Glucose-mediated mitochondrial reprogramming by cholesterol export at TM4SF5-enriched mitochondria-lysosome contact sites.

BACKGROUND: Transmembrane 4 L six family member 5 (TM4SF5) translocates subcellularly and functions metabolically, although it is unclear how intracellular TM4SF5 translocation is linked to metabolic contexts. It is thus of interests to understand how the traffic dynamics of TM4SF5 to subcellular endosomal membranes are correlated to regulatory roles of metabolisms. METHODS: Here, we explored the metabolic significance of TM4SF5 localization at mitochondria-lysosome contact sites (MLCSs), using in vitro cells and in vivo animal systems, via approaches by immunofluorescence, proximity labelling based proteomics analysis, organelle reconstitution etc. RESULTS: Upon extracellular glucose repletion following depletion, TM4SF5 became enriched at MLCSs via an interaction between mitochondrial FK506-binding protein 8 (FKBP8) and lysosomal TM4SF5. Proximity labeling showed molecular clustering of phospho-dynamic-related protein I (DRP1) and certain mitophagy receptors at TM4SF5-enriched MLCSs, leading to mitochondrial fission and autophagy. TM4SF5 bound NPC intracellular cholesterol transporter 1 (NPC1) and free cholesterol, and mediated export of lysosomal cholesterol to mitochondria, leading to impaired oxidative phosphorylation but intact tricarboxylic acid (TCA) cycle and ß-oxidation. In mouse models, hepatocyte Tm4sf5 promoted mitophagy and cholesterol transport to mitochondria, both with positive relations to liver malignancy. CONCLUSIONS: Our findings suggested that TM4SF5-enriched MLCSs regulate glucose catabolism by facilitating cholesterol export for mitochondrial reprogramming, presumably while hepatocellular carcinogenesis, recapitulating aspects for hepatocellular carcinoma metabolism with mitochondrial reprogramming to support biomolecule synthesis in addition to glycolytic energetics.

Benzo[a]pyrene represses synaptic vesicle exocytosis by inhibiting P/Q-type calcium channels in hippocampal neurons.

Humans are exposed to the common carcinogen benzo[a]pyrene (BaP) by ingesting contaminated foods and water or inhaling polluted air. Given the enriched lipids and reduced antioxidative properties in the brain and the accumulation of BaP in the brain due to its high lipophilicity, the brain is susceptible to BaP-induced toxicity. Exposure to BaP leads to impairments in learning and memory, increased anxiety behavior, and neuronal death. It induces protein dysfunctions in neuronal compartments that play essential roles in neuronal activity or physiology. However, the neurotoxicity of BaP on presynaptic terminals, which is crucial to neurotransmission by releasing synaptic vesicles that contain neurotransmitters, has not yet been investigated. In the present study, we investigated the toxicity of BaP at presynaptic terminals in living hippocampal neurons. These neurons were sourced from transgenic mice pups (postnatal 1-day, a total of 12 pups, equal numbers for each sex) that endogenously express synaptic vesicle-fused pHluorin, which is a green fluorescent protein that enables monitoring of synaptic vesicle dynamics. We observed that BaP suppressed synaptic vesicle exocytosis by inhibiting presynaptic Ca2+ entry via P/Q-type Ca2+ channels. Together with molecular docking simulation, we speculate that BaP and metabolites may bind to the P/Q Ca2+ channels. These results suggest the toxic mechanism of BaP exposure-induced abnormal behavior that provides a basis to evaluate the risk assessment of BaP-induced neurotoxicity.

Albumin-binding photosensitizer capable of targeting glioma via the SPARC pathway.

BACKGROUND: Malignant glioma is among the most lethal and frequently occurring brain tumors, and the average survival period is 15 months. Existing chemotherapy has low tolerance and low blood-brain barrier (BBB) permeability; therefore, the required drug dose cannot be accurately delivered to the tumor site, resulting in an insufficient drug effect. METHODS: Herein, we demonstrate a precision photodynamic tumor therapy using a photosensitizer (ZnPcS) capable of binding to albumin in situ, which can increase the permeability of the BBB and accurately target glioma. Albumin-binding ZnPcS was designed to pass through the BBB and bind to secreted protein acidic and rich in cysteine (SPARC), which is abundant in the glioma plasma membrane. RESULTS: When the upper part of a mouse brain was irradiated using a laser (0.2 W cm- 2) after transplantation of glioma and injection of ZnPcS, tumor growth was inhibited by approximately 83.6%, and the 50% survival rate of the treatment group increased by 14 days compared to the control group. In glioma with knockout SPARC, the amount of ZnPcS entering the glioma was reduced by 63.1%, indicating that it can target glioma through the SPARC pathway. CONCLUSION: This study showed that the use of albumin-binding photosensitizers is promising for the treatment of malignant gliomas.

GPCR Agonist-to-Antagonist Conversion: Enabling the Design of Nucleoside Functional Switches for the A2A Adenosine Receptor.

Modulators of the G protein-coupled A2A adenosine receptor (A2AAR) have been considered promising agents to treat Parkinson's disease, inflammation, cancer, and central nervous system disorders. Herein, we demonstrate that a thiophene modification at the C8 position in the common adenine scaffold converted an A2AAR agonist into an antagonist. We synthesized and characterized a novel A2AAR antagonist, 2 (LJ-4517), with Ki = 18.3 nM. X-ray crystallographic structures of 2 in complex with two thermostabilized A2AAR constructs were solved at 2.05 and 2.80 Å resolutions. In contrast to A2AAR agonists, which simultaneously interact with both Ser2777.42 and His2787.43, 2 only transiently contacts His2787.43, which can be direct or water-mediated. The n-hexynyl group of 2 extends into an A2AAR exosite. Structural analysis revealed that the introduced thiophene modification restricted receptor conformational rearrangements required for subsequent activation. This approach can expand the repertoire of adenosine receptor antagonists that can be designed based on available agonist scaffolds.

Design, Synthesis and Biological Evaluation of 1,3,5-Triazine Derivatives Targeting hA1 and hA3 Adenosine Receptor.

Adenosine mediates various physiological activities in the body. Adenosine receptors (ARs) are widely expressed in tumors and the tumor microenvironment (TME), and they induce tumor proliferation and suppress immune cell function. There are four types of human adenosine receptor (hARs): hA1, hA2A, hA2B, and hA3. Both hA1 and hA3 AR play an important role in tumor proliferation. We designed and synthesized novel 1,3,5-triazine derivatives through amination and Suzuki coupling, and evaluated them for binding affinities to each hAR subtype. Compounds 9a and 11b showed good binding affinity to both hA1 and hA3 AR, while 9c showed the highest binding affinity to hA1 AR. In this study, we discovered that 9c inhibits cell viability, leading to cell death in lung cancer cell lines. Flow cytometry analysis revealed that 9c caused an increase in intracellular reactive oxygen species (ROS) and a depolarization of the mitochondrial membrane potential. The binding mode of 1,3,5-triazine derivatives to hA1 and hA3 AR were predicted by a molecular docking study.

Artificial intelligence assessment for early detection and prediction of renal impairment using electrocardiography.

PURPOSE: Although renal failure is a major healthcare burden globally and the cornerstone for preventing its irreversible progression is an early diagnosis, an adequate and noninvasive tool to screen renal impairment (RI) reliably and economically does not exist. We developed an interpretable deep learning model (DLM) using electrocardiography (ECG) and validated its performance. METHODS: This retrospective cohort study included two hospitals. We included 115,361 patients who had at least one ECG taken with an estimated glomerular filtration rate measurement within 30 min of the index ECG. A DLM was developed using 96,549 ECGs of 55,222 patients. The internal validation included 22,949 ECGs of 22,949 patients. Furthermore, we conducted an external validation with 37,190 ECGs of 37,190 patients from another hospital. The endpoint was to detect a moderate to severe RI (estimated glomerular filtration rate < 45 ml/min/1.73m2). RESULTS: The area under the receiver operating characteristic curve (AUC) of a DLM using a 12-lead ECG for detecting RI during the internal and external validation was 0.858 (95% confidence interval 0.851-0.866) and 0.906 (0.900-0.912), respectively. In the initial evaluation of 25,536 individuals without RI patients whose DLM was defined as having a higher risk had a significantly higher chance of developing RI than those in the low-risk group (17.2% vs. 2.4%, p < 0.001). The sensitivity map indicated that the DLM focused on the QRS complex and T-wave for detecting RI. CONCLUSION: The DLM demonstrated high performance for RI detection and prediction using 12-, 6-, single-lead ECGs.

Structure-activity relationships of novel quinazoline derivatives with high selectivity for HER2 over EGFR.

The gene amplification of human epidermal growth factor receptor 2 (HER2) plays an essential role in the proliferation and progression of several cancers. However, HER2 inhibitors such as lapatinib strongly suppress wild-type EGFR, resulting in severe adverse effects. Therefore, there is an unmet need for highly selective HER2 inhibitors. In this study, we describe the design and synthesis of novel quinazoline derivatives that exhibit enhanced selectivity for HER2 over wild-type EGFR. Structure-activity relationship analysis indicated that the selectivity for HER2 over EGFR depends on the aniline moiety at C-4 and the substituents at C-6 in the quinazoline derivatives. Compound 7c with an IC50 of 8 nM for HER2 exhibited significantly higher selectivity for HER2 over EGFR, with a 240-fold improvement over lapatinib. In addition, the synthesized compounds exhibited anti-proliferative activity in the nanomolar range against SKBR3, a human breast cancer cell line that overexpresses HER2.

N-terminus-independent activation of c-Src via binding to a tetraspan(in) TM4SF5 in hepatocellular carcinoma is abolished by the TM4SF5 C-terminal peptide application.

Active c-Src non-receptor tyrosine kinase localizes to the plasma membrane via N-terminal lipid modification. Membranous c-Src causes cancer initiation and progression. Even though transmembrane 4 L six family member 5 (TM4SF5), a tetraspan(in), can be involved in this mechanism, the molecular and structural influence of TM4SF5 on c-Src remains unknown. Methods: Here, we investigated molecular and structural details by which TM4SF5 regulated c-Src devoid of its N-terminus and how cell-penetrating peptides were able to interrupt c-Src activation via interference of c-Src-TM4SF5 interaction in hepatocellular carcinoma models. Results: The TM4SF5 C-terminus efficiently bound the c-Src SH1 kinase domain, efficiently to the inactively-closed form. The complex involved protein tyrosine phosphatase 1B able to dephosphorylate Tyr530. The c-Src SH1 domain alone, even in a closed form, bound TM4SF5 to cause c-Src Tyr419 and FAK Y861 phosphorylation. Homology modeling and molecular dynamics simulation studies predicted the directly interfacing residues, which were further validated by mutational studies. Cell penetration of TM4SF5 C-terminal peptides blocked the interaction of TM4SF5 with c-Src and prevented c-Src-dependent tumor initiation and progression in vivo. Conclusions: Collectively, these data demonstrate that binding of the TM4SF5 C-terminus to the kinase domain of inactive c-Src leads to its activation. Because this binding can be abolished by cell-penetrating peptides containing the TM4SF5 C-terminus, targeting this direct interaction may be an effective strategy for developing therapeutics that block the development and progression of hepatocellular carcinoma.

Subtle Chemical Changes Cross the Boundary between Agonist and Antagonist: New A3 Adenosine Receptor Homology Models and Structural Network Analysis Can Predict This Boundary.

Distinguishing compounds' agonistic or antagonistic behavior would be of great utility for the rational discovery of selective modulators. We synthesized truncated nucleoside derivatives and discovered 6c (Ki = 2.40 nM) as a potent human A3 adenosine receptor (hA3AR) agonist, and subtle chemical modification induced a shift from antagonist to agonist. We elucidated this shift by developing new hA3AR homology models that consider the pharmacological profiles of the ligands. Taken together with molecular dynamics (MD) simulation and three-dimensional (3D) structural network analysis of the receptor-ligand complex, the results indicated that the hydrogen bonding with Thr943.36 and His2727.43 could make a stable interaction between the 3'-amino group with TM3 and TM7, and the corresponding induced-fit effects may play important roles in rendering the agonistic effect. Our results provide a more precise understanding of the compounds' actions at the atomic level and a rationale for the design of new drugs with specific pharmacological profiles.

Recognition of nuclear export signals by CRM1 carrying the oncogenic E571K mutation.

The E571K mutation of CRM1 is highly prevalent in some cancers, but its mechanism of tumorigenesis is unclear. Glu571 of CRM1 is located in its nuclear export signal (NES)-binding groove, suggesting that binding of select NESs may be altered. We generated HEK 293 cells with either monoallelic CRM1WT/E571K or biallelic CRM1E571K/E571K using CRISPR/Cas9. We also combined analysis of binding affinities and structures of 27 diverse NESs for wild-type and E571K CRM1 with structure-based bioinformatics. While most NESs bind the two CRM1 similarly, NESs from Mek1, eIF4E-transporter, and RPS2 showed >10-fold affinity differences. These NESs have multiple charged side chains binding close to CRM1 position 571, but this feature alone was not sufficient to predict different binding to CRM1(E571K). Consistent with eIF4E-transporter NES binding weaker to CRM1(E571K), eIF4E-transporter was mislocalized in tumor cells carrying CRM1(E571K). This serves as proof of concept that understanding how CRM1(E571K) affects NES binding provides a platform for identifying cargoes that are mislocalized in cancer upon CRM1 mutation. Finally, we showed that large affinity changes seen with some NES peptides (of Mek1 and RPS2) do not always translate to the full-length cargoes, suggesting limitations with current NES prediction methods. Therefore, comprehensive studies like ours are imperative to identify CRM1 cargoes with real pathogenic potential.

pCRM1exportome: database of predicted CRM1-dependent Nuclear Export Signal (NES) motifs in cancer-related genes.

MOTIVATION: The consensus pattern of Nuclear Export Signal (NES) is a short sequence motif that is commonly identified in protein sequences, whether the motif acts as an NES (true positive) or not (false positive). Finding more plausible NES functioning regions among the vast array of consensus-matching segments would provide an interesting resource for further experimental validation. Better defined NES should also allow meaningful mapping of cancer-related mutation positions, leading to plausible explanations for the relationship between nuclear export and disease. RESULTS: Possible NES candidate regions are extracted from the cancer-related human reference proteome. Extracted NES are scored for reliability by combining sequence-based and structure-based approaches. The confidently identified NES candidate motifs were checked for overlap with cancer-related mutation positions annotated in the COSMIC database. Among the ∼700 cancer-related sequences in the COSMIC Cancer Gene Census, 178 sequences are predicted to have possible NES motifs containing cancer-related mutations at their key positions. These lists are organized into our database (pCRM1exportome), and other protein sequences in the human reference proteome can also be retrieved by their UniProt IDs. AVAILABILITY AND IMPLEMENTATION: The database is freely available at http://prodata.swmed.edu/pCRM1exportome. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Water-Soluble Phthalocyanines Selectively Bind to Albumin Dimers: A Green Approach Toward Enhancing Tumor-Targeted Photodynamic Therapy.

Targeted delivery of therapeutic agents is of particular interest in the field of cancer treatment. However, there is an urgent need for developing clinically promising targeting approaches that can be readily administered in a green manner. Methods: Five phthalocyanine derivatives bearing different anionic and cationic groups were designed and synthesized. Then, their binding affinity with albumin were studied using gel assays, optical spectra and computational simulation. Finally, in vitro and in vivo fluorescence imaging and photodynamic therapy (PDT) evaluations were carried out. Results: The two positively charged compounds could selectively bind to albumin dimer over albumin monomer, while the three negatively charged phthalocyanines could bind to both albumin monomer and dimer. Following systemic administration, the phthalocyanines show improved tumor accumulation via transport by natural albumin. PDT evaluations indicate that one of the positively charged compounds, ZnPcN4, shows outstanding phototherapeutic efficacy against tumors in preclinical models. Conclusion: Our findings demonstrate that the use of water-soluble phthalocyanines as photosensitizers and in vivo albumin as a natural carrier may provide a green and efficient approach for tumor-targeted imaging and therapy.

Importance of protein dynamics in the structure-based drug discovery of class A G protein-coupled receptors (GPCRs).

Demand for novel GPCR modulators is increasing as the association between the GPCR signaling pathway and numerous diseases such as cancers, psychological and metabolic disorders continues to be established. In silico structure-based drug design (SBDD) offers an outlet where researchers could exploit the accumulating structural information of GPCR to expedite the process of drug discovery. The coupling of structure-based approaches such as virtual screening and molecular docking with molecular dynamics and/or Monte Carlo simulation aids in reflecting the dynamics of proteins in nature into previously static docking studies, thus enhancing the accuracy of rationally designed ligands. This review will highlight recent computational strategies that incorporate protein flexibility into SBDD of GPCR-targeted ligands.

Structural prerequisites for CRM1-dependent nuclear export signaling peptides: accessibility, adapting conformation, and the stability at the binding site.

Nuclear export signal (NES) motifs function as essential regulators of the subcellular location of proteins by interacting with the major nuclear exporter protein, CRM1. Prediction of NES is of great interest in many aspects of research including cancer, but currently available methods, which are mostly based on the sequence-based approaches, have been suffered from high false positive rates since the NES consensus patterns are quite commonly observed in protein sequences. Therefore, finding a feature that can distinguish real NES motifs from false positives is desired to improve the prediction power, but it is quite challenging when only using the sequence. Here, we provide a comprehensive table for the validated cargo proteins, containing the location of the NES consensus patterns with the disordered propensity plots, known protein domain information, and the predicted secondary structures. It could be useful for determining the most plausible NES region in the context of the whole protein sequence and suggests possibilities for some non-binders of the annotated regions. In addition, using the currently available crystal structures of CRM1 bound to various classes of NES peptides, we adopted, for the first time, the structure-based prediction of the NES motifs bound to the CRM1's binding groove. Combining sequence-based and structure-based predictions, we suggest a novel and more straight-forward approach to identify CRM1-binding NES sequences by analysis of their structural prerequisites and energetic evaluation of the stability at the CRM1's binding site.

In Vivo Albumin Traps Photosensitizer Monomers from Self-Assembled Phthalocyanine Nanovesicles: A Facile and Switchable Theranostic Approach.

Albumin is a promising candidate as a biomarker for potential disease diagnostics and has been extensively used as a drug delivery carrier for decades. In these two directions, many albumin-detecting probes and exogenous albumin-based nanocomposite delivery systems have been developed. However, there are only a few cases demonstrating the specific interactions of exogenous probes with albumin in vivo, and nanocomposite delivery systems usually suffer from tedious fabrication processes and potential toxicity of the complexes. Herein, we demonstrate a facile "one-for-all" switchable nanotheranostic (NanoPcS) for both albumin detection and cancer treatment. In particular, the in vivo specific binding between albumin and PcS, arising from the disassembly of injected NanoPcS, is confirmed using an inducible transgenic mouse system. Fluorescence imaging and antitumor tests on different tumor models suggest that NanoPcS has superior tumor-targeting ability and the potential for time-modulated, activatable photodynamic therapy.

Thyroid dysfunction in children with leukemia over the first year after hematopoietic stem cell transplantation.

Background Thyroid function in children with leukemia during the first year after hematopoietic stem cell transplantation (HSCT) was investigated. Methods The medical records of 186 subjects [111 boys and 75 girls; lymphoid=75, myeloid=111; median age at HSCT was 10.7 (0.8-21.8) years old] were reviewed retrospectively. Results In children with leukemia, T3 decreased at 1 month (p<0.001) and recovered 9 months to the levels before HSCT. TSH decreased at 1 month (p<0.001), recovered at 3 months and increased at 12 months (p<0.001) to the levels before HSCT. The incidence of euthyroid sick syndrome (ESS, 23.2%, 15.5%, 5.9%, 5.2%, 3.9%, p for trend <0.001) decreased and subclinical hypothyroidism (SH, 0%, 3.9%, 14.8%, 22.1%, 21.3%, p for trend <0.001) increased at 1, 3, 6, 9 and 12 months after HSCT. Out of 55 patients developing ESS during 3 months after HSCT, 54 recovered to normal thyroid function within 5 months without medication. Among the total 186 subjects, 21 patients have been treated with levothyroxine. Both height and weight standard deviation scores continued to decrease over 1 year after HSCT. Conclusions In children with leukemia, one-quarter had ESS at 1 month and one-fifth had SH at 12 months and continued growth impairments were observed during 1 year after HSCT. Most of the ESS patients recovered to normal within 5 months without medication. More long-term follow-up of thyroid function and growth in children with leukemia after HSCT is crucial.

Understanding G Protein-Coupled Receptor Allostery via Molecular Dynamics Simulations: Implications for Drug Discovery.

Unraveling the mystery of protein allostery has been one of the greatest challenges in both structural and computational biology. However, recent advances in computational methods, particularly molecular dynamics (MD) simulations, have led to its utility as a powerful and popular tool for the study of protein allostery. By capturing the motions of a protein's constituent atoms, simulations can enable the discovery of allosteric hot spots and the determination of the mechanistic basis for allostery. These structural and dynamic studies can provide a foundation for a wide range of applications, including rational drug design and protein engineering. In our laboratory, the use of MD simulations and network analysis assisted in the elucidation of the allosteric hotspots and intracellular signal transduction of G protein-coupled receptors (GPCRs), primarily on one of the adenosine receptor subtypes, A2A adenosine receptor (A2AAR). In this chapter, we describe a method for calculating the map of allosteric signal flow in different GPCR conformational states and illustrate how these concepts have been utilized in understanding the mechanism of GPCR allostery. These structural studies will provide valuable insights into the allosteric and orthosteric modulations that would be of great help to design novel drugs targeting GPCRs in pathological states.

Ultraslow Water-Mediated Transmembrane Interactions Regulate the Activation of A2A Adenosine Receptor.

Water molecules inside a G-protein coupled receptor (GPCR) have recently been spotlighted in a series of crystal structures. To decipher the dynamics and functional roles of internal water molecules in GPCR activity, we studied the A2A adenosine receptor using microsecond molecular-dynamics simulations. Our study finds that the amount of water flux across the transmembrane (TM) domain varies depending on the receptor state, and that the water molecules of the TM channel in the active state flow three times more slowly than those in the inactive state. Depending on the location in solvent-protein interface as well as the receptor state, the average residence time of water in each residue varies from ∼O(10(2)) ps to ∼O(10(2)) ns. Especially, water molecules, exhibiting ultraslow relaxation (∼O(10(2)) ns) in the active state, are found around the microswitch residues that are considered activity hotspots for GPCR function. A continuous allosteric network spanning the TM domain, arising from water-mediated contacts, is unique in the active state, underscoring the importance of slow water molecules in the activation of GPCRs.

A novel pyrazole derivative protects from ovariectomy-induced osteoporosis through the inhibition of NADPH oxidase.

Osteoclast cells (OCs) are differentiated from bone marrow-derived macrophages (BMMs) by activation of receptor activator of nuclear factor κB (NF-κB) ligand (RANKL). Activation of NADPH oxidase (Nox) isozymes is involved in RANKL-dependent OC differentiation, implicating Nox isozymes as therapeutic targets for treatment of osteoporosis. Here, we show that a novel pyrazole derivative, Ewha-18278 has high inhibitory potency on Nox isozymes. Blocking the activity of Nox with Ewha-18278 inhibited the responses of BMMs to RANKL, including reactive oxygen species (ROS) generation, activation of mitogen-activated protein (MAP) kinases and NF-κB, and OC differentiation. To evaluate the anti-osteoporotic function of Ewha-18278, the derivative was applied to estrogen-deficient ovariectomized (OVX) ddY mice. Oral administration of Ewha-18278 (10 mg/kg/daily, 4 weeks) into the mice recovered bone mineral density, trabecular bone volume, trabecular bone length, number and thickness, compared to control OVX ddY mice. Moreover, treatment of OVX ddY mice with Ewha-18278 increased bone strength by increasing cortical bone thickness. We provide that Ewha-18278 displayed Nox inhibition and blocked the RANKL-dependent cell signaling cascade leading to reduced differentiation of OCs. Our results implicate Ewha-18278 as a novel therapeutic agent for the treatment of osteoporosis.

Structure-Activity Relationships of Neplanocin A Analogues as S-Adenosylhomocysteine Hydrolase Inhibitors and Their Antiviral and Antitumor Activities.

On the basis of the potent inhibitory activity of neplanocin A (1) against S-adenosylhomocysteine (AdoHcy) hydrolase, we analyzed the comprehensive structure-activity relationships by modifying the adenine and carbasugar moiety of 1 to find the pharmacophore in the active site of the enzyme. The introduction of 7-deazaadenine instead of adenine eliminated the inhibitory activity against the AdoHcy hydrolase, while 3-deazaadenine maintained the inhibitory activity of the enzyme, indicating that N-7 is essential for its role as a hydrogen bonding acceptor. The substitution of hydrogen at the 6'-position with fluorine increased the inhibitory activity of the enzyme. The one-carbon homologation at the 5'-position generally decreased the inhibitory activity of the enzyme, indicating that steric repulsion exists. A molecular docking study also supported these experimental data. In this study, 6'-fluoroneplanocin A (2) was the most potent inhibitor of AdoHcy hydrolase (IC50 = 0.24 µM). It showed a potent anti-VSV activity (EC50 = 0.43 µM) and potent anticancer activity in all the human tumor cell lines tested.