Wade Wilson, a former special forces operative working as a mercenary, meets an escort named Vanessa at a local bar, and their meeting ends up becoming a relationship. A year later, Wilson proposes to her, but he is then diagnosed with liver, lung, prostate and brain cancer. Despite Vanessa's love, he detests the thought of her watching him waste away, and he leaves her in the middle of the night.
A recruiter from a covert organization approaches Wilson and promises him abilities that will cure his cancer. Wilson reluctantly agrees. He is taken to a remote laboratory where he meets Ajax and Angel Dust, and instantly resents them. Ajax injects a mutation-activating serum into Wilson and subjects him to daily torture to activate it. When Wilson's body fails to respond, Ajax asphyxiates him in an oxygen chamber, causing him to develop an accelerated healing factor that cures him but leaves him disfigured. Wilson finds a way to destroy the lab and escapes his confines. He battles Ajax, but relents when Ajax promises a cure. Ajax then impales him with rebar and leaves him for dead.
Wilson survives and attempts to return to Vanessa, but is afraid of her reaction to his disfigurement after people on the streets appear to be disgusted by his appearance. After consulting his best friend Weasel, Wilson begins the task of tracking down Ajax to get the cure. He becomes a masked vigilante, adopts the alter ego "Deadpool", and resides with an elderly blind woman, Al. Following a string of leads, Deadpool tracks Ajax to a convoy on an expressway. He executes the numerous convoy guards, subdues Ajax, and demands the cure. He is interrupted by Colossus and Negasonic Teenage Warhead. They try to convince him to join the X-Men, and Ajax escapes. They handcuff Deadpool, but he frees himself by severing his own hand, which regenerates overnight.
Ajax and Angel Dust go to Weasel's bar and learn about Vanessa. Weasel warns Deadpool that she is in danger, and they go to the club where she works to warn her, but Deadpool hesitates. Ajax and Angel Dust abduct Vanessa and tell Deadpool to meet them at a decommissioned helicarrier in a scrapyard. Deadpool persuades Colossus and Negasonic to help him rescue Vanessa. They travel to the scrapyard and battle Ajax, Angel Dust and their team of mercenaries. As Colossus and Negasonic fight Angel Dust, Deadpool kills most of the mercenaries and engages Ajax in hand-to-hand combat atop the helicarrier. Negasonic accidentally destroys the equipment stabilizing the helicarrier, tipping it and scattering Ajax and Deadpool. As Colossus carries Negasonic and Angel to safety, Deadpool saves Vanessa and injures Ajax. When Deadpool demands his cure, Ajax laughs and reveals there is none. Deadpool kills Ajax in response. Vanessa is initially angry with Wade for abandoning her, but they get back together after she learns why he did not return sooner.
- Team Griffin/Eeveelution, Peter Griffin, Glenn Quagmire, Cleveland Brown, Joe Swanson, Blythe Baxter, Zoe Trent, Pepper Clark, Penny Ling, Vinnie Terrio, Sunil Nevla, Minka Mark, Russell Ferguson, Buttercream Sunday, Mitzi, Dolphy, Knockout, Rachel, Puffer, Mighty Joe, Daylight, Jenny, Pete, Kevin, Morley, Blitzer, Belle, Dudley Puppy, Kitty Katswell, Joy, Sadness, Anger, Fear, Disgust, Judy Hopps, and Nick Wilde guest star in this film.
- Dawn Bellwether, Diamond Tiara, Silver Spoon, Alter, Samantha and Slate will work for Francis Freeman in this film.
- It's revealed that Hiatt Grey's Engines are old friends with Deadpool.
- Thomas & Friends: The Great Race, My Little Pony: Friendship is Magic season 6, My Little Pony: Equestria Girls - Friendship Games, Littlest Pet Shop season 4, Zootopia, and Deadpool were all released on 2016.
- Family Guy and Deadpool are both made by 20th Century Fox.
- The storyline continues in Thomas and Twilight Sparkle's Adventures of X-Men: Apocalypse.