Disclaimer: Everything that follows are my own personal thoughts and should in know way be regarded as the truth, the right of freedom of speech and my comic fandom are the reasons for the points i am raising that is all.
Since the marketers have been beating us down with a stick by reminding us that this Spiderman is darker and gritter, my first intuition for the next villain is...
And my reasoning is this, because this film series will be most mirror to the tone a pacing of Nolan's Batman (or at-least that's the feeling i'm getting) they should follow the pattern. Starting with the Lizard presents a great opportunity to dig deep into some of Peter's personal troubles. The first time when the Lizard was introduce peter didn't really want to fight him because of the man that was inside and they did a great job with that in Spectacular Spider-man television show (which they should of gotten back on the air instead of Ultimate Spider man). Now for a sequel you could do the outer exposure of Spider-mans impact on the world much like The Dark Knight and then you could also go the Iron Man 2 route with the fact that there are more and more people trying to get their hands on his kind of technology. They could introduce Brock as Peter's best friend from their childhood who returns after a significant and suspicious absence.
The Film's Premises could be: 2 years after the events of the first film (and year after the game, if they count that in the mythos). Oscorp has been working on a secret project based on some of the recently found documents of Richard Parker and Curt Connors documents for something that will make Spider-man seem obsolete. Meanwhile Peter's childhood best friend returns from a suspicious 24 12 year absence. Returning with an unknown agenda and getting a job as a intern at Oscorp Eddie becomes attach and hellbent on finding out the secret project being done at Oscorp while simultaneous slowly deteriorating they already strained relationship between Gwen and Peter as a new love enters his life Liz Allen. Peter must face the ultimate outcome that occurs when Brock gets his curious hands on the mysterious project.
The other Logical choice is none other than...
Now at first glance he seems to much of a colorful and campy villain, but giving the right motive Mysterio could be a formidable and potential mind blowing adversary. One of the Great things about Mysterio in the Ultimate Spider-man was he was clever enough to get away. He could be the villain that makes Spider-man question the motives of regular villains. Mysterio could play the joker angel of a villain that doesn't have a clear motive, thus it being difficult for Spider-man to catch him and therefore having him doubt if he can ever bring himself to become a true hero, exposing Spider-man to the fact that not all evil is black and white.
Premises: In the events following the first film new villains have risen to take down Spider-man but no have proven themselves but given Spider-man nothing more than a headache, making him seen invincible and becoming a true hero, but with the rise of his ego comes the terrifying threat of the bugle who taint Spider-man's image as a little boy in costume with no clear idea as to what he is doing. Taking advantage of the rising doubt in Spider-man is the villainous Mysterio and villain unlike anything Spider-man has faced, a villain that has no clear motives. Digger deeper into his past Spider-man has to face the deadly link between Mysterio Oscorp and his family legacy... wondering if he is a hero the world needs.
and the only other villains I can think of are Electro and Vulture. But giving the brief preview of the person talking to Curt Connors in the prison cell i can safely assume that they are going with the Vulture.