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League One side Rovers were beaten 2-1 by Barrow of the National League at the Memorial Stadium on Sunday.
They have only won once in their last eight matches in all competitions.
"We are giving really sloppy goals away and for me we're becoming a soft touch in our own box," the 38-year-old Rovers manager told BBC Points West.
"It might be coming to the end of the road for one or two if they don't start improving. We have to invest in the team now, I think.
"It's come to the stage now where this group's done very, very well but if we've got ambition to get into the Championship in the next two or three years we have to move it forward and invest wisely in the team.
"I know the areas where I need to strengthen now - it's not rocket science."
Clarke's side are next in action on Saturday when they host Bury in League One.
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Bristol Rovers manager Darrell Clarke says it could be the "end of the road" for some of his players after their shock FA Cup second-round defeat.