Outcome of primary hemophagocytic lymphohistiocytosis: a report on 143 patients from the Italian Registry.

Primary hemophagocytic lymphohistiocytosis (pHLH) is a severe, life-threatening hyperinflammatory syndrome caused by defects in genes of the granule-dependent cytotoxic pathway. Here we investigated the clinical presentation and outcome in a large cohort of 143 patients with pHLH diagnosed in the last 15 years and enrolled in the Italian registry. The median age at diagnosis was 12 months (IQR 2-81), and ninety-two patients (64%) fulfilled the HLH-2004 criteria. Out of 111 patients who received first-line combined therapy (HLH-94, HLH-2004, Euro-HIT protocols), 65 (59%) achieved complete response (CR) and 21 (19%) partial response (PR). Thereafter, 33 patients (30%) reactivated, and 92 (64%) received HSCT, 78 of whom (85%) survived and were alive at a median follow-up from diagnosis of 67 months. Thirty-six patients (25%) died before HSCT and 14 (10%) after. Overall, 93 patients (65%) were alive after a median follow-up of 30 months. Unadjusted predictors of non-response were age.

A Nationwide Study of GATA2 Deficiency in Italy Reveals Novel Symptoms and Genotype-phenotype Association.

GATA2 deficiency is a rare disorder encompassing a broadly variable phenotype and its clinical picture is continuously evolving. Since it was first described in 2011, up to 500 patients have been reported. Here, we describe a cohort of 31 Italian patients (26 families) with molecular diagnosis of GATA2 deficiency. Patients were recruited contacting all the Italian Association of Pediatric Hematology and Oncology (AIEOP) centers, the Hematology Department in their institution and Italian societies involved in the field of vascular anomalies, otorhinolaryngology, dermatology, infectious and respiratory diseases. Median age at the time of first manifestation, molecular diagnosis and last follow-up visit was 12.5 (age-range, 2-52 years), 18 (age-range, 7-64 years) and 22 years (age-range, 3-64), respectively. Infections (39%), hematological malignancies (23%) and undefined cytopenia (16%) were the most frequent symptoms at the onset of the disease. The majority of patients (55%) underwent hematopoietic stem cell transplantation. During the follow-up rarer manifestations emerged. The clinical penetrance was highly variable, with the coexistence of severely affected pediatric patients and asymptomatic adults in the same pedigree. Two individuals remained asymptomatic at the last follow-up visit. Our study highlights new (pilonidal cyst/sacrococcygeal fistula, cholangiocarcinoma and gastric adenocarcinoma) phenotypes and show that lymphedema may be associated with null/regulatory mutations. Countrywide studies providing long prospective follow-up are essential to unveil the exact burden of rarer manifestations and the natural history in GATA2 deficiency.

Severe Lactic Acidosis Caused by Thiamine Deficiency in a Child with Relapsing Acute Lymphoblastic Leukemia: A Case Report.

Lactic acidosis is characterized by an excessive production of lactic acid or by its impaired clearance. Thiamine deficiency is an uncommon cause of lactic acidosis, especially in countries where malnutrition is rare. We describe the case of a 5-year-old boy who presented with a central nervous system relapse of acute lymphoblastic leukemia. During the chemotherapy regimen, the patient developed drug-induced pancreatitis with paralytic ileus requiring prolonged glucosaline solution infusion. In the following days, severe lactic acidosis (pH 7.0, lactates 253 mg/dL, HCO3- 8 mmol/L) was detected, associated with hypoglycemia (42 mg/dL) and laboratory signs of acute liver injury. Due to the persistent hypoglycemia, the dextrose infusion was gradually increased. Lactates, however, continued to raise, so continuous venovenous hemodiafiltration was started. While lactates initially decreased, 12 h after CVVHDF suspension, they started to raise again. Assuming that it could have been caused by mitochondrial dysfunction due to vitamin deficiency after prolonged fasting and feeding difficulties, parenteral nutrition and thiamine were administered, resulting in a progressive reduction in lactates, with the normalization of pH during the next few hours. In the presence of acute and progressive lactic acidosis in a long-term hospitalized patient, thiamine deficiency should be carefully considered and managed as early as possible.

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.

Autoimmune Lymphoproliferative Syndrome (ALPS) Disease and ALPS Phenotype: Are They Two Distinct Entities?

Autoimmune lymphoproliferative syndrome (ALPS) is an inherited disorder of lymphocyte homeostasis classically due to mutation of FAS, FASL, and CASP10 genes (ALPS-FAS/CASP10). Despite recent progress, about one-third of ALPS patients does not carry classical mutations and still remains gene orphan (ALPS-U, undetermined genetic defects). The aims of the present study were to compare the clinical and immunological features of ALPS-FAS/CASP10 versus those of ALPS-U affected subjects and to deepen the genetic characteristics of this latter group. Demographical, anamnestic, biochemical data were retrieved from medical record of 46 ALPS subjects. An enlarged panel of genes (next-generation sequencing) was applied to the ALPS-U group. ALPS-U subjects showed a more complex phenotype if compared to ALPS-FAS/CASP10 group, characterized by multiorgan involvement (P = 0.001) and positivity of autoimmune markers (P = 0.02). Multilineage cytopenia was present in both groups without differences with the exception of lymphocytopenia and autoimmune neutropenia that were more frequent in ALPS-U than in the ALPS-FAS/CASP10 group (P = 0.01 and P = 0.04). First- and second-line treatments were able to control the symptoms in 100% of the ALPS-FAS/CASP10 patients, while 63% of ALPS-U needed >2 lines of treatment and remission in some cases was obtained only after target therapy. In the ALPS-U group, we found in 14 of 28 (50%) patients 19 variants; of these, 4 of 19 (21%) were known as pathogenic and 8 of 19 (42%) as likely pathogenic. A characteristic flow cytometry panel including CD3CD4-CD8-+TCRαß+, CD3+CD25+/CD3HLADR+, TCR αß+ B220+, and CD19+CD27+ identified the ALPS-FAS/CASP10 group. ALPS-U seems to represent a distinct entity from ALPS-FAS/CASP10; this is relevant for management and tailored treatments whenever available.

Long-term proliferation of immature hypoxia-dependent JMML cells supported by a 3D in vitro system.

Juvenile myelomonocytic leukemia (JMML) is a rare clonal stem cell disorder that occurs in early childhood and is characterized by the hyperactivation of the RAS pathway in 95% of the patients. JMML is characterized by a hyperproliferation of granulocytes and monocytes, and little is known about the heterogeneous nature of leukemia-initiating cells, as well as of the cellular hierarchy of the JMML bone marrow. In this study, we report the generation and characterization of a novel patient-derived three-dimensional (3D) in vitro JMML model, called patient-derived JMML Atypical Organoid (pd-JAO), sustaining the long-term proliferation of JMML cells with stem cell features and patient-specific hallmarks. JMML cells brewed in a 3D model under different microenvironmental conditions acquired proliferative and survival advantages when placed under low oxygen tension. Transcriptomic and microscopic analyses revealed the activation of specific metabolic energy pathways and the inactivation of processes leading to cell death. Furthermore, we demonstrated the pd-JAO-derived cells' migratory, propagation, and self-renewal capacities. Our study contributes to the development of a robust JMML 3D in vitro model for studying and defining the impact of microenvironmental stimuli on JMML disease and the molecular mechanisms that regulate JMML initiating and propagating cells. Pd-JAO may become a promising model for compound tests focusing on new therapeutic interventions aimed at eradicating JMML progenitors and controlling JMML disease.

PAX5 fusion genes are frequent in poor risk childhood acute lymphoblastic leukaemia and can be targeted with BIBF1120.

BACKGROUND: Despite intensive risk-based treatment protocols, 15% of paediatric patients with B-Cell Precursor Acute Lymphoblastic Leukaemia (BCP-ALL) experience relapse. There is urgent need of novel strategies to target poor prognosis subgroups, like PAX5 translocated. METHODS: We considered 289 childhood BCP-ALL cases consecutively enrolled in Italy in the AIEOP-BFM ALL2000/R2006 protocols and we performed extensive molecular profiling, integrating gene expression, copy number analyses and fusion genes discovery by target-capture NGS. We developed preclinical strategies to target PAX5 fusion genes. FINDINGS: We identified 135 cases without recurrent genetic rearrangements. Among them, 59 patients (43·7%) had a Ph-like signature; the remaining cases were identified as ERG-related (26%), High-Hyperdiploid-like (17%), ETV6::RUNX1-like (8·9%), MEF2D-rearranged (2·2%) or KMT2A-like (1·5%). A poor prognosis was associated with the Ph-like signature, independently from other high-risk features. Interestingly, PAX5 was altered in 54·4% of Ph-like compared to 16·2% of non-Ph-like cases, with 7 patients carrying PAX5 fusions (PAX5t), involving either novel (ALDH18A1, IKZF1, CDH13) or known (FBRSL1, AUTS2, DACH2) partner genes. PAX5t cases have a specific driver activity signature, extending to multiple pathways including LCK hyperactivation. Among FDA-approved drugs and inhibitors, we selected Dasatinib, Bosutinib and Foretinib, in addition to Nintedanib, known to be LCK ligands. We demonstrated the efficacy of the LCK-inhibitor BIBF1120/Nintedanib, as single agent or in combination with conventional chemotherapy, both ex vivo and in patient-derived xenograft model, showing a synergistic effect with dexamethasone. INTERPRETATION: This study provides new insights in high-risk Ph-like leukaemia and identifies a potential therapy for targeting PAX5-fusion poor risk group. FUNDING: Ricerca Finalizzata-Giovani Ricercatori (Italian Ministry of Health), AIRC, Transcall, Fondazione Cariparo.

Sirolimus Restores Erythropoiesis and Controls Immune Dysregulation in a Child With Cartilage-Hair Hypoplasia: A Case Report.

Cartilage-hair hypoplasia (CHH) is a syndromic immunodeficiency characterized by metaphyseal dysplasia, cancer predisposition, and varying degrees of anemia. It may present as severe combined immunodeficiency in infancy, or slowly progress until fully manifesting in late adolescence/adulthood. No targeted treatment is currently available, and patients are usually managed with supportive measures, or are offered a bone marrow transplant if the clinical phenotype is severe and a suitable donor is available. We report the case of a young girl presenting with transfusion-dependent erythropoietic failure and immunological features resembling autoimmune lymphoproliferative syndrome who responded well to empirical sirolimus. She later developed a marked growth delay, which was ultimately attributed to metaphyseal dysplasia. A diagnosis of CHH was reached through whole-genome sequencing (WGS), after a less sensitive genetic diagnostic strategy failed. The patient eventually underwent a haploidentical bone marrow transplant due to progressive combined immunodeficiency manifested as cryptococcal meningoencephalitis. This case illustrates the potential role of sirolimus in correcting anemia and partially controlling the immune aberrations associated with CHH, and serves as a reminder of the invaluable role of WGS in diagnosing patients with complex and atypical presentations.

Response to upfront azacitidine in juvenile myelomonocytic leukemia in the AZA-JMML-001 trial.

Allogeneic hematopoietic stem cell transplantation (HSCT) is the only curative therapy for most children with juvenile myelomonocytic leukemia (JMML). Novel therapies controlling the disorder prior to HSCT are needed. We conducted a phase 2, multicenter, open-label study to evaluate the safety and antileukemic activity of azacitidine monotherapy prior to HSCT in newly diagnosed JMML patients. Eighteen patients enrolled from September 2015 to November 2017 were treated with azacitidine (75 mg/m2) administered IV once daily on days 1 to 7 of a 28-day cycle. The primary end point was the number of patients with clinical complete remission (cCR) or clinical partial remission (cPR) after 3 cycles of therapy. Pharmacokinetics, genome-wide DNA-methylation levels, and variant allele frequencies of leukemia-specific index mutations were also analyzed. Sixteen patients completed 3 cycles and 5 patients completed 6 cycles. After 3 cycles, 11 patients (61%) were in cPR and 7 (39%) had progressive disease. Six of 16 patients (38%) who needed platelet transfusions were transfusion-free after 3 cycles. All 7 patients with intermediate- or low-methylation signatures in genome-wide DNA-methylation studies achieved cPR. Seventeen patients received HSCT; 14 (82%) were leukemia-free at a median follow-up of 23.8 months (range, 7.0-39.3 months) after HSCT. Azacitidine was well tolerated and plasma concentration--time profiles were similar to observed profiles in adults. In conclusion, azacitidine monotherapy is a suitable option for children with newly diagnosed JMML. Although long-term safety and efficacy remain to be fully elucidated in this population, these data demonstrate that azacitidine provides valuable clinical benefit to JMML patients prior to HSCT. This trial was registered at www.clinicaltrials.gov as #NCT02447666.

Thrombotic Microangiopathy Associated with Macrophage Activation Syndrome: A Multinational Study of 23 Patients.

OBJECTIVE: To describe the clinical characteristics, treatment, and outcomes of a multinational cohort of patients with macrophage activation syndrome (MAS) and thrombotic microangiopathy (TMA). STUDY DESIGN: International pediatric rheumatologists were asked to collect retrospectively the data of patients with the co-occurrence of MAS and TMA. Clinical and laboratory features of patients with systemic juvenile idiopathic arthritis (sJIA)-associated MAS and TMA were compared with those of an historical cohort of patients with sJIA and MAS. RESULTS: Twenty-three patients with MAS and TMA were enrolled: 17 had sJIA, 2 systemic lupus erythematosus, 1 juvenile dermatomyositis, 1 mixed connective tissue disease, and 2 undifferentiated connective tissue disease. Compared with the historical cohort of MAS, patients with sJIA with coexistent MAS and TMA had higher frequencies of renal failure and neurologic involvement, hemorrhage, jaundice, and respiratory symptoms, as well as more severe anemia and thrombocytopenia, higher levels of alanine aminotransferase, lactate dehydrogenase, bilirubin and D-dimer, and lower levels of albumin and fibrinogen. They also required admission to the intensive care unit more frequently. Among patients tested, complement abnormalities and reduced ADAMTS13 activity were observed in 64.3% and 44.4% of cases, respectively. All patients received glucocorticoids. Treatment for TMA included plasma-exchange, eculizumab, and rituximab. CONCLUSIONS: The possible coexistence of MAS and TMA in rheumatic diseases may be underrecognized. This association should be considered in patients with MAS who develop disproportionate anemia, thrombocytopenia, and lactate dehydrogenase increase, or have multiorgan failure.

Effect of Blinatumomab vs Chemotherapy on Event-Free Survival Among Children With High-risk First-Relapse B-Cell Acute Lymphoblastic Leukemia: A Randomized Clinical Trial.

Importance: Blinatumomab is a CD3/CD19-directed bispecific T-cell engager molecule with efficacy in children with relapsed or refractory B-cell acute lymphoblastic leukemia (B-ALL). Objective: To evaluate event-free survival in children with high-risk first-relapse B-ALL after a third consolidation course with blinatumomab vs consolidation chemotherapy before allogeneic hematopoietic stem cell transplant. Design, Setting, and Participants: In this randomized phase 3 clinical trial, patients were enrolled November 2015 to July 2019 (data cutoff, July 17, 2019). Investigators at 47 centers in 13 countries enrolled children older than 28 days and younger than 18 years with high-risk first-relapse B-ALL in morphologic complete remission (M1 marrow, <5% blasts) or with M2 marrow (blasts ≥5% and <25%) at randomization. Intervention: Patients were randomized to receive 1 cycle of blinatumomab (n = 54; 15 µg/m2/d for 4 weeks, continuous intravenous infusion) or chemotherapy (n = 54) for the third consolidation. Main Outcomes and Measures: The primary end point was event-free survival (events: relapse, death, second malignancy, or failure to achieve complete remission). The key secondary efficacy end point was overall survival. Other secondary end points included minimal residual disease remission and incidence of adverse events. Results: A total of 108 patients were randomized (median age, 5.0 years [interquartile range {IQR}, 4.0-10.5]; 51.9% girls; 97.2% M1 marrow) and all patients were included in the analysis. Enrollment was terminated early for benefit of blinatumomab in accordance with a prespecified stopping rule. After a median of 22.4 months of follow-up (IQR, 8.1-34.2), the incidence of events in the blinatumomab vs consolidation chemotherapy groups was 31% vs 57% (log-rank P < .001; hazard ratio [HR], 0.33 [95% CI, 0.18-0.61]). Deaths occurred in 8 patients (14.8%) in the blinatumomab group and 16 (29.6%) in the consolidation chemotherapy group. The overall survival HR was 0.43 (95% CI, 0.18-1.01). Minimal residual disease remission was observed in more patients in the blinatumomab vs consolidation chemotherapy group (90% [44/49] vs 54% [26/48]; difference, 35.6% [95% CI, 15.6%-52.5%]). No fatal adverse events were reported. In the blinatumomab vs consolidation chemotherapy group, the incidence of serious adverse events was 24.1% vs 43.1%, respectively, and the incidence of adverse events greater than or equal to grade 3 was 57.4% vs 82.4%. Adverse events leading to treatment discontinuation were reported in 2 patients in the blinatumomab group. Conclusions and Relevance: Among children with high-risk first-relapse B-ALL, treatment with 1 cycle of blinatumomab compared with standard intensive multidrug chemotherapy before allogeneic hematopoietic stem cell transplant resulted in an improved event-free survival at a median of 22.4 months of follow-up. Trial Registration: ClinicalTrials.gov Identifier: NCT02393859.

Underlying CTLA4 Deficiency in a Patient With Juvenile Idiopathic Arthritis and Autoimmune Lymphoproliferative Syndrome Features Successfully Treated With Abatacept-A Case Report.

BACKGROUND: Functional variants of the cytotoxic T-lymphocyte antigen-4 (CTLA4) could contribute to the pathogenesis of disorders characterized by abnormal T-cell responses. CASE PRESENTATION: We report a case of a 13-year-old girl who first presented with polyarticular juvenile idiopathic arthritis poorly responsive to treatment. During the following years the patient developed cytopenias, chronic lymphoproliferation, high values of T-cell receptor αß+ CD4- CD8- double-negative T cells and defective Fas-mediated T cells apoptosis. Autoimmune lymphoproliferative syndrome was diagnosed and therapy with mycophenolate mofetil was started, with good hematological control. Due to the persistence of active polyarthritis, mycophenolate mofetil was replaced with sirolimus. In the following months the patient developed hypogammaglobulinemia and started having severe diarrhea. Histologically, duodenitis and chronic gastritis were present. Using the next generation sequencing-based gene panel screening, a CTLA4 mutation was detected (p.Cys58Serfs*13). At the age of 21 the patient developed acute autoimmune hemolytic anemia; steroid treatment in combination with abatacept were started with clinical remission of all symptoms, even arthritis. CONCLUSIONS: Targeted immunologic screening and appropriate genetic tests could help in the diagnosis of a specific genetically mediated immune dysregulation syndrome, allowing to select those patients who can take advantage of target therapy, as in the case of abatacept in CTLA4 deficiency.

Hemolysis and Neurologic Impairment in PAMI Syndrome: Novel Characteristics of an Elusive Disease.

PSTPIP1-associated myeloid-related proteinemia inflammatory (PAMI) syndrome is a rare early-onset autoinflammatory disease associated with various hematologic findings, including chronic neutropenia and pancytopenia. We report a unique case of PAMI syndrome in a toddler with transfusion-dependent hemolytic anemia, hepatosplenomegaly, failure to thrive, developmental delay, and multiple malformations. Because of acute inflammatory-driven decompensation, anakinra was started with dramatic improvement of both the hematologic and neurologic involvement. A customized next-generation sequencing panel later identified a de novo pathogenic variant in the PSTPIP1 gene, confirming the diagnosis. Our case illustrates the broad spectrum of phenotypes associated with PAMI syndrome, which should be considered in any case of unexplained cytopenias associated with autoinflammatory stigmata. It is also one of the few reports of neurologic involvement in PSTPIP1-associated inflammatory diseases. Increased awareness of this rare disease and early performance of genetic testing can correctly diagnose PAMI syndrome and prevent disease complications.

Unusual Late-onset Enteropathy in a Patient With Lipopolysaccharide-responsive Beige-like Anchor Protein Deficiency.

In recent years, monogenic causes of immune dysregulation syndromes, with variable phenotypes, have been documented. Mutations in the lipopolysaccharide-responsive beige-like anchor (LRBA) protein are associated with common variable immunodeficiency, autoimmunity, chronic enteropathy, and immune dysregulation disorders. The LRBA protein prevents degradation of cytotoxic T-lymphocyte antigen 4 (CTLA4) protein, thus inhibiting immune responses. Both LRBA and CTLA4 deficiencies usually present with immune dysregulation, mostly characterized by autoimmunity and lymphoproliferation. In this report, we describe a patient with an atypical clinical onset of LRBA deficiency and the patient's response to abatacept, a fusion protein-drug that mimics the action of CTLA4.

Bortezomib-Responsive Refractory Anti-N-Methyl-d-Aspartate Receptor Encephalitis.

BACKGROUND: Anti-N-methyl-d-aspartate receptor encephalitis is a central nervous system inflammatory autoimmune disease affecting adults and children. The use of first- and second-line immunotherapies is supported. Recent reports suggest the efficacy of bortezomib in severe anti-N-methyl-d-aspartate encephalitis in adult patients not responsive to second-line treatment; there are no data about pediatric patients. PATIENT DESCRIPTION: We describe an eight-year-old child with anti-N-methyl-d-aspartate encephalitis not responsive to first- and second-line treatments who experienced marked clinical improvement after bortezomib administration. DISCUSSION: Bortezomib is a selective and reversible inhibitor of the 26S proteasome, which is used to treat oncologic and rare autoimmune disorders in pediatric patients. As observed in adult patients, bortezomib administration induced anti-N-methyl-d-aspartate antibody titer decline and clinical improvement with an acceptable risk profile. CONCLUSION: This is the first report of the use of bortezomib in children with anti-N-methyl-d-aspartate encephalitis; it could be a useful therapeutic option in children with refractory anti-N-methyl-d-aspartate encephalitis.