Characterization of an influenza A (H3N2) virus resistant to the cyclopentane neuraminidase inhibitor RWJ-270201

The novel influenza virus neurarninidase (NA) inhibitor, (1S,2S,3R,4R)-3-[( 1S)-(acetylamino)-2-ethylbutyl]-4-[(aminoiminomethyl)amino]-2-hydroxy-cyclo pentanecarboxylic acid (RWJ-270201, BCX-1812), is a potent inhibitor of inf luenza A and B viruses in cell culture and in infected mice. A mouse-adapte d strain of influenza A/Shangdong/09/93 (H3N2) virus was serially passaged in the presence of 1 muM compound. After the fourth passage, breakthrough o f resistant virus occurred. By the tenth passage, a twice plaque purified i solate was obtained which could replicate in 10 muM inhibitor. The 50% effe ctive concentration (EC50) values for RWJ-270201 against wild-type and resi stant viruses, determined by using a cytopathic effect inhibition assay, we re 0.007 and 23 muM, respectively. Cross-resistance to zanamivir and oselta mivir carboxylate was observed. The hemagglutinin (HA) and NA genes of the virus were sequenced to determine the mutation(s) which conferred drug resi stance. No differences were found between the resistant and wild-type virus es in the NA gene. However, a point mutation resulting in a single amino ac id change (Lys189Glu) was found in the resistant viral HA. The wild-type an d resistant viruses were compared for virulence in BALB/c mice. The resista nt virus was approximately tenfold less virulent than the wild-type virus b ased upon virus challenge dose. Mice infected with a lethal dose of the res istant virus could still be effectively treated with RWJ-270201. Thus, the HA mutation may allow for the spread of the virus in cell culture in the pr esence of the NA inhibitor, but not in mice. (C) 2001 Elsevier Science B.V. All rights reserved.