Massie, who angered Trump by leading a push to release Justice Department files tied to the late sex offender Jeffrey Epstein, was defeated by Ed Gallrein, a former Navy SEAL backed by the president and bolstered by heavy outside spending by pro-Israel groups.
The contest offered the latest evidence of Trump's hold over Republicans.
This followed the ouster in a primary on Saturday of another Trump critic, Senator Bill Cassidy of Louisiana, and losses for dissenting state lawmakers in Indiana primaries on May 5.
Gallrein embraced the role Trump gave him and focused his pitch to voters on his personal history and unwavering loyalty to the president.
"Massie got Trumped. Donald Trump is the sun and the moon and the stars in the Republican Party in Kentucky," said Kentucky-based Republican strategist TJ Litafik.
But rather than signal wider appeal for the president, Massie's defeat underscored Trump's dominance among party activists who shape primary outcomes, even as his sagging approval ratings and high gas prices raise questions about the party's chances with the broader electorate in November's midterm elections.
Massie incurred Trump's ire by successfully leading a push to release Justice Department files tied to Epstein, and his criticism of the Iran war.
The contest between the libertarian-leaning Massie and Gallrein was the most expensive US House primary in history, costing more than $US32 million ($A45 million) in ad spending.
Other primaries are unfolding on Tuesday across Alabama, Georgia, Idaho, Kentucky, Oregon and Pennsylvania, helping to shape the battlefield for November's elections, when Democrats aim to take control of the House and potentially the Senate despite Republican gains in a national redistricting fight.
with AP
