ALBERT

Description: The primary immunodeficiencies (PID) are rare inherited diseases characterised by severe dysfunction of adaptive and/or innate immunity. Over 200 distinct PIDs have been described, with 20 of them accounting for 〉 90% cases. The 3 most common PIDs are severe combined immunodeficiency (SCID), Wiskott Aldrich syndrome (WAS) and chronic granulomatous disease (CGD). Many have been successfully treated by allogeneic hematopoietic stem cell transplantation (Allo-HSCT), however, almost all published series to date have focussed on paediatric patients. In the largest published series, the median age at transplant is 〈 1 year for SCID, 12.7 years for CGD, and 3 years for WAS. Early Allo-HSCT is preferred for PID patients, but not always possible. Delayed diagnosis and 'milder' clinical phenotypes may delay treatment with curative intent. With improved supportive care PID patients are more likely to survive to early adulthood without Allo-HSCT, despite serious co-morbidities. Furthermore, for several PIDs, controversy surrounding optimal timing of Allo-HSCT remains due to rarity of disease and lack of experience. Furthermore, Allo-HSCT for older adolescents and adults with PID has previously been avoided due to severe TRM and poor outcomes. Here we report the largest group of consecutive adult PID patients described to date (n=27) who underwent Allo-HSCT in the adult HSCT programme of University College London. The mean age was 24 years (range 17-50) at transplant. Of these, 11 patients had X-linked or autosomal recessive CGD and 16 had other inherited PID, including CVID (1 with T-NHL, 1 with HLH, 1 with GLILD) (n=3), autoimmune LPD (1 Fas mutation) (n=2), X-linked LPD (n=1), DCML +/- Gata2 mutation (n=2), Gata2 Defic (n=1), Cg-SCID (n=1), CID with CD27 Defic (1 with HL and DLBCL, 1 with HL) (n=2), NK Defic (n=1), AR IL12 Recb Defic (n=1), XIAP with Crohn's and HLH (n=1) and Rag2 compound heterozygote with red cell aplasia (n=1). Stem cell donors were matched unrelated (MUD) (n=14), mismatched unrelated (MMUD) (n=5, all 1 Ag mismatch) or matched related donors (MRD) (n=8). Stem cell source was PBSC (n=20) and BM (n=7). Reduced intensity conditioning was used in 26 patients including Flu/Mel/Alemtuzumab (n=17), Flu/Bu/Alemtuzumab (n=8) and Flu/Bu/ATG (n=1). One patient underwent myeloablative conditioning with Flu/Cy/TBI. Additional GVHD prophylaxis with cyclosporine was used in all patients. Reverse isolation, antimicrobial and antifungal prophylaxis were used to reduce the risk of infectious complications. Acute and chronic GVHD incidence and severity and time to engraftment, lineage specific chimerism, immune reconstitution and discontinuation of immunoglobulin replacement therapy were recorded. Overall survival (OS) at 2 years for all patients was 85.6%, and by donor was 75% for MRDs (n=8) and 89.5% for MMUD/MUD (n=19) with a mean follow up of 35 months (range: 7 to 11 years 1 month). Analysis of outcome by diagnosis demonstrated OS at 2 years of 81.8% for CGD patients (n=11) and 87.5% for other PIDs (n=16). TRM was low with only four deaths observed at a median follow-up of 22 months (n=27). 2 patients died of multi organ failure and sepsis prior to engraftment, one at 7 months post-HSCT of granulomatous meningitis and grade III aGVHD and one at 28 months post-HSCT of sepsis in the context of chronic extensive GVHD. 8 patients developed grades I-II aGVHD (6 skin only, 1 involving the gut and 1 progressing to chronic lung GVHD). GVHD had resolved in all but one patient at last follow up. For all patients, neutrophil and platelet engraftment occurred after a median of 12 and 14 days respectively. Peripheral blood chimerism was available for 22 of the 23 surviving patients at last follow up. Multilineage full donor chimerism was observed in 8 (36%) with mixed chimerism observed in at least one cell lineage tested for the remainder. At 1-year post transplant, 80% of those tested had achieved a normal lymphocyte count (1.0-2.8 x 109/L), 47% had normalization of absolute CD3+ count (0.7-2.1 x 109/L), 40% had a normal CD4+ count (0.3-1.4 x109/L) and 87% had normalisation of their CD8+ cell count (0.2-0.9 x109/L). Allo-HSCT is well tolerated in this patient group and should be considered as an alternative therapeutic option in PID patients not transplanted in childhood where an appropriate donor is available. Triggers for referral should include life threatening infections, malignancy, severe autoimmunity and refractory disease. Disclosures Fielding: Baxalta: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees.

Description: Re-activation of latent EBV infection is a significant risk following T-cell depleted (TCD) allogeneic HSCT. The clinical course can range from asymptomatic viraemia to the development of post-transplantation lymphoproliferative disease (PTLD) and death. Since alemtuzumab depletes both T and B cells, it has been proposed that the risk of PTLD will be reduced compared to other TCD protocols. In an attempt to determine the risk of EBV viraemia or PTLD following allogeneic HSCT employing alemtuzumab-containing conditioning protocols, we reviewed outcomes in 91 consecutive adult patients treated at our institution. Of these, 84 patients had 〉6 months follow-up and are included in this analysis. Two strategies of TCD were employed in this cohort: in vivo TCD with alemtuzumab (median dose 100mg) as part of a reduced intensity regimen (fludarabine based n=51); and in vitro TCD of the PBSC graft with 20 mg alemtuzumab ‘in the bag’ following conventional myeloablative conditioning (n=33). We have shown previously that alemtuzumab antibody levels persist for longer periods (up to 8 weeks ) following this in vivo depletion strategy compared to the in vitro approach [Morris et al. Blood 2003 102:404–6]. The median follow-up is 337 days and median age was 43 years (range 16–67 years). Diagnoses were AML/MDS (n=39), NHL (n=16), Hodgkins lymphoma (n=11), MPD (n=7), CLL (n=5) and other (n=6). 41/84 had HLA-identical sibling donors, 2 mismatched sibling donors, 21 matched unrelated donors and 20 mismatched unrelated donors. Monitoring for EBV load was performed by TaqMan whole blood real time PCR on a weekly basis for 100 days post transplantation and then as seen for follow-up. Positive results defined as 〉200 copies/ml of EBV DNA. All patients with high levels of EBV viraemia (EBV DNA level 〉 40,000 copies/ml) underwent CT imaging, bone marrow examination and lumbar puncture followed by withdrawal of immunosuppression and a single infusion of 375mg/m2 Rituximab as per institution policy. Of 82 patients where serostatus was available, 92.7 % were anti-EBV IgG positive. The 12-month cumulative incidence (CI) of EBV reactivation was 34.5% (39.2% following in vivo TCD versus 27.3% following in vitro TCD, p=0.2). Median peak viral load was 1,922 copies/ml (range 205–6,210,000). The 12 month CI of high-level EBV viraemia (〉40,000 copies/ml) was 11.9% (7.84% following in vivo TCD versus 18.2% following in vitro TCD, p=0.19). Patients re-activated EBV at a median of 86 days post transplant (range 21–380 days). The median duration of viraemia was 42 days (range 1–460 days). The 12-month CI of recurrent EBV re-activation (〉1 episode) was 21.4% (15.5% following in vivo TCD versus 25.5% following in vitro TCD, p=0.4). Of 10 patients with high-level EBV viraemia, 1 patient had PTLD as confirmed by histology. 8 of 10 patients received a single dose of Rituximab and this was associated with a rapid decline in EBV DNA load to undetectable levels in all cases. No adverse events secondary to Rituximab were noted. The patient with PTLD was given a second dose of Rituximab and attained a CR that persists at 334 days post treatment. 12-month non-relapse mortality was 10% in patients who had EBV reactivation and 27.7% in patients without EBV reactivation (p=0.06). Baseline and post-transplantation characteristics of sex, age, diagnosis, in vivo vs. in vitro alemtuzumab, donor type, CMV viraemia and presence of past or active acute GVHD were not related to re-activation of EBV by uni- or multivariate analysis in this patient cohort. In summary, this report demonstrates that TCD using alemtuzumab-containing conditioning protocols is associated with a high frequency of EBV viraemia but low risk of PTLD or non-relapse death in a cohort of patients treated pre-emptively with Rituximab for high EBV DNA loads.

Description: Key Points We describe the first successful use of gene therapy in a severely affected adult with WAS. Gene therapy is a viable strategy for adult WAS patients with severe chronic disease complications where allogeneic transplantation presents.

Description: In previous studies we and others have shown that the production of new T cells from the thymus declines after the third decade and that while patients below the age of 30 reconstitute primarily new T cells, those over thirty reconstitute primarily by expansion of pre-existing mature T cells. However little attention has been paid to the γδ subset of T cells, which form an important component of mucosal immune protection and which represent approximately 5% of peripheral T cells. Two major subsets of γδ T cells are defined by the expression of Vδ1 versus Vδ2, with Vδ1+ cells predominating in the fetal circulation and in mucosal sites, while Vδ2+ cells predominate in adult life and in the peripheral circulation. In light of the differerential preponderance of the two subsets in the fetal versus adult circulation, we have examined the reconstitution of these two subsets of γδ T cells following hemopoietic stem cell transplantation in a cohort of 28 patients sampled at 3 monthly intervals to ask whether both subsets recover adequately from adult stem cells. In 44 normal individuals, the median levels of Vδ1 and Vδ2 cells are 12.46 (0.22 to 167.8) and 32.78 (4.48 to 190.1) cells/mm3 respectively. In patients under 30, the reconstitution of the Vδ1 and Vδ2 subsets follow similar kinetics, reaching a plateau at 9 months post transplant with comparable numbers of Vδ1 and Vδ2 cells (note that the normal ratio of Vδ1 to Vδ2 is 0.38, so in the patients there is a significant increase in the proportion of Vδ1 cells in the peripheral circulation). In patients over 30 years of age, there is an even more significant disparity in the reconstitution of the two subsets. The Vδ1 subset recovers with similar kinetics as is seen in the patients under 30, although to slightly lower final levels. The Vδ2 subset, however, shows very little recover y, reaching a plateau at 6 months at the bottom of the normal range for up to 2 years post transplant. In these patients the ratio between Vδ1 and Vδ2 is inverted with an increasingly greater proportion of Vδ1 cells at longer times after transplant, with ratios in excess of 10 by 24 months post transplant, a 30-fold increase in the normal proportion of Vδ1 + T cells. Given that the Vδ1 subset shows a very restricted repertoire compared to the Vδ2 subset, the overall capacity of the circulating γδ T cell population to recognise and respond to antigen will be significantly compromised in older transplant recipients.

Description: Key Points Allo-HSCT with RIC is safe and effective in younger adults with severe PID. Referral triggers should include severe infections, autoimmunity, malignancy, and disease progression despite conservative management.

Description: Reactivation of cytomegalovirus (CMV) remains a serious complication after allogeneic stem cell transplantation, but the role of γδ T cells is undefined. We have studied the immune reconstitution of Vδ2negative (Vδ2neg) γδ T cells, including Vδ1 and Vδ3 subsets and Vδ2positive (Vδ2pos) γδ T cells in 40 patients during the first 24 months after stem cell transplantation. Significant long-term expansions of Vδ2neg but not Vδ2pos γδ T cells were observed during CMV reactivation early after transplantation, suggesting direct involvement of γδ T cells in anti-CMV immune responses. Similarly, significantly higher numbers of Vδ2neg γδ T cells were detected in CMV-seropositive healthy persons compared with seronegative donors; the absolute numbers of Vδ2pos cells were not significantly different. The expansion of Vδ2neg γδ T cells appeared to be CMV-related because it was absent in CMV-negative/Epstein-Barr virus-positive patients. T-cell receptor-δ chain determining region 3 spectratyping of Vδ2neg γδ T cells in healthy subjects and patients showed restricted clonality. Polyclonal Vδ2neg cell lines generated from CMV-seropositive healthy donors and from a recipient of a graft from a CMV-positive donor lysed CMV-infected targets in all cases. Our study shows new evidence for role of γδ T cells in the immune response to CMV reactivation in transplantation recipients.