For a three-act plot analysis, put on your screenwriter’s hat. Moviemakers know the formula well: at the end of Act One, the main character is drawn in completely to a conflict. During Act Two, she is farthest away from her goals. At the end of Act Three, the story is resolved.
Act I
"Act I" (actually Acts I and II) is about the initial schemes and dreams that ends up destroying everything for the main characters (which will happen in "Act III," which is actually Act IV and Act V). Vittoria and the Duke of Brachiano want love, and Flamineo wants power and status. Flamineo tries to get this by arranging an adulterous meeting between his sister and the Duke and by messing with Camillo's plans to renew the affection of his wife. Then, he amps it up by actually killing Camillo and abetting the Duke's designs. (The Duke also has Isabella killed.) By the end of "Act I", it seems like the bad guys are going to get what they want… This act ends with the murders of Camillo and Isabella—the calamitous betrayals created by those wicked desires.
Act II
"Act II" (actually, Act III and Act IV) is all about the complications—revenge in the works. We see Vittoria's trial, where she manages to seem sort of innocent, even though she isn't—and we witness her sentencing to a "house of convertites" (or "house of penitent whores"). We also see Francisco send his fake love-letter to Vittoria, and its inability to cause the Duke's permanent jealousy. Vittoria and the Duke just escape, eloping to Padua, and Flamineo joins them there. Meanwhile, Francisco and Monticelso (now the Pope) work on revenge, and end up hiring Lodovico as their assassin… Wheels are moving, but they're not grinding out any conclusions. Those will all come in "Act III."
Act III
In "Act III" (actually, Act V), in Padua, we finally see how the villains' desires lead to horrifying reactions. It all comes back in their faces: Francisco and Lodovico get revenge—Brachiano dies a horrible death by poisoning and strangling, and Lodovico and Gasparo stab Flamineo and Vittoria to death (along with the maid, Zanche). Their murders are complementary to the murders and betrayals committed in the beginning. Even Lodovico gets punished—the avenger is punished for killing Vittoria and Flamineo, but poetically for all the bloodshed and crime he's supposed to have committed in the past. "Evil can only create more evil" seems to be the message (which is pretty much what Giovanni says when he arrives to clean up the scene of the crime and arrest Lodovico). The only people who aren't punished are the people who orchestrated the revenge—Monticelso and Francisco. They didn't get their hands dirty in any direct physical way, and escape punishment for taking the law into their own hands (since this was apparently just, given how guilty their enemies really were).