La Liga said the decision had been made "due to the uncertainty that has arisen in Spain over the past few weeks".
The plan had come in for heavy criticism, with claims it would affect the competition's integrity. Villarreal would have been the designated home team, but it was much more likely to have felt like an extra home game for Barca, given their global fanbase.
Players protested against the move in the current round of La Liga matches by refusing to move for the first few seconds of each game.
"La Liga deeply regrets that this project, which represented a historic and unparalleled opportunity for the international expansion of Spanish soccer, will not be able to move forward," a statement read on Tuesday night.
"Holding an official match outside our borders would have been a decisive step in the global growth of the competition, strengthening the international presence of clubs, the positioning of players, and the visibility of Spanish soccer in a strategic market such as the United States.
"The project fully complied with all federative regulations and did not affect the integrity of the competition, as confirmed by the competent institutions responsible for ensuring compliance, which opposed it for other reasons.
"In an increasingly competitive global landscape, where leagues such as the Premier League or competitions like the UEFA Champions League continue to expand their reach and ability to generate revenue, initiatives like this are essential to ensure the sustainability and growth of Spanish soccer.
"Renouncing such opportunities hinders the generation of new income, limits clubs' capacity to invest and compete and reduces the international projection of the entire Spanish soccer ecosystem."
The announcement comes after earlier this month Como backed the move to play their Serie A game against AC Milan in Australia next February, saying that "sometimes sacrifice is essential", pointing to the financial gulf that has opened up between Serie A and the Premier League.
Thibaut Courtois had earlier in the day pulled no punches in his assessment of the Miami match.
"It adulterates the competition, totally," the Real Madrid goalkeeper said.
"They (La Liga) put it in because it's in their interest to do that and it adulterates the competition, plus it doesn't comply with the players' agreement.
"We have to play at home and away. It's not the same to play a team at home and away."
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