Atletico Madrid, Inter Milan and AC Milane decided to drop out of the Super League on Wednesday, leaving the new competition on the verge of extinction before it even started.
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Only three clubs were still officially involved - Real Madrid, Barcelona and Juventus.
The moves by Atletico, Inter and then AC Milan came a day after the six Premier League clubs gave up on the controversial breakaway competition.
Atletico said the decision was made after their board of directors met on Wednesday.
The Spanish club said it "decided to formally communicate to the Super League and the rest of the founding clubs its decision not to formalise its participation in the project."
Atletico said the "circumstances" that allowed them to join the new league on Monday "no longer existed today."
"For the club, harmony is essential for everyone involved in the (Atletico) family, especially our fans," it said. "The first team squad and its coach showed satisfaction with the club's decision, understanding that sporting merits must prevail over any other criteria."
Inter said the club was committed to delivering the best soccer experience for fans because "innovation and inclusion have been part of our DNA since our foundation."
"Our commitment with all stakeholders to improve the football industry will never change," the Italian club said.
"Inter believe that football, like any sector of activity, must have an interest in constantly improving its competitions in order to continue to excite fans of all ages all over the world, within a framework of financial sustainability."
Seven-time European champions AC Milan issued a statement saying: "We accepted the invitation to participate in the Super League project with the genuine intention to deliver the best possible European competition for football fans around the world and in the best interest of the club and our own fans," Milan said.
"Change is not always easy, but evolution is necessary for progress, and the structures of European football have evolved and changed over the decades.
"However, the voices and the concerns of fans around the world have clearly been expressed about the Super League, and AC Milan must be sensitive to the voice of those who love this wonderful sport."
Australian Associated Press
