It happened in the eastbound lanes of the freeway near Lincoln Alexander Parkway.
Ontario Provincial Police said a passenger in the vehicle was pronounced dead and the driver is in critical condition.
The OPP is investigating the fatal collision.
An image shared from the scene by the OPP shows a small sedan hit the guide rail.  It is not yet clear what caused the collision.
The eastbound lanes of Highway 403 are closed at Lincoln Alexander Parkway as police investigate.