Noninvasive diagnosis of renal-allograft rejection by measurement of messenger RNA for perforin and granzyme B in urine

Background: Acute rejection is a serious and frequent complication of renal transplantation, and its diagnosis is contingent on the invasive procedure of allograft biopsy. A noninvasive diagnostic test for rejection could imp rove the outcome of transplantation. Methods: We obtained 24 urine specimens from 22 renal-allograft recipients with a biopsy-confirmed episode of acute rejection and 127 samples from 63 recipients without evidence of acute rejection. RNA was isolated from the u rinary cells. Messenger RNA (mRNA) encoding the cytotoxic proteins perforin and granzyme B and a constitutively expressed cyclophilin B gene were meas ured with the use of a competitive, quantitative polymerase-chain-reaction assay, and the level of expression was correlated with allograft status. Results: The log-transformed mean (+/-SE) levels of perforin mRNA and granz yme B mRNA, which encode cytotoxic proteins, but not the levels of constitu tively expressed cyclophilin B mRNA, were higher in the urinary cells from the 22 patients with a biopsy-confirmed episode of acute rejection than in the 63 recipients without an episode of acute rejection (perforin, 1.4+/-0. 3 vs. -0.6+/-0.2 fg per microgram of total RNA; P<0.001; and granzyme B, 1. 2+/-0.3 vs. -0.9+/-0.2 fg per microgram of total RNA; P<0.001). Analysis in volving the receiver-operating-characteristic curve demonstrated that acute rejection can be predicted with a sensitivity of 83 percent and a specific ity of 83 percent with the use of a cutoff value of 0.9 fg of perforin mRNA per microgram of total RNA, and with a sensitivity of 79 percent and a spe cificity of 77 percent with the use of a cutoff value of 0.4 fg of granzyme B mRNA per microgram of total RNA. Sequential urine samples were obtained from 37 patients during the first nine days after transplantation, and meas urements of the levels of mRNA that encoded cytotoxic proteins identified t hose in whom acute rejection developed. Conclusions: Measurement of mRNA encoding cytotoxic proteins in urinary cel ls offers a noninvasive means of diagnosing acute rejection of renal allogr afts. (N Engl J Med 2001; 344:947-54.) Copyright (C) 2001 Massachusetts Med ical Society.