DISCUSSION
This study analyzed lymphocyte recovery after allo-HSCT in pediatric
patients with malignant and non-malignant diseases up to 180 days after
HSCT. Over this 6-month period, lymphocyte reconstitution was gradual,
and each subtype showed different reconstitution rates on D+100. Our
study reiterates that lymphocyte count after HSCT impacts patient
survival and is associated with pre-transplant factors such as age,
source of stem cells, donor, serotherapy, conditioning, disease, and
clinical outcomes like CMV and aGvHD. 3,6–8
On D+100, 89% of patients achieved ALC recovery while CD3+CD8+ and
CD16+CD56+ lymphocytes showed the highest recovery rates (68% and 47%,
respectively). As natural killer (NK) lymphocytes and TCD8+ clonal
expansion propel lymphocyte reconstitution in the first 3 months after
HSCT, ALC recovery in our cohort occurred possibly due to these
subtypes. 3,17 Kim et al (2013) also obtained this
pattern of reconstitution in children and similar median counts for the
same period, except for the higher percentage of NK recovered patients
owing to a lower cut-off value based on the age-matched reference
values. 18
In this study, patients <8 years showed better recovery for
both ALC on D+30 and CD3+CD4+ on D+100. This finding contrast with those
of Bartelink et al (2013), which indicated no association between IR and
age in a cohort that also included umbilical cord blood transplants.19 Previous studies have showed an earlier recovery
trend in younger patients, through rapid recovery of CD3+CD4+, CD19+ and
ALC in patients <5 years and CD3+CD8+, CD19+ and CD16+CD56+ in
patients <10 years. 7,18 A consensus on this
matter is hampered because published studies have used distinct
lymphocyte recovery criteria according to stem cell sources and
age-matched reference values. Our results are inconclusive due to the
small number of patients. However, there is evidence that children
rapidly recover naïve TCD4+ compared with adults, supporting the
association between age and greater pre-HSCT thymic activity as an
influencing factor in IR. 20
The potential deleterious effect on thymus caused by MAC regimen was
attenuated owing to higher ALC recovery on D+30 when compared with that
in the RIC regimen. In children, this impact might be reduced because
the thymus is fully functional until puberty, after which thymopoiesis
progressively diminishes. 21 Although MAC is related
to a better CD19+ recovery compared with RIC regimen in adults, our
study showed no association between the type of conditioning regimen and
lymphocyte subtype recovery. 22 Patient and transplant
heterogeneity led to a variety of preparatory regimens, masking possible
correlations between IR and the intensity of these regimens. Moreover,
individual conditions (age and weight) may have also altered
pharmacokinetics and pharmacodynamics of regimens leading to different
exposures and effects on IR. 22,23
Patients who received BM grafts achieved higher ALC recovery on D+30.
Although PBSC mobilized by granulocyte-colony stimulating factor contain
more cells contributing to lymphocyte count (CD34+, T lymphocytes and
NK) and promote CD3+CD4+ rapid recovery, 93% of the PBSC recipients in
this cohort were haplo-HSCT. 24–26 Therefore, donor
type may have an impact on early lymphocyte recovery, regardless of the
stem cell source.
We observed that serotherapy interfered with ALC recovery on D+180. The
same was noted in a pediatric study that indicated inadequate ALC
recovery up to 1 year after HSCT. 18 In our cohort,
lymphocyte subtypes were not associated with serotherapy. In a large
pediatric cohort of malignant and non-malignant diseases, Admiraal et al
(2015) reported that only high concentrations of ATG after BM or PBSC
transplant reduced TCD4+ reconstitution, whereas in umbilical cord blood
transplants even low concentrations of ATG exerted a deleterious effect.
Therefore, CD3+CD4+ and serotherapy revealed no association in our study
possibly because of the inclusion of only BM and PBSC transplants, the
reduced number of patients who used serotherapy, and the inter-patient
variability of doses and periods of ATG administration.27
Viral infections and lymphocyte reconstitution have a complex
correlation as opportunistic infections are related to both the cause
and the effect of delayed IR. 28,29 Patients who
recovered ALC and CD3+CD8+ on D+100 had a higher CI of CMV infection,
which occurred on a median 37 days after transplant in our patients.
Literature shows that impairment of CD3+CD8+ early reconstitution
contributes to CMV reactivation, while high counts of CD3+CD4+ and
CD16+CD56+ represent protective factors in adults.30,31 But after infection, a clonal expansion of TCD8+
lymphocytes occurs in response to the CMV antigenic stimulus, resulting
in an oligoclonal repertoire of memory T cells. 32 The
high CD3+CD8+ count observed in patients with CMV infection, described
also by Janecsko et al (2016), possibly denotes its specific CMV
expansion, which could be confirmed in a larger cohort using molecular
tests. 30
The development of aGvHD ≥grade 2 resulted in lower CD3+CD4+
reconstitution on D+100. Previous reports showed that patients with GvHD
had low counts of CD3+CD4+ between the 1st and
3rd months after HSCT, especially naïve TCD4+ cells.33,34 The
intense immunodepression is due to the damage that the disease causes in
thymic function and BM microenvironment – essential factors for
lymphopoiesis – but can be aggravated by GvHD treatment with
corticosteroids.35 Considering the early development
of aGvHD in this cohort (median day +28), the poor CD3+CD4+ recovery on
D+100 may reflect the disease itself and its treatment.
In our study, haploidentical
transplants had a higher CI of CMV infection and aGvHD but OS was
similar to other types of transplants. The inadequate ALC recovery on
D+30 and the higher CMV infection risk in haplo-HSCT may have resulted
from the immunosuppressive effect of GvHD prophylaxis with PT-Cy, CsA
and MMF 36–38 We also observed that CD19+ counts on
D+100 were lower in haplo-HSCT than in MSD and unrelated transplants.
Although major outcomes after haplo-HSCT are comparable to those of MSD
and matched unrelated donors, more studies are needed to better
understand IR in the haplo-HSCT settings, especially for pediatric
patients with non-malignant diseases39.
In accordance with a previous study, our data suggest that ALC≥500/µL on
D+100 is a good predictor of OS for pediatric patients undergoing
allo-HSCT. 12 Several cell subsets play a crucial role
in protecting from infections, however sophisticated exams are not
readily available in resource-limited countries. These countries need
simple biomarkers to demonstrate if transplanted patients have achieved
an acceptable T cell reconstitution. In our study, we observed a trend
towards inadequate CD3+CD4+ recovery on D+100 in patients dying from
severe infections, but the reduced number of patients was insufficient
to reveal a significant effect on OS. However, there is robust evidence
in the literature suggesting that this is the most relevant parameter
for survival as it is related to mortality from infections.33,40
Our retrospective study has important limitations related to the
irregularity in immune surveillance by flow cytometry after HSCT.
Moreover, the relatively small number of patients and disease
heterogeneity did not allow us to reach any definitive conclusions.
Nevertheless, these real-world data can be extrapolated to other
pediatric HSCT centers and also form the basis for future collaborative
studies.
This study exposes the reality in developing countries, ranging from
limited financial resources for laboratory tests, to the failure to
follow-up due to long travel distances to the reference center.
Concurrently, as developed countries move towards predictive medicine in
IR monitoring, through greater regularity and specificity of tests,
herein we still endeavor to implement cost-effective strategies that may
help our patients. To better estimate the time to reach immunocompetence
after allo-HSCT in resource-limited countries, we need to standardize IR
monitoring with the available tests. Although only studies compiling all
data in a prospective way will be able to address this question,
monitoring ALC counts on a monthly basis after transplant and measuring
lymphocyte subtypes on D+100 and D+180 are easy to perform and could be
implemented in clinical practice to identify patients at risk for
inadequate IR.
In conclusion, this study summarized the results of IR after HSCT for
malignant and non-malignant diseases in one of the main pediatric
transplant centers in Latin America. Lymphocyte reconstitution was
progressive up to D+180, mainly due to CD3+CD8+ counts. Overall, younger
age, BM grafts, MAC regimen, non-malignant disease, non-use of
serotherapy, and non-haploidentical donor were associated to adequate
lymphocyte recovery. Multiple factors influence IR and our study
suggests that ALC≥500/µL on D+100 may be a predictor of pediatric
survival after HSCT, preferably concomitant with lymphocyte
subpopulations monitoring.