An ambulance carrying Ashya pulled out of a hospital in Malaga around 0615 GMT and headed to the airport, from where he is expected to fly to the capital of the Czech Republic.
The doe-eyed boy has been in the middle of a week-long legal saga that began when his desperate parents took him out of a British hospital against medical advice, triggering a cross-border manhunt that saw them briefly jailed in Spain.
The case dominated British news, with public opinion shifting from outrage to sympathy as it became clear parents Brett King, 51, and his wife Neghemeh King, 45, had taken their son abroad to avoid treatment they believed would turn him into a "vegetable."
The parents spent four days in a Spanish jail and although they were released after British prosecutors dropped their case, they could not move him from a hospital in Spain until his status as a ward of court, imposed after they were imprisoned, was lifted.
Ashya, who has a brain tumour, is now headed to a Czech centre specializing in proton beam therapy, the treatment his parents sought for him but that was unavailable to them in Britain.
Proton beam therapy, which is more precise than traditional radiotherapy, allows doctors to deliver higher doses of energy to a tumour while better sparing surrounding healthy tissue.
According to the PTC, the procedure costs about 1.8 million kroner (65,000 euros, $84,000) in the Czech Republic, compared with 108,000 euros in the United States.
The Kings have said they will sell an apartment in Malaga to fund Ashya's treatment.
Ashya recently underwent brain tumour surgery in Southampton, in southern England, but his parents took him from the hospital there after disagreeing with his treatment.
The lead paediatrician at Southampton hospital did not believe that the alternative therapy would help rid the boy of his medallublastoma cancer.
The Kings' legal troubles prompted an outpouring of public support in Britain, where tens of thousands of people signed a petition calling for the boy to be reunited with his parents.
The case even gained the attention of Prime Minister David Cameron, who upon learning that the case against the Kings had been dropped, tweeted: "It's important this little boy gets treatment and the love of his family."