title: Biological and structural evaluation of 10R- and 10S-methylthio-DDACTHF reveals a new role for sulfur in inhibition of glycinamide ribonucleotide transformylase.
abstract: Glycinamide ribonucleotide transformylase (GAR Tfase) is a folate-dependent enzyme in the de novo purine biosynthesis pathway, which has long been considered a potential target for development of anti-neoplastic therapeutics. Here we report the biological and X-ray crystallographic evaluations of both independent C10 diastereomers, 10S- and 10R-methylthio-DDACTHF, bound to human GAR Tfase, including the highest-resolution apo GAR Tfase structure to date (1.52 Å). Both diastereomers are potent inhibitors (Ki = 210 nM for 10R, and Ki = 180 nM for 10S) of GAR Tfase and exhibit effective inhibition of human leukemia cell growth (IC₅₀ = 80 and 50 nM, respectively). Their inhibitory activity was surprisingly high, and these lipophilic C10-substituted analogues show distinct advantages over their hydrophilic counterparts, most strikingly in retaining potency in mutant human leukemia cell lines that lack reduced folate carrier protein activity (IC₅₀ = 70 and 60 nM, respectively). Structural characterization reveals a new binding mode for these diastereoisomers, in which the lipophilic thiomethyl groups penetrate deeper into a hydrophobic pocket within the folate-binding site. In silico docking simulations of three other sulfur-containing folate analogues also indicate that this hydrophobic cleft represents a favorable region for binding lipophilic substituents. Overall, these results suggest sulfur and its substitutions play an important role in not only the binding of anti-folates to GAR Tfase but also the selectivity and cellular activity (growth inhibition), thereby presenting new possibilities for the future design of potent and selective anti-folate drugs that target GAR Tfase.
authors.affiliation: Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, United States.
authors.forename: Stephen
authors.initials: S
authors.lastname: Connelly
authors.forename: Jessica K
authors.initials: JK
authors.lastname: DeMartino
authors.forename: Dale L
authors.initials: DL
authors.lastname: Boger
authors.forename: Ian A
authors.initials: IA
authors.lastname: Wilson
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D001059:Apoproteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D019731:Carbon-Nitrogen Ligases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D020128:Inhibitory Concentration 50; D007938:Leukemia / Q000188:drug therapy / Q000201:enzymology; D008958:Models, Molecular; D008968:Molecular Conformation; D062105:Molecular Docking Simulation; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D051027:Phosphoribosylaminoimidazolecarboxamide Formyltransferase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051026:Phosphoribosylglycinamide Formyltransferase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D013237:Stereoisomerism; D013329:Structure-Activity Relationship; D013763:Tetrahydrofolates / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology
chemical_list: C503367:10-methylthio-DDACTHF; D000964:Antimetabolites, Antineoplastic; D001059:Apoproteins; D004791:Enzyme Inhibitors; D009363:Neoplasm Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D013763:Tetrahydrofolates; D051026:Phosphoribosylglycinamide Formyltransferase; D051027:Phosphoribosylaminoimidazolecarboxamide Formyltransferase; D019731:Carbon-Nitrogen Ligases; C494823:GART protein, human