Villeneuve was set to make his Le Mans return in June, but the Canadian has now been benched for the WEC’s blue riband event.
It is unclear if the 52-year-old winner of 11 grands prix will return for the remaining three rounds of the season at Monza, Fuji and Bahrain.
Villeneuve made a highly-anticipated debut in the World Endurance Championship this year with the Vanwall Hypercar squad, joining Tom Dillmann and Esteban Guerreri 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 Francesco Castellacci's 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.
The decision means that Villeneuve will not have the chance to complete the Triple Crown which consists of the Indianapolis 500, Le Mans 24 Hours and either the Monaco Grand Prix or the F1 world title, depending on personal views.
Villeneuve won the Indy 500 in 1995 before moving to F1 where he took secured the championship title at the second attempt with Williams. He continued in grand prix racing until 2006, making 163 starts in total.
He previously made two appearances at Le Mans with the factory Peugeot team in 2007-08, finishing second in 2008 in the Peugeot 908 LMP1 he shared with Marc Gene and Nicolas Minassian.
Vautier, the 2021 Sebring 12 Hours winner with JDC-Miller Motorsports, has been racing for LMP2 squad Algarve Pro Racing in the European Le Mans Series this season.
The Frenchman raced in IMSA for JDC-Miller from 2018 until the conclusion of its relationship with Cadillac at the end of last season, and was not renewed for the team's switch to a customer Porsche 963 that made its debut in the most recent round at Laguna Seca.
The squad that previously operated under the ByKolles name is making its first visit to Le Mans since 2020, having been absent from the first two seasons of the Hypercar regulations that replaced LMP1.