Travel from Rome to Venice by train?

The high speed trains take approx. 3h 45 min form Rome to Venice and stop only in Florence and Bologna along the way.

The high speed train does not go through either Siena or Spello. If you want to visit either one, it's a detour on a different train. You could potentially visit Siena by getting off the high speed train in Florence and then taking a bus back to Siena. Then to proceed to Venice, you'd go back to Florence by bus and take the high speed train again from Florence to Venice.

Another way to get to Siena is to take a Regionale train to Chiusi, then another train from Chiusi to Siena (not much different in time, but cheaper than going to Florence with the high speed). Yet another alternative is to take a bus from Roma to Siena with the Sena Bus company (departures every couple of hours from the Rome Tiburtina station).

Spello is in Umbria, and to get there you need to take a train toward Foligno-Ancona.

The high speed trains also stop at Padua, but that is right outside of Venice. You can't "hop-on-hop-off" these trains as they have all reserved seats. If you do want to stop off for a few hours along the way, you will need separate tickets.

If you take an early train, you can have at least one, perhaps two, memorable meals in Bologna and still get to Venezia that night. A quick look at the sights of Bologna is more than half a day, in my opinion.

It is not clear if you want to overnight, or just break your journey for a few hours.
Bologna or Verona would be the obvious choices for a night or two.
Siena involves connecting trains, so I would advise against it.
As others have said, you will require 2 separate train bookings.