ALBERT

Description: Isolated congenital asplenia (ICA) is the only known human developmental defect exclusively affecting a lymphoid organ. In 2013, we showed that private deleterious mutations in the protein-coding region of RPSA, encoding ribosomal protein SA, caused ICA by haploinsufficiency with complete penetrance. We reported seven heterozygous protein-coding mutations in 8 of the 23 kindreds studied, including 6 of the 8 multiplex kindreds. We have since enrolled 33 new kindreds, 5 of which are multiplex. We describe here 11 new heterozygous ICA-causing RPSA protein-coding mutations, and the first two mutations in the 5′-UTR of this gene, which disrupt mRNA splicing. Overall, 40 of the 73 ICA patients (55%) and 23 of the 56 kindreds (41%) carry mutations located in translated or untranslated exons of RPSA. Eleven of the 43 kindreds affected by sporadic disease (26%) carry RPSA mutations, whereas 12 of the 13 multiplex kindreds (92%) carry RPSA mutations. We also report that 6 of 18 (33%) protein-coding mutations and the two (100%) 5′-UTR mutations display incomplete penetrance. Three mutations were identified in two independent kindreds, due to a hotspot or a founder effect. Finally, RPSA ICA-causing mutations were demonstrated to be de novo in 7 of the 23 probands. Mutations in RPSA exons can affect the translated or untranslated regions and can underlie ICA with complete or incomplete penetrance.

Description: Evans syndrome (ES) is a rare severe autoimmune disorder characterized by the combination of autoimmune hemolytic anemia and immune thrombocytopenia. In most cases, the underlying cause is unknown. We sought to identify genetic defects in pediatric ES (pES), based on a hypothesis of strong genetic determinism. In a national, prospective cohort of 203 patients with early-onset ES (median [range] age at last follow-up: 16.3 years ([1.2-41.0 years]) initiated in 2004, 80 nonselected consecutive individuals underwent genetic testing. The clinical data were analyzed as a function of the genetic findings. Fifty-two patients (65%) received a genetic diagnosis (the M+ group): 49 carried germline mutations and 3 carried somatic variants. Thirty-two (40%) had pathogenic mutations in 1 of 9 genes known to be involved in primary immunodeficiencies (TNFRSF6, CTLA4, STAT3, PIK3CD, CBL, ADAR1, LRBA, RAG1, and KRAS), whereas 20 patients (25%) carried probable pathogenic variants in 16 genes that had not previously been reported in the context of autoimmune disease. Lastly, no genetic abnormalities were found in the remaining 28 patients (35%, the M− group). The M+ group displayed more severe disease than the M− group, with a greater frequency of additional immunopathologic manifestations and a greater median number of lines of treatment. Six patients (all from the M+ group) died during the study. In conclusion, pES was potentially genetically determined in at least 65% of cases. Systematic, wide-ranging genetic screening should be offered in pES; the genetic findings have prognostic significance and may guide the choice of a targeted treatment.

Description: Introduction: CD38 is a type II transmembrane glycoprotein expressed on immature T and B lymphocytes (thymocytes and hematogones), NK cells, activated T-cells, plasma-cells, monocytes and red blood cells. This ectoenzyme is a ribonucleosyl cyclase (Cyclic ADP ribose hydrolase) involved in the regulation of calcium fluxes. CD38 is absent from quiescent lymphocytes. The lowest levels are present on erythrocytes while CD38 is brightly expressed by hematogones and plasma-cells. The latter has led to the development of daratumumab, a therapeutic monoclonal antibody used in the treatment of multiple myeloma. Because of the wide distribution of this antigen, other diseases could be considered for such a therapy. Among them, acute lymphoblastic leukemia (ALL) could represent an interesting target. We thus investigated the level of expression of CD38 in a cohort of 128 samples from B-lineage ALL (113 at diagnosis and 15 at relapse). We also compared it to that of the normal counterpart of these blasts, hematogones, because of the potential use of CD38 as a leukemia associated immunophenotype (LAIP). Moreover, in 15 paired samples of diagnosis/relapse, we examined the stability of this expression during disease evolution. Patients, Material and Methods: A total of 62 females and 66 males were included in the cohort, with a median age of 20 years old (range 4 months to 90 years). There were 57 children (below 15 yo), 13 adolescents and young adults (15-25 yo) and 58 adults. CD38 expression was investigated in 45 bone marrow (BM) samples and in 83 peripheral blood (PB) samples. The median level of blasts was 62% in BM and 48% in PB. According to EGIL classification, there were 19 B-I, 66 B-II, 38 B-III, 2 B-IV and intracytoplasmic mu chain was not investigated in 3 cases. In parallel, 26 samples of BM with morphologically evidence hematogones were used to compare the level of expression of CD38 on these cells. Immunophenotyping panels comprised CD38 antibodies conjugated to allophycocyanin and all samples were analyzed on a Canto II flow cytometer (BD biosciences, San Jose, CA). Results: CD38 was always present on hematogones. It was expressed by 122 of the 128 B-ALL samples tested (95,3%). Partial expression, between 20 and 70% of the blasts was noted for 9 patients (7%) while 113 patients (88%) had more than 70% positive blasts. For 102 patients (79,7%), the whole population was stained (100% of the blasts). Among the 15 samples obtained at relapse, CD38 was always expressed, partially only in one case. The mean fluorescence intensity (MFI), expressed using the flow cytometer arbitrary units (AI) ranged between 510 and 1396 AI (median 878 AI) in the group of patients with 20-70% CD38+ blasts (n=9) and between 317 and 26466 AI (median 4704 AI) for the 113 patients with more than 70% CD38+ blasts (n=113). The level of fluorescence was always higher on hematogones compared to blasts, with a median MFI of 14915 AI (range 3834-34501 AI). CD38 expression is also used as a LAIP, to discriminate minimal residual disease (MRD) from regenerating hematogones. In our department, a threshold of 6300 AI (median of hematogones - 2 SD) is used to define the LAIP of CD38low blasts. In the cohort reported here, 82 patients presented this LAIP (64%). Conversely, high expression of CD38 with an MFI 〉10 000 AI was present for 23 of the 128 patients. For the 15 patients with paired samples of diagnosis and relapse, 9 had low levels of CD38 (LAIP). In none of these cases modulation of CD38 was observed, this LAIP remaining stable over time. Conclusion: CD38 has been reported by several authors as one of the best LAIP marker for the detection of MRD in B-ALL, allowing a good follow-up of the patients because of its stability. Moreover, this study confirms that CD38 could be a valuable therapeutic target in most of B-ALL cases, being expressed in over 95% of the cases. Used together with chemotherapy, daratumumab could thus be useful to treat B-ALL both in first line or at relapse. Disclosures Moreau: Amgen: Honoraria; Bristol-Myers Squibb: Honoraria; Novartis: Honoraria; Celgene: Honoraria; Takeda: Honoraria; Janssen: Honoraria, Speakers Bureau.

Description: Background Cranial radiotherapy (CRT) is associated with early and late side effects. Intrathecal (IT) and systemic chemotherapy could successfully replace CRT in most protocols for standard risk ALL. However, in medium and high risk ALL patients (pts) its omission is still debatable. Aim We investigated the long-term outcome, the occurrence of second malignant neoplasms (SMN) and the incidence of late toxicities in pts randomized for receiving or not CRT in the EORTC 58832 study. Methods From 1983 to 1989, ALL children under 18 years (yrs) were included in EORTC Children Leukemia Group BFM-oriented studies, either 58831, for standard risk pts (Riehm-Langerman Risk Factor (RF) 〈 1.2), or 58832, for medium risk (RF 1.2-1.69) and high risk pts (RF ≥1.7). Pts with central nervous system (CNS) involvement at diagnosis were ineligible. The present report focusses on pts included in the 58832 trial (randomized for receiving or not prophylactic CRT). Prophylactic CNS therapy consisted of 4 high-dose methotrexate (HD-MTX) injections (2500 mg/m2) during consolidation and 7 IT MTX injections scheduled during the treatment period. Pts still in complete remission (CR) after the end of late intensification were randomized for receiving prophylactic CRT (standard arm) or not (experimental arm) before the start of continuation therapy. Dose of CRT was age dependent: 24 Gy (〉 2 yrs), 20 Gy (1-2 yrs) and 16 Gy (〈 1 yr). Endpoints were: disease-free survival (DFS) (event: relapse, death in CR), incidence of SMN, event-free survival (EFS) (event: relapse, death in CR, SMN), incidence of late toxicities, and overall survival (OS) from randomization. Results 788 pts were included in the 58831/58832 study. Among them, 189 were randomized in the 58832 study to receive CRT (n=93) or No CRT (n=96). A total of 6 pts did not meet eligibility criteria, 2 had an early relapse, 3 had an early protocol violation and 2 refused allocated treatment. Finally, 176 randomized pts were included in the analyses: 84 in the CRT group and 92 in the No CRT group. The median follow-up was 20 yrs (range 4-32 yrs). Omission of CRT did not increase the 25-yr incidence of isolated CNS relapse, any CNS relapse or non-CNS relapse (4.8 vs 6.5; 11.9 vs 8.7 and 16.7 vs 21.8 in the CRT vs No CRT arms, respectively). No relapses occurred after 10 yrs. The 25-yr DFS rates were similar in both arms: 70.2% with CRT and 67.4% without CRT; No CRT vs CRT hazard ratio (HR)=1.08, 95% CI (0.63, 1.83). CRT was associated with an increase of the 25-yr SMN incidence: 13.2% with CRT and 3.9% without CRT. In the CRT arm, 9 pts (10.7%) developed SMN: 2 acute myeloid leukemias (AML), 1 non-Hodgkin lymphoma, 1 thyroid carcinoma, 4 meningiomas and 1 malignant histiocytosis. One SMN (meningioma) occurred after a CNS combined relapse. Three pts developed second SMN (meningiomas): 1 after an AML and 2 after a first meningioma. In the No CRT arm, 3 pts (3.3%) had SMN: 1 pleomorphic xanthoastrocytoma, 1 melanoma and 1 adenocarcinoma of the ileum. One SMN occurred after a bone marrow (BM) relapse. The 25-yr EFS rates were similar in both arms: 60.3% with CRT and 63.2% without CRT, HR=0.90, 95% CI (0.55, 1.46). CRT was also associated with an increase of late CNS and endocrine toxicities. Five pts (19.2% of the pts with available data) developed leukoencephalopathy in the CRT arm, versus 2 pts (8.7%) in the No CRT arm. Noteworthy, 1 of those 2 pts received CRT for a BM relapse, while the other received total body irradiation for a CNS relapse. Stroke was observed in 2 pts (7.7%) who received CRT. In contrast, there was no clear increase of the incidence of cognitive disturbance after CRT: 33.3% in the CRT arm vs 25.0% in the No CRT arm. Regarding endocrine toxicities, GH deficiency, hypothyroidism and precocious puberty were more frequent in the CRT arm: 53.1% vs 29.6%, 27.8% vs 0% and 29.4% vs 0%, respectively. Finally, the 25-yr OS rates were similar in both arms: 78.5% with CRT and 78.1% without CRT, HR=1.00, 95% CI (0.53, 1.88). Conclusion In medium and high risk pts without CNS involvement at diagnosis and treated with HD-MTX in the EORTC trial 58832 (1983-1989), omission of CRT did not increase the risk of CNS or non-CNS relapse. On long-term evaluation, CRT was associated with a higher incidence of SMN, late CNS and endocrine toxicities. These long-term results indicate that prophylactic CRT can be safely omitted in childhood medium and high risk ALL pts receiving IT and systemic chemotherapy (including HD-MTX) as CNS prophylaxis. Table Table. Disclosures No relevant conflicts of interest to declare.

Description: Background Hemophagocytic lymphohistiocytosis (HLH) is an inflammatory condition caused by uncontrolled proliferation of activated lymphocytes and macrophages secreting an excess of inflammatory cytokines. When untreated, primary HLH is invariably fatal. Treatment requires the achievement of remission of HLH prior to allogeneic hematopoietic stem cell transplantation. Despite significant treatment progress, pre-HSCT mortality remains a challenge. In the Etoposide-based HLH-94 and HLH-2004 studies pre-HSCT mortality was 27% and 19%, respectively. A better understanding of the pathophysiology of primary HLH has opened new avenues for targeted immunotherapy. Based on our previous observation concerning the use of Antithymoglobulin in HLH, we propose a new therapeutic strategy with Alemtuzumab in association with steroids and cyclosporine A (CSA) as first line treatment in primary HLH. In contrast to ATG, Alemtuzumab does not activate T lymphocytes while killing them. Therefore, we expect a better tolerance and efficacy of Alemtuzumab. This may have a positive impact not only on survival until HSCT, but also on overall survival and quality of life, especially with regard to long-term neurological sequelae. Methods 24 consecutive treatment naïve patients with genetically confirmed primary HLH had received first line Alemtuzumab in association to steroids and CSA from 01/2009 to 06/2015 in the Unit for Pediatric Immunology in Necker Hospital Paris, as well as two additional patients in 10/2016 and 10/2018 respectively, who could not be included in the prospective trial. From 06/2015 to 06/2019, 29 patients have been enrolled in a multicenter, open, phase I/II, non-comparative, non randomized study (NCT02472054). Patients with lymphohistiocytic activation syndrome who had not received any specific treatment prior to enrollment except steroids and CSA were included. Treatment consisted in intravenous administration of Alemtuzumab in association to Methylprednisolone and CSA. The primary outcome measures is the number of surviving patients until HSCT, secondary outcome measures the number of complete remissions following treatment at Day (D)14, D21, D28. To assess the efficacy of the Alemtuzumab, the time of delay between the first administration of Alemtuzumab and complete remission will be determined. Alemtuzumab Pharmacokinetics will be done. All adverse events are reported. Results Retrospective analysis of 26 patients (pilot study): The median age of patients was 1.9 months (birth - 7 years), 6 patients were neonates. When Alemtuzumab was started, out of 26 patients 12 (46.1%) required intensive care, 8 (30.7%) mechanical ventilation, 13 (50%) had neurological involvement, 9 (34.6%) hepatocellular insufficiency. One 2-month-old Munc13-4 patient died at H+48 after two administrations of Alemtuzumab (total dose 1.5mg/kg) for hepatic failure and acute renal failure. A second patient with Perforin deficiency did not respond neither to three courses of Alemtuzumab (cumulative dose 6.5mg/kg) nor repeated Etoposid, 40mg/kg ATG, or Ruxulotinib. He died at D+65. The 24 remaining patients survived until HSCT (survival 92.3%). As shown in the figure, two patients required additional treatment. Overall 22 patients achieved CR, 2 PR at the time of HSCT. The prospective study enrolled 29 patients from 06/2015 to 06/2019. Median age at onset of HLH was 0.5 years (range 0.02 to 17.2 years), one patient withdrawed consent. 12 patients received one course, 13 two, 2 three and one patient 4 courses of Alemtuzumab. 24 patients with a genetic confirmed HLH predisposition reached the primary endpoint with 22 surviving until HSCT (91,6%). One patient is still awaiting HSCT. The three remaining patients are one CA-EBV patient and a newborn with secondary HLH due to fulminant HSV hepatitis, who both died, as well as a patient with predominant neurological HLH without genetic diagnosis who is in sustained remission without any specific treatment. Detailed results from the completed study will be presented. Conclusions This is the first report on Alemtuzumab as first line approach in the treatment of primary HLH. Our results in more than 50 pediatric patients treated in a pilot study and prospective trial indicate that Alemtuzumab allows controlling HLH activity with a favorable safety and tolerability profile in a very fragile population. 92.3% and 91.6% of patients respectively survived to HSCT. Figure Disclosures No relevant conflicts of interest to declare. Off Label Disclosure: Alemtuzumab (Campath) has been used in a prospective trial to evaluate its efficacy as first line treatment in Familial Lymphohistiocytosis.