The Darkholders started out as Thulsa Doom's followers. After Doom's death at the hands of Kull, they used the Darkhold to create the first vampire, Varnae. Varnae turned some members into vampires. Other vampires were Nosferatu, Verdelet, Saracen, Maracen, and Boy.
Some of the first vampires, such as Boy, Maracen, Saracen, Verdelet, and presumably Lamia became known as the Ancient. It is unknown whether all or any of those were themselves Darkholders.
Surviving Darkholders managed to make off with the Darkhold before Atlantis sank.
The Modern Age
The Darkholders resurfaced in modern times. Led by DeGuzman, they secured a temporary alliance with Lilith, despite the opposition of other leaders. Lilith herself kept an open perspective on the use of the cult for her own purposes.
Under Lilith's instructions, they attacked Victoria Montesi, who was a reluctant heir to the Montesi responsibility of safeguarding the Darkhold, and seen by Lilith as a threat to it. As the assassins failed to kill Montesi, they were given to Lilith, who had the Lilin Doc reshape them to be faster and stronger. Their next attack on the Darkhold Redeemers was repelled as well.
After Lilith's defeat, the Darkholders sent hitmen to Washington to kill the Darkhold Redeemers
The Red Slayer
They later took interest in Carnage. Barry Gleason orchestrated Cletus Kasady's meeting with them. It was revealed that he was planned as a sacrifice to awaken Chthon, based on the prophecy in the Darkhold that stated "When the Red Slayer spills blood on sacred stone, he who sleeps shall wake and what walked will walk once again." This plan was eventually thwarted.