Josh Keaton

Peter Parker/Spider-Man (voice)

Lacey Chabert

Gwen Stacy (voice)

James Arnold Taylor

Harry Osborn (voice)

Alan Rachins

Norman Osborn (voice)

Deborah Strang

Aunt May Parker (voice)

Ben Diskin

Eddie Brock (voice)

Dee Bradley Baker

Curt Connors/Lizard (voice)

Robert Englund

Adrian Toomes/Vulture (voice)

Crispin Freeman

Max Dillon/Electro (voice)

Jeff Bennett

Montana/Shocker (voice)

Daran Norris

J. Jonah Jameson (voice)

Joshua LeBar

Flash Thompson (voice)

Peter MacNicol

Otto Octavius/Doctor Octopus (voice)

Alanna Ubach

Liz Allen (voice)

Grey DeLisle

Sally Avril (voice), Betty Brant (voice)

Clancy Brown

Alex O'Hirn/Rhino (voice)