Hematopoietic stem cell transplantation for inborn errors of immunity: 30-year single-center experience.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) represents an effective treatment for a variety of inborn errors of immunity (IEI). We report the experience of children affected by IEI who received allo-HSCT over a period of 32 years at IRCCS Istituto Giannina Gaslini, Genoa, Italy. HSCTs were performed in 67 children with IEI. Kaplan-Meier estimates of overall survival (OS) rate at 5 years in the whole group of patients was 83.4% after a median follow-up of 4 years. Median age at transplant was 2.5 years. Eight allo-HSCTs were complicated by either primary or secondary graft failure (GF), the overall incidence of this complication being 10.9%. Incidence of grade 3-4 acute GvHD (aGvHD) was 18.7%, significantly lower in the haploidentical transplant cohort (p = 0.005). Year of transplant (≤2006 vs. >2006) was the main factor influencing the outcome. In fact, a significant improvement in 5-year OS was demonstrated (92.5% >2006 vs. 65% ≤2006, p = 0.049). Frequency of severe aGvHD was significantly reduced in recent years (≤2006 61.5%, vs. >2006 20%, p = 0.027). A significant progress has been the introduction of the TCR αß/CD19-depleted haploidentical platform, which was associated with the absence of severe aGvHD. However, it was associated with 23.5% incidence of GF. All but one patient experiencing GF in the this specific cohort were successfully retransplanted. In summary, allo-HSCT is confirmed to be an effective treatment for children with IEI, even in the absence of an HLA-matched donor.

Haploidentical Stem Cell Transplantation After TCR-αß+ and CD19+ Cells Depletion In Children With Congenital Non-Malignant Disease.

Haploidentical hematopoietic stem cell transplantation (haplo-HSCT) represents a valuable alternative for children with nonmalignant disease and ex vivo negative selection of TCR-αß+ cells is an emerging graft manipulation option that carries several potential advantages in terms of reduced risk of graft-versus-host disease (GvHD) and improved immune reconstitution. We report all consecutive patients with a diagnosis of nonmalignant disease who received a TCR-αß+ and CD19+depleted haplo-HSCT at "IRCCS Istituto Giannina Gaslini" from 2013 to 2019; the conditioning regimen was myeloablative or non-myeloablative, depending on underlying disease; all patients received antithymocyte globulin and rituximab. No post-transplantation GvHD prophylaxis was given in presence of a TCR-αß+ cell dose in the graft lower than the threshold of 1 × 105/kg of the recipient's weight. Among 20 HSCTs, engraftment occurred in 17 (85%) after a median of 14 and 12 days from graft infusion for neutrophils and platelets, respectively. Primary graft failure was diagnosed in 3 (15%) patients, and 2 (10%) experienced secondary rejection; all of these patients underwent a second HSCT. The cumulative incidence of a-GvHD and c-GvHD was 15% (2 = grade 1, 1 = grade 4) at 90 days and 5% (1 = grade 1) at 7 months, respectively. Cytomegalovirus reactivation requiring pre-emptive treatment was observed in 9 patients (45%). One patient developed a JC virus-related progressive multifocal leukoencephalopathy, successfully managed with donor-derived virus-specific T-cell infusions. A complete immunological recovery was reached in most patients within 6 months. After a median follow-up of 4 years, 18 patients are alive, with a cumulative survival probability of 90%. Haplo-HSCT after ex vivo TCR-αß+/CD19+ negative selection may be considered a good option for children with nonmalignant diseases because it ensures a high engraftment rate with an acceptable risk of graft failure, very low incidence of significant GvHD, and good immune reconstitution with low frequency of severe virus-related disease. However, the control of viral infection/reactivation should be kept high to promptly provide pre-emptive treatments and approaches of antiviral adoptive immunotherapy.

Late-term fetuses with reduced umbilical vein blood flow volume: An under-recognized population at increased risk of growth restriction.

OBJECTIVES: To investigate the umbilical vein and uterine arteries blood flow volume (UV-Q, UtA-Q) in late-term pregnancies. STUDY DESIGN: This was a prospective observational cohort study of singleton pregnancies ≥40 + 0 weeks in which UV-Q and UtA-Q, both absolute and normalized for estimated fetal weight (EFW) values, were evaluated in relation to AC drop of ≥20 percentiles from 20 weeks to term, Doppler signs of fetal cerebral blood flow redistribution and composite adverse perinatal outcome. The presence of neonatal hypoglycaemia and the need of formula milk supplementation were also examined. RESULTS: The study population comprised 200 women. Fetuses with AC drop (n = 34) had a significantly lower UV-Q and UV-Q/EFW than fetuses without AC drop (n = 166): median UV-Q 184 ml/min (IQR 143-225) vs 233 ml/min (IQR 181-277), p = 0.0006; median UV-Q/EFW 55 ml/min/kg (IQR 42-66) vs 63 ml/min/kg (IQR 48-74), p = 0.03. Fetuses with cerebral blood flow redistribution (n = 48) had a significantly lower UV-Q and UV-Q/EFW than those without (n = 134): median UV-Q 210 ml/min (IQR 155-263) vs 236 ml/min (IQR 184-278), p = 0.04; median UV-Q/EFV 58 ml/min/kg (IQR 45-70) vs 65 ml/min/kg (IQR 50-76), p = 0.04. There was a significant moderate correlation between middle cerebral artery pulsatility index (MCA-PI) and UV-Q and UV-Q/EFW (Spearman Rho -0.20 and -0.20; p = 0.008 and p = 0.006). CONCLUSIONS: The umbilical vein blood flow volume might have a potential role to identify fetuses with stunted growth in late-term pregnancies.

sFlt-1/PlGF ratio in hypertensive disorders of pregnancy in patients affected by COVID-19.

OBJECTIVES: To analyze soluble Fms-like tyrosine Kinase 1 (sFlt-1) and Placental Growth Factor (PlGF) ratio concentrations in COVID-19 pregnant patients with and without Hypertensive Disorders of Pregnancy (HDP), compared with non COVID-19 pregnant patients with HDP and a control group. STUDY DESIGN: We recruited and obtained a complete follow-up of 19 COVID-19 pregnant patients with HDP and of 24 COVID-19 normotensive pregnant patients. Demographic, clinical and sFlt-1/PlGF ratio findings were compared with a group of 185 non COVID-19 pregnant patients with HDP and 41 non COVID normotensive patients. Findings were based on univariate analysis and on a multivariate adjusted model, and a case by case analysis of COVID-19 pregnant patients with an abnormal sFlt-1/PlGF ratio > 38 at recruitment. MAIN OUTCOME MEASURES: sFlt-1/PlGF ratio. RESULTS: We confirmed a significant higher prevalence of HDP in women affected by COVID-19 compared to control population. sFlt-1/PlGF ratio was found high in HDP patients, with and without of Sars-Cov2 infection. COVID-19 patients with worse evolution of the disease showed greater rates of obesity and other comorbidities. sFlt/PlGF ratio proved not to be helpful in the differential diagnosis of the severity of this infection. CONCLUSIONS: COVID-19 pregnant patients showed a higher prevalence of HDP compared to non COVID-19 controls, as well as higher comorbidity rates. In spite of the possible common endothelial target and damage, between Sars-Cov-2 infection and HDP, the sFlt1/PlGF ratio did not correlate with the severity of this syndrome.

First trimester placental volume is reduced in hypertensive disorders of pregnancy associated with small for gestational age fetus.

INTRODUCTION: Trophoblastic invasion and placental growth are critical for pregnancy outcome. The placental volume can be assessed by 3 D ultrasound using Virtual Organ Computer-aided Analysis (VOCAL). Epidemiological and clinical data suggest that there are two different clinical phenotypes of hypertensive disorders of pregnancy (HDP) that coexist at any gestational age: HDP associated to fetal growth impairment and HDP associated to appropriate for gestational age fetal growth. The aim of this study was to determine whether placental volume in the first trimester of pregnancy differs between women with HDP associated or not to fetal growth impairment and uncomplicated pregnancies. METHODS: This is a retrospective cross-sectional study of prospectively recruited data in which maternal characteristics, Doppler velocimetry of uterine arteries, and three-dimensional (3 D) volume of the placenta were collected at 11 + 1 - 13 + 6 gestational weeks. The placental quotient (PQ) was calculated as placental volume/crown rump length. RESULTS: In a 2-year period, we prospectively collected first trimester data of 1322 women. For the purposes of this cross-sectional study, 57 women that delivered a SGA fetus, 34 that developed HDP-AGA, and six that developed HDP-SGA, respectively, were included in the study as cases. The control group was made of 117 uncomplicated pregnancies. The PQ was higher in women with uncomplicated pregnancies (PQ median 16.36 cm3/cm) than in all other study groups (PQ in SGA: 13.02 cm3/cm, p < .001; PQ in HDP-AGA: 12.65 cm3/cm, p = .002; and PQ in women with HDP-SGA: 8.33 cm3/cm [IQR 6.50-10.13], p < .001). The lowest PQ was observed in women with HDP-SGA and was significantly lower than PQ in either women with SGA or those with HDP-AGA (p = .02 and p = .04, respectively). The mean uterine artery pulsatility index was the highest in women with HDP-SGA (median 2.30) compared to all other groups (uncomplicated pregnancies 1.48, p < .0001; women with SGA 1.59, p = .001; and women with HDP-AGA 1.75, p = .009). DISCUSSION: Our findings suggest that HDP associated with SGA is characterized by impaired placental growth and perfusion as soon as in the first trimester of pregnancy. The role of PQ, isolated or in association with other biophysical parameters, to predict HDP with fetal growth impairment remains to be evaluated.

Folic acid versus 5- methyl tetrahydrofolate supplementation in pregnancy.

Folate (vitamin B9) is widely accepted to protect against fetal neural tube defects. The main sources of dietary folate are folic acid-fortified foods and folic acid-containing dietary supplements. However, folic acid is inactive in the human body and must be converted by the liver into the active molecule 5-methyltetrahydrofolate (5-MTHF). 5-MTHF functions as a methyl donor in many metabolic reactions, including the conversion of homocysteine into methionine, the biosynthesis of glycine from serine, and the biosynthesis of DNA precursor molecules. Therefore, folate is fundamental for growth, especially in the embryonic and fetal stages. Prescription of folic acid to women in the preconception period and during pregnancy is a consolidated practice. However, it can pose health risks in certain conditions, such as megaloblastic anemia, where it will conceal megaloblastic anemia due to vitamin B12 deficiency and in cases of reduced hepatic transformation of folic acid (e.g. due to genetic variants or during some pharmacotherapies). Some of these risks can be avoided by supplementation with 5-MTHF rather than folic acid. Because 5-MTHF does not require activation, it is immediately available to mother and fetus and does not accumulate in blood like folic acid does in cases of reduced hepatic transformation. This paper reviews the advantages and disadvantages of folate supplementation with folic acid versus 5-MTHF, with a focus on maternal and fetal health.

Continuous Femoral Catheter for Postoperative Analgesia After Total Knee Arthroplasty.

INTRODUCTION: Postoperative pain management in the total knee replacement (TKR) represent a fundamental step for a positive outcome, allowing rapid mobilization, already on the first day. Further, continuous peripheral nerve block techniques have been reported to allow effective and safe control of acute postoperative pain, ensuring the implementation and completion of an accurate and intensive joint rehabilitation program. AIM: The aim of this study was to assess early mobility and compliance of patients that underwent TKR surgery using the femoral block. METHODS: For the study, all patients that underwent TKR from 2015 to 2018 with ASA score between II-III was evaluated. Patients underwent vital parameters monitoring and were treated initially with midazolam (0.05mg / kg) e.v. combined sciatic block + femoral perineural catheter positioning using a peripheral nerve stimulation-assisted technique. RESULTS: Intraoperatively, satisfactory analgesia was guaranteed in all patients, associated with a complete muscle relaxation of the affected limb. High patient compliance, associated with good control of acute postoperative pain was obtained in the first 24 hours after surgery (VAS 0- 1). The ROM outcomes were good in all rehabilitation stages, managing to reach 90 degrees bending on the seventh day with an average KSS score of 88-90. Patient satisfaction and impact on quality of life were assessed with the SF-36 showing average scores of 78. CONCLUSION: Continuous femoral nerve blocking ensures good postoperative analgesia in TKR allowing an early joint mobilization, a rapid functional recovery of the knee and increasing patient compliance during the post-operative rehabilitation program.

Motivating Cord Blood Donation with Information and Behavioral Nudges.

Umbilical cord blood is a source of hematopoietic stem cells essential to treat life-threatening diseases, such as leukemia and lymphoma. However, only a very small percentage of parents donate upon delivery. The decision to donate the cord blood occurs at a very specific time and when parents likely experience emotional, informational, and decisional overloads; these features of cord blood donation make it different from other pro-social activities. In collaboration with an OB-GYN clinic in Milan, Italy, we conducted the first randomized controlled trial that applies tools from behavioral science to foster cord blood donation, and quantified the gains that informational and behavioral "nudges" can achieve. We found that information and "soft" commitments increased donations; approaching expecting parents closer to the delivery date and providing them with multiple reminders, moreover, had the strongest impact. However, a significant portion of women who expressed consent to donate could not do so because of organizational constraints. We conclude that simple, non-invasive behavioral interventions that address information gaps and procrastination, and that increase the salience of the activity can substantially enhance altruistic donations of cord blood, especially when coupled with organizational support.

Evaluation of Chimerism Dynamics after Allogeneic Hematopoietic Stem Cell Transplantation in Children with Nonmalignant Diseases.

It is recognized that chimerism following hematopoietic stem cell transplantation (HSCT) is a dynamic process. The aims of this study were to describe the evolution of chimerism in children with nonmalignant diseases who underwent allogeneic HSCT, and to analyze the risk factors influencing chimerism status. A total of 101 HSCTs were performed in 85 patients with nonmalignant diseases. The donor was unrelated in 62.4% of HSCTs. Reduced-intensity conditioning (RIC) regimen was administered in 48.5% of patients. Acute graft-versus-host disease (aGVHD) occurred in 51.7% and chronic GVHD (cGVHD) in 39.7% of patients. Analysis of chimerism was performed through amplification of 9 specific short tandem repeats by polymerase chain reaction at engraftment and 1, 6, and 12 months after HSCT. Upon first evaluation, complete chimerism (CC) was detected in 34.7% and mixed chimerism (MC) in 55.4%, whereas graft failure occurred in 9.9% of patients. Severe aGVHD was associated with CC (P = .031). The last chimerism evaluation showed CC in 72.1%, stable MC in 12.8%, and progressive MC in 3.5%. CC was associated with a higher incidence of aGVHD (P = .016) and cGVHD (P = .022), whereas the RIC regimen was associated with graft failure (P = .026). One- and 3-year overall survival (OS) was 87.4% and 80.5%, respectively, with a lower OS at 3 years in patients with CC compared with those with MC (P = .008). aGVHD and cGVHD represent factors favoring CC, thus close, careful follow-up of chimerism is recommended in patients affected by nonmalignant disease.

Monitoring of Busulphan Concentrations in Children Undergone Hematopoietic Stem Cell Transplantation: Unicentric Experience over 10 years.

BACKGROUND AND OBJECTIVES: The aim of this report is to describe the experience in the management of busulphan-based conditioning regimen administered before hematopoietic stem cell transplantation (HSCT) in children. METHODS: We report the values of the first dose AUC (area under the concentration-time curve, normal target between 3600 and 4800 ng·h/mL) in children treated with oral and intravenous busulphan, and we analyze the impact of some clinical variables in this cohort of patients. RESULTS: 82 children treated with busulphan before HSCT were eligible for the study: 57 received oral busulphan with a mean AUC of 3586 ng·h/mL, while 25 received intravenous busulphan with a mean AUC of 4158 ng·h/mL. Dose adjustment was based on first dose AUC. The dose was increased in 36 children (43.9%) and decreased in 26 patients (31.7%). Age at HSCT (P = 0.015), cumulative dose of busulphan as mg/m2 (P < 0.001), busulphan dose prescribed as mg/Kg (P = 0.001), intravenous busulphan administration (P < 0.001), type of stem source cells (P = 0.016), and type of HSCT (P = 0.03) were associated with AUC levels. No statistically significant differences were found between transplant-related toxicity, acute and chronic graft versus host disease, engraftment, and AUC levels. CONCLUSIONS: We concluded that older age at HSCT, intravenous administration of busulphan, cumulative, and prescribed dose of busulphan are associated with higher AUC levels. The absence of significant correlations between toxic events, graft failure, and AUC suggests the efficacy of busulphan concentrations monitoring in our patients.

Allogeneic hematopoietic stem cell transplantation in congenital disorders: A single-center experience.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the treatment of choice for a variety of congenital disorders. We report the experience of children affected by congenital diseases other than bone marrow failure syndromes who received allo-HSCT over a period of 25 years at G. Gaslini Paediatric Research Institute. HSCTs were performed in 57 children with congenital diseases (25 with congenital immunodeficiencies, 10 with severe combined immunodeficiencies, and 22 with metabolic diseases). Overall survival rate at 3 years in the whole group of patients was 76.9%, with a trend in favor of better outcome in children with metabolic diseases and in those who received cord blood cells (85.9%) vs bone marrow cells (72.4%).

Italian Advisory Board: sFlt-1/PlGF ratio and preeclampsia, state of the art and developments in diagnostic, therapeutic and clinical management.

Extensive research has been published, showing the usefulness of angiogenic markers in both diagnosis and subsequent prediction and management of preeclampsia and placenta-related disorders. Recent evidence provides a helpful cut off for the Elecsys ratio sFlt-1 to PlGF, that predicts preeclampsia development in women with sign and symptoms, before its clinical onset in the short term. In Europe, no accordance exists for the use of such kind of test in clinical practice; only German guidelines have recently taken it into account, as a diagnostic aid for preeclampsia, in conjunction with other clinical findings. This panel of Italian experts recently met, in order to review the literature and to promote the evaluation of the clinical utility of sFlt-1/PlGF ratio at the Italian country level, as regards: prediction of preeclampsia during the first trimester, prediction or exclusion of new onset or recurrence in patients with risk factors for preeclampsia, triage of patients suffering from gestational hypertension, evaluation of disease severity, prediction of adverse maternal and fetal outcomes.

An unusual pattern of B-cell immunological reconstitution after allogeneic stem cell transplantation: a possible correlation with CMV reactivation?

IR after HSCT is a slow process that involves several components of the immune response, and, in allogeneic setting, it can be delayed by GvHD and immuno-suppressive therapy. Our study on IR post-HSCT included a child with FA who underwent MUD transplantation. To evaluate B, T and NK cell reconstitution and to investigate the differentiation of B lymphocyte repertoire, this patient was carefully monitored at various time points by IgHCDR3 (third complementarity determining region of the immunoglobulin heavy chain) fingerprinting and by FACS analysis. IgHCDR3 fingerprinting showed a strong oligoclonality of IgM and IgG profiles from day +60 to +180 post-transplant. CMV reactivation was present at the same time points and overlapped the clonal pattern shown in IgHCDR3 fingerprinting. Immunophenotype analysis showed early repopulation of T and NK cells following HSCT, whereas B cells increased first at one yr post-transplant. The overlapping of virus reactivation and B-cell clonal expansion seems to suggest that B lymphocytes may be involved in the CMV immunological response, at least in the early time points after HSCT when the immune repertoire is still reconstituting.

Unrelated hematopoietic stem cell transplantation for Cernunnos-XLF deficiency.

Cernunnos-XLF deficiency is a rare CI characterized by a defective DNA DSB repair mechanism. Its clinical manifestations are growth retardation, dysmorphic features, malformations, and severe B- and T-cell lymphopenia. BM failure may complicate the clinical picture. To date, there have been no described patients with CSy undergoing allogeneic HSCT. We report a case of CSy treated successfully with unrelated allogeneic HSCT after a reduced-intensity conditioning regimen. Two yr after HSCT, the patient maintains full donor engraftment, normal hematopoiesis, and progressively improving immune competence, thus suggesting that HSCT may be the treatment of choice for CSy.

V(H)3 and V(H)6 immunoglobulin M repertoire reconstitution after hematopoietic stem-cell transplantation in children.

BACKGROUND: Immune reconstitution after hematopoietic stem-cell transplantation (HSCT) occurs gradually. Thus, a variable period of immunodeficiency may be present, leading to immunomediated complications, such as graft-versus-host disease (GVHD) and opportunistic infections. METHODS: To better understand the kinetics of B-cell repertoire reconstitution in children, 49 pediatric patients were analyzed before and after transplantation by immunoglobulin (Ig) HCDR3 fingerprinting, which is a molecular technique that analyzes one of the hypervariable segments of the Ig heavy chain, which provides the amino acid residues that are essential to interact with antigens. RESULTS: In healthy donors, the CDR3 fingerprinting profile shows 16 to 20 bands, and each band corresponds to a particular length of CDR3. This situation is considered polyclonal. Patients analyzed just after transplantation show strong oligoclonality, because only a few CDR3 bands are detected within the first 3 to 6 months. CONCLUSIONS: The authors' data show a significant lag in diversification of the B-cell repertoire, which reaches the polyclonal situation of normal healthy donors approximately 6 months after HSCT. This period may vary depending on the type of transplant (autologous vs. allogeneic) and on the immunosuppressive therapy related to GVHD.

B-cell repertoire reconstitution after hematopoietic stem cell transplantation in children evaluated by immunoglobulin heavy chain third complementarity determining region fingerprinting.

Immunodeficiency after hematopoietic stem cell transplantation leads to a high risk of opportunistic infections. We evaluated B-lymphocyte reconstitution in 36 children by heavy chain third complementarity determining region (CDR3)-fingerprinting and immunophenotypic analysis. The time necessary to return to the normal immunoglobulin heavy chain-CDR3 polyclonal situation was basically related to the type of transplant and this process did not recapitulate fetal ontogenesis.

Prospective molecular monitoring of BCR/ABL transcript in children with Ph+ acute lymphoblastic leukaemia unravels differences in treatment response.

Children with Philadelphia-chromosome-positive (Ph+) acute lymphoblastic leukaemia (ALL) represent a subgroup at very high risk for treatment failure, despite intensive chemotherapy. However, recent retrospective studies showed that Ph+ childhood ALL is a heterogeneous disease with regard to treatment response. We have prospectively monitored, by reverse transcription polymerase chain reaction (RT-PCR) during follow-up, the presence of the BCR/ABL fusion transcript in Ph+ ALL children diagnosed in the Italian multicentre Associazione Italiana Ematologia Oncologia Pediatrica ALL-AIEOP-95 therapy protocol. To our knowledge, this is the first report on the evaluation of minimal residual disease (MRD) in childhood Ph+ ALL prospectively enrolled in an intensive, Berlin-Frankfurt-Munster (BFM)-type treatment protocol. Twenty-seven of 36 (75.0%) Ph+ patients consecutively enrolled into the high-risk group of the AIEOP-ALL protocol between May 1995 and October 1999 were successfully analysed. Twenty were good responders to the pre-phase of prednisone/intrathecal methotrexate treatment (PGR) and seven were poor responders (PPR). Within the PPR group, the RT-PCR monitoring constantly showed positivity for the BCR/ABL fusion transcript and all the patients died of disease progression. In contrast, highly sensitive qualitative RT-PCR monitoring revealed heterogeneity within the PGR group of Ph+ childhood ALL patients. Three different subgroups could be defined, according to the clearance of Ph+ cells within the first 5 months of treatment. This provides useful information on the capability of chemotherapy to reduce the leukaemic clone, with prognostic implications.