Ronald P. Pelletier

The impact of mycophenolate mofetil dosing patterns on clinical outcome after renal transplantation

Abstract: Background:  Mycophenolate mofetil (MMF) has proven to be a very effective drug for the prevention of acute rejection following renal transplantation when dosed as prescribed at 2 or 3 g/d. However, circumstances arise in clinical transplantation where the dose must be lowered, either to avoid drug toxicity or because of concurrent infection. The impact on the incidence of acute rejection and graft survival when the MMF dose must be lowered has not previously been investigated.

Impact of acute rejection and early allograft function on renal allograft survival

Both acute rejection and the function of a renal allograft early after transplantation correlate with long-term graft survival. In this study we assessed the relationship between these two factors in 843 adult recipients of first cadaveric renal grafts, transplanted at a single institution and followed for a minimum of 3.5 years. Patients were divided into four groups according to (1) history of acute rejection (AR) during the first 6 months after transplantation, and (2) concentration of serum creatinine at 6 months after transplantation (SCr(6mo) < or > or = 2 mg/dl). Death censored allograft survival was not significantly different among patients without AR and with low SCr(6mo) (group 1, n=376), patients without AR but with an elevated SCr(6mo) (group 2, n=117), and patients with AR but low SCr(6mo) (group 3, n=185). In contrast, graft survival was significantly worse in patients with AR and an elevated SCr(6mo) (group 4, n=165) compared with patients in the other three groups (Cox, P<0.0001). The elevated SCr(6mo) in group 4 patients was not necessarily the consequence of AR, as 32% of patients in group 4 had a SCr at 10 days after transplantation (SCr(10d)), before they had AR, that was equal to or higher than the SCr(6mo). Based on this observation we investigated the implications of the SCr(10d) concentration for graft prognosis. The SCr(10d) correlated weakly with graft survival (Cox, P=0.05). However, an elevated SCr(10d) correlated with other potential risk factors for graft survival including: Older donors (P<0.0001), male recipients (P<0.0001), and heavier recipients (P<0.0001, all by multivariate regression); and posttransplant factors such as, increasing numbers of AR (P<0.0001), higher posttransplant blood pressure (P<0.0001), and lower doses of cyclosporine (P<0.0001, all by multivariate regression). In conclusion, graft dysfunction predicts poor graft survival only when associated with AR. Similarly, AR predicts a poor renal allograft survival only when associated with graft dysfunction. The SCr(10d) is an indicator of risk factors from both the donor and recipient, and an elevated SCr(10d) predicts a higher risk of acquiring additional risk factors early after transplantation.

Treatment with anti-vascular cell adhesion molecule 1 monoclonal antibody induces long-term murine cardiac allograft acceptance.

Daily in vivo treatment of murine H-2d --> H-2b cardiac allograft recipients with 400 micrograms/day i.p. of M/K-2, a mAb to the endothelial adhesion molecule VCAM-1, resulted in prolongation of graft survival. The surviving allografts showed little of the histologic changes observed in acutely rejecting control allografts. When antibody treatment was discontinued after 20 days, grafts continued to function for at least 40 more days. This was approximately 30 days after mAb was no longer detectable by ELISA in the circulation, or by immunoperoxidase staining at the graft site. The most notable feature of grafts that survived 60 days was the presence of mild interstitial fibrosis. Endothelial reactivity was minimal with the mAbs MECA-32 and M/K-2, which have been used in previous studies to visualize the extensive endothelial inflammation that develops during untreated acute rejection. There was a mild cellular infiltrate containing T cells, but few macrophages. However, infiltrating T cells appeared to be inactive in that IL-2R+ cells were immunohistologically undetectable and mRNA for IL-2, IL-4, or IFN-gamma was undetectable by polymerase chain reaction. In general, the immunologic conditions in these long-term grafts differed from those seen in normal cardiac tissue, cardiac isografts, or cardiac allografts. These data demonstrate that M/K-2 mAb can suppress cardiac allograft rejection and induce long-term graft acceptance. This graft survival appears to be associated with the development of a unique state of immunity at the graft site.

Multicenter evaluation of a novel endothelial cell crossmatch test in kidney transplantation.

Background. Despite their clinical importance, clinical routine tests to detect anti-endothelial cell antibodies (AECA) in organ transplantation have not been readily available. This multicenter prospective kidney transplantation trial evaluates the efficacy of a novel endothelial cell crossmatch (ECXM) test to detect donor-reactive AECA associated with kidney allograft rejection. Methods. Pretransplant serum samples from 147 patients were tested for AECA by a novel flow cytometric crossmatch technique (XM-ONE) using peripheral blood endothelial progenitor cells as targets. Patient enrolment was based on acceptance for transplantation determined by donor lymphocyte crossmatch results. Results. Donor-reactive AECA were found in 35 of 147 (24%) patients. A significantly higher proportion of patients with a positive ECXM had rejections (16 of 35, 46%) during the follow-up of at least 3 months compared with those without AECA (13 of 112, 12%; P<0.00005). Both IgG and IgM AECAs were associated with graft rejections. Mean serum creatinine levels were significantly higher in patients with a positive ECXM test at 3 and 6 months posttransplant. Conclusions. XM-ONE is quick, easy to perform on whole blood samples and identifies patients at risk for rejection and reduced graft function not identified by conventional lymphocyte crossmatches.

Evidence for a genetic predisposition towards acute rejection after kidney and simultaneous kidney-pancreas transplantation.

Background. In vitro production of tumor necrosis factor-&agr; (TNF-&agr;), interferon-&ggr; (IFN-&ggr;), interleukin 10 (IL-10), and transforming growth factor-&bgr; (TGF-&bgr;) correlate with their respective genetic polymorphisms. We analyzed the relationship between these genetic polymorphisms and posttransplant outcome. Methods. Using DNA polymerase chain reaction (PCR) technology, polymorphisms for TNF- &agr;, IFN-&ggr;, IL-10, and TGF-&bgr; were determined for 82 kidney (K) and 19 simultaneous kidney-pancreas (SKP) recipients. These results were analyzed with regard to the incidence of acute rejection (AR), and the timing and severity of the first AR episode. Results. A high TNF-&agr; production phenotype correlated with recurrent acute rejection (AR) episodes (P <0.026). Compared with the low TNF-&agr; production phenotype, more patients with the high production phenotype had a post-AR serum creatinine >2.0 mg/dl, but this was not statistically significant (64 vs. 35%, P =0.12). There was no relationship between TNF-&agr; genotype and the time to first AR episode or incidence of graft loss. IFN-&ggr; production phenotype showed no correlation with any of these clinical outcome parameters. There was an increase in AR incidence as the IL-10 production phenotype increased (low, intermediate, high), but only in low TNF-&agr; producer phenotypes (P =0.023). Conclusions. Patients with a polymorphic cytokine genotype putatively encoding for high in vivo TNF-&agr; production, and to a lesser extent IL-10 cytokine genotypes putatively encoding for higher levels of in vivo IL-10 production, had a worse clinical outcome regarding AR episodes. These data support the hypothesis that the strength of alloimmune responsiveness after transplantation in part is genetically determined.

Clinical Significance of MHC‐Reactive Alloantibodies that Develop after Kidney or Kidney–pancreas Transplantation

The purpose of this study was to determine the relationships between acute rejection, anti‐major histocompatibility complex (MHC) class I and/or class II‐reactive alloantibody production, and chronic rejection of renal allografts following kidney or simultaneous kidney‐pancreas transplantation. Sera from 277 recipients were obtained pretransplant and between 1 month and 9.5 years post‐transplant (mean 2.6 years). The presence of anti‐MHC class I and class II alloantibodies was determined by flow cytometry using beads coated with purified MHC molecules. Eighteen percent of recipients had MHC‐reactive alloantibodies detected only after transplantation by this method. The majority of these patients produced alloantibodies directed at MHC class II only (68%). The incidence of anti‐MHC class II, but not anti‐MHC class I, alloantibodies detected post‐transplant increased as the number of previous acute rejection episodes increased (p = 0.03). Multivariate analysis demonstrated that detection of MHC class II‐reactive, but not MHC class I‐reactive, alloantibodies post‐transplant was a significant risk factor for chronic allograft rejection, independent of acute allograft rejection. We conclude that post‐transplant detectable MHC class II‐reactive alloantibodies and previous acute rejection episodes are independent risk factors for chronic allograft rejection. Implementing new therapeutic strategies to curtail post‐transplant alloantibody production, and avoidance of acute rejection episodes, may improve long‐term graft survival by reducing the incidence of chronic allograft rejection.

Survival Benefit with Kidney Transplants from HLA-Incompatible Live Donors

BACKGROUND A report from a high-volume single center indicated a survival benefit of receiving a kidney transplant from an HLA-incompatible live donor as compared with remaining on the waiting list, whether or not a kidney from a deceased donor was received. The generalizability of that finding is unclear. METHODS In a 22-center study, we estimated the survival benefit for 1025 recipients of kidney transplants from HLA-incompatible live donors who were matched with controls who remained on the waiting list or received a transplant from a deceased donor (waiting-list-or-transplant control group) and controls who remained on the waiting list but did not receive a transplant (waiting-list-only control group). We analyzed the data with and without patients from the highest-volume center in the study. RESULTS Recipients of kidney transplants from incompatible live donors had a higher survival rate than either control group at 1 year (95.0%, vs. 94.0% for the waiting-list-or-transplant control group and 89.6% for the waiting-list-only control group), 3 years (91.7% vs. 83.6% and 72.7%, respectively), 5 years (86.0% vs. 74.4% and 59.2%), and 8 years (76.5% vs. 62.9% and 43.9%) (P<0.001 for all comparisons with the two control groups). The survival benefit was significant at 8 years across all levels of donor-specific antibody: 89.2% for recipients of kidney transplants from incompatible live donors who had a positive Luminex assay for anti-HLA antibody but a negative flow-cytometric cross-match versus 65.0% for the waiting-list-or-transplant control group and 47.1% for the waiting-list-only control group; 76.3% for recipients with a positive flow-cytometric cross-match but a negative cytotoxic cross-match versus 63.3% and 43.0% in the two control groups, respectively; and 71.0% for recipients with a positive cytotoxic cross-match versus 61.5% and 43.7%, respectively. The findings did not change when patients from the highest-volume center were excluded. CONCLUSIONS This multicenter study validated single-center evidence that patients who received kidney transplants from HLA-incompatible live donors had a substantial survival benefit as compared with patients who did not undergo transplantation and those who waited for transplants from deceased donors. (Funded by the National Institute of Diabetes and Digestive and Kidney Diseases.).

Quantifying the Risk of Incompatible Kidney Transplantation: A Multicenter Study

Incompatible live donor kidney transplantation (ILDKT) offers a survival advantage over dialysis to patients with anti‐HLA donor‐specific antibody (DSA). Program‐specific reports (PSRs) fail to account for ILDKT, placing this practice at regulatory risk. We collected DSA data, categorized as positive Luminex, negative flow crossmatch (PLNF) (n = 185), positive flow, negative cytotoxic crossmatch (PFNC) (n = 536) or positive cytotoxic crossmatch (PCC) (n = 304), from 22 centers. We tested associations between DSA, graft loss and mortality after adjusting for PSR model factors, using 9669 compatible patients as a comparison. PLNF patients had similar graft loss; however, PFNC (adjusted hazard ratio [aHR] = 1.64, 95% confidence interval [CI]: 1.15–2.23, p = 0.007) and PCC (aHR = 5.01, 95% CI: 3.71–6.77, p < 0.001) were associated with increased graft loss in the first year. PLNF patients had similar mortality; however, PFNC (aHR = 2.04; 95% CI: 1.28–3.26; p = 0.003) and PCC (aHR = 4.59; 95% CI: 2.98–7.07; p < 0.001) were associated with increased mortality. We simulated Centers for Medicare & Medicaid Services flagging to examine ILDKTs effect on the risk of being flagged. Compared to equal‐quality centers performing no ILDKT, centers performing 5%, 10% or 20% PFNC had a 1.19‐, 1.33‐ and 1.73‐fold higher odds of being flagged. Centers performing 5%, 10% or 20% PCC had a 2.22‐, 4.09‐ and 10.72‐fold higher odds. Failure to account for ILDKTs increased risk places centers providing this life‐saving treatment in jeopardy of regulatory intervention.

De novo Thrombotic Microangiopathy in Renal Allograft Biopsies—Role of Antibody‐Mediated Rejection

The most common cause of thrombotic microangiopathy (TMA) in renal allografts is thought to be calcineurin inhibitor toxicity. Antibody‐mediated rejection (AMR) can also cause TMA, but its true impact on de novo TMA is unknown. In a retrospective review of renal allograft biopsies from January 2003 to December 2008 at our institution, we determined the prevalence of TMA in patients with C4d positive (n = 243) and C4d negative (n = 715) biopsies. Over 90% of patients received cyclosporine in both groups. De novo TMA was seen in 59 (6.1%) patients; most of them (55%) with C4d positive biopsy. Among patients with C4d positive biopsies, 13.6% had TMA, as compared to only 3.6% patients with C4d negative biopsies (p < 0.0001). Incidence of graft loss between C4d positive and C4d negative TMA groups was not significantly different, but 70% of patients with C4d positive TMA who received plasmapheresis had slightly lower graft loss rate. In biopsies with AMR‐associated TMA, glomerulitis and peritubular capillaritis were significantly more prominent. AMR is the most common cause of TMA in renal allografts in our patient population. It is important to recognize AMR‐related TMA because plasmapheresis treatment may be beneficial.

Murine Renal Allografts: Spontaneous Acceptance Is Associated with Regulated T Cell-Mediated Immunity

It was shown >20 yr ago that mice will spontaneously accept renal allografts in the absence of immunosuppression, but the mechanism responsible for this is not understood. We transplanted DBA/2 (H-2d) kidneys into nephrectomized C57BL/6 (H-2b) mice, and the allografts were spontaneously accepted for >60 days without immunosuppression. In contrast, nonimmunosuppressed cardiac and skin allografts in the same strain combination are rejected within approximately 10 days. The accepted renal allografts have a prominent leukocytic infiltrate, suggesting an ongoing, local immune response. At 60 days post-transplant, the recipients of accepted renal allografts display DBA/2-reactive alloantibodies. They also display DBA/2-reactive delayed-type hypersensitivity responses that are actively counter-regulated by DBA/2-induced TGF-β production, but not by IL-10 production. These data suggest that a donor-reactive, cell-mediated immune mechanism involving TGF-β is associated with the spontaneous acceptance of renal allografts in mice.