Hot favourite Teofilo has been ruled out of Saturday's 2000 Guineas at Newmarket after suffering a setback.
Teofilo (r) beat Holy Roman Emperor in the Dewhurst Stakes
Last season's champion two-year-old, trained in Ireland by Jim Bolger, was found to be sore at the back of a knee.
The Galileo colt, who is named after former Cuban boxer Teofilo Stevenson, won all five starts in 2006, including the Dewhurst Stakes at Newmarket.
He was 2-1 favourite to begin his pursuit of Triple Crown glory with victory in the season's first Classic.
Bookmakers say his withdrawal has cost backers an estimated £2.5 million in lost bets.
One Irish punter stood to land 50,000 euros if he had won the Guineas after placing a 2,000 euro (£1,365) bet at 25-1 before he won the Futurity Stakes in August 2006.
Bolger said the horse had a gallop at 0630 BST on Thursday, and moved with his usual fluency, but an old knee problem flared up later.
"My staff and I have made every effort to have Teofilo fit and well for the first Classic of 2007," he said.
"But we must now accept that a cautious approach is the correct way to go as more serious damage could be done if subjected to a race at this time.
The County Carlow trainer had hoped Teofilo might become the first horse since the great Nijinsky in 1970 to land the treble of 2,000 Guineas, Epsom Derby and St Leger.
He named his star colt after the triple Olympic heavyweight champion as he felt the horse had similar fighting qualities.
His absence marks the latest blow for ante-post punters ahead of this year's 2000 Guineas.
2000 GUINEAS BETTING
4-1 US Ranger
13-2 Strategic Prince
7-1 Major Cadeaux
10-1 Diamond Tycoon
(Odds: Stan James 03/05/07)
The three-year-old had drifted alarmingly in the betting 10 days ago, before Bolger said he had recovered from the knee problem.
Before that, Guineas second favourite Holy Roman Emperor was retired to stud so he could replace the 2006 winner George Washington, who had fertility problems.
George Washington, trained by Aidan O'Brien at his Ballydoyle yard in Tipperary, will return to racing and is set to run at Royal Ascot in June.
Tony Kenny, spokesman for bookmakers William Hill, said Teofilo's withdrawal was "a massive body blow" for punters.
"We need superstars in the game to keep the interest in the sport, and fingers crossed we'll see Teofilo come back to full fitness for this year's Derby," said Kenny.
Newmarket managing director Lisa Hancock acknowledged the favourite's absence was a blow for the meeting.
"His problems have well publicised, but it's terribly disappointing the horse isn't there," she said.
Meanwhile, Bolger will seek consolation on Sunday in the 1000 Guineas for fillies with favourite Finsceal Beo (Irish for 'living legend').
He said an update on Teofilo's injury would be issued within a fortnight.