A police spokesman said the blast was probably caused by gas and that there had been about 15 people in the building, which included an office of the International Air Transport Association (IATA) and an art gallery.
"There is an unknown number of injured and trapped people," said Jirina Fikejzova, Prague emergency services spokeswoman. A Reuters witness at the site said about a dozen people were being treated by emergency services.
The blast blew out windows in neighbouring buildings, including Prague's landmark Cafe Slavia.
A fire department spokeswoman said there have not been reports of any deaths. Neighbouring buildings were being evacuated, the spokeswoman said.