In response to fans of Antonblast and Pizza Tower attempting to start an arbitrary rivalry between the two games, Anton creator Tony Grayson tweeted "Why not both?" with an illustration of Anton and Peppino kissing.

The title card for the "Gnome Forest" level has become the subject of the "Lario" meme, which features an edited version of the card with the word "Lario" above the characters' heads as a mashup of "Sneaky Snitch" by Kevin MacLeod and "Superstar" by Young Thug. This meme would go onto be referenced within Pizza Tower itself, with Gnome Forest's secret music being renamed "Everybody Wants To Be A Secret" after the meme became popular.

If the player idles on the main menu for about 40 seconds without inputting anything, Peppino will eventually jumpscare them, causing the game to close itself.

Gnome Forest was originally supposed to have the player switch between Peppino and Gustavo & Brick constantly, with them being in a race to deliver more pizzas. This was scrapped in favour of a stage that almost exclusively features Gustavo & Brick so that players could more easily get used to their controls.

In the Slum area of the hub world, there is a peaceful green cheeseslime named Snotty. If one is to kill Snotty, a permanent tombstone will be placed in the hub world, but if he is kept alive, he will be added to the Crumbling Tower of Pizza stage as a rescue, and the save file with which the game was completed will get a stamp saying "Snotty Approved". Saving Snotty does not contribute to the game's completion rating, and if one kills Snotty after the save file is Snotty Approved, the stamp will not be taken away.

The Noise was designed to be an obvious parody of the 1980s Domino's Pizza claymation anti-mascot "The Noid". The game's developer McPig has claimed that if The Noise wasn't based on The Noid, the boss would've had a parody of Vocaloid Hatsune Miku instead, in reference to Domino's App feat. Hatsune Miku, an app that became a niche YouTube Poop meme in the early 2010s.

Before release, fans claimed to hear the phrase "Door stuck" in the music track "Peppino's Sauce Machine", hailing from a Counter-Strike video released in 2007. This would lead to a semi-official remix from composer Mr. Sauceman titled "Peppino's Door Problem", which mixes the "DOOR STUCK!" audio from the video into "Peppino's Sauce Problem".

During development of Pizza Tower, multiple demo builds were released - among these was a build known as the "Peter Griffin Experience", built off of the 2018 "early test build". This demo replaced every single sprite of Peppino with a highly compressed edit of a stock image of Peter Griffin from Family Guy made to resemble Peppino, and replaced one of the game's musical tracks with a fan-made Family Guy remix.

After this build, the Peter Griffin "arms resting" pose would appear in some builds as a taunt, albeit as an actual sprite and not an edit, and a video would be posted by developer McPig showcasing the taunt, accompanied by the first note of the Family Guy theme song, under the name "family", likely referencing a a meme video that plays the first note and ends. This taunt was removed for unknown reasons in the final game.

During development of the game, Peppino was stated to be a war veteran who suffers from PTSD. Following the release of the game, this was retconned, with game creator McPig stating that Peppino probably merely imagined his participation in conflict.

Two background characters featured in the Pizzascare level, a pink-clothed Peppino and a dark-skinned blue-clothed heterochromatic Noise, are fan characters known as Peddito and the Doise created by a fan named Robbie1scool. The Doise would be included as The Noise's exclusive boss in the Noise Update.

One of the Pizza Grannies on the hub world claims that the player can type "/pizza" to order a pizza from Peppino's Pizzeria as an "Instant Pizza Gaming Promotion". This is a reference to a promotion done for EverQuest II, where the player could type "/pizza" into the game chat to open up the Pizza Hut website.

The game was originally planned to feature a "Heat Meter" that kept track of how many enemies the player was able to defeat in a row. More consecutive kills would build up the player's "heat," while getting hit and spending too much time without attacking enemies would lose heat. Higher heat levels would additionally make enemies more aggressive, increase the amount of points lost from taking damage, and grant Peppino new attacks. The development team compared this mechanic to a similar one found in God Hand.

The Heat Meter was fully implemented in a demo released to Patreon backers in April of 2021, but was simplified in later builds before ultimately being removed in October of 2022 due to mixed audience responses; several animations associated with it were also reconfigured for other purposes. Despite this, graphics for the Heat Meter and code for the more aggressive enemy behavior are still present in the final game. The Pizza Slug, Shrimp Thug, and Tribe Cheese enemies would also have their high-aggression modes enabled by default.

When one defeats Spoiler:The Doise, The Noise's exclusive boss fight, he is Spoiler:killed by what appears to be a fake version of Peddito. If one rematches the Doise on the hub world, The Noise will simply find his bloody corpse and look on in confusion. Fitting with this joke, in the final boss, the Doise does not have a phase in the boss rush, instead just having his corpse thrown on the ground by Pizzaface, giving the player some time to breathe, then disappearing.

Two of Peppino's sprite animations are references to non-verbal Italian gestures:
• One of his idle animations has him bite his hands, a threatening (and sometimes sexually loaded) gesture known as "si t’ancagliu" or "when I catch you, I'll kill you".
• The pose he does on the P-rank screen is "l'ombrello" or "the umbrella", an Italian equivalent to the middle finger.

Pizza Tower shifted from a puzzle/exploration-based game to a fast-paced game during development in response to videos of people speedrunning the SAGE 2019 demo.

Before the Pizza Priest and Stupid Rat were created, some transformations had unique objects associated with them, two that would've taken a similar role to the priest are in the game's files:
• A tombstone, which would turn Peppino back from a ghost
• A pile of corn, which would distract Mort and free Peppino from him, referencing how corn was used as a health pick-up in Mort the Chicken
and some blocks that could only be broken by transformations, like the rat, also remain in the files
• A barrel block
• A hot metal block, which would be destroyed by both knight and rocket transformations
• A ball block
• A Gustavo and Brick block

When replaying through an already-completed save file, one can change the Peppino's Final Judgement ranking screen. This was not always the case, as originally attempting to change the judgement would show a screen of an ominous Peppino, drawn with details than his sprites but less detailed than his battle introduction portraits, with the caption "THE JUDGEMENT WAS FINAL. PLEASE TRY A NEW SAVE FILE." This screen would eventually be implemented in the Halloween update, albeit without its text, when one attempts to exit a newly introduced bonus level through the ghost transformation.

The Noise, Pepperman, and The Vigilante were all planned to be playable at some point - Vigilante and Noise were both intended to be regular playable characters, while Pepperman was intended as a one-level only character on Refrigerator-Refrigerador-Freezerator, much like Gustavo & Brick. All 3 of them have code and animations left over in the game, of varying compatability. The Noise would later be re-added in a future update due to all of his animations being completed.

The Captain Goblin on the Crust Cove level originally was called Captain Bomb and had a more humanoid design. The only animations made for him are simply the same image rotated and modified.

Pizzahead's third phase was originally going to be, instead of a boss rush, Pizzahead growing desperate and starting to violently attack Peppino directly through kicks and punches rather than cartoon gags. This was scrapped as the development team didn't want Pizzahead to be perceived as an angry character as much as just an evil clown.