The decision will be a massive blow to the ride-hailing app business, which has faced mounting criticism over issues including its apparent failure to vet drivers.
Its current licence expires on 30 September, although it has 21 days to appeal, during which time it is allowed to continue operating.
TfL said in a statement: "TfL's regulation of London's taxi and private hire trades is designed to ensure passenger safety. Private hire operators must meet rigorous regulations, and demonstrate to TfL that they do so, in order to operate. TfL must also be satisfied that an operator is fit and proper to hold a licence.
"TfL has concluded that Uber London Limited is not fit and proper to hold a private hire operator licence. TfL considers that Uber's approach and conduct demonstrate a lack of corporate responsibility in relation to a number of issues which have potential public safety and security implications."
Tom Elvidge, general manager of Uber in London, said TfL and the Mayor of London Sadiq Khan had "caved in to a small number of people who want to restrict consumer choice".
He continued: If this decision stands, it will put more than 40,000 licensed drivers out of work and deprive Londoners of a convenient and affordable form of transport.
"To defend the livelihoods of all those drivers, and the consumer choice of millions of Londoners who use our app, we intend to immediately challenge this in the courts."
Uber has looked to improve its image through marketing over the last 18 months, appointing Bartle Bogle Hegarty to handle its advertising in March 2016 and Manning Gottlieb OMD to its media.
BBH's work has attempted to counter Uber’s bad publicity with emotionally warm ads. A June spot marked the brand’s UK TV debut with an ad that told the story of Grace and Miles on their first date.
Meanwhile, The London Taxi Drivers’ Association unsurprisingly welcomed TfL's decision, which it said was "long over overdue in the interest of public safety".
Steve McNamara, the LTDA’s general secretary, said: "The mayor has made the right call not to relicense Uber. Since it first came onto our streets Uber has broken the law, exploited its drivers and refused to take responsibility for the safety of passengers.
"We expect Uber will again embark on a spurious legal challenge against the mayor and TfL, and we will urge the court to uphold this decision. This immoral company has no place on London’s streets."
One of the issues TfL has with Uber is the possible use of Greyball, which it described as "software that can be used to block regulatory bodies from gaining full access to the app and prevent officials from undertaking regulatory or law enforcement duties".
Uber asserts that an "independent review" found that the software has "never been used or considered in the UK for the purposes cited by TfL" and that it had informed TfL of this.
However, TfL said Uber’s "approach to explaining the use of Greyball" demonstrated a lack of corporate responsibility.
A spokesman for Addison Lee said: "Today’s decision is a matter for Tfl and Uber. Addison Lee continues to operate as normal."
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