Emergency crews were called to the northbound lanes of Highbury Avenue, just past Commissioners Road, at around 11:14 p.m. to reports of a single-vehicle rollover.
Upon arrival, officials located a Chrysler Sebring with two adult female occupants.
Police say the driver was pronounced dead at the scene, while the passenger was transported to hospital with unknown injuries, where she remains.
The names of the victims have not been released, but police say they are not from London.
Officers from the Forensic Identification Section and members of the Traffic Management Unit were called to the scene to investigate and determine the cause of the collision.
Highbury Avenue was shut down between Commissioners Road and Hamilton Road. Southbound lanes have since re-opened, while northbound lanes are expected to re-open around 7:30 a.m.
The London Police Service is asking anyone who may have witnessed the collision to call 519-661-5670.