Trump was referencing De Niro's recent attack on him at the Tony Awards, where the actor launched an expletive at the president.
"I'm gonna say one thing. F**k Trump," De Niro said.
His statement received a standing ovation from the crowd.
De Niro then introduced singer Bruce Springsteen.
"Bruce, you can rock the house like nobody else and even more importantly in these perilous times, you rock the vote, always fighting for, in your own words, truth, transparency and integrity in government. Boy, do we need that now."
Not long after, De Niro also issued an apology to Canadians for the "idiotic behaviour of my president".
De Niro said Trump's remarks about Canada were a "disgrace" and apologised to Prime Minister Justin Trudeau and others who attended the Group of Seven summit of leaders in Canada.
Trump called Trudeau "dishonest" and "weak" following the summit on Saturday. Trump advisers also ripped Trudeau, branding him a back-stabber.
De Niro made his comments about Trump's remarks about Trudeau at a launch ceremony for a new restaurant and hotel complex in Toronto.
Trump's tweets seemed to reference De Niro's performance in the 1980 film Raging Bull, in which he portrayed Jake LaMotta, an Italian American middleweight boxer.
De Niro won a Best Actor Academy Award for the role.
- With AP
