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Nature Structural & Molecular Biology Nov 2023In mammals, the kidney plays an essential role in maintaining blood homeostasis through the selective uptake, retention or elimination of toxins, drugs and metabolites....
In mammals, the kidney plays an essential role in maintaining blood homeostasis through the selective uptake, retention or elimination of toxins, drugs and metabolites. Organic anion transporters (OATs) are responsible for the recognition of metabolites and toxins in the nephron and their eventual urinary excretion. Inhibition of OATs is used therapeutically to improve drug efficacy and reduce nephrotoxicity. The founding member of the renal organic anion transporter family, OAT1 (also known as SLC22A6), uses the export of α-ketoglutarate (α-KG), a key intermediate in the Krebs cycle, to drive selective transport and is allosterically regulated by intracellular chloride. However, the mechanisms linking metabolite cycling, drug transport and intracellular chloride remain obscure. Here, we present cryogenic-electron microscopy structures of OAT1 bound to α-KG, the antiviral tenofovir and clinical inhibitor probenecid, used in the treatment of Gout. Complementary in vivo cellular assays explain the molecular basis for α-KG driven drug elimination and the allosteric regulation of organic anion transport in the kidney by chloride.
Topics: Animals; Organic Anion Transport Protein 1; Chlorides; Kidney; Biological Transport; Anions; Ketoglutaric Acids; Mammals
PubMed: 37482561
DOI: 10.1038/s41594-023-01039-y -
Neurobiology of Disease Aug 2023Loss of dopaminergic midbrain neurons perturbs l-serine and d-serine homeostasis in the post-mortem caudate putamen (CPu) of Parkinson's disease (PD) patients. However,...
Loss of dopaminergic midbrain neurons perturbs l-serine and d-serine homeostasis in the post-mortem caudate putamen (CPu) of Parkinson's disease (PD) patients. However, it is unclear whether the severity of dopaminergic nigrostriatal degeneration plays a role in deregulating serine enantiomers' metabolism. Here, through high-performance liquid chromatography (HPLC), we measured the levels of these amino acids in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys and MPTP-plus-probenecid (MPTPp)-treated mice to determine whether and how dopaminergic midbrain degeneration affects the levels of serine enantiomers in various basal ganglia subregions. In addition, in the same brain regions, we measured the levels of key neuroactive amino acids modulating glutamatergic neurotransmission, including l-glutamate, glycine, l-aspartate, d-aspartate, and their precursors l-glutamine, l-asparagine. In monkeys, MPTP treatment produced severe denervation of nigrostriatal dopaminergic fibers (⁓75%) and increased the levels of serine enantiomers in the rostral putamen (rPut), but not in the subthalamic nucleus, and the lateral and medial portion of the globus pallidus. Moreover, this neurotoxin significantly reduced the protein expression of the astrocytic serine transporter ASCT1 and the glycolytic enzyme GAPDH in the rPut of monkeys. Conversely, concentrations of d-serine and l-serine, as well as ASCT1 and GAPDH expression were unaffected in the striatum of MPTPp-treated mice, which showed only mild dopaminergic degeneration (⁓30%). These findings unveil a link between the severity of dopaminergic nigrostriatal degeneration and striatal serine enantiomers concentration, ASCT1 and GAPDH expression. We hypothesize that the up-regulation of d-serine and l-serine levels occurs as a secondary response within a homeostatic loop to support the metabolic and neurotransmission demands imposed by the degeneration of dopaminergic neurons.
Topics: Mice; Animals; Serine; 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Dopamine; Corpus Striatum; Mesencephalon; Amino Acids; Putamen; Homeostasis
PubMed: 37451474
DOI: 10.1016/j.nbd.2023.106226 -
ACS Omega Jul 2023Organosilicon and organotin compounds have been widely used in many fields, such as organic synthesis, materials science, and biochemistry, because of their unique...
Organosilicon and organotin compounds have been widely used in many fields, such as organic synthesis, materials science, and biochemistry, because of their unique physical and electronic properties. Recently, two novel compounds containing C-Si or C-Sn bonds have been synthesized. These compounds can be used for late modification of drug-like molecules such as probenecid, duloxetine, and fluoxetine derivatives. However, the detailed reaction mechanisms and the influencing factors that determine selectivity are still unclear. Moreover, several questions remain that are valuable to investigate further, such as (1) the influence of the solvent and the lithium salt on the reaction of the Si/Sn-Zn reagent, (2) the stereoselective functionalization of C-O bonds, and (3) the differences between silylation and stannylation. In the current study, we have explored the above issues with density functional theory and have found that stereoselectivity was most likely caused by the oxidative addition of cobalt to the C-O bond of alkenyl acetate with chelation assistance and that transmetalation was most likely the rate-determining step. For Sn-Zn reagents, the transmetalation was achieved by anion and cation pairs, whereas for Si-Zn reagents, the process was facilitated by Co-Zn complexes.
PubMed: 37426225
DOI: 10.1021/acsomega.3c02177 -
Biomedicines May 2023Probenecid is an old uricosuric agent used in clinics to treat gout and reduce the renal excretion of antibiotics. In recent years, probenecid has gained attention due... (Review)
Review
Probenecid is an old uricosuric agent used in clinics to treat gout and reduce the renal excretion of antibiotics. In recent years, probenecid has gained attention due to its ability to interact with membrane proteins such as TRPV2 channels, organic anion transporters, and pannexin 1 hemichannels, which suggests new potential therapeutic utilities in medicine. Some current functions of probenecid include their use as an adjuvant to increase the bioavailability of several drugs in the Central Nervous System (CNS). Numerous studies also suggest that this drug has important neuroprotective, antiepileptic, and anti-inflammatory properties, as evidenced by their effect against neurological and neurodegenerative diseases. In these studies, the use of probenecid as a Panx1 hemichannel blocker to reduce neuroinflammation is highlighted since neuroinflammation is a major trigger for diverse CNS alterations. Although the clinical use of probenecid has declined over the years, advances in its use in preclinical research indicate that it may be useful to improve conventional therapies in the psychiatric field where the drugs used have a low bioavailability, either because of a deficient passage through the blood-brain barrier or a high efflux from the CNS or also a high urinary clearance. This review summarizes the history, pharmacological properties, and recent research uses of probenecid and discusses its future projections as a potential pharmacological strategy to intervene in neurodegeneration as an outcome of neuroinflammation.
PubMed: 37371611
DOI: 10.3390/biomedicines11061516 -
Cells May 2023Controversial reports have suggested that SARS-CoV E and 3a proteins are plasma membrane viroporins. Here, we aimed at better characterizing the cellular responses...
Controversial reports have suggested that SARS-CoV E and 3a proteins are plasma membrane viroporins. Here, we aimed at better characterizing the cellular responses induced by these proteins. First, we show that expression of SARS-CoV-2 E or 3a protein in CHO cells gives rise to cells with newly acquired round shapes that detach from the Petri dish. This suggests that cell death is induced upon expression of E or 3a protein. We confirmed this by using flow cytometry. In adhering cells expressing E or 3a protein, the whole-cell currents were not different from those of the control, suggesting that E and 3a proteins are not plasma membrane viroporins. In contrast, recording the currents on detached cells uncovered outwardly rectifying currents much larger than those observed in the control. We illustrate for the first time that carbenoxolone and probenecid block these outwardly rectifying currents; thus, these currents are most probably conducted by pannexin channels that are activated by cell morphology changes and also potentially by cell death. The truncation of C-terminal PDZ binding motifs reduces the proportion of dying cells but does not prevent these outwardly rectifying currents. This suggests distinct pathways for the induction of these cellular events by the two proteins. We conclude that SARS-CoV-2 E and 3a proteins are not viroporins expressed at the plasma membrane.
Topics: Cricetinae; Animals; SARS-CoV-2; Cricetulus; COVID-19; Cell Membrane; CHO Cells
PubMed: 37296595
DOI: 10.3390/cells12111474 -
Journal of Xenobiotics May 2023Accurate predictions of drug uptake transporter involvement in renal excretion of xenobiotics require determination of in vitro transport kinetic parameters under...
Accurate predictions of drug uptake transporter involvement in renal excretion of xenobiotics require determination of in vitro transport kinetic parameters under initial-rate conditions. The purpose of the present study was to determine how changing the incubation time from initial rate to steady state influences ligand interactions with the renal organic anion transporter 1 (OAT1), and the impact of the different experimental conditions on pharmacokinetic predictions. Transport studies were performed with Chinese hamster ovary cells expressing OAT1 (CHO-OAT1) and the Simcyp Simulator was used for physiological-based pharmacokinetic predictions. Maximal transport rate and intrinsic uptake clearance (CL) for PAH decreased with increasing incubation time. The CL values ranged 11-fold with incubation times spanning from 15 s (CL, initial rate) to 45 min (CL, steady state). The Michaelis constant (K) was also influenced by the incubation time with an apparent increase in the Km value at longer incubation times. Inhibition potency of five drugs against PAH transport was tested using incubation times of either 15 s or 10 min. There was no effect of time on inhibition potency for omeprazole or furosemide, whereas indomethacin was less potent, and probenecid (~2-fold) and telmisartan (~7-fold) more potent with the longer incubation time. Notably, the inhibitory effect of telmisartan was reversible, albeit slowly. A pharmacokinetic model was developed for PAH using the CL value. The simulated plasma concentration-time profile, renal clearance, and cumulative urinary excretion-time profile of PAH agreed well with reported clinical data, and the PK parameters were sensitive to the time-associated CL value used in the model.
PubMed: 37218810
DOI: 10.3390/jox13020016 -
Mediators of Inflammation 2023Neuropathic pain is a complex sort of pain that is detrimental to individuals' health, both physically and mentally, but merely a small portion of them could witness... (Review)
Review
BACKGROUND
Neuropathic pain is a complex sort of pain that is detrimental to individuals' health, both physically and mentally, but merely a small portion of them could witness pain alleviation. Mirogabalin, by distinctive binding characteristics of voltage-gated calcium channels, has won approval from the Japanese authority as a third member of gabapentinoids in Japan. Our review was aimed at encompassing the bulk of clinical research on mirogabalin, which included clinical trials, special considerations, coadministration studies, case reports, and cost-effectiveness studies.
METHODS
A review was carried out on a series of platforms, such as PubMed, MEDLINE, and Scopus, up to December 2021 using the keywords as follows: "mirogabalin OR mirogabalin besylate OR Tarlige OR DS-5565" AND "neuropathic pain OR Neuropathy."
RESULTS
Mirogabalin demonstrated analgesic activity and manageable adverse reactions and provides a new alternative for individuals with PHN or DPNP in 3 phase II and 4 III trials. Mirogabalin alleviated pain markedly in comparison with placebo. Administration of mirogabalin on a long-term basis is a flexible dosage regimen for patients with PHN. It is noteworthy that mirogabalin should be administrated cautiously when combined with probenecid and cimetidine on account of a slight increase in pharmacodynamics effects of mirogabalin.
CONCLUSION
The development of mirogabalin allows further optimization of individual treatment strategies so as to provide more therapeutic choices in this medical domain.
Topics: Humans; Neuralgia; Bridged Bicyclo Compounds; Analgesics; Cimetidine
PubMed: 37152369
DOI: 10.1155/2023/4893436 -
Zhongguo Ying Yong Sheng Li Xue Za Zhi... Sep 2022To investigate whether probenecid (PROB) could improve the proliferation and migration ability of rats' pulmonary artery smooth muscle cells induced by platelet-derived...
OBJECTIVE
To investigate whether probenecid (PROB) could improve the proliferation and migration ability of rats' pulmonary artery smooth muscle cells induced by platelet-derived growth factor-BB (PDGF-BB).
METHODS
Primary pulmonary artery smooth muscle cells (PASMCs) of SD rats were cultured , and were randomly divided into control group (CON group), PDGF-BB group (10 ng/ml PDGF-BB treatment for 24 h) and PDGF-BB+PROB group (10 ng/ml PDGF-BB and 200 μmol/L PROB treatment for 24 h, PROB is a specific blocker of pannexin-1). CCK-8 method was used to select the suitable intervention concentrations of PROB and PDGF-BB, and to detect the proliferation of PASMCs in each group. The migration ability of PASMCs was detected by Transwell assay and cell scratch test. Immunofluorescence cytochemistry and Western blot were used to detect the protein expressions and distribution of osteopontin (OPN) and proliferating cell nuclear antigen (PCNA) in PASMCs.
RESULTS
Compared with CON group, the migration and proliferation ability of PASMCs in PDGF-BB group were enhanced (<0.05). After treated with PROB, the migration and proliferation ability of PASMCs in PDGF-BB+PROB group were decreased significantly (<0.05). Compared with CON group, the expression and protein levels of OPN and PCNA in PDGF-BB group were increased significantly (<0.05), while the expression and protein levels of OPN and PCNA in PDGF-BB+PROB were decreased significantly (<0.05).
CONCLUSION
Probenecid inhibits the migration and proliferation of PDGF-BB-induced PASMCs by blocking Pannexin-1.
Topics: Rats; Animals; Becaplermin; Proliferating Cell Nuclear Antigen; Probenecid; Proto-Oncogene Proteins c-sis; Pulmonary Artery; Cell Proliferation; Rats, Sprague-Dawley; Myocytes, Smooth Muscle; Cells, Cultured
PubMed: 37088767
DOI: 10.12047/j.cjap.6326.2022.101 -
European Thyroid Journal Aug 2023Thyroid hormone (TH) transport represents a critical first step in governing intracellular TH regulation. It is still unknown whether the full repertoire of TH...
OBJECTIVE
Thyroid hormone (TH) transport represents a critical first step in governing intracellular TH regulation. It is still unknown whether the full repertoire of TH transporters has been identified. Members of the solute carrier (SLC) 22 family have substrates in common with the known TH transporters of the organic anion-transporting peptide family. Therefore, we screened the SLC22 family for TH transporters.
METHODS
Uptake of 1 nM of iodothyronines or sulfated iodothyronines in COS1 cells expressing SLC22 proteins was performed.
RESULTS
We first tested 25 mouse (m) SLC22 proteins for TH uptake and found that the majority of the organic anion transporter (OAT) clade were capable of 3,3',5-triiodothyronine and/or thyroxine (T4) transport. Based on phylogenetic tree analysis of the mouse and human (h) SLC22 family, we selected eight hSLC22s that grouped with the newly identified mouse TH transporters. Of these, four tested positive for uptake of one or more substrates, particularly hSLC22A11 showed robust (3-fold over control) uptake of T4. Uptake of sulfated iodothyronines was strongly (up to 17-fold) induced by some SLC22s, most notably SLC22A8, hSLC22A9, mSLC22A27 and mSLC22A29. Finally, the zebrafish orthologues of SLC22A6/8 drOatx and drSlc22a6l also transported almost all (sulfated) iodothyronines tested. The OAT inhibitors lesinurad and probenecid inhibited most SLC22 proteins.
CONCLUSIONS
Our results demonstrated that members of the OAT clade of the SLC22 family constitute a novel, evolutionary conserved group of transporters for (sulfated) iodothyronines. Future studies should reveal the relevance of these transporters in TH homeostasis and physiology.
Topics: Humans; Mice; Animals; Phylogeny; Zebrafish; Sulfates; Thyroid Hormones; Membrane Transport Proteins; Organic Anion Transporters
PubMed: 37074673
DOI: 10.1530/ETJ-23-0023 -
Journal of Cancer 2023Pannexin 1 (PANX1) is expressed in many tissue types including tissues of neural origin. Neuroblastoma (NB) is a neural crest-derived malignancy mainly occurring in...
Pannexin 1 (PANX1) is expressed in many tissue types including tissues of neural origin. Neuroblastoma (NB) is a neural crest-derived malignancy mainly occurring in children. The majority of NB patients present with high-risk disease for which current therapies are ineffective. Here, we show that while PANX1 is expressed in NB of all stages, high expression in high-risk NB is associated with a reduced survival probability. PANX1 channel inhibition using probenecid (PBN) or carbenoxolone (CBX) reduced the proliferation of our panel of high-risk NB cell lines. We show that expression of the Y10F PANX1 mutant, which cannot be phosphorylated on tyrosine 10 and acts in a dominant-negative manner, curtailed NB cell proliferation. Furthermore, PBN and CBX treatment halted the growth of NB spheroids and in some cases triggered the regression of established NB spheroids. Finally, both drugs reduced the progression of high-risk NB . Together our data indicate that PANX1 channels regulate human NB malignant properties and that the use of PBN or CBX may provide a new therapeutic approach for high-risk NB.
PubMed: 37056395
DOI: 10.7150/jca.79552