"Andre is reluctantly unable to play," tournament director Bruce Flory said in a statement.
"He had every intention of playing, but just didn't feel physically right in his practice sessions here.
"We hate to lose any player, especially one who was to make his final appearance here, but with the stellar field we have."
The 36-year-old Agassi an eight-time grand slam winner with 60 career titles, had been hoping to end his career in a blaze of glory over the summer. He plans to wrap up two decades of professional tennis at the U.S. Open starting August 28.
But after a training session in the Cincinnati heat, the veteran said he would not be able to play.
Agassi's summer farewell tour has been frustrating, with a 2-2 record since a third-round defeat at Wimbledon by Spain's Rafael Nadal.
