Influenza is an infection of the upper respiratory tract caused by influenza virus. Neuraminidase (NA) plays an essential role in replication and infection of influenza virus. It is considered as a suitable target for antivirus agents development. Several potent NA inhibitors (NAIs) developed by using structure-based rational design have been widely used in clinic for treatment of influenza. However, the emergence of NA inhibitor-resistant virus mutations significantly limit their effectiveness. The molecular basis of virus resistance to NAIs has been a focus of intensive research. This review highlights recent progress in rational design and resistance mechanism of NAIs. We hope this review will be useful to not only researchers who are interested in developing novel NAIs but also clinical pharmacists.