Spacey, 59, was charged in December with one count of felony, indecent assault and battery.
The alleged incident is reported to have occured on July 7, 2016, after the former House of Cards actor gave a teenage boy excessive alcohol.
During the pre-trial hearing, Spacey remained silent before his attorney, Alan Jackson, entered his client's 'not guilty' plea.
Formor Boston TV presenter Heather Unruh accused Spacey of sexual assault against her son - who was 18 at the time - in Nantucket's Club Car.
Spacey could face up to five years in prison if convicted of the crime.
The charges were announced last month and Spacey is set to reappear in court on March 4.