The City Hall-funded initiative, due to launch in September, will target people across Europe, the United States and Asia in an effort to counter what the mayor has described as a coordinated campaign of falsehoods damaging London's international reputation.
The campaign will showcase the capital's "rich heritage, world-class experiences, culture, creativity and role as a centre for innovation and trade".
Khan said London had become the subject of a growing wave of misleading content online.
He said: "We are facing a relentless and unprecedented attack of lies and hatred from those wanting to damage our capital's standing and our hugely important tourism industry.
"Disinformation about London has become a truly global scourge.
"It's a money-making industry pushing lies about our capital and preying on people's fears around the world, so we must fight back on a global scale.
"That's why we're creating this major new campaign to challenge these false narratives and demonstrate once again why there's no better place in the world than London."
A report published earlier this year by the Greater London Authority found a sharp rise in online posts portraying London as dangerous or in decline.
According to the study, social media activity promoting such narratives increased by between 150 and 200 per cent between March 2024 and March 2026, while migration-related posts referencing London surged by more than 350 per cent.
The report also found that, during some months, more than 15,000 posts on X written in Japanese falsely claimed the capital was lawless and under Islamic governance.
Earlier this year, BBC News highlighted a series of AI-generated videos falsely depicting a taxpayer-funded water park in Croydon as part of a wider trend of fabricated content about the city.
Critics have questioned the mayor's approach.
Conservative politician Susan Hall said Khan "should spend more time trying to fix things as opposed to saying 'la la la la la', I'm not listening, everything's fine'".
Meanwhile, Reform UK's London mayoral candidate Laila Cunningham argued: "If the mayor of London wants to encourage more tourism, he should deal with the crime he has allowed to spiral out of control instead of suggesting that victims of crime are liars."
