A total of 24 people died on the scene and three more later in hospital, said Alvaro Escobar, a spokesman for the prosecutor's office in Cundinmarca department.
The bus was heading from Bogota to the city of Cali, some 500 kilometers (310 miles) to the southwest, highway police commander Francisco Patino said.
The bus veered off the road and overturned, Colombian TV footage showed.
The vehicle was part of a caravan of four that were traveling to Cali, returning people from a corporate convention in Bogota.
People on some of the other buses said the driver of the one that crashed had complained about the brakes on the first leg of the journey but that the bus company failed to replace the vehicle.
Earlier this month eight people died and 11 were injured when a bus flipped over in Ibague, 200 kilometers (120 miles) west of Bogota, apparently after speeding.