Lazio ran out 3-0 winners in the Serie A meeting, thanks to goals from Felipe Caicedo, Ciro Immobile and Danilo Cataldi.
The result is a blow for the Giallorossi in the race for the Champions League places - although they remain ahead of their city rivals in the standings.
Eusebio Di Francesco's side went behind early on to Caicedo's finish, although they did have chances to equalise prior to Immobile's strike - the substitute converting from the penalty spot after Federico Fazio had been adjudged to have fouled Joaquin Correa.
Aleksandar Kolarov was sent off in the final minutes after picking up a second yellow card.
Roma are next in action on Wednesday in Portugal, as they face Porto in the second leg of their Champions League last-16 tie. The Giallorossi lead 2-1 after the first leg.
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