1. Donor selection for unrelated cord blood transplants. Gluckman E, Rocha V. Curr Opin Immunol. 2006; 18:565-70.

OVERVIEW

Hematopoietic stem cell transplantation (HSCT) from an unrelated cord blood (UCB) is now standard practice for the treatment of hematological disorders. The number of unrelated cord blood transplants is increasing, with more than 8000 patients reported worldwide.

Umbilical cord blood transplantation (UCBT) has extended the availability of allogeneic HSCT to patients who would otherwise not be eligible for this curative approach. The number of UCBTs has increased dramatically, because several studies have demonstrated that results from HLA- mismatched UCBTs were comparable to those from HLA- matched unrelated bone marrow transplants in children and in adults.

In comparison with other sources of HSCT, UCB offers logistic and clinical advantages, such as: faster availability of banked cryopreserved UCB units; extension of the donor pool owing to tolerance of 1-2 HLA mismatches; lower incidence and severity of acute GVHD; lower risk of transmitting infections by latent viruses; lack of donor attrition; lack of risk to the donor; and higher frequency of rare haplotypes.

One of the major complications of UCBT is delayed engraftment owing to the low number of cells infused, which is 10-times less than a BM collection. Study of the role of cell dose and HLA compatibility to guide transplant centers in the selection of the best unit for transplant is extremely important.

IMPACT OF CELL DOSE AND HLA MATCHING ON OUTCOMES

The Eurocord group has recently analyzed the impact of diagnosis, cell dose and HLA incompatibilities in patients receiving a single UCBT in 925 patients with malignant disease and 279 patients with non-malignant disease. Donor-recipient histocompatibility was determined by serology or antigen typing for HLA- and HLA-B and by DNA typing for HLA-DRB1.

Patients with malignant diseases. The median number of nucleated cells (NCs) infused was 3.1 x 10⁷ NC/kg (range 2-5 x 10⁷ NC/kg). At day 100, the cumulative incidence (CI) of neutrophil recovery was 77.4% and of platelet recovery to 20,000/µl was 54.7%. Engraftment was associated with the number of cells infused (p<0.0001). HLA was a second factor, with a difference between 0-1 (81%), 2 (75%) and 3-4 (63%) HLA incompatibilities (p=0.037). There was no correlation between engraftment and the type of HLA mismatches (class I or class II, or HLA-A, -B or –DRB1).

The CI of aGVHD grade II-IV was 33% and the CI of aGVHD grade III-IV was 17.3%; it was not associated with the number of cells infused or with the number of HLA disparities. The CI of cGVHD was 28.6% and it was increased in the group who received a low cell count or a 3-4 HLA-mismatched transplant.

The CI of relapse was 23.1% at 100 months and it was not associated with the number of cells infused, whereas increasing the number of HLA disparities decreased the relapse incidence.

The CI of overall survival (OS) and disease-free survival (DFS) at 100 months was 33.4% and 30.8%, respectively. Patients who received <3 x 10⁷ NC/kg at freezing of cord blood unit or ≤2.1 x 10⁷ NC/kg at thawing had decreased probability of survival The number of HLA incompatibilities had no influence on OS or DFS. This was explained by the positive impact of the number of HLA mismatches on relapse and the negative impact on engraftment.

In summary for patients with malignant diseases: Cell dose was the most important factor for outcome; a minimum cell dose of 3 x 10⁷ NC/kg at collection and of 2 x 10⁷ NC/kg at infusion must be targeted. The number of HLA mismatches increased the risk of delayed engraftment and led to a higher incidence of TRM and cGVHD; however, it decreased the risk of relapse, resulting in an absence of the role of HLA mismatches for OS and DFS. The type of HLA mismatch did not influence outcomes, but matching for HLA-DRB1 seems better for patients receiving a graft that has two HLA incompatibilities. Increasing cell dose abrogated the effect of HLA mismatches, but not for grafts with 3 or 4 HLA incompatibilities.

Patients with non-malignant diseases. The median number of NCs infused was 6.4 x 10⁷ NC/kg (range 0.8-66 x 10⁷NC/kg). At day 100, the CI of neutrophil recovery was 69.3% and CI of platelet recovery was 50%. Both neutrophil and platelet recoveries were associated with the median number of cells infused (p<0.000055). HLA was also an important factor associated with neutrophil recovery, with a statistical difference between 0-1 and ≥2 HLA mismatches (p=0.046). The role of HLA mismatches was abrogated by increasing cell dose, except in the group of patients who received a 3-4 HLA-mismatched transplant. There was no correlation between neutrophil recovery and the class of HLA mismatches (class I or class II, or HLA-A, -B or –DRB1).

The CI of aGVHD grade II-IV was 31.8% and grade III-IV 18%, and was only associated with the number of HLA incompatibilities (p=0.0029). The CI of cGVHD was 24% and was also associated with the number of HLA incompatibilities (p=0.01).

The CI of OS at 100 months was 49% and it was influenced by cell dose and by the number of HLA mismatches. The group who received an UCBT with <3.5 x 10⁷ NC/kg at infusion and a 2-3 HLA-mismatched transplant had <20% survival. Increasing cell dose partially abrogated the effect of HLA mismatches.

In summary for patients with malignant diseases: Patients with non-malignant diseases must receive a higher cell dose to obtain engraftment than patients with malignant disorders; it should not be below 4.9 x 10⁷ NC/kg at collection and 3.5 x 10⁷ NC/kg at infusion. In non-malignant disease, HLA mismatches played a major role for engraftment, GVH and survival; it is partially abrogated by increasing cell dose, and two HLA-DRB1 mismatches seemed to have an adverse effect.

OTHER FACTORS

The role of CD34+ cells: Several authors have shown that the number of CD34+ cells in the transplant correlated with engraftment. Unfortunately, this measurement cannot be used for comparative studies between centers because of the absence of standardization of the counting method.

Colony forming units (CFU-GM) is another method for counting the stem cell content. However, there is too much inter-laboratory variability.

Role of allelic typing in UCBT outcomes. An analysis of 122 patient-CB donor combinations using high-resolution (HR) typing for both HLA class I and class II loci showed no statistical association between the number of HLA mismatches and neutrophil recovery; however, based on HLA HR typing in the rejection direction (donor alleles not shared by the recipient), 1 or 2 HLA-A HR disparities were associated with reduced incidence of engraftment (p=0.04). An analysis of the influence of single alleles on the subtyping level in GVHD direction only for the HLA-B (comparing 0 mismatches versus 1 and 2 locus mismatches), indicated that incompatibilities in the HLA-B locus were associated with the incidence of a GVHD grade II-IV (p=0.04). In this study, there was no benefit of additional HR typing for HLA-A, -B, -C, -DR and –DQ for outcomes after UCBT.

Current recommendations for choosing the bests CB unit. In order of preference: 6/6 matched CB and >3 x 10⁷NC/kg; 5/6 matched CB and >4 x 10⁷ NC/kg; 4/6 matched CB and >5 x 10⁷ NC/kg. Use of units with <3 x 10⁷ NC/kg and/or 3-4 HLA mismatches is not recommended; in this case a double CBT might be considered. (See Annotated Bibliography, Category III, Multi-Cord Transplants)

2. Unrelated donor hematopoietic cell transplantation: marrow or umbilical cord blood? Grewal SS, Barker JN, Davies SM, Wagner JE. Blood 2003; 101:4233-4244.

The authors report that the single most important factor influencing time to hematopoietic recovery appears to be the nucleated cell count of the graft relative to the recipient size. They suggest that a minimum acceptable nucleated cell dose should be 1.5 X 10⁷ nucleated cells/kg to reduce time to myeloid recovery and increase the probability of engraftment.

3. Umbilical cord blood transplantation: where do we stand? Wadlow RC, Porter DL. Biol Blood Marrow Transplant 2002; 8:637-647.

The authors state that available data suggest that nucleated cell dose should be the primary criterion for donor selection, and they point out that recently recommended doses include 1.5 X 10⁷ and 2.0 X 10⁷ nucleated cells/kg recipient body weight.

4. Transplantation of unrelated donor umbilical cord blood in 102 patients with malignant and nonmalignant diseases: influence of CD34 cell dose and HLA disparity on treatment-related mortality and survival. Wagner JE, Barker JN, DeFor TE, Baker KS, Blazar BR, Eide C et al. Blood 2002; 100:1611-1618.

Data on 102 patients (median age 7.4 years) indicated that CD34 cell dose was the one factor consistently identified as significantly associated with the rate of engraftment, treatment-related mortality and survival. They concluded that there is a high probability of survival in recipients of cord blood grafts that are disparate in no more than 2 HLA antigens when the grafts contain at lease 1.7 X 10⁵ CD34+ cells/kg recipient body weight.5. Cell dose and speed of engraftment in placental/umbilical cord blood transplantation: graft progenitor cell content is a better predictor than nucleated cell quantity. Migliaccio AR, Adamson JW, Stevens CE, Dobrila NL, Carrier CM, Rubinstein P. Blood 2000; 96:2717-2722.

The posttransplant course of 204 patients who received grafts evaluated for hematopoietic colony-forming cell (CFC) content were analyzed using univariate and multivariate life-table techniques. Neutrophil and platelet engraftment were shown to correlate with the cell dose, whether estimated as total nucleated cell (TNC) or CFC per kilogram of recipient’s weight. The CFC content of umbilical cord blood units was associated more rigorously with the major covariates of posttransplantation survival than the TNC.

5. Principles and tools for selection of umbilical cord blood and unrelated adult donor grafts. Lee SJ, Kamani N, Confer D. Biol Blood and Marrow Transplant 2008;14(Suppl 1):112-119.

The impact of HLA matching on major outcomes in UCB transplantation continues to be refined. TNC was previously thought to be the sole determinant of outcome with partially HLA matched UCB transplantation, but HLA matching, particularly at low TNC dose, appears to play an important role. Relatively small sample sizes limit the consistency of findings from cord blood studies, but the consensus supports consideration of both TNC and HLA matching in the selection of optimal UCB units.

UCB graft cell dose is a critical determinant of hematopoietic recovery and survival after UCBT, but there is increasing evidence that HLA match is also a key factor in outcome. Current data suggest that HLA match is critically important in the setting of a low cell dose. The complete elucidation of the impact of graft dose and match on UCBT outcome will require the analysis of larger cohorts of patients.

The impact of a 1 versus 2 antigen mismatch, the “trade-off” between HLA-match and cell dose in unit selection, the importance of allele-level matching at HLA-A and B, the match vector, and whether HLA-C or DQB1 should be considered in the selection of UCB units for transplantation cannot be fully discerned at the current time.

Currently, the NMDP recommends typing all loci (A, B, C, DRB1 and DQB) at the allele level. Matching should follow the current standard of intermediate level for HLA-A and -B, and allele level for HLA-DRB1. All recommendations are based on selection of a unit with an appropriate cell dose. Currently available data would consider this to be a unit that has >2.5-3.0 x 107 TNC/kg recipient body weight.

Tools for locating the optimal adult donor or UCBA UNIT: NMDP provides software to transplant centers (TRANS Link®) that allows real-time searching of the complete NMDP inventory of adult donors and UCB units. TRANS Link produces a donor/UCB list that is sorted with the best of potential donors/UCB at the top. An NMDP innovation named HapLogic^sm predicts the likelihood of allele-level matching based on calculated HLA haplotype frequencies within major racial and ethnic populations. Another NMDP software product is the Multi-Cord Tool which evaluates HLA matching between the proposed recipient and the UCB units as well as matching between UCB units. The tool was developed to simplify the UCB selection process for double cord transplantation.

[For more details, also see the editorial by Kamani et al (Annotated Bibliography, V. HLA matching, citation #6]

6. State of the art review: HLA matching and outcome of unrelated donor umbilical cord blood transplants. (Editorial) Kamani N, Spellman S, Hurley CK, Barker J, Smith FO, Oudshoorn M, et al Biol Blood and Marrow Transplant 2008;14:1-6.

This editorial summarizes current data and provides guidance in selecting UCB units for unrelated donor UCBT based on HLA matching. Of note is the fact that much of the data on the impact of HLA match on UCBT is currently preliminary or inconclusive so that guidelines will be refined over time. Also to be noted is that studies by various investigators often result in conflicting conclusions.

When reviewing the CB transplant literature, it is important to note that the majority of UCBTs reported have utilized matching at the antigen level for HLA-A and HLA-B loci and at the allele level for HLA-DRB1 loci.

Impact of HLA matching on engraftment after UCBT. In the largest analysis done to date on 562 UCBT recipients, Rubinstein et al have shown that there is a progressive delay in myeloid engraftment by day 42 with increasing HLA mismatching. However, the authors could not detect a significant difference between those who received grafts mismatched at 1 or 2 antigens.

A study by Eapen et al compared outcomes of 503 UCBTs and 282 UBMTs in children <16 years with acute leukemia to assess the influence of both cell dose and HLA matching on outcome. These authors found that higher cell doses resulted in a higher probability of both neutrophil and platelet recovery in 1 antigen-mismatched UCBT but had no effect in 2-antigen mismatched transplants, suggesting that cell dose may not be able to overcome the adverse impact of mismatching in the setting of 4 out of 6 matched UCBT. However, an EBMT study was unable to demonstrate a statistically significant interaction between the cell dose and HLA disparities for either neutrophil or platelet recovery.

In summary, whereas cell dose is important for engraftment in UCBT, HLA match also appears to impact the engraftment rates.

Impact of HLA matching on GVHD. A Eurocord study showed that whereas the degree of matching did not impact overall GVHD risk, the risk of grade III-IV GVHD was higher in recipients of UCB where class I and II disparities coexisted between donor and recipient. A COBLT study indicated that HLA matching impacted grade II-IV risk with a significantly higher risk of aGVHD in recipients of 4/6 matched UCB compared to 5-6/6 matched UCB. A study by Eapen et al failed to show a correlation between HLA match and aGVHD or cGVHD risk among recipients of UCB transplants.

Role of HLA compatibility on survival after UCBT. Only a few reports of UCBT have analyzed the effects of HLA mismatching on patient survival. Studies by Rubinstein et al and Wagner et al indicated an association of HLA-mismatching with the outcome of UCBT. In contrast, a study of 550 UCBTs by Eurocord failed to find an effect of HLA disparity on 3-year survival. In the Eurocord analysis of patients with malignant diseases, increasing HLA disparity resulted in lower rates of engraftment and higher treatment-related mortality and cGVHD, but this did not translate into a lower disease-free survival or overall survival because of a lower relapse rate. HLA disparity did impact survival for patients with nonmalignant diseases.

High-resolution HLA matching. The impact of high-resolution HLA matching in UCBT has been investigated in a number of papers. Studies by Ohnuma et al and Cornetta et al suggested that high-resolution HLA typing may help select UCB units better suitable for a favorable outcome. However, Kogler et al retrospectively typed 122 UCBT donor and recipient pairs by high resolution at HLA-A, -B, -C, -DRB1, and –DQB1 and failed to show any association with survival in univariate analysis.

Summary. Although UCB graft cell dose is a critical determinant of hematopoietic recovery and survival after UCBT, there is increasing evidence that HLA match is also a key factor in UCBT outcome with mismatch adversely impacting on both engraftment and survival. It is not currently known how cell dose and HLA match interact and it is not clear whether the same criteria will apply to both pediatric and adult recipients.

All recommendations are based on selection of a unit with an appropriate cell dose. Currently available data would consider this to be a unit that has >2.5-3.0 x 10⁷ total precryopreserved nucleated cells per kg recipient body weight. The interaction between cell dose and HLA matching is especially critical in adults undergoing UCBT and underlines the need for drastic increases in the unrelated cord blood inventory.

7. Hematopoietic stem-cell transplants using umbilical-cord blood. Gluckman E. N Engl J Med 2001; 344:1860-1861.

Eurocord data indicate that recipients of <1 X 10⁷ nucleated cells/kg had a transplant related mortality of 75%, whereas patients infused with at least 3 X 10⁷ nucleated cells/kg had a transplant related mortality of only 30%.

8. Current status of umbilical cord blood hematopoietic stem cell transplantation. Gluckman E. Exp Hematol 2000; 28:1197-1205.

Among other conclusions in this review is the recommendation that umbilical cord blood units should be selected on the basis > 4 X 10⁷/kg recipient body weight before thawing.

(NOTE: This article suggests a higher TNC number than is suggested by a number of other authors. The evaluation of the effects of cryopreservation, storage and thawing are still uncertain, and are likely to be variable among cord blood banks and transplant centers. These factors need further study before definitive conclusions on appropriate cell dose can be reached.)

9. Pre-transplant prognostic factors for patients with high-risk leukemia undergoing an unrelated cord blood transplantation. Iori AP, Cerretti R, De Felice L, Screnci M, Mengarelli A, Romano A, Caniglia M, Cerilli L, Gentile G, Moleti ML, Giona F, Agostini F, Pasqua I, Perrone MP, Pinto MR, Grapulin L, Testi AM, Martino P, De Rossi G, Mandelli F, Arcese W. Bone Marrow Transplant. 2004;33:1097-105.

This is a report of long-term outcomes and pre-transplant prognostic factor analysis for 42 consecutive patients (median age 12, range 1-42) with high-risk leukemia prospectively undergoing an unrelated cord blood transplants in two centers participating in the same transplant program.

At 90 days after transplant, all the evaluable patients recovered hematopoiesis, which was of full donor origin in 92%. The authors suggested that the exclusion of methotrexate from their regimen for GVHD prophylaxis might have contributed to facilitate the hematopoietic reconstitution. However, because of prolonged aplasia, G-CSF was administered to 13 of 39 (33%) evaluable patients and 12 recovered hemopoiesis, which was of donor origin in 10.

The authors were unable to find any statistically significant factor affecting myeloid engraftment, but patients who received a higher (p=0.07) number of CFU-GM reconstituted full chimeric hematopoiesis significantly faster than the other patients.

To prevent GVHD, only cyclosporine and low dose 6-MPr were used. The incidence of either advanced acute (II-IV grade: 21%; III-IV grade: 9.5%) or extensive chronic (20%) was considerably lower than that observed in children grafted from an unrelated HLA matched bone marrow donor, and compares favorably with that reported from other studies in patients, mainly pediatric, undergoing an unrelated cord blood transplant. They did not find a relation between the number of HLA disparities between donor and recipient and the incidence and severity of either acute or chronic GVHD.

Relapse occurred in only nine patients, six of whom were transplanted in advanced phase, for a cumulative relapse incidence of 25% at 4 years. Thus the graft-versus-leukemia effect does not seem to be impaired after cord blood transplantation despite the low incidence of GVHD.

Life threatening infections were less than expected: only two patients in the series died of bacterial sepsis.

Despite a significant correlation with the number of CD34+ cells and total nucleated cells infused, the number of CFU-GM emerged as the principal variable predicting patient outcomes.

In conclusion, the authors suggested that a cord blood unit for transplant should be selected on the basis of its proliferative potential, but pointed out that, at present, there is limited availability of this information in the cord blood banks and there is an absence of standardized methods for in vitro assays.