Overall cohort description and HSCT characteristics
98 patients were included in this study, of whom 66 (67%) were male and 32 (33%) female. Patient characteristics are summarized in Table 1. Transplants were done in the period 2007-2010 for 29 (30%) patients, while 69 (70%) patients were transplanted in the period 2011-2018. Median age at HSCT was 10 months (IQR 3-44). Median follow-up was 6 years (IQR 4-9). Donors included 46 (47%) matched unrelated donors and 52 (53%) related donors (16% matched sibling donors and 36% haploidentical donors). Stem cell sources were bone marrow (BM) in 47 (48%), umbilical cord blood in 6 (6%), and peripheral blood stem cells (PBSC) in 45 (46%) HSCTs. A busulfan-based myeloablative conditioning regimen was used for 41% of HSCTs.
Phenotypes with associated genotypes are listed in Table 2. The most represented diagnoses were in the categories of SCID and PIRD, with 43 (44%) and 25 (26%) patients, respectively. The underlying genetic condition was known at the time of the transplant in 85% of cases. There was a statistically significant difference between the two time periods of HSCT (p=0.005): from 2007 to 2010 we categorized the specific molecular defect prior to HSCT in 20/29 (69%) patients, while from 2011 to 2018 in 63/69 (91%), likely reflecting increased genetic testing availability over time. Of the 15 patients without genetic diagnosis at the time of the first transplant, as noted in Table 2, we identified the underlying molecular defect after the transplant for 5 of them, with a median time of 14 months (IQR 10-45). No differences in age at HSCT between the two eras (p=0.19). There were no correlations at both univariate and multivariate analysis between identification of the genetic mutation and gender (p=0.21; OR=1.00, 95% Confidence Interval (CI), 0.99-1.00, p=0.91), age at HSCT (p=0.79; OR=0.45, 95% CI, 0.14-1.47, p=0.19), donor relation (p=0.98; OR=0.84, 95% CI, 0.20-3.53, p=0.81), stem cell source (p=0.99; PBSC OR=0.92, 95% CI, 0.22-3.80, p=0.92), and conditioning regimen (p=0.96; NMA OR=1.09, 95% CI, 0.27-4.33, p=0.90).
Overall survival and genetic identification
The OS at 5 years post-HSCT was 87% (95% CI, 0.78-0.92). Twelve (12%) patients died, at a median of 185 days after transplant (IQR 75-472). There was a correlation between the identification of the genetic condition before transplant and OS at 5-years or at last follow-up post-HSCT (p=0.0002), with 58% (95% CI, 0.30-0.79) of patients alive in the group of unknown genetic defect versus 93% (95% CI, 0.84-0.97) in the group with known genetic defect (Figure 1).
Between the two time periods of HSCT: 79% (95% CI, 0.60-0.90) of patients in the first period group (2007-2010) vs 91% (95% CI, 0.82-0.96) in the second period group (2011-2018) survived (p=0.15). A statistical difference in OS by known versus unknown genetic defects was present in both periods. In the first timeframe, 5-year OS was 55% (95% CI, 0.20-0.80) for patients who did not have a known genetic defect versus 90% (95% CI, 0.65-0.97) with a known genetic defect (p=0.032), while in the second time period 67% (95% CI, 0.20-0.90) of patients who did not have a known genetic defect versus 94% (95% CI, 0.84-0.98) with known genetic defect (p=0.013) were alive, suggesting that knowledge of genetic mutation, rather than advances in HSCT technique or supportive care, is driving the differences in survival.
The mortality rate was 40% (95% CI, 0.16-0.68) in the group without genetic diagnosis versus 7% (95% CI, 0.02-0.15) in the group with genetic diagnosis (p<0.001). In the unknown genotype group, death was attributed to infection in 50% of cases, organ toxicity in 33%, and progressive underlying disease (i.e., HLH reactivation) in the remaining 17%; while in the known group, death was due to infection in 83% of cases and organ toxicity in 17%. There was a significant difference in the 1-year cumulative incidence of organ-toxicity-related death between the two groups, 13% (95% CI, 0.02-0.40) in the unknown versus 1% (95% CI, 0.00-0.06) in the known (p=0.012). Detailed characteristics of deceased patients are provided in supplementary Table S1.
Event-free survival and genetic identification
Twenty patients (20%) required a second HSCT, mostly due to graft failure (95%), at a median time of 177 days after the first one (IQR 77-583), and of these 15 (75%) were alive at last follow-up. The EFS at last follow-up post-HSCT was 74% (95% CI, 0.64-0.82). As shown in Figure 2, comparing the two groups with and without genetic diagnosis, EFS at 5-years post-HSCT was influenced by the identification of the underlying molecular defect (p=0.006): EFS for the group without a genetic diagnosis was 44% (95% CI, 0.18-0.68), while for the group with a genetic diagnosis was 76% (95% CI, 0.64-0.84).
There was no difference in EFS at 5-years post-HSCT (p=0.62) by era of transplant: in the first period (2007-2010) the EFS was 66% (95% CI, 0.45-0.80), while in the second period (2011-2018) was 73% (95% CI, 0.60-0.82). In the first period, for the group without a genetic diagnosis the EFS at 5-years post-HSCT was 44% (95% CI, 0.14-0.72) versus 75% (95% CI, 0.50-0.88) for the group with (p=0.12); in the second period the EFS at 5-years post-HSCT was 50% (95% CI, 0.11-0.80) for the group without a genetic diagnosis versus 75% (95% CI, 0.61-0.85) for the group with (p=0.03).
Genetic identification and complications post-HSCT
There was a correlation between the identification of the genetic mutation and graft failure/rejection, seen in 47% (95% CI, 0.21-0.73) of patients without genetic diagnosis versus 19% (95% CI, 0.12-0.29) with (p=0.021). As shown in Figure 3, five patients (33%) with unknown genetic defect received a second transplant, at a median time of 57 days after the first one (IQR 50-361). Regarding patients with a genetic diagnosis, 15 (18%) received a second transplant, at a median time of 175 days after the first one (IQR 112-489).
After the first HSCT, mortality was 5% (95% CI, 0.01-0.12) for the known genetic group vs 20% (95% CI, 0.4-0.48) for the unknown group (p=0.022); while, after the second HSCT, mortality was 13% (95% CI, 0.16-0.40) for the known genetic group vs 60% (95% CI, 0.15-0.95) for the unknown group (p=0.048). There was no difference between the two time periods for graft failure/rejection (p=0.25).
Other considerations, outside the scope of this study, include the potential for related donors to be carriers of unknown genotype at time of transplant, thereby leading to unsuccessful HSCT due to potential functional insufficiency of donor cells, although in our cohort we found no significant differences in either OS at 5 years post HSCT, graft failure or EFS for patients with unknown genotype among those who received a matched sibling donor (possible carriers) versus a matched unrelated donor (not carrier) versus a haploidentical donor (probable carrier for autosomal recessive disorders; possible carrier for female donors to patients with X-linked disorders).
Genetic identification and GVHD
As shown in Table 3, of the patients with GVHD, 18 developed acute grade II-IV, at a median of 22 days post-HSCT (IQR 16-63) for a 100-day cumulative incidence of 18% (95% CI 0.11-0.26). Of five patients with chronic GVHD (cGVHD), 2 were extensive in nature, for a 3-year cumulative incidence of 15% (95% CI 0.01-0.46). No differences were observed between the two time-periods for either acute GVHD (aGVHD) (p=0.43) or cGVHD (p=0.60). No differences between the two groups, with genetic diagnosis and without, were observed regarding aGVHD (p=0.30) or cGVHD (p=0.91).
In our cohort, we analyzed cell reconstitution when all criteria were available and satisfied. We observed a T-cell reconstitution in 19/54 (35%) at 6 months, in 48/69 (69%) at 12 months, and in 66/80 (83%) at last follow-up. Median time for T-cell reconstitution was 245 days (IQR 160-538). There were no correlations between identification of the genetic mutation and the incidence of T-cell reconstitution at 6-months post-HSCT (p=0.15), at 12-months post-HSCT (p=0.72), or at last follow-up (p=0.14).
B-cell reconstitution was documented in 55 patients (56%), at a median time of 365 days post-HSCT (IQR 219-730), and it was in patients with genetic defect known prior to HSCT: 4 (27%) patients with unknown genetic defect versus 51 (61%) with known genetic defect (p=0.012). As shown in Table 3, no significant correlations were detected between the identification of the genetic diagnosis or lack thereof and development of post-HSCT infections, including viral (p=0.40), bacterial (p=0.42), or fungal (p=0.86).
SCID versus non-SCID
OS at 5-years or at last follow-up post-HSCT was correlated with identified genetic cause in both SCID and non-SCID disorders (p=0.002), in both time periods. In the SCID group, 5-year survival was 56% (95% CI, 0.15-0.84) of patients without a genetic diagnosis, versus 94% (95% CI, 0.80-0.98) with genetic diagnosis; in the non-SCID group, 57% (95% CI, 0.17-0.84) of patients without a genetic diagnosis versus 91% (95% CI, 0.79-0.97) with a genetic diagnosis were alive after 5 years. We did not find any SCID versus non-SCID influence of identification of the genetic mutation prior to transplant: incidence of aGVHD (non-SCID p=0.27; SCID p=0.82) versus cGVHD (non-SCID p=N.A.; SCID p=0.91). However, there was a significant correlation between the identification of the genetic defect and graft failure for non-SCID patients (p=0.025), with 43% (95% CI, 0.10-0.81) of patients with unknown genetic defect versus 11% (95% CI, 0.03-0.23) of patients with genetic diagnosis; in contrast, no such correlation was found for SCID patients (p=0.29), (Supplementary Table S2).
Predominant hematopoietic cell (HC) vs Combined HC and non-HC immune dysfunction
In patients with a known genetic defect at the time of the transplant, 43 (52%) had a genetic defect predominantly affecting the HC compartment, while 40 (48%) had defects in genes that resulted in combined HC and non-HC immune dysfunction (supplementary Table S3). As shown in Figure 4, there was no significant difference in OS at 5 years or at last follow-up post-HSCT based on the function of the genes affected in this cohort (p=0.56): 38 (90%; 95% CI, 0.77-0.96) patients with genetic diagnoses affecting HC and non-HC immune function vs. 44 (92%; 95% CI, 0.78-0.98) patients with genetic diagnoses affecting predominantly HC cells survived. EFS was not influenced by the function of the genes affected in this cohort (p=0.23): EFS was 69% (95% CI, 0.50-0.81) for the combined HC and non-HC group versus 78% (95% CI, 0.62-0.88) for HC predominant group.