This seems to work, using a negative look-ahead.

use strict;
use warnings;

my @words = qw{
   test
   tesi
   stoat
   trout
   };

my $regex = qr
   {(?x)
       ^
       (.)
       .*?
       (?!\1)
       .
       $
   };

print
   m{$regex}
   ? qq{    Matched: $_\n}
   : qq{Not matched: $_\n}
   for @words;
[download]

Prints

Not matched: test
    Matched: tesi
    Matched: stoat
Not matched: trout
[download]

I hope this is helpful.

Cheers,

JohnGG