The England international will be out for the match at Allianz Arena along with goalkeeper Roman Burki
The club also confirmed that both players had tested negative for Covid-19.
The two clubs will face off at the Allianz Arena in Munich having both lost their most recent Bundesliga matches after starting the new season off with a win.
The two sides have made a habit of facing one another in the German Super Cup, with Wednesday’s match the seventh meeting between the sides in the last 11 years.
Dortmund have won three of those fixtures, including last year’s, with Sancho and Burki’s absence set to deal them a blow in this season’s encounter.
Sancho has recorded 35 goals and 45 assists across his first 101 outings for Lucien Favre’s side, leading to strong interest this summer from Manchester United.
Dortmund, though, have rebuffed all of the Red Devils’ efforts to sign Sancho and on Tuesday, club captain Marco Reus appeared to rule out any move for the England international this year.
“For us players that was great news. We were all happy that he will be with us for the season,” Reus told Sky Germany.
“Also because he delivers points for the team. Goals, assists – in the end, that translates to a lot of points.
“He is an essential player for us. If he will be as great of a player as [Cristiano] Ronaldo, only time will tell. He needs a little bit of time, many experiences.”
Speaking to Dortmund’s official website, Reus outlined the challenge that his side will face against last season’s treble winners.
“They are very, very good. If you win the Champions League, especially in the way they did, it’s impressive. We are playing against the best team in Europe. It’s a good game to see where we stand,” said Reus.
“I think both sides will put a strong team out. There’s a title at stake, and there has been a lot of prestige on the game for decades. No one wants to leave the pitch as a loser, so we will do everything we can to win tomorrow.”