Villeneuve was set to make his Le Mans return in June as part of his pursuit to complete the Triple Crown of Motorsport, but has now been benched for the WEC's blue riband event.
It is unclear if the 52-year-old will return for the remaining three rounds of the season at Monza, Fuji and Bahrain - although he is named on the entry list for Monza.
Villeneuve made a highly-anticipated debut in the World Endurance Championship this year with the Vanwall Hypercar squad, joining Tom Dillmann and Esteban Guerrieri in its Vandervell 680 non-hybrid LMH car.
However, he struggled for competitiveness in the opening leg of the season, with a crash in the Sebring Prologue and another race-ending incident with a Ferrari GTE car at Spa compounding his misery.
He was also involved in another shunt at Portimao in April, although that followed a brake failure in his Vanwall that left him unable to stop the car in time to avoid hitting the barriers.
Villeneuve won the Indy 500 in 1995 before moving to F1, winning the championship in his just second season in 1997 with Williams.
He previously made two appearances at Le Mans with the factory Peugeot team in 2007-08, finishing second in 2008 in the Peugeot 908 he shared with Marc Gene and Nicolas Minassian.
Villeneuve's place at Vanwall will be taken by former JDC-Miller driver Vautier, who raced at the Circuit de la Sarthe last year in an Oreca 07 entered by ARC Bratislava in the LMP2 class.
Vautier had been racing with JDC-Miller in IMSA since 2018 and scored his maiden overall win at Sebring last year together with team-mates Loic Duval and Sebastien Bourdais.
However, he wasn't retained by JDC-Miller following its move from Cadillac to Porsche customer machinery in 2023, with the 33-year-old instead signing with Algarve Pro in the European Le Mans Series.
Vautier wasn't named as part of Algarve Pro's Le Mans line-up in the LMP2 class, but will get to race at La Sarthe for a second season running after joining Vanwall.
Dillmann and Guerrieri will be his team-mates for what will be the first Le Mans outing since 2020 for the squad that previously operated under the ByKolles name.
