After being saved by Scarlet Spider following Arnim Zola's takeover of the S.H.I.E.L.D. Tri-Carrier, Spider-Man must work with him to liberate Manhattan and his fellow students from HYDRA's clutches. Even when they save Agent Venom and Iron Spider, they must also undo transformations of the HYDRA Goblins, avoid Doctor Octopus and Kraven the Hunter, and defeat Arnim Zola.
After learning that Norman Osborn injected himself with an Anti-Goblin serum to prevent further transformations into Green Goblin following the last encounter, Doctor Octopus teams up with Baron Mordo and uses the Siege Perilous to pull the Ultimate Green Goblin from Miles Morales' universe into theirs so that HYDRA can be his master. Now Spider-Man and Miles have to work together with Doctor Strange to keep the demonic Goblin from using the Siege Perilous to destroy reality.
With the Siege Perilous destroyed in the aftermath of the fight with the Ultimate Green Goblin, Miles Morales is forced to stay in Spider-Man's reality until S.H.I.E.L.D. can find a way to get him and his world's Goblin back to their home, and Spider-Man works to keep him safe. After Norman Osborn becomes Iron Patriot, Spider-Man must help him when he is targeted by Doctor Octopus and Vulture.
After Spider-Man and the Web Warriors are told by Vulture that there is a spy among the other heroes, he and the Web Warriors work to find out who it is while Nova and Nick Fury are away on a secret mission. They test out the new training area that was designed by Leo Fitz and Jemma Simmons, only to discover that the robots were sabotaged. Dr. Connors once again becomes Lizard and now sports a contagious bite that turns others into lizards. When everyone ends up infected, it's up to Spider-Man to stop the Lizard and cure everyone before the infection spreads out of ...
As Spider-Man and Iron Spider pursue Vulture, their pursuit brings them to a nearby island where they discovered that Doctor Octopus has captured Sandman and has cloned him from his sand samples in order to find the "Ultimate Sandman" that would join his Sinister Six.
Spider-Man and Agent Venom have been cracking down at defeating possible Sinister Six recruits like Shocker, Grizzly, and Beetle. When Doctor Octopus (now paired up with Dr. Michael Morbius) has made an entirely new symbiotic that's corrosive to the original one, it takes control of Harry Osborn after he and and his team intervenes. The Anti-Venom symbiotic controls Harry into destroying Spider-Man and Agent Venom as the new Anti-Venom. Now Spider-Man and Agent Venom must work with Iron Patriot to free Harry from the Anti-Venom symbiotic.
While answering a distress signal that leads them to an abandoned mall in Queens during a drought, Spider-Man, Kid Arachnid, Iron Spider, and Scarlet Spider find a vacuum-sealed room where they unknowingly release Hydro-Man. When Hydro-Man causes the Spider-Jet to do an emergency landing in a Queens neighborhood where Aunt May is, the Web Warriors must find a way to defeat Hydro-Man as Scarlet Spider works to keep Aunt May out of harm's way.