Luton will definitely begin life in Coca-Cola League Two on minus 30 points next month after the Football Association rejected an appeal.
The Hatters were docked 10 points by the FA as well as fined £50,000 for breaching regulations over payments to agents.
This was followed by a 20-point penalty from the Football League after the club exited administration without agreeing a Company Voluntary Agreement.
The club were attempting to wipe out the FA's penalty but a hearing in London has refused to overturn the decision.
An FA statement said: "The deduction of 10 points was a heavy sanction, as it was intended to be, but was not excessive as a reflection of the seriousness of the breaches and the need to deter such conduct within football clubs.
"It is highly unfortunate for Luton Town and their loyal fans that shortly after the FA regulatory commission reached its decision the Football League quite separately imposed a 20-point deduction for entirely different actions by the club."