RESUMEN
Protein arginine methyltransferase 5 (PRMT5) is an enzyme that can symmetrically dimethylate arginine residues in histones and nonhistone proteins by using S-adenosyl methionine (SAM) as the methyl donating cofactor. We have designed a library of SAM analogues and discovered potent, cell-active, and selective spiro diamines as inhibitors of the enzymatic function of PRMT5. Crystallographic studies confirmed a very interesting binding mode, involving protein flexibility, where both the cofactor pocket and part of substrate binding site are occupied by these inhibitors.
RESUMEN
We previously reported the discovery of substituted benzimidazole fusion inhibitors with nanomolar activity against respiratory syncytial virus (Andries, K.; et al. Antiviral Res. 2003, 60, 209-219). A lead compound of the series was selected for preclinical evaluation. This drug candidate, JNJ-2408068 (formerly R170591, 1), showed long tissue retention times in several species (rat, dog, and monkey), creating cause for concern. We herein describe the optimization program to develop compounds with improved properties in terms of tissue retention. We have identified the aminoethyl-piperidine moiety as being responsible for the long tissue retention time of 1. We have investigated the replacement or the modification of this group, and we suggest that the pKa of this part of the molecules influences both the antiviral activity and the pharmacokinetic profile. We were able to identify new respiratory syncytial virus inhibitors with shorter half-lives in lung tissue.
Asunto(s)
Antivirales/síntesis química , Bencimidazoles/síntesis química , Piperidinas/síntesis química , Piridinas/síntesis química , Virus Sincitiales Respiratorios/efectos de los fármacos , Proteínas Virales de Fusión/antagonistas & inhibidores , Animales , Antivirales/farmacocinética , Antivirales/farmacología , Bencimidazoles/farmacocinética , Bencimidazoles/farmacología , Células HeLa , Humanos , Riñón/metabolismo , Hígado/metabolismo , Pulmón/metabolismo , Masculino , Piperidinas/farmacocinética , Piperidinas/farmacología , Piridinas/farmacocinética , Piridinas/farmacología , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad , Distribución TisularRESUMEN
A cell-based assay was used to discover compounds inhibiting respiratory syncytial virus (RSV)-induced fusion in HeLa/M cells. A lead compound was identified and subsequent synthesis of >300 analogues led to the identification of JNJ 2408068 (R170591), a low molecular weight (MW 395) benzimidazole derivative with an EC(50) (0.16 nM) against some lab strains almost 100,000 times better than that of ribavirin (15 microM). Antiviral activity was confirmed for subgroup A and B clinical isolates of human RSV and for a bovine RSV isolate. The compound did not inhibit the growth of representative viruses from other Paramyxovirus genera, i.e. HPIV2 and Mumps Virus (genus Rubulavirus), HPIV3 (genus Respirovirus), Measles virus (genus Morbillivirus) and hMPV. Efficacy in cytopathic effect inhibition assays correlated well with efficacy in virus yield reduction assays. A concentration of 10nM reduced RSV production 1000-fold in multi-cycle experiments, irrespective of the multiplicity of infection. Time of addition studies pointed to a dual mode of action: inhibition of virus-cell fusion early in the infection cycle and inhibition of cell-cell fusion at the end of the replication cycle. Two resistant mutants were raised and shown to have single point mutations in the F-gene (S398L and D486N). JNJ 2408068 was also shown to inhibit the release of proinflammatory cytokines IL-6, IL-8 and Rantes from RSV-infected A549 cells.