Survival in Allosensitized Children After Listing for Cardiac Transplantation

PEDIATRIC TRANSPLANTATION

Survival in Allosensitized Children After Listing for Cardiac Transplantation Brian Feingold, MD,a Pam Bowman, RN,a Adriana Zeevi, PhD,b Alin L. Girnita, MD,b Eric S. Quivers, MD,a Susan A. Miller, MD,a and Steven A. Webber, MBChBa Background: Little is known about the effect of pre-transplant alloantibody in the pediatric cardiac transplant population. Methods: All cardiac listings (n ⫽ 298) at Children’s Hospital of Pittsburgh from January 1990 through February 2006 were reviewed to determine the impact of allosensitization on transplantation outcomes. Analysis focused on: (1) wait list outcomes; (2) survival from the time of listing, regardless of subsequent transplantation; (3) post-transplant graft and patient survival; and (4) post-transplant freedom from graft vasculopathy. Institutional policy required a negative, prospective crossmatch for candidates with panel-reactive antibody ⬎20%. Results: Alloantibody data were available for 252 (85%) listings. Median time to transplantation was greater for sensitized vs non-sensitized subjects (2.7 months vs 1.3 months; p ⫽ 0.02). At 1 year after listing, sensitized subjects had a higher incidence of death (22% vs 8.4%; p ⫽ 0.055). Survival at all time-points after listing (regardless of transplantation) was worse for sensitized subjects (p ⫽ 0.04). Although no statistically significant differences in post-transplant graft or patient survival were noted, pre-transplant allosensitization was associated with decreased freedom from graft vasculopathy (hazard ratio [HR] 2.76, 95% confidence interval [CI] 1.18 to 6.45; p ⫽ 0.019). Conclusions: A policy requiring a negative, prospective crossmatch for highly sensitized candidates is associated with longer wait list time and higher mortality after listing. The development of graft vasculopathy appears to be influenced by the presence of pre-transplant alloantibody. J Heart Lung Transplant 2007;26:565–71. Copyright © 2007 by the International Society for Heart and Lung Transplantation.

The presence of pre-formed antibodies against non–self human leukocyte antigens (HLA) has been associated with hyperacute, acute and chronic rejection in renal and adult cardiac transplant patients.1– 8 However, only sparse data exist regarding the impact of preformed alloantibodies after listing and transplantation in the pediatric cardiac transplant population. Allosensitization is being increasingly recognized in pediatric candidates with congenital heart disease, especially when prior surgeries have incorporated homograft material.9 –12 We sought to evaluate the impact of alloantibodies on pre-transplant survival, post-transplant graft and patient survival and post-transplant graft

From the aDivision of Pediatric Cardiology and Cardiopulmonary Transplantation, Children’s Hospital of Pittsburgh, Pittsburgh, Pennsylvania; and bDepartment of Pathology, University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania. Submitted September 19, 2006; revised February 11, 2007; accepted March 12, 2007. Reprint requests: Brian Feingold, MD, Division of Pediatric Cardiology and Cardiopulmonary Transplantation, Children’s Hospital of Pittsburgh, 3705 Fifth Avenue, Pittsburgh, PA 15213. Telephone: 412-692-5541. Fax: 412-692-6991. E-mail: [email protected] Copyright © 2007 by the International Society for Heart and Lung Transplantation. 1053-2498/07/$–see front matter. doi:10.1016/ j.healun.2007.03.015

vasculopathy in a retrospective cohort of children listed for cardiac transplantation. METHODS Data Collection After institutional review board approval, all subjects listed for heart transplantation from January 1990 through February 2006 at Children’s Hospital of Pittsburgh were identified from the transplantation database at the Division of Cardiology. Patients listed for multiorgan transplantation were excluded. Data abstracted from the medical record of each subject included: demographics; listing diagnosis; blood type; history of prior sensitizing events (transfusion, prior transplantation, mechanical circulatory support); active wait list time; wait list outcome (death, transplantation, delisting, alive waiting); and post-transplant outcomes (death, re-transplantation, chronic graft vasculopathy). All pre-transplant complement-dependent cytotoxicity panel-reactive antibody (CDC-PRA) results were collected. Results of the retrospective cytotoxicity donorspecific crossmatch were also recorded. Determination of Allosensitization Historically, PRA was performed by the modified AMOS technique13,14 until 1997 when anti-human globulin 565

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(AHG) was added to the assay to enhance sensitivity.15 In 2001, the enzyme-linked immunosorbent assay (ELISA)16 began to be used in combination with CDCPRA for determination of sensitization status. For purposes of this analysis, only dithiothreitol (DTT)-treated PRA results were collected. Subjects were categorized as being “sensitized” with any pre-transplant CDC-PRA ⬎10%. Allosensitization status was routinely assessed at evaluation/listing and at the time of transplantation. A negative prospective crossmatch (or, more recently, a negative “virtual” crossmatch) was required for transplantation of patients with CDC-PRA ⬎20%. Determination of Graft Vasculopathy Each subject’s serial coronary angiography reports were reviewed chronologically from the time of transplantation for determination of the presence or absence of graft vasculopathy (CAD). We routinely assessed for CAD in subjects weighing ⱖ12 kg by selective coronary angiography biannually, beginning 1 year after transplantation. All coronary angiograms were compared with the most recent prior angiography (if applicable) and were reviewed by the catheterizing physician and an experienced adult interventional cardiologist. Any luminal irregularities or attenuation/obliteration of distal vessels were considered evidence of CAD. For subjects who died and in whom an autopsy was performed, the presence of CAD on autopsy was also considered evidence of CAD. A subject’s earliest positive angiography (or findings of CAD on autopsy) was considered time of “failure.” In those without evidence of CAD, the duration of follow-up was limited to the time of last negative angiogram. Subjects in whom CAD was not assessed (post-transplant follow-up of ⬍1 year or weight ⬍12 kg) were excluded from CAD analysis. Statistical Analysis Continuous data are presented as mean ⫾ standard deviation or median (interquartile range, IQR). Categorical data are presented as count and percentage of total. Group characteristics were compared using the 2-sample t-test, Mann–Whitney U-test, chi-square test or Fisher’s exact test. Survival estimates were computed using the Kaplan–Meier technique and compared using the log-rank test. For the multivariate analyses, univariate predictors (p ⱕ 0.1) were entered into a stepwise Cox proportional hazard model. Because our focus was to describe the impact of pre-transplant allosensitization on transplant-related outcomes, multivariate analyses were performed only when sensitization status was a significant univariate predictor of survival. All statistical analyses were conducted at ␣ ⫽ 0.05, employing 2-sided tests using STATA v9 software (StataCorp LP, College Station, TX).

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RESULTS A total of 298 listings (in 277 subjects) were identified. Pre-transplant allosensitization status was not available for 46 (15%) listings, which were excluded from further analysis. Two-hundred sixteen (86%) were non-sensitized and 36 (14%) were sensitized. Table 1 shows the distribution of listings by CDC-PRA. There were 24 listings (10%) in 23 subjects with CDC-PRA ⬎20%. Characteristics at listing by sensitization status are shown in Table 2. There were no significant differences in age at listing, gender or race. The sensitized group had a higher proportion of subjects with diagnoses of congenital heart disease and re-transplantation, but this did not reach statistical significance. The sensitized group was more likely to have a history of prior blood transfusion (p ⫽ 0.03) and prior sternotomy (p ⫽ 0.001), whereas no differences in prior extracorporeal membrane oxygenation (ECMO) or ventricular assist device (VAD) exposure was noted. Finally, a larger proportion of subjects in the non-sensitized group were de-listed (p ⫽ 0.02). Because these subjects disproportionately “add” survival time to the non-sensitized group, all de-listed subjects were excluded in the Kaplan–Meier analyses of wait list outcomes and survival after listing for transplantation. De-listed subjects remained in the competing risks analysis, however, as the goal of this analysis was to depict all possible outcomes after listing. Wait List Outcomes The median active wait list time was 2.6 months (IQR 0.9 to 12.2) for the sensitized group and 1.3 months (IQR 0.5 to 3.6) for the non-sensitized group (p ⫽ 0.008). In those who were transplanted, the active wait list time was greater for sensitized vs non-sensitized subjects (2.7 months, IQR 0.9 to 7.0 vs 1.3 months, IQR 0.5 to 3.7; p ⫽ 0.02). Further analysis of wait list survival by Kaplan–Meier analysis (censored at transplantation and last follow-up) showed no significant difference in survival between the groups (Figure 1). Because each of these analyses failed to simultaneously account for all possible wait list outcomes (death, transplantation, de-listing and alive waiting), a Table 1. Distribution of Listings by CDC-PRA PRA 0% 1–10% 11–20% 21–30% 31–50% 51–80% 81–100% Total

n 170 46 12 7 5 7 5 252

% of 252 67% 18% 5% 3% 2% 3% 2% 100%

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Table 2. Listed Characteristics of Subjects by Allosensitization Status

Age at listing (years) Female gender Race White Non-white Listing diagnosis CM CHD Re-transplant Other Blood type O A B AB History of ECMOa VADb Transfusionc Sternotomyd Thoracotomye Outcomes Transplantation Died listed De-listed Alive waiting Wait list time (mo)f List to Tx time (mo)

Non-sensitized (n ⫽ 216) 5.0 (0.5–13.9) 93 (43)

Sensitized (n ⫽ 36) 2.7 (0.7–11.9) 16 (44)

181 (84) 35 (16)

34 (94) 2 (6)

107 (50) 90 (42) 13 (6) 6 (3)

12 (33) 21 (58) 3 (8) —

91 (42) 88 (41) 27 (12) 10 (5)

15 (42) 14 (39) 2 (5) 5 (14)

25 (12) 13 (6) 113 (73) 91 (43) 26 (17)

3 (9) 2 (6) 30 (91) 26 (72) 6 (25)

NS NS 0.03 0.001 NS

145 (67) 19 (9) 48 (22) 4 (2) 1.3 (0.5–3.6) 1.3 (0.5–3.7)

23 (64) 9 (25) 2 (6) 2 (6) 2.6 (0.9–12.2) 2.7 (0.9–7.0)

NS 0.009 0.02 NS 0.008 0.02

p-value NS NS NS

NS

NS

Data presented as median (inter-quartile range) or count (%). CHD, congenital heart disease; CM, cardiomyopathy; ECMO, extracorporeal membrane oxygenation; mo, months; Tx, transplant; VAD, ventricular assist device. a n ⫽ 211 vs 35; bn ⫽ 212 vs 35; cn ⫽ 154 vs 33; dn ⫽ 210 vs 36; e n ⫽ 157 vs 24; fn ⫽ 168 vs 34.

competing risks analysis of all listed subjects was also performed. Figure 2 shows the cumulative incidence of each of the competing outcomes stratified by allosensitization status. These graphs illustrate that the sensitized group had a higher proportion of subjects alive and waiting early after listing than the non-sensitized group. Also, death occurred more frequently early after listing in the sensitized group and accounted for a larger relative proportion of outcomes at 12 months after listing (22.2 ⫾ 6.9% vs 8.4 ⫾ 1.9%; Z-test, p ⫽ 0.055). Finally, fewer sensitized subjects received transplantation at 12 months (p ⫽ NS). Survival After Listing for Transplantation In contrast to the analyses of wait list outcomes presented above, Figure 3 illustrates survival after listing, regardless of subsequent transplantation status. This analysis gives a broader measure of outcome after listing by following subjects until the time of most recent follow-up rather than being confined only to wait list

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follow-up. Survival of the sensitized group was worse than that of the non-sensitized group (p ⫽ 0.04), with much of the difference occurring within 2 years of listing. Non-sensitized status, no prior sternotomy or pretransplant ECMO, cardiomyopathy and transplantation were found to be univariate predictors of survival after listing. Multivariate Cox proportional hazards modeling (with transplantation as a time-dependent co-variate) revealed pre-transplant sensitization (hazard ratio [HR] 1.89, 95% confidence interval [CI] 1.08 to 3.31; p ⫽ 0.025) and a history of pre-transplant ECMO (HR 3.27, 95% CI 1.87 to 5.69; p ⬍ 0.001) to be independent predictors of death after listing for cardiac transplantation. Survival After Transplantation A total of 168 listings (56%) in 155 subjects resulted in transplantation. Characteristics of the transplanted population are shown in Table 3. Retrospective crossmatch data were available for 148 (88%) transplantations, with 3 (2%) positive crossmatches. Although the limited number of subjects with a positive crossmatch prohibited meaningful statistical analyses, ages at listing and transplantation were 5.2, 8.5 and 11.3 months and 5.5, 10.1 and 11.3 months, respectively. Two of these subjects died during follow-up (at 9 and 41 months after transplantation) of non-cardiac causes (overwhelming respiratory syncytial virus infection and sequelae of mitochondrial disorder diagnosed after transplantation). Neither subject had evidence of significant cellular or humoral rejection on autopsy. Both subjects had autopsy evidence of non-occlusive graft vasculopathy. Stratified Kaplan–Meier analyses for post-transplant survival are shown in Figure 4. These curves illustrate graft (Figure 4a) and patient (Figure 4b) survival after transplantation in sensitized and non-sensitized subjects. Neither analysis showed a significant difference between the groups.

Figure 1. Wait list survival (censored for transplantation). *De-listed subjects excluded.

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Figure 2. Competing outcomes after listing for pediatric cardiac transplantation. Percentages indicate outcomes at 12 months after listing.

Post-transplant Graft Vasculopathy One-hundred eighteen subjects (70%) underwent at least one assessment for graft vasculopathy. Greater freedom from development of CAD in the non-sensitized group (p ⫽ 0.04) is illustrated by the survival curves depicted in Figure 5. Female gender, non-white race, re-transplantation, history of VAD, sensitization status, history of previous sternotomy and age at transplantation were found to be univariate predictors of shorter CAD-free survival. Multivariate Cox proportional hazards modeling revealed pre-transplant sensitization (HR 2.76, 95% CI 1.18 to 6.45; p ⫽ 0.019), listing diagnosis of re-transplantation (HR 3.52, 95% CI 1.03 to 12.09; p ⫽ 0.045) and pre-transplant VAD (HR 5.57, 95% CI 1.76 to 17.69; p ⫽ 0.004) to be independent predictors of earlier onset of CAD after transplantation. Sensitized Subjects Outcomes for subjects with CDC-PRA 11% to 20% and ⬎20% are shown in Table 4. Although each outcome

Figure 3. Survival after listing (regardless of transplantation). *Delisted subjects excluded.

was worse for the more highly sensitized group, due to the relatively small number of patients, no statistical differences were found. DISCUSSION We have shown that sensitized pediatric cardiac transplant candidates have longer waiting times and an increased likelihood of death while awaiting transplantation, as has been described previously in renal transplantation.17,18 Given the fact that patients may be denied listing precisely because they are highly allosensitized, the disparity in pre-transplant outcomes is likely to be greater than these data illustrate. Not only are wait list outcomes worse for sensitized subjects, but survival after listing, regardless of subsequent transplantation, is also worse. Table 3. Transplanted Subject Characteristics and CAD Outcomes Non-sensitized Sensitized (n ⫽ 145) (n ⫽ 23) p-value Post-Tx follow-up (mo) 66.1 (20.8–105.9) 42.3 (7.0–64.3) 0.04 Transplantation rate 67% 64% NS Female gender 65 (45) 12 (52) NS Race NS White 125 (86) 21 (91) Non-white 20 (14) 2 (9) Listing diagnosis NS CM 71 (49) 8 (35) CHD 60 (41) 13 (57) Re-transplant 11 (8) 2 (8) Other 3 (2) — Post-Tx CADa 47 (46) 7 (47) NS Time to CADa (mo) 69.9 (38.7–97.7) 41.3 (13.0–70.9) 0.02 Data presented as median (inter-quartile range) or count (%); CAD, graft vasculopathy; CHD, congenital heart disease; CM, cardiomyopathy; mo, months; Tx, transplant. a n ⫽ 103 vs 15.

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Table 4. Sensitized Subjects

Died (listed) Died (all follow-up) Transplanted De-activated CADa

PRA 11–20% (n ⫽ 12) 2 (17%) 4 (33%) 9 (75%) — 2 (33%)

PRA ⬎20% (n ⫽ 24) 7 (29%) 14 (58%) 14 (58%) 2 (8%) 5 (56%)

Data presented as count (%). a n ⫽ 6 (PRA 11–20%) vs 9 (PRA ⬎20%).

Figure 4. (a) Graft survival after transplantation. (b) Patient survival after transplantation.

Despite the appearance of modest graft and patient survival advantages for the non-sensitized group late after transplantation, neither proved to be statistically significant. In the adult renal and cardiac transplant literature, the presence of alloantibodies has been shown to impact post-transplant graft and patient survival.1– 8 One explanation for the lack of statistical difference between the groups is the insufficient number of subjects (particularly in the sensitized group) in the late phase of the Kaplan–Meier analyses. Our find-

Figure 5. Freedom from graft vasculopathy.

ing that pre-sensitized subjects developed graft vasculopathy earlier than non-sensitized subjects is consistent with this explanation because such an observation should ultimately be associated with worse late posttransplant survival. Even after accounting for the impact of listing diagnosis, gender, race, age at transplantation, history of prior sternotomy and pre-transplant VAD exposure by multivariate analysis, pre-sensitization predicted a shorter time to development of CAD. The lack of divergence in the survival estimates of the groups early after transplantation might also be explained by the requirement for a negative prospective (or virtual) crossmatch in highly sensitized subjects. Thus, risk of donor-specific antibody was minimized and early deaths, which might have been expected in the sensitized group due to hyperacute or accelerated rejection, were not observed. Most data linking allosensitization to adverse pre- and post-transplant outcomes have been collected in adult renal and cardiac transplant candidates and recipients,2– 8,16,19 –21 with only sparse reports in pediatric cardiac patients.22–26 Focused study of the pediatric age group is important due to several differences from the adult cardiac transplant population including: listing diagnoses; immunologic maturity; use of homograft material in prior congenital heart disease repairs; and wait list times and survival. Although the current study did not assess acute rejection profiles by pre-transplant sensitization status, our group previously demonstrated that children with adverse first-year rejection profiles more commonly had pre-transplant anti-HLA antibodies detected by ELISA.22 An association with acute rejection was also shown in separate analyses of pre-transplant PRA data by Chartrand et al23 and by Mahle et al.24 Mahle and colleagues reported the experience of the Pediatric Heart Transplant Study (PHTS), a multi-institutional North American research consortium.24 They reported that an elevated PRA at listing was associated with longer time to transplantation and higher risk of death while waiting, as was found in the current study. The PHTS study also demonstrated shorter time to first acute rejection and worse post-transplant survival. Although

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pre-transplant PRA data were available for ⬎1,000 patients, limitations of the study included: non-uniformity of antibody detection techniques (including no data on use of DTT treatment of serum samples); varying crossmatch procedures; diverse pre- and posttransplant interventions for sensitized candidates; and varying policies toward transplantation of sensitized candidates (e.g., requirement for prospective crossmatch). The PHTS study did not show a difference in time to development of post-transplant CAD. By contrast, our prior study in pediatric patients22 suggested a relationship between pre-transplant sensitization and the development of graft vasculopathy. It is likely that our historic policy of requiring a negative prospective crossmatch in sensitized patients biased our results toward worse pre-transplant outcomes. Most pediatric donor hearts are procured from distant sites and prospective crossmatching is generally not feasible. Therefore, a policy mandating a prospective crossmatch will deprive many sensitized candidates of the chance of transplantation, potentially increasing wait list mortality, as was observed in the present study. More recently, we and others have identified suitable donors for sensitized patients on the basis of a “virtual” crossmatch. This requires full donor (and recipient) HLA typing to be available, along with knowledge of unacceptable antigens based on thorough evaluation of the anti-HLA antibodies present in the candidate. It is hoped that increased use of the “virtual” crossmatch may help more sensitized candidates achieve transplantation without increasing the risk of adverse post-transplant outcomes. This assumption remains to be studied in the cardiac transplant population. It should also be noted that very little work has addressed the impact of desensitization therapies on pre- and post-transplant outcomes in the pediatric heart transplant population.27 Various therapies (e.g., intravenous immunoglobulin, cyclophosphamide, plasmapharesis, rituximab) have shown promise in adult renal and cardiac transplant candidates.28 –30 We have found intravenous immunoglobulin to have little effect on lowering PRA in children with congenital heart disease, most of whom had prior surgical repairs that incorporated homograft material.27 The optimal approach to desensitization in this challenging population warrants prospective evaluation in multi-center trials. Due to the retrospective nature of this study, our analysis was confined to the traditional methods employed for the detection of alloantibodies prior to transplantation. Inherent limitations of the CDC-PRA technique include its relative insensitivity for the detection of anti-HLA antibody and difficulty discerning the specificities of incompatible antibodies when a large number of alloantibodies are present. In contrast,

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newer solid-phase assays have greater sensitivity in the detection of alloantibodies and allow precise identification of the anti-HLA antibodies. Many of the historic outcome studies based on CDC-PRA will likely need to be replicated once the most contemporary methods are widely adopted. Finally, our study focused exclusively on the impact of pre-transplant allosensitization on outcomes without assessment of de novo alloantibodies. There is a growing body of literature, including one recent report in pediatric patients,26 showing adverse outcomes related to the development of alloantibodies after transplantation. In conclusion, we found that sensitization prior to transplantation is associated with increased wait list mortality, longer wait list times, increased all-cause mortality after listing (regardless of subsequent transplantation), and earlier development of post-transplant coronary artery disease. Our analysis did not show a difference in graft and patient survival, but was limited by the small number of patients late after transplantation. The nearly equivalent early mortality after transplantation we observed in the sensitized and nonsensitized groups likely reflects our historic policy of not intentionally transplanting across a positive prospective crossmatch. The alternative strategy of transplantation with the first available organ across a positive or predicted positive crossmatch may reduce wait list time and mortality, but at the expense of inferior post-transplant survival. This vexing dilemma becomes increasingly important as more highly sensitized children palliated with homografts are referred for transplantation. REFERENCES 1. Patel R, Terasaki PI. Significance of the positive crossmatch test in kidney transplantation. N Engl J Med 1969;280:735–9. 2. Iwaki Y, Terasaki PI. Sensitization effect. In: Terasaki PI, ed. Clinical transplants. Los Angeles, CA: UCLA Tissue Typing Laboratory; 1986:257. 3. Martin S, Taylor CJ. The immunologically sensitized renal transplant recipient: the impact of advances in technology on organ allocation and transplant outcome. Transplant Rev 1999; 13:40 –51. 4. Hardy MA, Suciu-Foca N, Reed E, et al. Immunomodulation of kidney and heart transplants by anti-idiotypic antibodies. Ann Surg 1991;214:522–30. 5. Lavee J, Kormos RL, Duquesnoy RJ, et al. Influence of panelreactive antibody and lymphocytotoxic crossmatch on survival after heart transplantation. J Heart Lung Transplant 1991;10: 921–30. 6. Monteiro F, Rodrigues H, Viggiani C, Fiorelli A, Stolf N, Kalil J. Influence of pretransplant allosensitization in cardiac transplant outcome. Transplant Proc 1999;31:2988. 7. Kobashigawa JA, Sabad A, Drinkwater D, et al. Pretransplant panel reactive-antibody screens. Are they truly a marker for poor outcome after cardiac transplantation? Circulation 1996; 94(suppl II):II-294 –7.

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8. Loh E, Bergin JD, Couper GS, Mudge GH. Role of panel reactive antibody cross reactivity in predicting survival after orthotopic heart transplantation. J Heart Lung Transplant 1994;13:194 –201. 9. Shaddy RE, Hunter DD, Osborn KA, et al. Prospective analysis of HLA immunogenicity of cryopreserved valved allografts used in pediatric heart surgery. Circulation 1996;94:1063–7. 10. Shaddy RE, Thompson DD, Osborn KA, Hawkins JA, Fuller TC. Persistence of human leukocyte antigen (HLA) antibodies after one year in children receiving cryopreserved valved allografts. Am J Cardiol 1997;80:358 –9. 11. Hawkins JA, Breinholt JP, Lambert LM, et al. Class I and class II anti-HLA antibodies after implantation of cryopreserved allograft material in pediatric patients. J Thorac Cardiovasc Surg 2000;119: 324 –30. 12. Breinholt JP, Hawkins JA, Lambert LM, Fuller TC, Profaizer T, Shaddy RE. A prospective analysis of the immunogenicity of cryopreserved nonvalved allografts used in pediatric heart surgery. Circulation 2000;102(suppl III):III-179 – 82. 13. Amos DB, Bashir H, Boyle W, MacQueen M, Tiilikainen A. A simple microcytotoxicity test. Transplantation 1969;7:220 –2. 14. Terasaki PI, Bernoco D, Park MS, Ozturk G, Iwaki Y. Microdroplet testing for HLA-A, -B, -C, and -D antigens. The Philip Levine Award Lecture. Am J Clin Pathol 1978;69:103–20. 15. Fuller TC, Fuller AA, Golder M, Rodey GE. HLA alloantibodies and the mechanism of the antiglobulin augmented lymphocytotoxicity procedure. Hum Immunol 1997;56:94 –105. 16. McKenna RM, Takemoto SK, Terasaki PI. Anti-HLA antibodies after solid organ transplantation. Transplantation 2000;69:319 –26. 17. Crew RJ, Ratner LE. Overcoming immunologic incompatibility: transplanting the difficult to transplant patient. Semin Dial 2005; 18:474 – 81. 18. Wolfe RA, Ashby VB, Milford EL, et al. Comparison of mortality in all patients on dialysis, patients on dialysis awaiting transplantation, and recipients of a first cadaveric transplant. N Engl J Med 1999;341:1725–30. 19. Rose EA, Pepino P, Barr ML, et al. Relation of HLA antibodies and graft atherosclerosis in human cardiac allograft recipients. J Heart Lung Transplant 1992;11(suppl):S120 –3.

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20. Hess ML, Hastillo A, Mohanakumar T, et al. Accelerated atherosclerosis in cardiac transplantation: role of cytotoxic B-cell antibodies and hyperlipidemia. Circulation 1983;68(suppl II): II-94 –101. 21. Petrossian GA, Nichols AB, Marboe CC, et al. Relation between survival and development of coronary artery disease and anti-HLA antibodies after cardiac transplantation. Circulation 1989;80(suppl III):III-122–5. 22. Di Filippo S, Girnita A, Webber SA, et al. Impact of ELISA-detected anti-HLA antibodies on pediatric cardiac allograft outcome. Human Immunol 2005;66:513– 8. 23. Chartrand C, Servando ES, Chartrand S. Risk factors for acute rejection after pediatric heart transplantation. Transplant Proc 2001;33:1732– 4. 24. Mahle WT, Naftel DC, Edens RE, et al. Panel-reactive antibody cross-reactivity and outcomes in the PHTS. J Heart Lung Transplant 2004;23(suppl):S167. 25. Jacobs JP, Quintessenza JA, Boucek RJ, et al. Pediatric cardiac transplantation in children with high panel reactive antibody. Ann Thorac Surg 2004;78:1703–9. 26. Xydas S, Yang JK, Burke EM, et al. Utility of post-transplant anti-HLA antibody measurements in pediatric cardiac transplant recipients. J Heart Lung Transplant 2005;1289 –96. 27. Quivers E, Miller S, Morell V, Zeevi A, Webber S. Intravenous immunoglobulin (IVIG): effective for reduction of alloantibody in sensitized heart transplantation (HT) candidates? Pediatr Transplant 2005;9(suppl 6):51. 28. Yadin O, Sekiya N, Ettinger R. Intravenous immunoglobulin (IVIg) reduces antibody responses in highly sensitized dialysis patients. J Am Soc Nephrol 1992;3:887. 29. Glotz D, Antoine C, Julia P, Pegaz-Fiornet B, et al. Intravenous immunoglobulins and transplantation for patients with anti-HLA antibodies. Transplant Int 2004;17:1– 8. 30. McIntyre JA, Higgins N, Britton R, et al. Utilization of intravenous immunoglobulin to ameliorate alloantibodies in a highly sensitized patient with a cardiac assist device awaiting heart transplantation: fluorescence-activated cell sorter analysis. Transplantation 1996;62:691–3.