Sandra Rosen-Bronson

The detection of donor-directed, HLA-specific alloantibodies in recipients of unrelated hematopoietic cell transplantation is predictive of graft failure

Donor-directed human leukocyte antigen (HLA)-specific allo-antibodies (DSAs) cause graft failure in animal models of hematopoietic stem cell transplantation (HCT). Archived pretransplantation sera from graft failure patients (n = 37) and a matched case-control cohort (n = 78) were tested to evaluate the role of DSAs in unrelated donor HCT. Controls were matched for disease, disease status, graft type, patient age, and transplantation year. Patients had acute myeloid leukemia, acute lymphoblastic leukemia, chronic myelogenous leukemia, or myelodysplastic syndrome; 98% received myeloablative conditioning regimens 100% received T-replete grafts, 97% received marrow, 95% HLA-mismatched, and 97% received calcineurin-based graft-versus-host disease prophylaxis. Among the 37 failed transplantations, 9 (24%) recipients possessed DSAs against HLA-A, B, and/or DP, compared with only 1 (1%) of 78 controls. Therefore, the presence of DSAs was significantly associated with graft failure (odds ratio = 22.84; 95% confidence interval, 3.57-infinity; P < .001). These results indicate that the presence of pretransplantation DSAs in recipients of unrelated donor HCT is associated with failed engraftment and should be considered in HCT donor selection.

Efficient cDNA expression vectors for stable and transient expression of HLA-DR in transfected fibroblast and lymphoid cells

cDNA expression vectors with several useful features were constructed. First, the long terminal repeat of Rous sarcoma virus was used as a promoter to obtain high levels of expression in various cells of human and mouse origin. Second, cis-linked expression units that confer resistance either to mycophenolic acid or the neomycin analog G418 were inserted to facitate the isolation of transfected cells expressing the cDNA of interest. Third, by replicating in simian COS cells, these vectors can be used for efficient transient expression. cDNA fragments encoding the DR alpha or DR beta chains of human class II major histocompatibility complex antigens were inserted into these vectors and high levels of cell surface HLA-DR antigen were obtained after cotransfection into mouse and human fibroblasts. These vectors were also successfully used to correct the inability of a class II-negative B cell line, derived from a patient with a congenital immunodeficiency, to present peptide antigen to DR-restricted T cells.

On the relative immunogenicity of DR alloantigens : T cell recognition of HLA-DR2a and HLA-DR2b

The HLA-DR2 haplotype encodes two highly polymorphic DR molecules, DR2a and DR2b. Because little is known regarding the relative immunogenicity of different HLA-DR molecules, we have studied the T-cell recognition of DR2a and DR2b molecules from the DRw15, Dw2 haplotype. A series of DR2-specific alloreactive T-cell clones were analyzed with murine L-cell transfectants expressing either the DR2a or the DR2b molecules as stimulator cells in proliferation assays. Somewhat surprisingly, both DR2a and DR2b were capable of stimulating DR2-specific T-cell clones with equal magnitude and similar frequency. In addition, DR2a and DR2b are functionally distinct, that is, no clone was identified which was stimulated by both DR2a and DR2b molecules.

Diverse usage of human T-cell receptor gene segments in HLA-DR1 allospecific T-cell clones

T-cell recognition of alloantigen involves both the MHC molecule and its associated peptide ligand. To understand the relationship between the specificity of alloantigen recognition and the structure of TCR molecules, we have investigated TCR gene utilization by sequencing TCR genes from well-defined allospecific T-lymphocyte clones. Alloreactive TLC consisted of a panel of clones primed to recognize DR1-related alloantigens. Our sequencing results revealed extensively diverse, but nonrandom, usage of TCR AV and BV gene segments and essentially no conservation in CDR3 or junctional sequences. Such observations are consistent with allospecific TCR that interact with MHC molecules on a generic level while recognizing specific peptides. They also reduce potential enthusiasm for anti-TCR therapy in allograft rejection.

Paired Kidney Donor Exchanges and Antibody Reduction Therapy: Novel Methods to Ameliorate Disparate Access to Living Donor Kidney Transplantation in Ethnic Minorities

BACKGROUND Currently ethnic minority patients comprise 60% of patients listed for kidney transplantation in the US; however, they receive only 55% of deceased donor renal transplants and 25% of living donor renal transplants. Ethnic disparities in access to kidney transplantation result in increased morbidity and mortality for minority patients with end-stage renal disease. Because these patients remain dialysis dependent for longer durations, they are more prone to the development of HLA antibodies that further delay the possibility of receiving a successful kidney transplant. STUDY DESIGN Two to 4 pretransplant and post-transplant plasma exchanges and i.v. immunoglobulin were used to lower donor-specific antibody levels to less than 1:16 dilution; cell lytic therapy was used additionally in some cases. Match pairing by virtual cross-matching was performed to identify the maximal exchange benefit. Sixty candidates for renal transplantation were placed into 4 paired kidney exchanges and/or underwent antibody reduction therapy. RESULTS Sixty living donor renal transplants were performed by paired exchange pools and/or antibody reduction therapy in recipients whose original intended donors had ABO or HLA incompatibilities or both (24 desensitization and 36 paired kidney exchanges). Successful transplants were performed in 38 ethnic minorities, of which 33 were African American. Twenty-two recipients were white. Graft and patient survival was 100% at 6 months; graft function (mean serum creatinine 1.4 g/dL) and acute rejection rates (20%) have been comparable to traditional live donor kidney transplantation. CONCLUSIONS Paired kidney donor exchange pools with antibody reduction therapy can allow successful transplant in difficult to match recipients. This approach can address kidney transplant disparities.

Donor transmission of pineoblastoma in a two-yr-old male recipient of a multivisceral transplant: a case report.

Zhao P, Strohl A, Gonzalez C, Fishbein T, Rosen‐Bronson S, Kallakury B, Ozdemirli M. Donor transmission of pineoblastoma in a two‐yr‐old male recipient of a multivisceral transplant: a case report.

Polymorphism of the DR1 Haplotype: Structural and Functional Analysis

HLA-DR haplotypes vary in the number of Dw subtypes defined by the mixed lymphocyte reaction (MLR). DR2 and DR4 have been shown to be quite complex (1,2) whereas DR1 is relatively homogeneous in populations of Northern European extraction (3). In Caucasians, almost all DR1 individuals express a DR1,DQw5,Dwl haplotype. The DR1 haplotype in American blacks is always associated with DQw5, but approximately 50% of DR1 positive American blacks express a Dw allele undefined by mixed lymphocyte typing (Dw blank or Dw-) (4). Undefined Dw specificities associated with DR1 have been reported in Nigerians (5) and in HLAB14-related haplotypes (6–8). It is not clear whether the DR1,Dw-haplotype found in American blacks, which is not associated with HLA-B14, is the same variant described by others. The structure and function of the American black DR1,DQw5,Dw-haplotype have been studied and compared to the Caucasian DR 1 haplotype using alloproliferative T-cell clones and cDNA sequencing.

DR3 Heterogeneity in American Blacks

The heterogeneity of DR3-associated haplotypes in American Blacks is striking when compared with that in the Caucasoid populations. In American Blacks, only 7.7% of DR3-positive individuals are Dw3. Three DR3-associated haplotypes are generally observed in the American Black population. Two appear to be unique to Blacks and are 1) A30, Bw42, Dw-, DRw18, DRw52a, DQw4, and 2) Bw53 or Bw71, Dw3v, DRw 17, DRw52b, DQw2. HLA-D typing responses for these haplotypes are Dw- (DNV>70) and Dw3v (DNV 35–55), respectively. The third DR3 haplotype is identical to the Caucasian Al, B8, Dw3, DRw17, DRw52a, DQw2.

Sensitization and Preformed Donor Specific Antibody in Intestinal Transplantation: survival and rejection outcomes

Introduction: Sensitization has historically been shown to decrease allograft survival in intestinal transplantation (ITx). Contemporary data increasingly demonstrates that donor specific antibody (DSA) is responsible for increased risk of rejection and graft loss in sensitized ITx recipients. Methods: Beginning in 2008 recipients were screened for preformed DSA prior to transplant at our institution. All ITx performed at our institution from 2008 through 2016 were included in this study and evaluated with the use of a prospectively maintained database. DSA prior to transplant was determinedwith the use of solid phase assays andwas considered positivewith amedian fluorescence intensity (MFI) >1000. Standard immunosuppression (IS) consisted of IL2 blockade induction with maintenance IS of tacrolimus, sirolimus, and prednisone. Sensitized recipients generally received thymoglobulin induction and IVIG followed by standard maintenance IS. Results: 144 ITx transplants were performed during this period (69 pediatric and 75 adult) including 101 isolated intestine, 26 liver/intestine, 13 multivisceral transplants and 4 modified multivisceral transplants. The median follow up was 56.4 months. There were 3 comparison groups: non-sensitized recipients (n=74), sensitized recipients without preformed DSA (n=57) and sensitized recipients with preformed DSA (n=13). The 13 recipients with preformed DSA included 11 isolated intestine transplants, 1 multivisceral transplant and 1 modified multivisceral transplant and 9 were adults. The mean PRA +/− SD in the sensitized group without preformed DSA was 77 +/− 28 and 36 +/− 30 in the sensitized group with preformed DSA (p<0.001). Recipients with a positive T cell and/or B cell flow crossmatch was 7% in the sensitized group without preformed DSA and 54% in the sensitized group with preformed DSA (p=0.007); no patient in the non-sensitized group had a positive flow crossmatch. In the sensitized group without preformed DSA compared to the non-sensitized group, there was no statistical difference in 1-, 3, and 5-year patient survival (72%, 65%, and 65% versus 86%, 79%, and 75% p=0.065), 1-, 3-, and 5-year graft survival (70%, 65%, and 59% versus 84%, 78%, and 69% p=0.163), and 1-, 3-, and 5-year freedom from rejection (66%, 66%, and 51% versus 77%, 77%, and 71% p=0.160). In the sensitized group with preformed DSA compared to the non-sensitized group, there was a statistically lower 1-, 3-, and 5-year patient survival (49%, 49%, and 49% versus 86%, 79%, and 75% p=0.015), 1-, 3-, and 5-year graft survival (49%, 39%, and 39% versus 84%, 78%, and 69% p=0.017), and 1-, 3-, and 5-year freedom from rejection (25%, 25%, and 25%versus 77%, 77%, and 71% p=0.0001). Conclusions: Sensitized intestine recipients with preformed DSA have an increased risk of rejection, graft loss and mortality. Avoidance of preformed DSA in sensitized recipients may improve outcomes following intestine transplant. 205.2