and Slc22a7, in mice (Pácha, Kateřina, & and Soták, 2021), further investigations are warranted to elucidate the precise role of DBP in regulating transporters within the solute carrier (SLC) superfamily and to clarify the detailed molecular mechanisms underlying its effect on renal drug excretion. 5 Discussion and prospect Mammalian biological clock oscillation is a self-sustaining and intricate physiological system regulated through a series of positive and negative transcriptional and translational feedback loops. Within this complex network, genes interact and coordinate to sustain the stability and accuracy of circadian rhythms, which are essential for orchestrating physiological processes in organisms. In this review, we comprehensively summarize the core components and underlying molecular mechanisms of the circadian system. Special emphasis is placed on the pivotal role of DBP as a primary output clock gene integrated within the central feedback loop that significantly influences physiological rhythms. Furthermore, we explored the pharmacokinetics of DBP, particularly its involvement in absorption, distribution, metabolism, and excretion (ADME). We analyzed the potential molecular mechanisms by which DBP may regulate pharmacodynamic and pharmacotoxicological processes and examined its influence on peripheral tissues, particularly regarding drug-metabolizing enzymes and transporters. Additionally, we evaluated the impact of the rhythmic expression patterns of these enzymes, transporters, and hormone secretions within peripheral tissues and their interactions with metabolic signaling pathways. By elucidating the interplay between circadian biology and pharmacokinetics between circadian biology and pharmacokinetics, we aim to provide a theoretical framework for enhancing drug efficacy, minimizing tolerance, and optimizing therapeutic strategies. Ultimately, this study aims to facilitate the advancement of chronotherapy by integrating circadian considerations into personalized treatment.

This work was supported by the National Natural Science Foundation of China, Grant Number 32201959; National Key R&D Program of China, Grant Number 2024YFD2402101; Research and Application of Processing and In-depth Product Development Technology of Gastrodia elata in the Origin Area, Grant Number 2023B03J1307; The Rural Science and Technology Commissioner Project for Assisting Towns and Villages in Guangdong Province, Grant Number KTP20240971; The Tibetan materials and medicine industrial cluster in the Tibet Autonomous Region, Grant Number Nongban Jicai [2025] No. 2;

Al-Waeli, H., Nicolau, B., Stone, L., Abu Nada, L., Gao, Q., Abdallah, M., Al Subaie, A. (2020). Chronotherapy of non-steroidal anti-inflammatory drugs may enhance postoperative recovery. Scientific reports, 10 (1), 468. Almazroo, O. A., Miah, M. K., & Venkataramanan, R. (2017). Drug metabolism in the liver. Clinics in liver disease, 21 (1), 1-20. Amador, A., Campbell, S., Kazantzis, M., Lan, G., Burris, T. P., & Solt, L. A. (2018). Distinct roles for REV-ERBα and REV-ERBβ in oxidative capacity and mitochondrial biogenesis in skeletal muscle. PLoS One, 13 (5), e0196787. Beesley, S., Noguchi, T., & Welsh, D. K. (2016). Cardiomyocyte circadian oscillations are cell-autonomous, amplified by β-adrenergic signaling, and synchronized in cardiac ventricle tissue. PloS one, 11 (7), e0159618. Bicker, J., Alves, G., Falcão, A., & Fortuna, A. (2020). Timing in drug absorption and disposition: The past, present, and future of chronopharmacokinetics. British Journal of Pharmacology, 177 (10), 2215-2239. Bozek, K., Relógio, A., Kielbasa, S. M., Heine, M., Dame, C., Kramer, A., & Herzel, H. (2009). Regulation of clock-controlled genes in mammals. PloS one, 4 (3), e4882. Cao, B.-B., Li, D., Xing, X., Zhao, Y., Wu, K., Jiang, F., Li, J.-D. (2018). Effect of cisplatin on the clock genes expression in the liver, heart and kidney. Biochemical and Biophysical Research Communications, 501 (2), 593-597. doi: https://doi.org/10.1016/j.bbrc.2018.05.056 Cao, Q. R., Kim, T. W., Choi, J. S., & Lee, B. J. (2005). Circadian variations in the pharmacokinetics, tissue distribution and urinary excretion of nifedipine after a single oral administration to rats. Biopharmaceutics & drug disposition, 26 (9), 427-437. Cao, X., Yang, Y., Selby, C. P., Liu, Z., & Sancar, A. (2021). Molecular mechanism of the repressive phase of the mammalian circadian clock. Proc Natl Acad Sci U S A, 118 (2). doi: 10.1073/pnas.2021174118 Cascorbi, I. (2013). P‐glycoprotein (MDR1/ABCB1). Pharmacogenomics of Human Drug Transporters: Clinical Impacts, 271-293. Chen, M., Guan, B., Xu, H., Yu, F., Zhang, T., & Wu, B. (2019). The Molecular Mechanism Regulating Diurnal Rhythm of Flavin-Containing Monooxygenase 5 in Mouse Liver. Drug Metab Dispos, 47 (11), 1333-1342. doi: 10.1124/dmd.119.088450 Chen, X., Yu, F., Guo, X., Su, C., Li, S.-S., & Wu, B. (2021). Clock gene Bmal1 controls diurnal rhythms in expression and activity of intestinal carboxylesterase 1. Journal of Pharmacy and Pharmacology, 73 (1), 52-59. Chiou, Y. Y., Yang, Y., Rashid, N., Ye, R., Selby, C. P., & Sancar, A. (2016). Mammalian Period represses and de-represses transcription by displacing CLOCK-BMAL1 from promoters in a Cryptochrome-dependent manner. Proc Natl Acad Sci U S A, 113 (41), E6072-e6079. doi: 10.1073/pnas.1612917113 Costello, H. M., Johnston, J. G., Juffre, A., Crislip, G. R., & Gumz, M. L. (2022). Circadian clocks of the kidney: function, mechanism, and regulation. Physiological reviews . Crislip, G. R., Johnston, J. G., Douma, L. G., Costello, H. M., Juffre, A., Boyd, K., Gumz, M. L. (2022). Circadian Rhythm Effects on the Molecular Regulation of Physiological Systems. Comprehensive Physiology, 12 (1), 2769-2798. doi: https://doi.org/10.1002/j.2040-4603.2022.tb00198.x Currie, G. M. (2018). Pharmacology, part 2: introduction to pharmacokinetics. Journal of nuclear medicine technology, 46 (3), 221-230. Dallmann, R., Okyar, A., & Lévi, F. (2016). Dosing-time makes the poison: circadian regulation and pharmacotherapy. Trends in molecular medicine, 22 (5), 430-445. Daniel, H. (2004). Molecular and integrative physiology of intestinal peptide transport. Annu. Rev. Physiol., 66, 361-384. Dong, D., Yang, D., Lin, L., Wang, S., & Wu, B. (2020). Circadian rhythm in pharmacokinetics and its relevance to chronotherapy. Biochemical Pharmacology, 178, 114045. Dyar, K. A., Ciciliot, S., Wright, L. E., Biensø, R. S., Tagliazucchi, G. M., Patel, V. R., Schiaffino, S. (2014). Muscle insulin sensitivity and glucose metabolism are controlled by the intrinsic muscle clock. Molecular Metabolism, 3 (1), 29-41. doi: https://doi.org/10.1016/j.molmet.2013.10.005 Fan, X.-l., Song, Y., Qin, D.-X., & Lin, P.-Y. (2023). Regulatory Effects of Clock and Bmal1 on Circadian Rhythmic TLR Expression. International Reviews of Immunology, 42 (2), 101-112. doi: 10.1080/08830185.2021.1931170 Fang, B., Everett, L. J., Jager, J., Briggs, E., Armour, S. M., Feng, D., Lazar, M. A. (2014). Circadian enhancers coordinate multiple phases of rhythmic gene transcription in vivo. Cell, 159 (5), 1140-1152. Firsov, D., & Bonny, O. (2018). Circadian rhythms and the kidney. Nature Reviews Nephrology, 14 (10), 626-635. Fukami, T., Yokoi, T., & Nakajima, M. (2022). Non-P450 Drug-Metabolizing Enzymes: Contribution to Drug Disposition, Toxicity, and Development. Annual Review of Pharmacology and Toxicology, 62 (Volume 62, 2022), 405-425. doi: https://doi.org/10.1146/annurev-pharmtox-052220-105907 Gachon, F., Olela, F. F., Schaad, O., Descombes, P., & Schibler, U. (2006). The circadian PAR-domain basic leucine zipper transcription factors DBP, TEF, and HLF modulate basal and inducible xenobiotic detoxification. Cell Metab, 4 (1), 25-36. doi: 10.1016/j.cmet.2006.04.015 Ge, W., Sun, Q., Yang, Y., Ding, Z., Liu, J., & Zhang, J. (2023). Circadian PER1 controls daily fat absorption with the regulation of PER1-PKA on phosphorylation of bile acid synthetase. Journal of Lipid Research, 64 (6). Guo, B., Yang, N., Borysiewicz, E., Dudek, M., Williams, J., Li, J., Bateman, J. (2015). Catabolic cytokines disrupt the circadian clock and the expression of clock-controlled genes in cartilage via an NFкB-dependent pathway. Osteoarthritis and cartilage, 23 (11), 1981-1988. Hayashi, Y., Ushijima, K., Ando, H., Yanagihara, H., Ishikawa, E., Tsuruoka, S.-i., Fujimura, A. (2010). Influence of a time-restricted feeding schedule on the daily rhythm of abcb1a gene expression and its function in rat intestine. The Journal of pharmacology and experimental therapeutics, 335 (2), 418-423. Huang, X. M., Chen, W. L., Yuan, J. P., Yang, Y. H., Mei, Q. H., & Huang, L. X. (2013). Altered diurnal variation and localization of clock proteins in the remnant kidney of 5/6 nephrectomy rats. Nephrology (Carlton), 18 (8), 555-562. doi: 10.1111/nep.12111 Ikeda, R., Tsuchiya, Y., Koike, N., Umemura, Y., Inokawa, H., Ono, R., Okubo, N. (2019). REV-ERBα and REV-ERBβ function as key factors regulating Mammalian Circadian Output. Scientific reports, 9 (1), 10171. Janoski, J. R., Aiello, I., Lundberg, C. W., & Finkielstein, C. V. (2024). Circadian clock gene polymorphisms implicated in human pathologies. Trends Genet . doi: 10.1016/j.tig.2024.05.006 Jemnitz, K., Heredi-Szabo, K., Janossy, J., Ioja, E., Vereczkey, L., & Krajcsi, P. (2010). ABCC2/Abcc2: a multispecific transporter with dominant excretory functions. Drug Metabolism Reviews, 42 (3), 402-436. doi: 10.3109/03602530903491741 Johnston, J. G., & Pollock, D. M. (2018). Circadian regulation of renal function. Free Radical Biology and Medicine, 119, 93-107. doi: https://doi.org/10.1016/j.freeradbiomed.2018.01.018 Joyner, M. J., & Casey, D. P. (2015). Regulation of increased blood flow (hyperemia) to muscles during exercise: a hierarchy of competing physiological needs. Physiological reviews . Lavery, D. J., Lopez-Molina, L., Margueron, R., Fleury-Olela, F., Conquet, F., Schibler, U., & Bonfils, C. (1999). Circadian expression of the steroid 15 alpha-hydroxylase (Cyp2a4) and coumarin 7-hydroxylase (Cyp2a5) genes in mouse liver is regulated by the PAR leucine zipper transcription factor DBP. Mol Cell Biol, 19 (10), 6488-6499. doi: 10.1128/mcb.19.10.6488 Lee, Y.-H., Alberta, J. A., Gonzalez, F. J., & Waxman, D. J. (1994). Multiple, functional DBP sites on the promoter of the cholesterol 7 alpha-hydroxylase P450 gene, CYP7. Proposed role in diurnal regulation of liver gene expression. Journal of Biological Chemistry, 269 (20), 14681-14689. Leliavski, A. (2014). Physiological functions of the adrenocortical circadian clock. Niedersächsische Staats-und Universitätsbibliothek Göttingen. Lin, C., Feng, S., DeOliveira, C. C., & Crane, B. R. (2023). Cryptochrome-Timeless structure reveals circadian clock timing mechanisms. Nature, 617 (7959), 194-199. doi: 10.1038/s41586-023-06009-4 Lin, Y., Wang, S., Zhou, Z., Guo, L., Yu, F., & Wu, B. (2019). Bmal1 regulates circadian expression of cytochrome P450 3a11 and drug metabolism in mice. Communications Biology, 2 (1), 378. doi: 10.1038/s42003-019-0607-z Lin, Y., Wang, S., Zhou, Z., Guo, L., Yu, F., & Wu, B. (2019). Bmal1 regulates circadian expression of cytochrome P450 3a11 and drug metabolism in mice. Commun Biol, 2, 378. doi: 10.1038/s42003-019-0607-z Lu, D., Zhao, M., Chen, M., & Wu, B. (2020). Circadian clock–controlled drug metabolism: implications for Chronotherapeutics. Drug Metabolism and Disposition, 48 (5), 395-406. Ma, K., Xiao, R., Tseng, H. T., Shan, L., Fu, L., & Moore, D. D. (2009). Circadian dysregulation disrupts bile acid homeostasis. PLoS One, 4 (8), e6843. doi: 10.1371/journal.pone.0006843 Manoogian, E. N. C., & Panda, S. (2017). Circadian rhythms, time-restricted feeding, and healthy aging. Ageing Res Rev, 39, 59-67. doi: 10.1016/j.arr.2016.12.006 Mekbib, T. F. (2020). The Role of REVERBα E3 Ligases in Circadian Clock Function, and Metabolism. Morehouse School of Medicine. Merlino, A. (2023). Metallodrug binding to serum albumin: Lessons from biophysical and structural studies. Coordination Chemistry Reviews, 480, 215026. doi: https://doi.org/10.1016/j.ccr.2023.215026 Messori, L., & Merlino, A. (2016). Cisplatin binding to proteins: A structural perspective. Coordination Chemistry Reviews, 315, 67-89. Mitsui, S., Yamaguchi, S., Matsuo, T., Ishida, Y., & Okamura, H. (2001). Antagonistic role of E4BP4 and PAR proteins in the circadian oscillatory mechanism. Genes Dev, 15 (8), 995-1006. doi: 10.1101/gad.873501 Morishita, Y., Miura, D., & Kida, S. (2016). PI3K regulates BMAL1/CLOCK-mediated circadian transcription from the Dbp promoter. Bioscience, Biotechnology, and Biochemistry, 80 (6), 1131-1140. doi: 10.1080/09168451.2015.1136885 Murakami, Y., Higashi, Y., Matsunaga, N., Koyanagi, S., & Ohdo, S. (2008). Circadian clock-controlled intestinal expression of the multidrug-resistance gene mdr1a in mice. Gastroenterology, 135 (5), 1636-1644. e1633. Nakamura, T., Yonezawa, A., Hashimoto, S., Katsura, T., & Inui, K.-i. (2010). Disruption of multidrug and toxin extrusion MATE1 potentiates cisplatin-induced nephrotoxicity. Biochemical pharmacology, 80 (11), 1762-1767. Neves, A. R., Albuquerque, T., Quintela, T., & Costa, D. (2022). Circadian rhythm and disease: Relationship, new insights, and future perspectives. J Cell Physiol, 237 (8), 3239-3256. doi: 10.1002/jcp.30815 Oda, M., Koyanagi, S., Tsurudome, Y., Kanemitsu, T., Matsunaga, N., & Ohdo, S. (2014). Renal Circadian Clock Regulates the Dosing-Time Dependency of Cisplatin-Induced Nephrotoxicity in Mice. Molecular Pharmacology, 85 (5), 715-722. doi: https://doi.org/10.1124/mol.113.089805 Oh, J. H., Lee, J. H., Han, D. H., Cho, S., & Lee, Y. J. (2017). Circadian Clock Is Involved in Regulation of Hepatobiliary Transport Mediated by Multidrug Resistance-Associated Protein 2. J Pharm Sci, 106 (9), 2491-2498. doi: 10.1016/j.xphs.2017.04.071 Ohdo, S. (2010). Chronotherapeutic strategy: Rhythm monitoring, manipulation and disruption. Advanced Drug Delivery Reviews, 62 (9), 859-875. doi: https://doi.org/10.1016/j.addr.2010.01.006 Oishi, K., Koyanagi, S., & Ohkura, N. (2011). Circadian mRNA expression of coagulation and fibrinolytic factors is organ-dependently disrupted in aged mice. Experimental gerontology, 46 (12), 994-999. Pácha, J., Balounová, K., & Soták, M. (2021). Circadian regulation of transporter expression and implications for drug disposition. Expert Opinion on Drug Metabolism & Toxicology, 17 (4), 425-439. Pácha, J., Kateřina, B., & and Soták, M. (2021). Circadian regulation of transporter expression and implications for drug disposition. Expert Opinion on Drug Metabolism & Toxicology, 17 (4), 425-439. doi: 10.1080/17425255.2021.1868438 Pan, X., Terada, T., Okuda, M., & Inui, K.-I. (2003). Altered diurnal rhythm of intestinal peptide transporter by fasting and its effects on the pharmacokinetics of ceftibuten. Journal of Pharmacology and Experimental Therapeutics, 307 (2), 626-632. Patke, A., Young, M. W., & Axelrod, S. (2020). Molecular mechanisms and physiological importance of circadian rhythms. Nat Rev Mol Cell Biol, 21 (2), 67-84. doi: 10.1038/s41580-019-0179-2 Pulido, R. S., Munji, R. N., Chan, T. C., Quirk, C. R., Weiner, G. A., Weger, B. D., Deng, A. (2020). Neuronal activity regulates blood-brain barrier efflux transport through endothelial circadian genes. Neuron, 108 (5), 937-952. e937. Rowland, M., & Tozer, T. N. (2010). Clinical Pharmacokinetics and Pharmacodynamics: Philadelphia, PA: Lippincott Williams. Saito, H., Terada, T., Shimakura, J., Katsura, T., & Inui, K.-i. (2008). Regulatory mechanism governing the diurnal rhythm of intestinal H+/peptide cotransporter 1 (PEPT1). American Journal of Physiology-Gastrointestinal and Liver Physiology, 295 (2), G395-G402. Saltin, B., & Mortensen, S. P. (2012). Inefficient functional sympatholysis is an overlooked cause of malperfusion in contracting skeletal muscle. J Physiol, 590 (24), 6269-6275. doi: 10.1113/jphysiol.2012.241026 Segers, A., & Depoortere, I. (2021). Circadian clocks in the digestive system. Nat Rev Gastroenterol Hepatol, 18 (4), 239-251. doi: 10.1038/s41575-020-00401-5 Smolensky, M. H., Siegel, R. A., Haus, E., Hermida, R., & Portaluppi, F. (2011). Biological rhythms, drug delivery, and chronotherapeutics. Fundamentals and applications of controlled release drug delivery, 359-443. Sopasakis, V. R., Liu, P., Suzuki, R., Kondo, T., Winnay, J., Tran, T. T., Farese, R. V. (2010). Specific roles of the p110α isoform of phosphatidylinsositol 3-kinase in hepatic insulin signaling and metabolic regulation. Cell Metabolism, 11 (3), 220-230. Stelzl, T. (2018). Glycosylation of the intestinal peptide transporter: structure and function. Technische Universität München. Su, W., Xie, Z., Guo, Z., Duncan, M. J., Lutshumba, J., & Gong, M. C. (2012). Altered clock gene expression and vascular smooth muscle diurnal contractile variations in type 2 diabetic db/db mice. Am J Physiol Heart Circ Physiol, 302 (3), H621-633. doi: 10.1152/ajpheart.00825.2011 Sukumaran, S., Almon, R. R., DuBois, D. C., & Jusko, W. J. (2010). Circadian rhythms in gene expression: Relationship to physiology, disease, drug disposition and drug action. Advanced drug delivery reviews, 62 (9-10), 904-917. Takahashi, J. S. (2017). Transcriptional architecture of the mammalian circadian clock. Nature Reviews Genetics, 18 (3), 164-179. Takahashi, J. S. (2018). Molecular architecture of the mammalian circadian clock. Investigative Ophthalmology & Visual Science, 59 (9), 6-6. Takiguchi, T., Tomita, M., Matsunaga, N., Nakagawa, H., Koyanagi, S., & Ohdo, S. (2007). Molecular basis for rhythmic expression of CYP3A4 in serum-shocked HepG2 cells. Pharmacogenet Genomics, 17 (12), 1047-1056. doi: 10.1097/FPC.0b013e3282f12a61 Tao, W., Wu, J., Zhang, Q., Lai, S.-S., Jiang, S., Jiang, C., Li, C.-J. (2015). EGR1 regulates hepatic clock gene amplitude by activating Per1 transcription. Scientific reports, 5 (1), 15212. Tian, X., Li, J., Zamek-Gliszczynski Maciej, J., Bridges Arlene, S., Zhang, P., Patel Nita, J., Brouwer Kim, L. R. (2007). Roles of P-Glycoprotein, Bcrp, and Mrp2 in Biliary Excretion of Spiramycin in Mice. Antimicrobial Agents and Chemotherapy, 51 (9), 3230-3234. doi: 10.1128/aac.00082-07 Tong, Y., Zeng, P., Zhang, T., Zhao, M., Yu, P., & Wu, B. (2019). The transcription factor E4bp4 regulates the expression and activity of Cyp3a11 in mice. Biochemical Pharmacology, 163, 215-224. Uney, K., Tras, B., Corum, O., Yildiz, R., & Maden, M. (2019). Pharmacokinetics of pentoxifylline and its 5-hydroxyhexyl metabolite following intravenous administration in cattle. Trop Anim Health Prod, 51 (2), 435-441. doi: 10.1007/s11250-018-1710-8 Vanni, L. M., NP, A., Di Maggio, T. J., & Cooney, M. F. (2024). Pharmacological Pain Management Strategies. Core Curriculum for Pain Management Nursing-E-Book, 241. Voigt, R. M., Forsyth, C. B., & Keshavarzian, A. (2019). Circadian rhythms: a regulator of gastrointestinal health and dysfunction. Expert review of gastroenterology & hepatology, 13 (5), 411-424. Walker, W. H., Walton, J. C., DeVries, A. C., & Nelson, R. J. (2020). Circadian rhythm disruption and mental health. Translational Psychiatry, 10 (1), 28. doi: 10.1038/s41398-020-0694-0 Wu, T., Fu, O., Yao, L., Sun, L., ZhuGe, F., & Fu, Z. (2012). Differential responses of peripheral circadian clocks to a short-term feeding stimulus. Molecular Biology Reports, 39 (10), 9783-9789. doi: 10.1007/s11033-012-1844-0 Yang, X., Wood, P. A., Ansell, C. M., Ohmori, M., Oh, E.-Y., Xiong, Y., Hrushesky, W. J. (2009). β-catenin induces β-TrCP-mediated PER2 degradation altering circadian clock gene expression in intestinal mucosa of ApcMin/+ mice. Journal of biochemistry, 145 (3), 289-297. Ye, R., Selby, C. P., Chiou, Y. Y., Ozkan-Dagliyan, I., Gaddameedhi, S., & Sancar, A. (2014). Dual modes of CLOCK:BMAL1 inhibition mediated by Cryptochrome and Period proteins in the mammalian circadian clock. Genes Dev, 28 (18), 1989-1998. doi: 10.1101/gad.249417.114 Yonezawa, A., Masuda, S., Yokoo, S., Katsura, T., & Inui, K.-i. (2006). Cisplatin and oxaliplatin, but not carboplatin and nedaplatin, are substrates for human organic cation transporters (SLC22A1–3 and multidrug and toxin extrusion family). Journal of Pharmacology and Experimental Therapeutics, 319 (2), 879-886. Yoshitane, H., Asano, Y., Sagami, A., Sakai, S., Suzuki, Y., Okamura, H., Fukada, Y. (2019). Functional D-box sequences reset the circadian clock and drive mRNA rhythms. Communications Biology, 2 (1), 300. doi: 10.1038/s42003-019-0522-3 Yu, F., Zhang, T., Zhou, C., Xu, H., Guo, L., Chen, M., & Wu, B. (2019). The Circadian Clock Gene Bmal1 Controls Intestinal Exporter MRP2 and Drug Disposition. Theranostics, 9 (10), 2754-2767. doi: 10.7150/thno.33395 Yu, Z., Yang, J., Xiang, D., Li, G., Liu, D., & Zhang, C. (2020). Circadian rhythms and bile acid homeostasis: a comprehensive review. Chronobiology international, 37 (5), 618-628. Yuan, J., Guo, J., Zhang, M., Wang, Q., Huang, H., & Chen, Y. (2015). Upregulation of D site of albumin promoter binding protein in the brain of patients with intractable epilepsy. Molecular Medicine Reports, 11 (4), 2486-2492. Zhang, S. L., Yue, Z., Arnold, D. M., Artiushin, G., & Sehgal, A. (2018). A circadian clock in the blood-brain barrier regulates xenobiotic efflux. Cell, 173 (1), 130-139. e110. Zhang, Y. K., Yeager, R. L., & Klaassen, C. D. (2009). Circadian expression profiles of drug-processing genes and transcription factors in mouse liver. Drug Metab Dispos, 37 (1), 106-115. doi: 10.1124/dmd.108.024174. Figure legend Figure 1: The molecular biological clock mechanism of mammals is formed by the transcription-translation feedback loop (Patke et al., 2020). Figure 2: Identification of DBP and E4BP4 sequences in D-box MOCCS2 was used to analyze the common sites of DBP and E4BP4. b. Frequency distribution of TTATGCAA (D-Box #1) and TTACGTAA (D-box#18) around the common site of DBP/E4BP4. c. The cumulative relative frequency curves of D-BOX #1 and #18 around the common site. d. Dual-luciferase determination was performed on all D-BOX #1-#18 sequences to analyze the transcriptional activities of DBP and E4BP4 (Yoshitane et al., 2019). Figure 3: Pharmacokinetic schematic diagram, including absorption, distribution, metabolism and excretion. a. The interactions between drugs in the internal organs of the body. b. The process of the administration site and the site of action in each link of pharmacokinetics (Currie, 2018). Information & Authors Information Version history Copyright This work is licensed under a Non Exclusive No Reuse License. Authors Metrics & Citations Metrics Article Usage 290views 97downloads Citations Download citation Hao Jiang, Xiaoqing Zheng, Ying Chen, et al. Influence of the Circadian Clock Gene DBP on Pharmacokinetic Parameters: A Comprehensive Review. Authorea. 09 June 2025. DOI: https://doi.org/10.22541/au.174947599.90931999/v1 DOI: https://doi.org/10.22541/au.174947599.90931999/v1 If you have the appropriate software installed, you can download article citation data to the citation manager of your choice. Simply select your manager software from the list below and click Download. For more information or tips please see 'Downloading to a citation manager' in the Help menu.