Application and challenge of pancreatic organoids in therapeutic research.

The in-vivo non-human primate animal and in-vitro cell disease models play a crucial part in the study of the mechanisms underlying the occurrence and development of pancreatic diseases, but with increasingly prominent limitations with in-depth research. Organoids derived from human pluripotent and adult stem cells resemble human in-vivo organs in their cellular composition, spatial tissue structure and physiological function, making them as an advantageous research tool. Up until now, numerous human organoids, including pancreas, have been effectively developed, demonstrating significant potential for research in organ development, disease modeling, drug screening, and regenerative medicine. However, different from intestine, liver and other organs, the pancreas is the only special organ in the human body, consisting of an exocrine gland and an endocrine gland. Thus, the development of pancreatic organoid technology faces greater challenges, and how to construct a composite pancreatic organoid with exocrine and endocrine gland is still difficult in current research. By reviewing the fundamental architecture and physiological role of the human pancreas, along with the swiftly developing domain of pancreatic organoids, we summarize the method and characteristics of human pancreatic organoids, and its application in modeling pancreatic diseases, as a platform for individualized drug screening and in regenerative medicine study. As the first comprehensive review that focus on the pharmacological study of human pancreatic organoid, the review hopes to help scholars to have a deeper understanding in the study of pancreatic organoid.

Anti-stroke biologics: from recombinant proteins to stem cells and organoids.

The use of biologics in various diseases has dramatically increased in recent years. Stroke, a cerebrovascular disease, is the second most common cause of death, and the leading cause of disability with high morbidity worldwide. For biologics applied in the treatment of acute ischaemic stroke, alteplase is the only thrombolytic agent. Meanwhile, current clinical trials show that two recombinant proteins, tenecteplase and non-immunogenic staphylokinase, are most promising as new thrombolytic agents for acute ischaemic stroke therapy. In addition, stem cell-based therapy, which uses stem cells or organoids for stroke treatment, has shown promising results in preclinical and early clinical studies. These strategies for acute ischaemic stroke mainly rely on the unique properties of undifferentiated cells to facilitate tissue repair and regeneration. However, there is a still considerable journey ahead before these approaches become routine clinical use. This includes optimising cell delivery methods, determining the ideal cell type and dosage, and addressing long-term safety concerns. This review introduces the current or promising recombinant proteins for thrombolysis therapy in ischaemic stroke and highlights the promise and challenges of stem cells and cerebral organoids in stroke therapy.

The clinical effect evaluation of multidisciplinary collaborative team combined with palliative care model in patients with terminal cancer: a randomised controlled study.

OBJECTIVE: To evaluate the clinical effect of a multidisciplinary collaboration team combined with a palliative care model in patients with terminal cancer. METHOD: A total of 84 patients diagnosed with terminal cancer in our hospital were included and randomly divided into an intervention group and a control group, with 42 cases in each group. Patients in the intervention group were treated by a multidisciplinary collaborative team combined with the palliative care model, and patients in the control group were treated by routine nursing intervention. The Self-rating Anxiety Scale (SAS) and the Self-rating Depression Scale (SDS) were used to evaluate negative emotions and anxiety and depression of patients before and after intervention. The Quality of Life Scale (European Organization for Research and Treatment of Cancer [EORTC] QLQ-C30) and Social Support Scale (SSRS) were used to evaluate the quality of life and social support of patients. This study has been registered in 13/01/2023 (ClinicalTrials.gov Identifier: NCT05683236). RESULT: The general data of the two groups were comparable. After intervention, the SAS (43.7 ± 7.4 vs. 54.2 ± 9.3) and SDS scores (38.4 ± 6.5 vs. 53.1 ± 8.4) of the intervention group were significantly lower than those of the control group. The total SSRS score, subjective support score, objective support score and utilisation of support of the intervention group were significantly higher than those of the control group (P < 0.05). The overall quality of life score of the intervention group was higher than that of the control group, and the difference was statistically significant (79.5 ± 4.5 vs. 73.2 ± 3.6, P < 0.05). The scores of each functional scale were significantly higher than those of the control group (P < 0.05). CONCLUSION: Compared with conventional nursing, the application of the multidisciplinary collaborative team combined with tranquilisation therapy in patients with terminal cancer can significantly reduce the anxiety and depression of patients, enable patients to obtain comprehensive social support, and effectively improve the quality of life of patients. TRIAL REGISTRATION: ClinicalTrials.gov Identifier NCT05683236, 13/01/2023, Retrospectively registered.

Metrnl deficiency retards skin wound healing in mice by inhibiting AKT/eNOS signaling and angiogenesis.

Meteorin-like (Metrnl) is a novel secreted protein with various biological activities. In this study, we investigated whether and how Metrnl regulated skin wound healing in mice. Global Metrnl gene knockout mice (Metrnl-/-) and endothelial cell-specific Metrnl gene knockout mice (EC-Metrnl-/-) were generated. Eight-mm-diameter full-thickness excisional wound was made on the dorsum of each mouse. The skin wounds were photographed and analyzed. In C57BL/6 mice, we observed that Metrnl expression levels were markedly increased in skin wound tissues. We found that both global and endothelial cell-specific Metrnl gene knockout significantly retarded mouse skin wound healing, and endothelial Metrnl was the key factor affecting wound healing and angiogenesis. The proliferation, migration and tube formation ability of primary human umbilical vein endothelial cells (HUVECs) were inhibited by Metrnl knockdown, but significantly promoted by addition of recombinant Metrnl (10 ng/mL). Metrnl knockdown abolished the proliferation of endothelial cells stimulated by recombinant VEGFA (10 ng/mL) but not by recombinant bFGF (10 ng/mL). We further revealed that Metrnl deficiency impaired VEGFA downstream AKT/eNOS activation in vitro and in vivo. The damaged angiogenetic activity in Metrnl knockdown HUVECs was partly rescued by addition of AKT activator SC79 (10 µM). In conclusion, Metrnl deficiency retards skin wound healing in mice, which is related to impaired endothelial Metrnl-mediated angiogenesis. Metrnl deficiency impairs angiogenesis by inhibiting AKT/eNOS signaling pathway.

The generation and properties of human cortical organoids as a disease model for malformations of cortical development.

As three-dimensional "organ-like" aggregates, human cortical organoids have emerged as powerful models for studying human brain evolution and brain disorders with unique advantages of human-specificity, fidelity and manipulation. Human cortical organoids derived from human pluripotent stem cells can elaborately replicate many of the key properties of human cortical development at the molecular, cellular, structural, and functional levels, including the anatomy, functional neural network, and interaction among different brain regions, thus facilitating the discovery of brain development and evolution. In addition to studying the neuro-electrophysiological features of brain cortex development, human cortical organoids have been widely used to mimic the pathophysiological features of cortical-related disease, especially in mimicking malformations of cortical development, thus revealing pathological mechanism and identifying effective drugs. In this review, we provide an overview of the generation of human cortical organoids and the properties of recapitulated cortical development and further outline their applications in modeling malformations of cortical development including pathological phenotype, underlying mechanisms and rescue strategies.

Hepatic Nampt Deficiency Aggravates Dyslipidemia and Fatty Liver in High Fat Diet Fed Mice.

Nicotinamide phosphoribosyltransferase (Nampt) is the rate-limiting enzyme in the salvage pathway of nicotinamide adenine dinucleotide (NAD) biosynthesis. Thus far, hepatic Nampt has not been extensively explored in terms of its effects on serum lipid stability and liver lipids metabolism. In this study, hepatocyte-specific Nampt knockout (HC-Nampt-/-) mice were generated by Cre/loxP system. Nampt mRNA expression was reduced in the liver, but not in other tissues, in HC-Nampt-/- mice compared with wild-type (WT) mice. Hepatic Nampt deficiency had no effect on body weight and fasting blood glucose, and it did not induce atherosclerosis in mice under both normal chow diet (NCD) and high fat diet (HFD). At baseline state under NCD, hepatic Nampt deficiency also did not affect liver weight, liver function index, including alanine aminotransferase, aspartate aminotransferase, albumin and alkaline phosphatase, and serum levels of lipids, including triglycerides (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and non-esterified fatty acids (NEFA). However, under HFD, deficiency of hepatic Nampt resulted in increased liver weight, liver function index, and serum levels of TG, TC, HDL-C, and NEFA. Meanwhile, histopathological examination showed increased fat accumulation and fibrosis in the liver of HC-Nampt-/- mice compared with WT mice. Taken together, our results show that hepatic Nampt deficiency aggravates dyslipidemia and liver damage in HFD fed mice. Hepatocyte Nampt can be a protective target against dyslipidemia and fatty liver.

Metrnl regulates cognitive dysfunction and hippocampal BDNF levels in D-galactose-induced aging mice.

Aging is one of the main risk factors for cognitive dysfunction. During aging process, the decrease of brain-derived neurotrophic factor (BDNF) and the impairment of astrocyte function contribute to the cognitive impairment. Metrnl, a neurotrophic factor, promotes neural growth, migration and survival, and supports neural function. In this study, we investigated the role of Metrnl in cognitive functions. D-galactose (D-gal)-induced aging model was used to simulate the process of aging. Cognitive impairment was assessed by the Morris water maze test. We showed that Metrnl expression levels were significantly increased in the hippocampus of D-gal-induced aging mice. Metrnl knockout did not affect the cognitive functions in the baseline state, but aggravated the cognitive impairment in the D-gal-induced aging mice. Furthermore, Metrnl knockout significantly reduced hippocampal BDNF, TrkB, and glial fibrillary acidic protein (GFAP) levels in the D-gal-induced aging mice. In the D-gal-induced aging cell model in vitro, Metrnl levels in the hippocampal astrocytes were significantly increased, and Metrnl knockdown and overexpression regulated the BDNF levels in primary hippocampal astrocytes rather than in neurons. We conclude that Metrnl regulates cognitive functions and hippocampal BDNF levels during aging process. As a neurotrophic factor and an endogenous protein, Metrnl is expected to become a new candidate for the treatment or alleviation of aging-related cognitive dysfunction.

Humanized cerebral organoids-based ischemic stroke model for discovering of potential anti-stroke agents.

Establishing a stoke experimental model, which is better in line with the physiology and function of human brain, is the bottleneck for the development of effective anti-stroke drugs. A three-dimensional cerebral organoids (COs) from human pluripotent stem cells can mimic cell composition, cortical structure, brain neural connectivity and epigenetic genomics of in-vivo human brain, which provides a promising application in establishing humanized ischemic stroke model. COs have been used for modeling low oxygen condition-induced hypoxic injury, but there is no report on the changes of COs in response to in vitro oxygen-glucose deprivation (OGD)-induced damage of ischemic stroke as well as its application in testing anti-stroke drugs. In this study we compared the cell composition of COs at different culture time and explored the cell types, cell ratios and volume size of COs at 85 days (85 d-CO). The 85 d-CO with diameter more than 2 mm was chosen for establishing humanized ischemic stroke model of OGD. By determining the time-injury relationship of the model, we observed aggravated ischemic injury of COs with OGD exposure time, obtaining first-hand evidence for the damage degree of COs under different OGD condition. The sensitivity of the model to ischemic injury and related treatment was validated by the proven pan-Caspase inhibitor Z-VAD-FMK (20 µM) and Bcl-2 inhibitor navitoclax (0.5 µM). Neuroprotective agents edaravone, butylphthalide, P7C3-A20 and ZL006 (10 µM for each) exerted similar beneficial effects in this model. Taken together, this study establishes a humanized ischemic stroke model based on COs, and provides evidence as a new research platform for anti-stroke drug development.

Pharmacological characterization of MT-1207, a novel multitarget antihypertensive agent.

Hypertension is a serious public health problem worldwide. MT-1207, chemically named 3-(4-(4-(1H-benzotriazole-1-yl)butyl)piperazine-1-yl) benzisothiazole hydrochloride, is a new chemical entity that has entered into clinical trial as antihypertensive agent in China. In this paper we report the pharmacological profile of MT-1207 regarding its acute, subacute, and long-term effects on hypertensive animal models, and its actions on isolated organs in vitro as well as its molecular targets. Blood pressure (BP) was measured in conscious animals; amlodipine was taken as a positive control drug. We showed that both single dose of MT-1207 (1.25-20 mg/kg, ig) in spontaneously hypertensive rats (SHR) and MT-1207 (0.25-6 mg/kg, ig) in two-kidney one-clip (2K1C) dogs dose-dependently decreased BP. MT-1207 quickly decreased BP within 5 min after administration; the hypotensive effect lasted for 8 and 12 h, respectively, in SHR and 2K1C dogs without reflex increase in heart rate. Multiple doses of MT-1207 (5 mg · kg-1 · d-1 in SHR; 2 mg · kg-1 · d-1 in 2K1C dogs, for 7 days) significantly decreased BP, slightly reduced heart rate, and both of them recovered after withdrawal. Long-term administration of MT-1207 (10 mg · kg-1 · d-1 for 4 months or more time) produced a stable BP reduction, improved baroreflex sensitivity, reduced renal and cardiovascular damage in SHR, and delayed stroke occurrence and death in stroke-prone SHR. In isolated rat aortic rings precontracted by adrenaline, KCl, noradrenaline or 5-hydroxytryptamine (5-HT), MT-1207 (10-9-10-4 M) caused concentration-dependent relaxation. In a panel of enzyme activity or radioligand binding assays of 87 molecular targets, MT-1207 potently inhibited adrenergic α1A, α1B, α1D, and 5-HT2A receptors with Ki < 1 nM. The antagonism of MT-1207 against these receptors was confirmed in isolated rabbit arteries. We conclude that MT-1207 is a novel and promising single-molecule multitarget agent for hypertension treatment to reduce hypertensive organ damage and stroke mortality.

P7C3-A20 alleviates fatty liver by shaping gut microbiota and inducing FGF21/FGF1, via the AMP-activated protein kinase/CREB regulated transcription coactivator 2 pathway.

BACKGROUND AND PURPOSE: Non-alcoholic fatty liver disease (NAFLD) is a worldwide public health problem with no established pharmacological therapy. Here, we explored the potential benefit of P7C3-A20, a novel aminopropyl carbazole compound with neuroprotective activity, in a NAFLD model, induced in mice by a high-fat diet (HFD). EXPERIMENTAL APPROACH: C57BL/6J mice were given a HFD (42% fat content) for 16 weeks to induce NAFLD. P7C3-A20 (20 mg·kg-1 ·day-1 ) was given by gavage for 2 weeks. Indirect calorimetry, histological analysis, immunoblotting, immunohistochemistry, and biomedical examinations were performed. Gut microbiota were determined using a 16S ribosomal RNA sequencing analysis. KEY RESULTS: P7C3-A20 treatment reduced body weight gain/adiposity, improved insulin resistance, promoted energy expenditure (O2 consumption/CO2 production), inhibited lipid oxidation, suppressed hepatic inflammation (Kupffer cell number and pro-inflammatory factors), decreased necroptosis/apoptosis (receptor-interacting protein kinase 3, cleaved caspase-3, and TUNEL), and alleviated liver fibrosis and injury. Mechanistically, P7C3-A20 stimulated FGF21 and FGF1 via activating liver kinase B1 (LKB1) and AMP-activated protein kinase (AMPK), which further resulted in a reduced nuclear translocation of CREB-regulated transcription coactivator 2 (CRTC2). In AMPKα2 knockout mice, the protection of P7C3-A20 against HFD-induced metabolism abnormalities and fat accumulation, as well as the elevation of blood FGF21 and FGF1, was abolished. P7C3-A20 increased the gut microbiota species richness. Moreover, it enhanced the proportions of Akkermansia, Lactobacillus, and Prevotellaceae, while reducing the proportions of Enterobacteriaceae, Escherichia, and Parasutterella. CONCLUSIONS AND IMPLICATIONS: P7C3-A20 increased levels of NAD+ and alleviated NAFLD through stimulating FGF21 and FGF1 in an LKB1/AMPK/CRTC2-dependent manner and shaping gut microbiota. LINKED ARTICLES: This article is part of a themed issue on Cellular metabolism and diseases. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.10/issuetoc.

Cerebral organoids transplantation improves neurological motor function in rat brain injury.

BACKGROUND AND PURPOSE: Cerebral organoids (COs) have been used for studying brain development, neural disorders, and species-specific drug pharmacology and toxicology, but the potential of COs transplantation therapy for brain injury remains to be answered. METHODS: With preparation of traumatic brain injury (TBI) model of motor dysfunction, COs at 55 and 85 days (55 and 85 d-CO) were transplanted into damaged motor cortex separately to identify better transplantation donor for brain injury. Further, the feasibility, effectiveness, and underlying mechanism of COs transplantation therapy for brain injury were explored. RESULTS: 55 d-CO was demonstrated as better transplantation donor than 85 d-CO, evidenced by more neurogenesis and higher cell survival rate without aggravating apoptosis and inflammation after transplantation into damaged motor cortex. Cells from transplanted COs had the potential of multilinage differentiation to mimic in-vivo brain cortical development, support region-specific reconstruction of damaged motor cortex, form neurotransmitter-related neurons, and migrate into different brain regions along corpus callosum. Moreover, COs transplantation upregulated hippocampal neural connection proteins and neurotrophic factors. Notably, COs transplantation improved neurological motor function and reduced brain damage. CONCLUSIONS: This study revealed 55 d-CO as better transplantation donor and demonstrated the feasibility and efficacy of COs transplantation in TBI, hoping to provide first-hand preclinical evidence of COs transplantation for brain injury.

Cerebral Organoids Repair Ischemic Stroke Brain Injury.

Stroke is the second leading cause of death and main cause of disability worldwide, but with few effective therapies. Although stem cell-based therapy has been proposed as an exciting regenerative medicine strategy for brain injury, there are limitations. The developed cerebral organoids (COs) represent a promising transplantation source for stroke that remains to be answered. Here, we transplanted COs at 55 days and explored the feasibility in the rat middle cerebral artery occlusion (MCAO) model of stroke. COs transplantation at 6 h or even 24 h after MCAO significantly reduces brain infarct volume and improves neurological motor function. Transplanted COs show the potential of multilineage differentiation to mimic in vivo cortical development, support motor cortex region-specific reconstruction, form neurotransmitter-related neurons, and achieve synaptic connection with host brain via in situ differentiation and cell replacement in stroke. Cells from transplanted COs show extensive migration into different brain regions along corpus callosum. The mechanisms underlying COs transplantation therapy are also associated with enhanced neurogenesis, synaptic reconstruction, axonal regeneration and angiogenesis, and decreased neural apoptosis with more survival neurons after stroke. Moreover, COs transplantation promotes predominantly exogenous neurogenesis in the transplantation periphery of ipsilateral cortex and predominantly endogenous neurogenesis in the hippocampus and subventricular zone. Together, we demonstrate the efficacy and underlying mechanisms of COs transplantation in stroke. This preliminary but promising study provides first-hand preclinical evidence for COs transplantation as a potential and effective intervention for stroke treatment.

Targeting NAMPT as a therapeutic strategy against stroke.

Stroke is the second and the leading most common cause of death in the world and China, respectively, but with few effective therapies. Nicotinamide phosphoribosyltransferase (NAMPT) is the rate-limiting enzyme for nicotinamide adenine dinucleotide (NAD) salvage synthesis in mammals, thereby influencing NAD-dependent enzymes and constituting a strong endogenous defence system against various stresses. Accumulating in-vitro and in-vivo studies have demonstrated the neuroprotective effect of NAMPT in stroke. Here, we review the direct evidence of NAMPT as a promising target against stroke from five potential therapeutic strategies, including NAMPT overexpression, recombinant NAMPT, NAMPT activators, NAMPT enzymatic product nicotinamide mononucleotide (NMN), and NMN precursors nicotinamide riboside and nicotinamide, and describe the relevant mechanisms and limitations, providing a promising choice for developing novel and effective therapeutic interventions against ischaemic and haemorrhagic stroke.

Influence of Microbiota on Intestinal Immune System in Ulcerative Colitis and Its Intervention.

Ulcerative colitis (UC) is an inflammatory bowel disease (IBD) with chronic and recurrent characteristics caused by multiple reasons. Although the pathogenic factors have not been clarified yet, recent studies have demonstrated that intestinal microbiota plays a major role in UC, especially in the immune system. This review focuses on the description of several major microbiota communities that affect UC and their interactions with the host. In this review, eight kinds of microbiota that are highly related to IBD, including Faecalibacterium prausnitzii, Clostridium clusters IV and XIVa, Bacteroides, Roseburia species, Eubacterium rectale, Escherichia coli, Fusobacterium, and Candida albicans are demonstrated on the changes in amount and roles in the onset and progression of IBD. In addition, potential therapeutic targets for UC involved in the regulation of microbiota, including NLRPs, vitamin D receptor as well as secreted proteins, are discussed in this review.

Organoid technology for brain and therapeutics research.

Brain is one of the most complex organs in human. The current brain research is mainly based on the animal models and traditional cell culture. However, the inherent species differences between humans and animals as well as the gap between organ level and cell level make it difficult to study human brain development and associated disorders through traditional technologies. Recently, the brain organoids derived from pluripotent stem cells have been reported to recapitulate many key features of human brain in vivo, for example recapitulating the zone of putative outer radial glia cells. Brain organoids offer a new platform for scientists to study brain development, neurological diseases, drug discovery and personalized medicine, regenerative medicine, and so on. Here, we discuss the progress, applications, advantages, limitations, and prospects of brain organoid technology in neurosciences and related therapeutics.

Neuroprotective Efficacy of an Aminopropyl Carbazole Derivative P7C3-A20 in Ischemic Stroke.

AIM: NAMPT is a novel therapeutic target of ischemic stroke. The aim of this study was to investigate the effect of a potential NAMPT activator, P7C3-A20, an aminopropyl carbazole derivative, on ischemic stroke. METHODS: In vitro study, neuron protection effect of P7C3-A20 was investigated by co-incubation with primary neurons subjected to oxygen-glucose deprivation (OGD) or oxygen-glucose deprivation/reperfusion (OGD/R) injury. In vivo experiment, P7C3-A20 was administrated in middle cerebral artery occlusion (MCAO) rats and infarct volume was examined. Lastly, the brain tissue nicotinamide adenine dinucleotide (NAD) levels were detected in P7C3-A20 treated normal or MCAO mice. RESULTS: Cell viability, morphology, and Tuj-1 staining confirmed the neuroprotective effect of P7C3-A20 in OGD or OGD/R model. P7C3-A20 administration significantly reduced cerebral infarction in MCAO rats. Moreover, brain NAD levels were elevated both in normal and MCAO mice after P7C3-A20 treatment. CONCLUSIONS: P7C3-A20 has neuroprotective effect in cerebral ischemia. The study contributes to the development of NAMPT activators against ischemic stroke and expands the horizon of the neuroprotective effect of aminopropyl carbazole chemicals.

Targeting Nicotinamide Phosphoribosyltransferase as a Potential Therapeutic Strategy to Restore Adult Neurogenesis.

Adult neurogenesis is the process of generating new neurons throughout life in the olfactory bulb and hippocampus of most mammalian species, which is closely related to aging and disease. Nicotinamide phosphoribosyltransferase (NAMPT), also an adipokine known as visfatin, is the rate-limiting enzyme for mammalian nicotinamide adenine dinucleotide (NAD) salvage synthesis by generating nicotinamide mononucleotide (NMN) from nicotinamide. Recent findings from our laboratory and other laboratories have provided much evidence that NAMPT might serve as a therapeutic target to restore adult neurogenesis. NAMPT-mediated NAD biosynthesis in neural stem/progenitor cells is important for their proliferation, self-renewal, and formation of oligodendrocytes in vivo and in vitro. Therapeutic interventions by the administration of NMN, NAD, or recombinant NAMPT are effective for restoring adult neurogenesis in several neurological diseases. We summarize adult neurogenesis in aging, ischemic stroke, traumatic brain injury, and neurodegenerative disease and review the advances of targeting NAMPT in restoring neurogenesis. Specifically, we provide emphasis on the P7C3 family, a class of proneurogenic compounds that are potential NAMPT activators, which might shed light on future drug development in neurogenesis restoration.

Discovery of Novel Inhibitors and Fluorescent Probe Targeting NAMPT.

Nicotinamide phosphoribosyltransferase (NAMPT) is a promising antitumor target. Novel NAMPT inhibitors with diverse chemotypes are highly desirable for development of antitumor agents. Using high throughput screening system targeting NAMPT on a chemical library of 30000 small-molecules, we found a non-fluorescent compound F671-0003 and a fluorescent compound M049-0244 with excellent in vitro activity (IC50: 85 nM and 170 nM respectively) and anti-proliferative activity against HepG2 cells. These two compounds significantly depleted cellular NAD levels. Exogenous NMN rescued their anti-proliferative activity against HepG2 cells. Structure-activity relationship study proposed a binding mode for NAMPT inhibitor F671-0003 and highlighted the importance of hydrogen bonding, hydrophobic and π-π interactions in inhibitor binding. Imaging study provided the evidence that fluorescent compound M049-0244 (3 µM) significantly stained living HepG2 cells. Cellular fluorescence was further verified to be NAMPT dependent by using RNA interference and NAMPT over expression transgenic mice. Our findings provide novel antitumor lead compounds and a "first-in-class" fluorescent probe for imaging NAMPT.