Changing the lists of symbols prettified by prettify-symbols-mode is more tricky than it should be; I think this is a bug in prettify-symbols-mode. Here is a test case:

(with-current-buffer (get-buffer-create "fl")
  (erase-buffer)
  (emacs-lisp-mode)
  (prettify-symbols-mode -1)
  (setq prettify-symbols-alist '(("A" . ?B)))
  (prettify-symbols-mode)
  (setq prettify-symbols-alist '(("A" . ?C)))
  (prettify-symbols-mode)
  (insert "A A A")
  (pop-to-buffer (current-buffer)))

Looking at this code, I expect “C C C” to be shown. Yet it's not the case; instead, “B B B” is shown.

On the other hand, this works:

(with-current-buffer (get-buffer-create "fl")
  (erase-buffer)
  (emacs-lisp-mode)
  (prettify-symbols-mode -1)
  (setq prettify-symbols-alist '(("A" . ?B)))
  (prettify-symbols-mode)
  (prettify-symbols-mode -1)
  (setq prettify-symbols-alist '(("A" . ?C)))
  (prettify-symbols-mode)
  (insert "A A A")
  (pop-to-buffer (current-buffer)))

I think the line indicated by !!! in the following code sample is wrong:

    (define-minor-mode prettify-symbols-mode
      (...)
      (if prettify-symbols-mode
          ;; Turn on
          (when (setq prettify-symbols--keywords (prettify-symbols--make-keywords))
            (font-lock-add-keywords nil prettify-symbols--keywords) ;; !!! HERE
            (setq-local font-lock-extra-managed-props
                        (append font-lock-extra-managed-props
                                '(composition
                                  prettify-symbols-start
                                  prettify-symbols-end)))
            (when prettify-symbols-unprettify-at-point
              (add-hook 'post-command-hook
                        #'prettify-symbols--post-command-hook nil t))
            (font-lock-flush))

Indeed, turning on prettify-symbols-mode twice in a row causes two entries to be added to font-lock-add-keywords. This is not an issue in general, except if one changes prettify-symbols-alist between two calls. In that case font-lock-keywords gets two distinct entries for prettify-symbols, and the second one is ignored.

The proposed fix is to move the following snippet up, so that it runs in all cases, instead of just running when prettify-symbols-mode is turned off.

    (when prettify-symbols--keywords
      (font-lock-remove-keywords nil prettify-symbols--keywords)
      (setq prettify-symbols--keywords nil))

Cheers,
Clément.