Ex-HSBC boss Michael Geoghegan, former Channel 4 chairman Luke Johnson and the stockbroker Peter Hargreaves are among those backing a Leave vote on 23 June.
Excessive regulation, they argue, poses a threat to London's pre-eminence in the global financial services industry.
The Remain side says EU exit will lead to job losses and reduced investment.
Earlier in the referendum campaign, those wanting to stay in the EU claimed the support of 36 FTSE 100 bosses - while employers group such as the CBI and EEF also back the UK's continued membership.
The debate around the economy has focused on the impact on the UK and its key industries of leaving the EU and what trade deals the UK will be able to negotiate when - as envisaged by Leave campaigners - the UK leaves the EU's single market.
The government has said leaving the EU will be a drag on economic growth and leave households thousands of pounds worse off than they otherwise would have been.
Leave campaigners have said this is scaremongering and now 110 business figures have signed a letter to the Evening Standard, arguing that far from damaging the City of London, exiting the EU would allow it to consolidate its position as the world's largest international financial centre.
Speaking in a "personal capacity", the signatories - including company chief executives, fund managers and economists - say they have a "material interest" in maintaining the City as a vibrant and competitive place to do business and warn the EU is going in the wrong direction and puts this at risk.
"The EU is now shackled to the euro, a project doing damage to the social and economic fabric of member countries, including high youth unemployment, " they write.
"Meanwhile there is scant evidence that the EU will foster or support the kind of innovation which is essential if Europeans are to compete with the rest of the world.
How trade and the UK's economy are affected by membership of the EU.
"Specifically, we worry that the EU's approach to regulation now poses a genuine threat to our financial services industry and to the competitiveness of the City of London."
They conclude: "Assuming good political leadership and an effective regulatory environment, we believe that the City is most likely to strengthen its lead as the world's largest international financial centre, and continue to make a major contribution to the UK economy and employment, outside the EU but with continued access to its capital markets."
Britain Stronger in Europe said Leave supporters had "given up" trying to sound credible on the economy amid the overwhelming evidence that the UK would get a "worse deal" if it left the EU's internal market.
Just hours before Vote Leave released the letter, UKIP leader Nigel Farage - also a supporter of EU exit - warned the campaign group they risked losing the referendum unless the terms of debate were shifted from the economy to immigration.
He challenged them to hit their Remain opponents where they were "weakest" on the issue of "open-door" immigration but questioned the ability of the senior Conservatives to do this given the government's record on failing to reduce migration levels.
More than 100 business figures have thrown their weight behind the campaign to leave the EU, arguing the City would "thrive and grow" outside the EU.