On-line Supplemental Material

Supportive Care

All patients were nursed in HEPA-filtered rooms during the early post-HSCT aplastic period. Antimicrobial prophylaxis was given following institutional policies, but which can be summarized as follows. Bacterial prophylaxis consisted in ciprofloxacin or levofloxacin during neutropenia or until the start of broad-spectrum antibiotics. Antifungal prophylaxis was administered according to the protocols at each site, with either fluconazole and a pre-emptive strategy, or using a mould-active antifungal agent (posaconazole, voriconazole or other systemic antifungal drugs) until engraftment or whenever the patient was given steroids for the treatment of GVHD. Non-absorbable oral antimicrobials were not used in any center. Prophylaxis against Pneumocystis jiroveci consisted of cotrimoxazole from day −10 to day −2 and then was restarted after engraftment for a minimum of 1 year or until immunosuppressive therapy was stopped. Pentamidine was used if cotrimoxazole was contraindicated. For prevention of Cytomegalovirus (CMV)-related disease, all institutions followed a preemptive approach with polymerase chain reaction (PCR) monitoring. Acyclovir was recommended for a minimum of 1 year post-HSCT or until immunosuppressive therapy was stopped. Galactomannan testing and CMV polymerase chain reaction analysis were performed twice weekly, and Epstein-Barr virus (EBV) PCR analysis was performed weekly until day +100 or indefinitely for those with active GVHD. Intravenous immunoglobulin was recommended whenever the total IgG blood level was < 300 mg/dL.

Definitions

The Hematopoietic Cell Transplantation Comorbidity-age Index (HCT-CI/age) [18] and the European Group for Blood and Marrow Transplantation (EBMT) risk score [20] were calculated following standard recommendations [19]. The revised Disease Risk Index (rDRI) was developed as a tool to assign patients into one of 4 risk groups based both on the risk of progression of the underlying disease and the status of this underlying disease at the time of transplantation [17]. Organ toxicities possibly/probably related with the conditioning-PTCy platform were graded according to the Common Terminology Criteria for Adverse Events [CTCAE v.4] v.4. Myeloid engraftment was defined as an absolute neutrophil count of 0.5 x 109/L for at least 3 consecutive days. Platelet engraftment was defined as a platelet count of 20 x 109/L or higher, without transfusion support, for 7 consecutive days. Patients who survived more than 28 days after transplantation and who failed to achieve myeloid engraftment were considered as primary graft failures, while secondary graft failure was defined as the loss of donor engraftment.

Serial samples of peripheral blood and/or bone marrow were analyzed for degrees of donor–recipient chimerism using PCR of informative minisatellite loci, as previously described in detail [27]. To evaluate lineage chimerism, lysis buffer was added to PB and a selection of CD3+ve and CD19+ve cells from PB was done using immunomagnetic beads. Complete donor chimerism was defined as the presence of at least 95% donor DNA in the sample analyzed both in myeloid and T cells, or in 100% unfractionated nucleated cells.

Cytomegalovirus (CMV) infection was defined as the presence of at least 2 consecutive (within a minimum interval of 48 hours) positive results of a polymerase chain reaction (PCR) test for CMV in peripheral blood (PB) with a viral load > 1000 IU/mL, while CMV disease was defined as the demonstration of CMV in biopsy or autopsy specimens from clinically involved visceral sites by culture and/or histology or if CMV was detected in samples from clinically-defined sites of disease [22]. Epstein-Bar virus (EBV) infection was defined as the presence of at least 2 consecutive (within a minimum interval of 7 days) positive results of a PCR test for EBV in peripheral blood (PB) with a viral load > 1000 e.g.c./mL, with or without evidence of EBV-related post-transplant lymphoproliferative disease (EBV-PTLD). BK polyomavirus-related hemorrhagic cystitis was defined by accepted criteria [21]. The staging of acute and chronic GVHD followed established guidelines, and severe GVHD referred to either grade III-IV acute or extensive chronic GVHD.

On-line Supplemental.Multivariate analysis of the 1-year NRM and OS.

Variables / 1-year OS / 1-year PFS / 1-year NRM / 1-year relapse
P / HR
(95% C.I.) / P / HR
(95% C.I.) / P / HR
(95% C.I.) / P / HR
(95% C.I.)
Response
Complete remission (50)
Other response (31) / 0.007 / 0.33 (0.14 – 0.73) / 0.05 / 0.49 (0.19 – 0.86) / 0.170 / NA / 0.79 / NA
HCT-CI-age
2
3 / 0.023 / 0.4 (0.18 – 0.88) / 0.053 / NA / 0.150 / NA / 0.98 / NA
Recipient age (years)
< 50
50 / 0.016 / 0.35 (0.15 – 0.82) / 0.02 / 0.41 (0.19 – 0.86) / 0.014 / 0.24 (1.3 – 12.9) / 0.79 / NA
Failed prior alloHSCT
No (67)
Yes (14) / 0.05 / 0.4 (0.16 – 0.9) / 0.01 / 0.34 (0.15 – 0.77) / 0.46 / NA / 0.14 / NA
EBMT score
3
4 / 0.92 / NA / 0.98 / NA / 0.67 / NA / 0.52 / NA
Stem cell source
PBSC
BM / 0.99 / NA / 0.84 / NA / 0.59 / NA / 0.72 / NA

Abbreviations: NA = Not Applicable, OS, overall survival, PFS: progression free survival; CI., cumulative incidence: SCT: stem cell transplantation; HCT-CI: hematopoietic cell transplantation comorbidity index; EBMT: europeran bone marrow transplantation; HR,Hazard ratio; 95% confidence interval (95% C.I.). PBSC: peripheral blood stem cell. BM: bone marrow