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Canoga Park, CA, United States

Sasaki N.,One Lambda Inc.
Clinical transplants | Year: 2010

Almost all the HLA-matching effects found by the 2000 analysis were confirmed by this study. The only HLA-matching effect found in the 2000 analysis that disappeared were those of "small matching effect" found in sub-populations of type I diabetes (PRA < 10%, donor age 20-35). The 2000 analysis found a lack of HLA matching effect in non-African American kidney transplant patients with type I diabetes between 1987 and 2000. The 2000 analysis found that a patients' ethnic group was a factor in graft survival; African American patients were found to have a significantly lower 10-year graft survival in the 5 or 6 mismatched group (27%) compared to Caucasian patients (40%). In addition, Asian patients (42%) had higher graft survival compared to that of Caucasian patients. In this study, we observe a similar pattern with death-censored graft analysis for all ethnic groups with 10-year graft survivals at 72.9% for Asians, 69.5% for Caucasians, and 49.3% for African Americans. There was an overall lack of HLA-matching effect on patient survival in the 2000 analysis. In our current analysis, the patient survivals remained virtually the same despite moderate increase in graft survival over the same period of time. The HLA-C locus mismatch was found to have additive effect to the 10-year graft survival trends observed in A and B mismatch cases. HLA-DQ mismatch on the other hand, showed limited HLA-matching effect and did not show the same additive effect as C. There are various possible issues in the DQ mismatch analysis, from the consistency of DQ typing results, lack of diversity in the DQ antigen, to the possibility of DQ mismatch having little effect on the graft survival. Utilizing kidney transplant cases performed from 1995 through 2000, the 2000 analysis projected 10-year survivals of 64% and 47% for the 0 ABDR mismatch and 5 or 6 ABDR mismatched cases respectively; the 2000 projection only missed actual death-censored survivals by 9% lower for the 0 mismatch and 17% lower for the 5 or 6 mismatch cases. Utilizing the transplant cases of 2005 through 2009, we projected their 10-year graft survivals for year 2020. The 10-year graft survival for 0 ABDR mismatched patients is expected to be over 85% and nearly 70% for 5 or 6 ABDR mismatched patients. The general upward trend of graft survival we have observed in the last 10 years has been dependent upon the development of novel transplant protocols and use of novel immunomodulatory reagents. This trend is likely to continue given the promise of new drugs and personalized healthcare. The decreasing range of the differences in the 10-year graft survival between best matched and worst matched HLA groups is also likely to continue. One interesting trend that is clearly evident is the increasing difference between the best and worst HLA-matching in terms of the associated graft half-life. The positive HLA-matching effect on long-term graft survival is clearly evident and should be taken into consideration for all kidney transplants.

Everly M.J.,One Lambda Inc.
Clinical transplants | Year: 2011

Based on our knowledge that donor specific anti-HLA antibodies (DSA) are a major cause of allograft loss, determining how to monitor patients for DSA and how to treat them is important. Current published studies indicate that patients with preformed DSA differ from those without. Approximately 15-18 percent of transplant patients will have preformed DSA, which increases risk for early antibody mediated rejection (AMR) and allograft loss. The fact that nearly all AMR episodes occur in the first 1-2 months, coupled with the finding that a reduction in preformed DSA intensity within the first few weeks post-transplant decreases the risk of AMR, makes early testing important. It has also been shown that clearance of DSA at 6 months and 1 year can result in a decreased risk of transplant glomerulopathy and therefore, these times may be prime testing points. This monitoring schedule differs slightly from that of the patients who do not have performed DSA (i.e. low risk patients). Low risk patients who develop de novo DSA are most likely to do so in the first 6 months. However, more frequent sampling in the early months does not improve predictability of acute rejections in low risk patients and therefore, it is not as essential. Rather, testing at 6 months and then annually or biannually, would be beneficial, as it would serve to identify the 5 percent of new patients who develop DSA annually. Once these patients are identified, studies have shown that preemptive treatment to a goal of antibody clearance can be used to improve graft function and survival. In addition to screening for new DSA, monitoring for clearance of DSA along with histologic reversal of rejection in patients with AMR is important. In sum, there is substantial evidence suggesting that all patients need to have some monitoring for DSA to identify new onset of DSA or clearance of DSA. Additionally, in all DSA scenarios, treatment of persistent DSA is important, as it can lead to improved allograft survival.

Ravindranath M.H.,Terasaki Foundation Laboratory | Pham T.,Terasaki Foundation Laboratory | El-Awar N.,One Lambda Inc. | Kaneku H.,Terasaki Foundation Laboratory | Terasaki P.I.,Terasaki Foundation Laboratory
Molecular Immunology | Year: 2011

HLA-E shares several peptide sequences with HLA-class Ia molecules. Therefore, anti-HLA-E antibodies that recognize the shared sequences may bind to HLA-class Ia alleles. This hypothesis was validated with a murine anti-HLA-E monoclonal antibody (mAb) MEM-E/02, which reacted with microbeads coated with several HLA-B and HLA-C antigens. In this report, the hypothesis was reexamined with another mAb 3D12, considered to be specific for HLA-E. The antibody binding is evaluated by measuring mean fluorescence index [MFI] with Luminex Multiplex Flow-Cytometric technology. The peptide-inhibition experiments are carried out with synthetic shared peptides, most prevalent to HLA-E and HLA-Ia alleles. The results showed that mAb 3D12 simulated MEM-E/02 in recognizing several HLA-B and HLA-C antigens. Both 3D12 and MEM-E/02 did not bind to HLA-A, HLA-F and HLA-G molecules. As observed with MEM-E/02, binding of 3D12 to HLA-E is inhibited by the peptides sequences 115QFAYDGKDY123 and 137DTAAQI142. Decrease in binding of mAb 3D12 to HLA class Ia, after heat treatment of antigen coated microbeads, supports the contention that the epitope may be located at the outside of the "thermodynamically stable" α-helix conformations of HLA-E. Several sequence and structure-based web-tools were employed to validate the discontinuous epitopes recognized by the mAbs. The scores obtained by these web-tools distinguished the shared peptide sequences that inhibited the mAb binding to HLA-E. Furthermore, ElliPro web tool points out that both mAbs recognize the conformational discontinuous epitopes (the shared inhibitory peptide sequences) in the secondary structure of the HLA-E molecule. The study favors the contention that the domain of the shared inhibitory peptide sequences may be the most immunogenic site of HLA-E molecule. It also postulates and clarifies that amino acid substitution on or near the binding domains may account for the lack of cross reactivity of 3D12 and MEM-E/02 with HLA-A, HLA-F and HLA-G molecules. © 2010 Elsevier Ltd.

Ravindranath M.H.,Terasaki Foundation Laboratory | Kaneku H.,Terasaki Foundation Laboratory | El-Awar N.,One Lambda Inc. | Morales-Buenrostro L.E.,National Institute of Medical science and Nutrition Salvador Zubiran | Terasaki P.I.,Terasaki Foundation Laboratory
Journal of Immunology | Year: 2010

Natural anti-HLA Abs found in sera of healthy nonalloimmunized males recognize HLA-Ia alleles parallel to those recognized by anti-HLA-E mAbs (MEM-E/02/06/07). Therefore, some of the HLA-Ia Abs seen in healthy males could be due to anti-HLA-E Abs cross-reacting with HLA-Ia. If anti-HLA-E Abs occur in healthy nonalloimmunized males, it can be assessed whether they evoke HLA-Ia reactivity as do mouse HLA-E mAbs. IgG and IgM Abs to HLA-E and HLA-Ia alleles are identified in sera of healthy males using microbeads coated with recombinant denatured HLA-E or a panel of rHLA-Ia alleles. The pattern of allelic recognition is comparable to that of anti-HLA-E mAbs. Sixty-six percent of the sera with HLA-E IgG have a high level of HLA-Ia IgG, whereas 70% of those with no anti-HLA-E Abs have no HLA-Ia Abs. HLA-E IgM/IgG ratios of sera are divided into four groups: IgMLow/IgGLow, IgMHigh/ IgGLow, IgMHigh/IgGHigh, and IgM Low/IgGHigh. These groups correspond to anti-HLA-Ia IgM/IgG ratio groups. When HLA-E IgM and IgG are absent or present in males, the IgM or IgG of HLA-Ia are similarly absent or present. The mean fluorescent intensity of HLA-Ia Abs correlates with that of anti-HLA-E Abs. Most importantly, HLA-E and HLA-Ia reactivities of the sera are inhibited by the shared, but cryptic, peptide sequences 117AYDGKDY123 and 137DTAAQIS143. Therefore, Abs to the H chain of HLA-E may be responsible for some of the HLA-Ia allele reactivity of the natural HLA-Ia Ab in human sera. Absence of any anti-HLA-Ia Abs in 112 nonvegans and the presence of the same in vegans suggest that dietary meat proteins might not have induced the natural allo-HLA Abs. Copyright © 2010 by The American Association of Immunologists, Inc.

Sasaki N.,One Lambda Inc.
Clinical transplants | Year: 2011

This study began with the 2010 UNOS data-set of 181,653 deceased donor kidney transplant cases and 92,577 living donor cases. Cases with ambiguous or missing HLA typing were excluded, and the remaining cases were split into subgroups by the number of previous transplants and ethnic groups of donor-patient pairs. 41,128 Caucasian donor-patient pairs that were primary living-donor transplant cases were used as the pilot population to identify potential epitope groups that have a negative effect on graft outcome. Sixty four of the most common HLA-A and -B antigens were selected. Amino acid sequences of the most frequently corresponding allele in the Caucasian population were used to build the starting theoretical epitope table. Amino acids of the 115 polymorphic positions, analyzed in one, two or three positions, resulted in 15,801,920 combinations. After eliminating combinations shared by no allele or by all 64 alleles, the table was trimmed to 1,635,044. Grouping combinations according to their antigen list (antigens that share the combinations/epitopes), 40,830 epitope groups were left. Based on the distances between amino acid positions of each epitope, and the requirement that each epitope must be shared by at least one allele, but not all 64, the number of theoretical epitopes was reduced to 39,670 and 3,703 epitope groups of unique antigen lists. The pilot population was composed of 41,128 primary living-donor transplants with Caucasian donor-patient pairs. For each of the 40,830 epitope groups in the non-distance-restricted table, 15-year death-censored graft survival was computed for epitope-group mismatches--i.e., cases with a BMQ0001 mismatch, with a BMQ0002 mismatch, etc.. Results were compared, using the log rank test, with average graft survival. Of the 3,703 epitope groups, 2,487 appeared in over 1000 cases, but only 88 of them had significant p-values, which ranged from 0.006 to 0.049, with 76 of the 88 significantly below average, 12 above average (Fig. 1). We then ran survival analyses taking the 76 epitope groups that were below average two at a time--i.e., cases with mismatches of the first and second epitope group, the first and third, first and fourth, etc. Of more than 2,500 pairs, 148 resulted in significantly (p < 0.01) lower than average survival in those primary living-donor cases. The effect of the 76 epitope-group mismatches that showed below-average results was then analyzed for other transplant populations--Caucasian donor-patient pair cases with deceased donors , Caucasian donor-patient pair cases with re-transplant living donors, Caucasian donor-patient pair cases with re-transplant deceased donors, and African-American donor-patient pair cases with primary living donors. None of these four populations exhibited any significant effect due to the 76 epitope- group mismatches. Likewise, the effect of the 148 epitope-group combination mismatches detailed in paragraph 5, above, was analyzed for the other four other transplant populations, detailed in paragraph 6. That significant effect was absent in all four. The analyses were repeated on the 40,830 epitope groups without the 27 angstrom distance constraint. Of the 40,830 epitope groups, 439 exhibited a significantly lower 15-year graft survival, with p-value ranging from 0.0053 to 0.0498. Again, none of these had any negatively significant effects on graft survival for the other four transplant populations. In the pilot population, the negative effect of the epitope group mismatches was clearly seen, but the significant differences did not carry across to the other four populations. That absence may be partially explained by the allele level differences in the HLA-A and -B typing of the donors and patients. Past studies indicate that the appearance of DSA has a negative effect on the graft outcome, so the mismatch of epitopes recognized by these DSA could also have similar negative effects. With the data available at present, and with the currently available assays for antibody detection, we are not able to analyze the impact of specific epitope mismatches. We will need the development of new methods of antibody detection that specifically indicate the exact epitope to which an antibody binds before we can continue this effort to determine the negative impact on graft survival due to epitope mismatches.

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