There is an unused Hammer Bro. left in the game's data. The file's name is "xcmdl_linesman_hbros_line_sbn" suggesting that the Hammer Bro. would've been the linesman at one point during development.
When the Shy Guy wins a trophy, a cutscene is shown where he accidentally stumbles and his mask comes off while retrieving it. Luigi sees the Shy Guy's true appearance, but that player never does.
One of Wario's Power Shots, the Ultra Hand Return, is based on a Nintendo toy invented by Gunpei Yokoi in 1966 called the Ultra Hand. The toy worked to grab objects from a distance, and Wario will use it to hit a ball that is out of reach.
In the intro is a tournament board with every character in the game. In the bottom left corner of the board you can see the team of Toad and Toadette, hinting that they were possibly planned to appear as playable characters at one point during development.