You can have a website with
www.mysite.it/sub-site but in this case, you have to create the sub-site folder before you deploy into this directory in JMS.
JMS does not create the directory but onlu deploy in existing directory.
Concerning the 500 Internal Error this may be due to a permission problem.
You say that Symbolic Link is allowed and that you were able to create a slave site using Symboli LInk.
Do you have creae a fresh slave site or replicae an exisiting website.
When creating a fresh slave site, do you have this error in a URL
www.mysite.it/multisites/sub-site/installation/index.php ?
May be you have to provide more permission to execute the scripts.
May be also that you are not allowed to execute scripts into the /multisites directory and children directory.
The most probable reason is a permission issue.
You can try to fix this kind of problem with "hello world" procedure.
Install the "hello.php" into the directory that you want to check the permission and in which you want to be sure that you are allowed to execute scripts.