Differentiating between Allograft Active Rejection and Allograft Injury

Long-term survival of a renal transplant depends on accurate and timely detection of allograft rejection and effective treatment. The standard for diagnosis of rejection is histology obtained via needle biopsy; however, the technique is rarely utilized for surveillance due to high cost, complicated logistics, and potential complications, as well as patient discomfort and inconvenience.

A possible noninvasive marker for diagnosis of graft rejection is donor-derived cell-free DNA (dd-cfDNA) detected in the blood of transplant recipients. Data from several single-center studies indicate that dd-cfDNA levels in blood, measured as a fraction of the total cell-free DNA (cfDNA), can discriminate rejection from non-rejection in heart, lung, liver, and kidney allografts.

To date, there are few data on dd-cfDNA in renal transplants. In renal transplantation, there are no existing biomarkers that adequately measure the status of active injury to the allograft. Roy D. Bloom, MD, and colleagues reported on the Circulating Donor-Derived Cell-Free DNA in Blood for Diagnosing Acute Rejection in Kidney Transplant Recipients (DART) study in the Journal of the American Society of Nephrology [2017;28:2221-2232]. The DART study is a multicenter study of kidney transplant recipients using an analytically validated dd-cfDNA test that used targeted amplification and sequencing of single-nucleotide polymorphisms to quantify donor and recipient DNA contributions, with no need for prior genotyping of donor or recipient DNA.

The study enrolled 384 renal transplant recipients from April 2015 until May 2016; participants were recruited from 14 clinical sites. Of the 384 patients, 245 were enrolled within 1 to 3 months of their kidney transplant and 139 at the time of a clinically indicated renal biopsy. A subset of 107 clinically indicated biopsies that had matched plasma dd-cfDNA results provided the core dataset used for the analyses of dd-cfDNA to discriminate rejection from no rejection status. The biopsy-based pathologists’ reports were used as the diagnostic standard.

The characteristics of the DART study population were representative of the population in the United States renal transplantation registry. Patients in the active rejection group were more likely to be black and have received a transplant from a deceased donor compared with the no rejection group and with the overall DART population. Those in the active rejection group were significantly younger than those in the no rejection group.

At the time of the data lock, 219 patients had at least one renal biopsy; 242 biopsies had sufficient specimens and associated pathologists’ reported results. Of those 242, 204 were performed for clinical suspicion of rejection, 34 for surveillance, and four for follow-up of treated rejection. Of the 34 surveillance biopsies, only one revealed rejection; the scenario of no clinical indication for biopsy was not included in the subsequent calculations for the performance characteristics for dd-cfDNA to discriminate active rejection.

Three subclasses of rejection were combined in the primary analyses: (1) T-cell-mediated rejection (TCMR); (2) acute/active antibody-mediated rejection (ABMR); and (3) chronic, active ABMR. A review of 59 pathologists’ biopsy reports confirmed diagnosis of active rejection: 58 cases of active rejection in 204 biopsies performed for clinical suspicion (most commonly an elevation in serum creatinine), and one case of active rejection in 34 surveillance biopsies.

The researchers included all dd-cfDNA results that were collected at the same time that a clinically indicated biopsy was performed to define the area under the curve-received operating characteristic (AUC-ROC) performance of dd-cfDNA. There were 27 biopsy specimens from 27 patients with active rejection and 80 specimens from 75 patients without active rejection. Samples with >1% dd-cfDNA occurred significantly more often (P<.01) in these types of rejection and subelements: acute/active ABMR; chronic, active ABMR; any or moderate microvascular inflammation; linear C4d staining in peritubular capillaries; and presence of donor-specific antibody.

There were significant differences between groups in the fraction of dd-cfDNA in blood plasma. In the active rejection group, the median level of dd-cfDNA was significantly higher than in the comparator group of biopsy specimens without active rejection (1.6% vs 0.3%; P<.001). There was variation in median levels of dd-cfDNA by type of active rejection: 2.9% (ABMR); 1.2% (TCMR only, types IB and IIA); and 0.2% (TCMR only type IA).

Positive predictive value for active rejection at a cutoff of 1.0% dd-cfDNA was 61%; negative predictive value was 84%. The AUC for discriminating ABMR from samples without ABMR was 0.87 (95% confidence interval, 0.75-0.97). Positive predictive value for ABMR at a cutoff of 1.0% dd-cfDNA was 44%; negative predictive value was 96%.

Limitations to the study cited by the researchers included the inability to estimate the performance of dd-cfDNA to discriminate active rejection or ABMR in patients who may have had subclinical rejection because there were only 34 surveillance biopsies and only one finding of active rejection. Further, the number of active rejections was limited. Finally, biopsy-matched blood samples were not collected for all biopsy specimens.

In summary, the researchers said, “This report sets the initial foundation for the performance characteristics of dd-cfDNA to detect active rejection and injury of the renal allograft beyond serum creatinine and without the need for a biopsy. The next steps of development include studies to validate these findings and to demonstrate the clinical utility of this new type of immune monitoring of the graft.”

Takeaway Points

  • Researchers reported on the DART (Circulating Donor-Derived Cell-Free DNA in Blood for Diagnosing Acute Rejection in Kidney Transplant Recipients) study of the use of donor-derived cell-free DNA (dd-cfDNA) to differentiate active rejection of allograft from allograft injury.
  • The positive and negative predictive values for active rejection at a cutoff of 1.0% dd-cfDNA were 61% and 84%, respectively.
  • Using dd-cfDNA to assess allograft rejection and injury is feasible; levels <1% reflect the absence of active rejection and levels >1% indicate a probability of active rejection.