Juventus have been docked 10 points following a new hearing into the club's past transfer dealings.
Juve were initially handed a 15-point penalty in January but Italy's highest sporting court overturned that decision in April and ordered the case to be re-examined.
The new ruling was announced on Monday, before the team's game away to Empoli.
The penalty drops Juve to seventh place in Serie A, outside the European qualification spots.
Napoli have already secured the Serie A title and, before Monday's ruling, were 17 points ahead of second-placed Juve.
In April, a tribunal at Italy's Olympic Committee (CONI) revoked Juve's 15-point penalty from the Italian Football Federation (FIGC) appeals court.
It asked the appeals court to revise punishments to the club and some key figures, including former vice-chairman Pavel Nedved.
Long bans given to former chairman Andrea Agnelli, ex-chief executive Maurizio Arrivabene and sporting directors Federico Cherubini and Fabio Paratici have now been upheld.
A Juventus statement said that the club "takes note" of the decision and "reserves the right to read the reasons to evaluate a possible appeal" to CONI.
It added: "What was established by the fifth instance of judgment in this matter, which began more than a year ago, arouses great bitterness in the club and in its millions of supporters who, in the absence of clear rules, find themselves extremely penalised with the application of sanctions that seem to take into account the principle of proportionality.
"While not ignoring the need for urgency, which Juventus has never shied away from during the proceedings, it is emphasised that these are facts that still have to be evaluated by a judge."
If Juve beat Empoli, they will be two points behind AC Milan, who occupy the final Champions League spot, with two games to play.
The club is also involved in criminal proceedings over its accounting practices, with 12 current and former officials, including Agnelli, potentially facing trial.
According to Italian news agencies, a high court hearing on 26 October will decide whether any eventual trial will be held in Turin, where the investigation has taken place, or in either Milan or Rome.
Why have Juventus been docked points?
The Old Lady were handed a points deduction after an investigation into the club's past transfer dealings spanning two years from 2019 to 2021 by the FIGC.
Juventus were accused of fixing their balance sheets by artificial gains of around 60m euros from club transfers, charges they were found guilty of by the FIGC's appeals court in January.
The club, however, denied any wrongdoing and took their case to a tribunal at the Olympic Committee, Italy's highest sporting court.
The tribunal did not rule on the merits of the case but instead examined the legal legitimacy of the punishments handed out by the FIGC to the club and directors.
Juventus were among several Serie A clubs acquitted of suspect transfer activity by an FIGC appeals court trial last year but the investigation into the club was reopened due to new evidence from a separate criminal probe into their finances.
The initial 15-point sanction was tougher than the nine-point deduction prosecutors had requested, and the club hoped no points deduction would be applied once the case had been re-examined.
Who are the individuals involved?
Agnelli and the rest of the club's board, including former midfielder Nedved, stepped down in November while the investigation was taking place.
At the time, a statement said the resignations were "considered to be in the best social interest to recommend that Juventus equip itself with a new board of directors to address these issues".
The chairman had presided over the club for 13 years, during which time Juventus won nine successive Serie A titles and reached two Champions League finals.
But last year they made a £220m loss - a record for an Italian club.
Agnelli and Arrivabene were banned from Italian football for two years, with Nedved receiving an eight-month suspension.
Cherubini and Paratici were suspended domestically for 16 months and two and a half years respectively.
Paratici left the club to become managing director at Tottenham before January's ruling but his suspension was extended worldwide by Fifa in March.
Juventus are also facing an investigation from Uefa, European football's governing body, over potential breaches of its club licensing and financial fair play regulations, which was announced in March.
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