Neil Patrick Harris

Peter Parker / Spider-Man (voice)

Lisa Loeb

Mary Jane Watson (voice)

Ian Ziering

Harry Osborn (voice)

Stan Lee

Frank Elson (voice)

Angelle Brooks

Indira 'Indy' Daimonji (voice)

Ed Asner

Officer Barr (voice)

Virginia Madsen

Silver Sable (voice)

Ethan Embry

Max Dillon (voice)

Michael Dorn

Kraven the Hunter (voice)

James Marsters

Sergei (voice)

Clancy Brown

Policeman (voice)

Michael Clarke Duncan

Wilson Fisk / The Kingpin (voice)

Rob Zombie

Dr. Curt Connors / The Lizard (voice)

Gina Gershon

Shikata (voice)

John C. McGinley

Richard Damien (voice)

Jane Lynch

Oscorp Executive (voice)

Tara Strong

Christina (voice)

Devon Sawa

Flash Thompson (voice)

Jeffrey Combs

Professor (voice)

Karen Maruyama

Head Protestor (voice)

Jeremy Piven

Roland Gaines (voice)

Keith David

Agent Mosely (voice)

Kathy Griffin

Roxanne Gaines (voice)

Jennifer Hale

Female News Announcer (voice)

Julie Nathanson

Sally Johnson (voice)

Rino Romano

Muang / Drip Den guy (voice)

Keith Carradine

J. Jonah Jameson (voice)

Eve

Cheyenne Tate / The Talon (voice)

Brendan Connolly

Aleksei (voice)

Jamie Hanes

Special Agents Roberts (voice)

Keith Szarabajka

Wesley (voice)

SuChin Pak

Alison Tomita (voice)

Harold Perrineau

Louis Wyler / Turbo Jet (voice)

Kevin Michael Richardson

Security Guard (voice)

James Horan

Henchman (voice)