AFAIK not exactly, under 64bit mode you can run 16bit protected mode but not 16bit real mode code. While this would be an issue for Win1.x/Win2/x compatibility, the majority of 16bit Windows applications however are made for Win3.x which tend to be made for 16bit protected mode.

This is basically why you can run 16bit Windows programs/games under Wine on 64bit Linux (though you may need to enable some flag via /proc or /sys i don't remember the name of).