The film picks up several years after the events of Underworld: Blood Wars. Selene (Kate Beckinsale), now a legendary figure among both Lycans and Vampires, has withdrawn from the ongoing conflict, living in seclusion with her daughter, Eve. However, the fragile peace between the two species is shattered when a new, powerful vampire coven emerges, led by the enigmatic and ruthless Viktorine, a vampire elder long thought to be dead.

The story begins with Viktorine making a dramatic entrance by attacking a secret council meeting of vampire elders, slaughtering them to assert her dominance. Her goal is to unify all vampire factions under her rule and wage war on both the Lycans and humans, who she believes are inferior and a threat to vampire supremacy. Her most dangerous weapon is a serum that enhances vampire abilities, making them nearly invincible.

Selene is drawn back into the conflict when her old ally, David, finds her and warns her of Viktorine’s rise to power. He reveals that Viktorine has already begun a campaign to hunt down Selene and Eve, fearing their bloodline’s unique powers could challenge her reign. Selene, torn between protecting her daughter and stopping Viktorine, reluctantly agrees to help.