The Spanish club revealed that goalkeeper Courtois has an injury to his left adductor muscle, while left-back Marcelo is dealing with a left hamstring issue.
Madrid - which hosts Eibar on Saturday (AEDT) before the trip to the Etihad Stadium next Wednesday - offered no indication as to how long the duo will be out.
Both played in Monday's (AEDT) 2-1 LaLiga defeat away to Real Betis, though Marcelo was replaced by Ferland Mendy before the hour mark.
Zinedine Zidane's side have won just once in their last five outings in all competitions.
They took the lead against last-16 opponent City at the Santiago Bernabeu on 27 February, Vinicius Junior opening the scoring after 60 minutes.
However, Gabriel Jesus equalised before a Kevin De Bruyne penalty sealed a crucial away victory for Pep Guardiola's side.