Number eight Thomas Waldrom, flanker Tom Johnson and wing Charlie Sharples have all been dropped from the 23-man match-day squad after starting in England's 20-14 defeat to Australia.
Loosehead prop Joe Marler has been ruled out with a knee ligament injury, opening the way for Alex Corbisiero's return after overcoming a knee injury of his own.
James Haskell and Ben Morgan have both been included in the squad for the first time this autumn, replacing Waldrom and Johnson.
London Irish centre Jonathan Joseph has taken Sharples' place in the match-day squad after shaking an ankle injury.
Lancaster will confirm his starting XV on Thursday.
Morgan and Haskell are vying for Waldrom's number eight jersey with Tom Wood seemingly certain to start at blindside flanker.
Corbisiero is still England's first-choice loosehead, despite only starting two matches this season due to injury, but he is fit again and expected to leapfrog Mako Vunipola into the number one jersey.
England's preferred tactic on the summer tour was to play with two specialist full-backs, which could see Mike Brown starting on the wing in place of Sharples.
Alternatively, England could move Manu Tuilagi out wide and start with Joseph at outside centre.
"We have made some changes this week to freshen up the squad before the challenge of South Africa," Lancaster said.
"Unfortunately that means one or two players miss out who were in the match day squad against Australia.
"But they have all added value with their contributions in training and the games and I am sure they will be looking to play well for their clubs and force their way back in.
"The changes however give opportunities to those who have been pushing hard and we feel will add energy and balance to the team ahead of the significant South African challenge."
Mouritz Botha has been added to the squad as provisional injury cover for Geoff Parling, who is nursing a knee problem but the England medics expect him to be fit.
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