title: Symmetrical 1-pyrrolidineacetamide showing anti-HIV activity through a new binding site on HIV-1 integrase.
abstract: OBJECTIVE
  To characterize the functional and pharmacological features of a symmetrical 1-pyrrolidineacetamide, N,N'-(methylene-di-4,1-phenylene) bis-1-pyrrolidineacetamide, as a new anti-HIV compound which could competitively inhibit HIV-1 integrase (IN) binding to viral DNA.
  
  
  METHODS
  A surface plasma resonance (SPR)-based competitive assay was employed to determine the compound's inhibitory activity, and the 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazolium bromide cell assay was used to qualify the antiviral activity. The potential binding sites were predicted by molecular modeling and determined by site-directed mutagenesis and a SPR binding assay.
  
  
  RESULTS
  1-pyrrolidineacetamide, N,N'-(methylene-di-4,1-phenylene) bis-1-pyrrolidineacetamide could competitively inhibit IN binding to viral DNA with a 50% inhibitory concentration (IC(50)) value of 7.29+/-0.68 micromol/L as investigated by SPR-based investigation. Another antiretroviral activity assay showed that this compound exhibited inhibition against HIV-1(IIIB) replication with a 50% effective concentration (EC(50)) value of 40.54 micromol/L in C8166 cells, and cytotoxicity with a cytotoxic concentration value of 173.84 micromol/L in mock-infected C8166 cells. Molecular docking predicted 3 potential residues as 1-pyrrolidineacetamide, N,N'-(methylene-di-4,1-phenylene)bis-1- pyrrolidineacetamide binding sites. The importance of 3 key amino acid residues (Lys103, Lys173, and Thr174) involved in the binding was further identified by site-directed mutagenesis and a SPR binding assay.
  
  
  CONCLUSIONS
  This present work identified a new anti-HIV compound through a new IN-binding site which is expected to supply new potential drug-binding site information for HIV-1 integrase inhibitor discovery and development.
authors.affiliation: Drug Discovery and Design Center, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 201203, China.
authors.forename: Li
authors.initials: L
authors.lastname: Du
authors.forename: Ya-xue
authors.initials: YX
authors.lastname: Zhao
authors.forename: Liu-meng
authors.initials: LM
authors.lastname: Yang
authors.forename: Yong-tang
authors.initials: YT
authors.lastname: Zheng
authors.forename: Yun
authors.initials: Y
authors.lastname: Tang
authors.forename: Xu
authors.initials: X
authors.lastname: Shen
authors.forename: Hua-liang
authors.initials: HL
authors.lastname: Jiang
mesh_terms: D000081:Acetamides / Q000138:chemical synthesis* / Q000494:pharmacology*; D019380:Anti-HIV Agents / Q000138:chemical synthesis* / Q000494:pharmacology*; D001665:Binding Sites; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics; D019427:HIV Integrase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019428:HIV Integrase Inhibitors / Q000138:chemical synthesis* / Q000494:pharmacology*; D015497:HIV-1 / Q000187:drug effects; D006801:Humans; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010957:Plasmids / Q000235:genetics; D011759:Pyrrolidines / Q000138:chemical synthesis* / Q000494:pharmacology*; D013329:Structure-Activity Relationship
chemical_list: C540111:1-pyrrolidineacetamide; D000081:Acetamides; D019380:Anti-HIV Agents; D004279:DNA, Viral; D019428:HIV Integrase Inhibitors; D011759:Pyrrolidines; D019427:HIV Integrase