Viktoria Plzen recorded only their second ever home win in the Champions League to clinch third place in Group G and claim a spot in the Europa League.
Jan Kovarik put the home side ahead in the 62nd minute but Cengiz Under equalised five minutes later with a clinical strike.
Tomas Chory made it 2-1 to Viktoria Plzen in the 72nd minute before Roma substitute Luca Pellegrini was sent off for two bookable offences.
The victory meant Plzen finished third in Group G.
Three-time defending champion Real Madrid, which lost 3-0 to CSKA Moscow, had already won the group and Roma was already assured of finishing second.
Madrid finished with 12 points, Roma nine and Plzen and CSKA seven each.
Plzen advanced to the Europa League courtesy of a better head-to-head record with CSKA, having beaten the Russian side in their second meeting after a draw in their first encounter. The Czech side needed to match CSKA's result to finish third.
Injury-plagued Roma, a semifinalist last season, extended its winless streak in all competitions to five matches. Edin Dzeko, Daniele De Rossi, Lorenzo Pellegrini and Stephan El Shaarawy were each out injured for Roma.
When Roma hosted Plzen in October, Dzeko scored a hat trick in a 5-0 victory.
Roma ended with 10 men when substitute Luca Pellegrini picked up two yellow cards.
