Ferrari's Sebastian Vettel set the pace as Mercedes struggled in second practice at the Mexican Grand Prix.
Vettel was 0.614 seconds quicker than the fastest Mercedes of Valtteri Bottas in fourth, with Hamilton a further 0.349secs adrift in fifth.
Red Bull's Max Verstappen was second, 0.115secs behind Vettel, with Vettel's team-mate of Charles Leclerc third.
Mercedes' pace suggested Hamilton will struggle to tie up the title on Sunday - he needs 14 points more than Bottas.
To secure his sixth title, Hamilton at the very least needs to finish on the podium and hope Bottas' result goes his way.