The greyhound track first opened in the city in 1936
A campaign has begun to keep Portsmouth Greyhound Stadium open until the end of the season in September.
The land in Tipner is earmarked for redevelopment as part of plan to build 1,600 homes and a new M275 junction.
The owners of the site, SEEDA and Tipner Regeneration Land, said the lease will not be renewed and the stadium will have to close by 1 April.
Several charities will lose out from race nights and a campaign is under way to save the track.
The greyhound track opened in 1936.
Cosham Rotary Club said it had a race night booked in July and would expect to make £1,000 to £1,500 from the event.
David Long, from the charity organisation, said that equated to about 15% of the group's annual fundraising.
Track manager Eric Graham said many charities would lose out if the track closed
Sue Cresdee, bookings clerk at the stadium, said: "It is devastating news.
"It is not just the 47 staff affected but it has a knock-on effect to the charities that use the track and businesses which rely on racing in Portsmouth.
"We knew the lease was going to end but we are just asking to go until the end of the season in September so we can fulfil our commitments."
The Tipner redevelopment scheme will include new homes - 30% affordable housing - and it could create up to 1,500 jobs, the city council has said.
In a statement, SEEDA said: "[It is] a much-needed regeneration scheme for Portsmouth that will create an important gateway to the city and deliver housing, jobs and a park and ride facility.
"This was acquired from the Greyhound Racing Association (GRA) two years ago and was leased back to GRA for a fixed period.
"The lease expires on 1 April 2010, after which the site will be cleared and secured."