The auditor general, Libero Milone and deputy Ferruccio Panicco, filed the suit in 2022, alleging they were sacked after discovering financial irregularities. Panico has since died.
The court decision was handed down on Monday and published on Wednesday.
Milone told Reuters that he had no comment on the ruling but that he and his lawyers were considering their options.
The panel of judges who heard the case rejected each separate claim for damages for lost salary and injury to their reputations. Milone and Panico's estate were ordered to pay legal costs.
Milone, a former chairman and CEO of Deloitte in Italy, was appointed by Pope Francis in 2015 as part of an effort to clean up Vatican finances and raise accounting procedures to international standards of accountability and transparency.
He was told to resign in 2017 by Cardinal Archbishop Angelo Becciu, who was then the No. 2 in the Vatican's Secretariat of State, its most important department.
Becciu told Reuters in 2017 that Milone "went against all the rules and was spying on the private lives of his superiors and staff, including me".
Milone denied this, saying he just was seeking information he had a right to see as auditor general. Milone has said Becciu blocked his attempts to instil financial transparency. Becciu denied this.
In a separate trial that ended last month Becciu, the most senior Catholic Church official ever to stand trial before a Vatican criminal court, was convicted of embezzlement and fraud and sentenced to five-and-a-half years in jail.
Becciu denied all charges in that case, as did the eight other defendants who were convicted. That case is headed for an appeal expected to begin later this year or in 2025.
($1 = 0.9180 euros)
(Reporting by Philip Pullella; Editing by Bernadette Baum)
By Philip Pullella
