The Leicester Tigers fly-half took a blow while training for the club and initial reports suggest he could be out of action for six weeks.
That would rule the potential 2013 Lion out of the opening two games of the Championship.
"Toby has had a bang on the knee and it's being treated. We'll just have to wait and see how it progresses," Leicester head coach Richard Cockerill told the Daily Mail.
England kick-off their campaign against Scotland on February 4 before tackling Italy the following week.
It is the latest blow for interim head coach Stuart Lancaster, who booted Danny Care out of his squad yesterday following drink driving allegations.
Lancaster is already without Flood's Tigers team-mate Louis Deacon while there are also doubts over Manu Tuilagi.
England will name a 32-man squad for the tournament on January 11.