Winner of the race in 2018, he would have been heading back to the Festival next month aiming to become the first horse since Kauto Star in 2009 to regain the blue riband event.
The 10-year-old had appeared right back to his best this season, winning at Aintree and only recently in the Denman Chase at Newbury.
He was also prominent in the betting for the Grand National, although his participation in that race had still to be confirmed.
Trainer Colin Tizzard's son and assistant Joe said: "Unfortunately we've had to scratch Native River from the Gold Cup this morning - he's picked up a suspensory tendon injury, which means his season is over."
The Tizzards do still have a valid Gold Cup contender in their yard, with Betfair Chase winner Lostintranslation.
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