We compared 43 patients with an HLA-DRB1 allele-mismatched (DRB1-MM) donor (5/6 match) with 86 patients with an HLA-A, -B, and -DRB1 matched unrelated donor (6/6 MUD). The effect of an additional HLA-C mismatch was also studied. The two groups were well matched for all major characteristics such as year of HSCT, age, disease, disease risk index (DRI), stem cell dose, and conditioning.

The 5-year overall survival was 52% and 62% (p=0.27), and relapse-free survival was 42% and 47% (p=0.18) in DRB1-MM and MUD patients, respectively. Non-relapse mortality at one year was 12% and 15%, respectively (p=0.55). The cumulative incidence of relapse at 5 years was 46% in DRB1-MM patients and 35% in MUD patients (p=0.03). However, additional HLA-C mismatch affected the result negatively in the DR-mismatch cohort but not in the MUD cohort.

The results suggest that an isolated HLA-DRB1 allele mismatch is acceptable but that additional mismatches may be detrimental.

We included all the patients with a haematological malignancy who were consecutively treated with hematopoietic stem cell transplantation (HSCT) at Karolinska University Hospital with an HLA-A and -B identical but HLA-DRB1 allele-mismatched unrelated donor between 1994 and 2015. Most patients had acute myeloid leukaemia (AML) (n=18) or acute lymphoid leukaemia (ALL) (n=10). Other diagnoses were myelodysplastic syndrome (MDS) in four patients, chronic myeloid leukaemia (CML) in 5, lymphoma in 5, and rhabdomyosarcoma in 1 (Table 1). For disease-risk index (DRI) classification, disease indication in combination with disease stage at time of HSCT (and cytogentic data for AML and MDS patients) were used to classify patients into the low (n=5, 12%), intermediate (IM, n=21, 49%) or high risk (n=17, 40%) patient cohorts according to Armand et al.9.

The study was approved by the Research Ethics Committee of Karolinska Institutet (DNR 425/97). The procedures were in accordance with the Helsinki Declaration.

As controls, we selected patients transplanted using an HLA-A, -B, and -DRB1 matched (at allele level) unrelated donor (MUD). The controls were matched for age, diagnosis, disease stage, year of HSCT, and conditioning. Two controls were selected for each study patient. By DRI, there were 11 low-risk patients (13%), 49 intermediate-risk patients (57%), and 26 high-risk patients (30%) in the control group.

All patients and donors were typed using high-resolution molecular typing (i.e. PCR-SSP) for both HLA class I and II genes at allele level (at two fields level). All patient-donor pairs have recently been retrospectively re-typed.

Conventional myeloablative conditioning (MAC) was given to 82 patients and consisted of cyclophosphamide (Cy), 60 mg/kg for two days, in combination with busulphan (Bu), 4 mg/kg for 4 days (n=32), or total-body irradiation (TBI) at 4 × 3 Gy (n=49). Reduced-intensity conditioning (RIC) consisted of fludarabine, 30 mg/m²/day for 5‒6 days, in combination with Bu, 4 mg/kg/day for two days (n=27), Cy at 120 mg/kg (n=3), TBI at 2 × 3 Gy and Cy at 30 mg/kg for two days (n=12), or treosulphan at 12 g/m² for 3 days (n=6).10.

Most patients (n=126) were treated with anti-T-cell serotherapy before HSCT, with the last dose being given on the day before infusion of the graft. Serotherapy consisted of Thymoglobulin (n=111) (Genzyme, Cambridge, MA, USA), 2 mg/kg/day for 2‒5 days, Alemtuzumab (n=8) (Genzyme), 30 mg, ATG-F (n=3) (Frezenius, Gräfelfing, Germany) at 5 mg/kg/day for five days, or OKT-3 (n=4) at 5 mg/day for 5 days11. The dose of Thymoglobulin has been gradually reduced during the years as a result of continuous development of treatment protocols11. The choice of ATG brand depended on availability and any possible allergy against rabbit.

Most patients (n=110) were given cyclosporine (CsA) and methotrexate (MTX) as prophylaxis against GVHD. During the first month, CsA levels in blood were kept at 150‒200 ng/mL. In the absence of GVHD, the aim was to continue CsA at four to five months. Tacrolimus (Tac) and sirolimus were given to 15 patients, and CsA + prednisolon and Tac + MTX to one patient each. Two patients received a T-cell-depleted graft.

Both acute and chronic GVHD were diagnosed on the basis of clinical symptoms and/or biopsies from the skin, liver, gastrointestinal tract, or oral mucosa according to standard criteria121314. The patients were treated for grade-I acute GVHD with prednisolone, starting at 2 mg/kg/day with tapering after the initial response. In more severe cases, ATG, methylprednisolone, Infliximab, MTX, mesenchymal stromal cells (MSCs), and―more recently―placenta-derived decidual stromal cells were used15161718.

The source of graft was peripheral blood stem cells (PBSCs) in 83 cases and bone marrow (BM) in 46 cases. Stem cells were mobilized with subcutaneous G-CSF daily for 4‒6 days before aphaeresis to obtain PBSCs19.

Supportive care has been described in detail previously2021.

Relapse was defined as recurrent appearance of disease after complete remission, or disease progression after partial remission or stable disease. Cytomegalovirus (CMV) infection was defined as > 1,000 CMV DNA copies/mL of whole blood detected by PCR and it was pre-emptively treated using ganciclovir or foscarnet22. CMV disease was defined by the presence of symptoms combined with the detection of CMV in the affected organ. The diagnosis of post-transplantation lymphoproliferative disease (PTLD) was made according to the histological criteria reported for B-cell lymphoproliferative states following transplantation23.

Overall survival at 5 years was 62% (95% CI 50‒72%) in MUD patients and 52% (36‒66%) in DRB1-mismatched (DRB1-MM) patients (HR=1.37, 95% CI 0.78‒2.38, p=0.27). In the DRB1-MM cohort, an HLA-C mismatch affected survival, which was 71% (47‒86%) in C-matched patients and 33% (14‒52%) in C-mismatched patients, (HR=3.61, 95% CI 1.38‒9.43, p<0.01) (Figure 1A). No effect of HLA-C mismatch was seen in the MUD cohort (Table 2). NRM was 15% (7‒23%) and 12% (4‒23%) at one year in the two groups (RH=0.73, 95% CI 0.42‒1.28, p=0.55), respectively. No significant effect of HLA-C mismatch on NRM was seen in the two groups (Figure 1B, Table 2). Causes of death in the MUD group were relapse in 18 cases (21%), infection in 12 cases (14%), and GVHD in three cases (3%). In the DRB-MM cohort, 15 patients (35%) died of relapse, 4 (9%) died of infection, and one died of GVHD.

Donor age (HR=1.04, 95% CI: 1.01‒1.08, p=0.02) was the only factor significantly associated with mortality in multivariate analysis, while having an HLA-DRB1 and HLA-C mismatched donor was borderline-significant (HR=1.88, 95% CI: 0.89‒3.94, p=0.08) (Table 3). A donor age of > 45 years showed the worst OS (n=18, 28%), and there was a correlation between younger donor age and better OS (donor age 26‒45 years: 63%; donor age 19‒25 years: 77%).

The cumulative incidence of relapse at 5 years was 35% (25‒46%) in the MUD patients and 46% (30‒61%) in the HLA-DRB1-MM patients (1.75, 1.03‒3.13, p=0.030). In the DRB1-MM cohort, relapse was 33% (14-54%) or 60% (34-79%) depending on whether an HLA-C match or an additional HLA-C mismatch was present (HR=2.21, 95% CI 0.93‒5.24, p=0.075) (Figure 1).

In multivariate analysis, there were correlations between relapse and an HLA-DRB1 and -C mismatched donor (HR=2.92, 95% CI 1.36‒6.26, p=0.006), DRI high (HR=2.10, 95% CI: 1.09‒3.70, p=0.025), and higher total nucleated cell (TNC) dose (HR=1.05, 95% CI: 1.01‒1.08, p=0.008).

RFS at 5 years was 47% (36‒58%) and 42% (27‒57%) in the two groups, respectively (HR= 0.71, 95% CI 0.44‒1.17, p=0.18). In the DRB1-MM cohort, HLA-C mismatch affected RFS: 58% (34‒76%) in HLA-C-matched patients and 25% (9‒44%) in HLA-C-mismatched patients (HR=2.44, 95% CI 1.08‒5.56, p=0.032) (Figure 1). No effect of HLA-C mismatch was seen in the MUD cohort (Table 2).

In multivariate analysis, there were correlations between lower RFS and having an HLA-DRB1 and -C mismatched donor (HR=2.32, 95% CI: 1.21‒4.45, p=0.012), DRI high (HR=1.71, 95% CI 1.02‒2.88, p=0.041), and high TNC dose (HR=1.04, 95% CI: 1.01‒1.07, p=0.004).

The cumulative incidences of acute GVHD of grades II‒IV were 41% (31‒52%) and 28% (16‒42%) in the MUD and DRB1-MM groups, respectively (HR=0.60, 95% CI 0.32‒1.14, p=0.12), and those of grades III‒IV were 6% (2‒12%) and 5% (1‒14%) (HR=0.64, 95% CI 0.13‒3.22, p=0.57). No significant effect of HLA-C mismatch was found (Figure 2).

In multivariate analysis, higher donor age was associated with a lower incidence of moderate-to-severe acute GVHD (HR=0.66, 95% CI 0.44‒0.96, p=0.03).

The incidences of chronic GVHD were 50% (37‒61%) and 34% (19‒50%) in the MUD and DRB1-MM groups, respectively (HR=0.73, 95% CI 0.38‒1.37, p=0.27). No significant effect of HLA-C mismatch was found in the DRB1-MM group, while an HLA-C mismatch increased the incidence of chronic GVHD in the MUD group (38% vs. 77%; HR=2.65, 95% CI 1.38‒5.07, p=0.005) (Figure 2). In multivariate analysis, a parous female donor to male recipient (HR=3.10, 95% CI 1.28‒7.52, p=0.012), higher patient age (HR=1.02, 95% CI: 1.01‒1.03, p=0.030), and lower donor age (HR=0.96, 95% CI 0.93‒0.99, p=0.042) were associated with chronic GVHD.

Graft failure/rejection occurred in two patients in the MUD group and in three patients in the DRB1-MM group.

No significant difference in infections was seen in the two groups. CMV reactivation occurred in 50% of the patients in both groups. CMV disease occurred in six patients (7%) in the MUD group and in four patients (9%) in the DRB1-MM group. Epstein-Barr virus-related post-transplantation lymphoproliferative disease was seen in 5% of the patients in both groups.

No effect of ATG dose and brand was seen on any of the results in this study.