Jackson, who leads the Bills with 256 rushing yards and two touchdowns, sprained his left knee last week in a win over Baltimore and will wear a brace. Spiller sprained his ankle in the third quarter against the Ravens and returned to practice Wednesday after missing the two previous days.
Last season against Cleveland, Spiller went out with a shoulder injury in the first half and third-stringer Tashard Choice ran for 91 yards in Buffalo's 24-14 win.
Also, the Bills will be without starting cornerback Stephon Williams and free safety Jairus Byrd.
Browns quarterback Brandon Weeden will back up starter Brian Hoyer. Weeden sprained his thumb in Week 2 and Hoyer is 2-0 in two starts. Jason Campbell is inactive.