Hanna-Barbera developed Josie and the Pussycats to compete with Filmation's successful The Archie Show, after their own earlier attempt at a teenage-music-band cartoon, Mysteries Five, evolved instead into Scooby-Doo, Where Are You!.

To create the show, Hanna-Barbera and Archie Comics collaborated to adapt the Josie comic book into a music-based property, adding new characters like Alan M. and Valerie while dropping others.

Judy Waithe was originally cast as the voice of Josie but was replaced before the show premiered because CBS disliked her readings of the 'In the Know' interstitial segments; she is still incorrectly credited on some episode prints.

Producers held a talent search interviewing over 500 finalists to find real singers to match the comic book characters, eventually casting Kathleen Dougherty as Josie's singing voice, Cherie Moor (later known as Cheryl Ladd) as Melody's, and Patrice Holloway as Valerie's.

When music producer Danny Janssen assembled the singing trio, he was unaware the original comic characters were all Caucasian; rather than remove Patrice Holloway, Hanna-Barbera changed the comic book character Pepper into a new African-American character named Valerie, making her the first regularly appearing Black female character on a Saturday morning cartoon.

Alexander Cabot III physically resembles Shaggy Rogers from Scooby-Doo, and in the crossover episode 'The Haunted Showboat' from The New Scooby-Doo Movies, Casey Kasem voiced both Alex Cabot and Shaggy Rogers simultaneously.

In the same crossover episode, Don Messick voiced both Sebastian the cat and Scooby-Doo at the same time.

Josie and the Pussycats in Outer Space closely parallels Lost in Space, with Alexandra's disruptive, self-serving role mirroring that of Dr. Zachary Smith, and her pet alien Bleep resembling the Robinsons' pet Debbie the Bloop.