The
Spider's Rune is located in the
Slumbering Sanctuary. You access the
Slumbering Sanctuary after you beat the Conjunctivitis boss in the
Insufferable Crypt. The
Ram Rune is required to get there.
Once you have the Ram Rune, you need to take this path:
Prisoners' Quarters > Toxic Sewers > Ancient Sewers > Insufferable Crypt > Slumbering Sanctuary
I found this map to be very helpful.