Abstract: Increased production of inflammatory cytokines and myeloid-derived suppressor cells occurs in patients with coronavirus disease 2019. These inversely correlated with perforin-expressing natural killer (NK) and CD3+ T cells. We observed a lower number of perforin-expressing NK cells in intensive care unit (ICU) patients compared with non-ICU patients, suggesting an impairment of the immune cytotoxic arm as a pathogenic mechanism.

Abstract: Interferon-γ (IFNγ) is the pivotal mediator in murine models of primary haemophagocytic lymphohistiocytosis (pHLH). Given the similarities between primary and secondary HLH (sec-HLH), including macrophage activation syndrome (MAS), we investigate the involvement of the IFNγ pathway in MAS by evaluating levels of IFNγ and of the induced chemokines, and their relation with laboratory parameters of MAS in systemic juvenile idiopathic arthritis (sJIA) patients with MAS and in a murine MAS model. Methods The Luminex multiplexing assay was used to assess serum levels of interleukin (IL)-1β, IL-6, IFNγ and of the IFNγ-induced chemokines CXCL9, CXCL10 and CXCL11 in patients with sec-HLH (n=11) and in patients with sJIA (n=54), of whom 20 had active MAS at sampling. Expression of IFNγ-induced chemokines was assessed in IL-6 transgenic mice in which MAS is induced by TLR4 stimulation with lipopolysaccharide. Results Levels of IFNγ and of IFNγ-induced chemokines were markedly elevated during active MAS and sec-HLH and were significantly higher in patients with MAS compared with active sJIA without MAS. Levels in patients with active sJIA without MAS were comparable to those of patients with clinically inactive sJIA. During MAS, ferritin and alanine transferase levels and neutrophil and platelet counts were significantly correlated with serum levels of IFNγ and CXCL9. In murine MAS, serum levels of ferritin were significantly correlated with mRNA levels of Cxcl9 in liver and spleen. Conclusions The high levels of IFNγ and of IFNγ-induced chemokines and their correlation with the severity of laboratory abnormalities of MAS suggest a pivotal role of IFNγ in MAS.

Abstract: Introduction: MAS is a severe, life-threatening complication of rheumatic diseases that occurs most frequently in patients with sJIA. The mainstay of treatment for MAS is high dose glucocorticoids (GCs); however, GCs do not always provide adequate control in all patients. Additional treatments are used without a standardized approach; however, morbidity and mortality remain high. Data from animal models of MAS and from observational studies in patients suggest that overproduction of IFNγ is a driver of the hyperinflammation observed in MAS; neutralization of IFNγ has been shown to revert the signs and symptoms of MAS in murine models, and high IFNγ levels are strongly correlated with laboratory parameters of disease severity in patients. Objective: To assess the efficacy and safety of intravenous (IV) infusions of emapalumab, a fully human anti-IFNγ monoclonal antibody, in patients with MAS associated with sJIA. Methods: This was a pilot, open-label, single-arm, phase 2 study (NCT03311854) that included patients with MAS (2016 ACR/EULAR criteria) associated with sJIA and who had not responded adequately to high dose IV GC and other treatments. Emapalumab was initiated at a dose of 6 mg/kg on Day 0 and continued at 3 mg/kg every 3 days until Day 15, and then twice weekly for an additional 2 weeks (i.e. until Day 28). As per protocol, 10 infusions were planned over the 4 weeks; however, treatment could be shortened if MAS remission was achieved earlier, or extended if required to achieve response. All patients were followed up for 4 weeks after the last emapalumab infusion and were offered to enter a long-term, follow-up study. Results: We report preliminary results from 14 patients with a median age of 11 (range 2-25) years who were enrolled in the trial (11 in Europe and 3 in the USA). All patients had failed high-dose GC; in addition, several patients had received cyclosporine A and/or anakinra within 10 days of being enrolled in the study. Treatment with emapalumab resulted in rapid IFNγ neutralization, as demonstrated by a decrease in the levels of the IFNγ-induced chemokine (C-X-C motif) ligand 9 (CXCL9), and subsequent deactivation of T cells, as indicated by the decrease in soluble interleukin-2 receptor levels. A progressive improvement in all clinical and laboratory parameters of MAS was observed. Emapalumab infusions were well tolerated by all patients, with no discontinuations. A cytomegalovirus reactivation was reported in 1 patient as a serious event related to emapalumab and resolved with antiviral therapy. During the study, GC doses were tapered in all patients. Administration of anakinra for the treatment of underlying sJIA was maintained/introduced during the study, as required. Conclusion: Emapalumab administration led to rapid neutralization of IFNγ, as indicated by a rapid decrease in CXCL9 levels, and was efficacious in controlling MAS in all patients. Emapalumab was well tolerated and had a favorable safety profile. These results support the pathogenic role of IFNγ in MAS/sJIA and the therapeutic value of IFNγ neutralization in MAS patients who have failed high-dose GCs and other treatments. Disclosures De Benedetti: Novimmune: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Sanofi: Consultancy, Research Funding; Roche: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Sobi: Consultancy, Research Funding; AbbVie: Consultancy, Research Funding. Grom: AB2 Bio: Consultancy; Novartis: Consultancy. Brogan: Sobi: Consultancy; Novartis: Consultancy; Roche: Consultancy; UCB: Consultancy. Eleftheriou: Sobi: Speakers Bureau. Papadopoulou: Sobi: Speakers Bureau. Quartier: Pfizer: Consultancy, Speakers Bureau; AbbVie: Consultancy, Speakers Bureau; Chugai-Roche: Consultancy, Speakers Bureau; Lilly: Consultancy, Speakers Bureau; Novartis: Consultancy, Speakers Bureau; Sobi: Consultancy, Speakers Bureau. Antón: Sobi: Consultancy; Novartis: Consultancy; Roche: Consultancy; Pfizer: Consultancy; GSK: Consultancy; AbbVie: Consultancy. Frederiksen: Sobi: Current Employment. Asnaghi: Sobi: Current Employment. Ballabio: Sobi: Consultancy, Ended employment in the past 24 months. de Min: Sobi: Consultancy, Ended employment in the past 24 months. OffLabel Disclosure: Emapalumab is an IFN-y blocking antibody indicated for the treatment of adult and pediatric (newborn and older) patients with primary hemophagocytic lymphohistiocytosis (HLH) with refractory, recurrent, or progressive disease or intolerance with conventional HLH therapy.

Abstract: Background: Primary HLH is a rare, life-threatening immune disorder characterized by a hyperinflammatory state. In patients with primary HLH, interferon gamma (IFNy) is often markedly elevated and is considered one of the key cytokine driving the hyperinflammatory state. The treatment goal of primary HLH is to stabilize the disease by controlling the associated hyperinflammation to bring patients to allogeneic hematopoietic stem cell transplantation, the only curative therapy. Conventional HLH therapy comprises immunotherapies (namely, dexamethasone and etoposide), which, unfortunately, predispose patients to infections and toxicity. Emapalumab is a fully human, anti-IFNy monoclonal antibody that neutralizes IFNy. Currently, there is no regulatory precedent or validated response criteria for efficacy assessment to guide clinical trials in primary HLH. In the pivotal study of emapalumab in primary HLH, objective response criteria were used to define the primary endpoint of overall response (Locatelli et al NEJM 2020). These response criteria were defined based on the Histiocyte Society HLH diagnostic criteria (Henter et al Pediatr Blood Cancer 2007), clinical considerations from the study's Scientific Steering Committee, and available experience reported with conventional HLH treatments. We now report on findings of a sensitivity analysis of overall response rate (ORR) to emapalumab using various assessment criteria. Methods: The open-label pivotal study included patients aged ≤18 years with a diagnosis of primary HLH and active disease (NCT01818492; Locatelli et al NEJM 2020). The initial dose of emapalumab was 1 mg/kg given intravenously every 3 days. Subsequent doses could be increased to 10 mg/kg if required, based on predefined laboratory and clinical response parameters, for a treatment duration of 8 weeks. In addition to emapalumab, all patients received dexamethasone, and a protocol amendment allowed for concomitant use of other HLH treatments if deemed appropriate by the investigator. ORR at end of treatment was analyzed as per the protocol definition in the 27 patients previously treated with conventional therapies. In addition, several pre-specified and post hoc sensitivity analyses were performed to pressure test the data; including: (i) a pre-specified analysis using a more conservative approach where any patient who received concomitant HLH therapies during the study was imputed as non-responder; (ii) a pre-specified analysis with physician-reported response rates recorded by the study investigators, based on their clinical judgement and previous experience in treating patients with primary HLH; and (iii) a post hoc sensitivity analysis using a previously published definition of overall response (Henter et al Pediatr Blood Cancer 2007). Results: 63% (95% confidence interval [CI], 0.42, 0.81) of 27 treatment-experienced patients had a response according to the pivotal study protocol definition of ORR (Fig. 1). A pre-specified sensitivity analysis on the primary endpoint where any patient who received concomitant HLH therapy and imputed as non-responders showed a magnitude of response similar to that observed in the protocol-defined primary analysis (59.3%; 95% CI 0.39, 0.78; n=22). Use of the response criteria defined by Marsh et al (Pediatr Blood Cancer 2013) in a retrospective analysis of 27 patients with primary HLH also resulted in a similar ORR to the protocol-defined primary endpoint in treatment-experienced patients (70.4%; 95% CI 0.50, 0.86). When platelet count was added to this analysis, the percentage of responders to emapalumab increased to 74.1% (95% CI 0.54, 0.89). The pre-specified analysis of physician-reported response rates was also in line with the primary analysis, with 70.4% (95% CI 0.50, 0.86) of 27 treatment-experienced patients deemed to have a response to emapalumab. Conclusion: The current analyses using different definitions of treatment response support the primary analysis results by having a numerically comparable point estimate to the primary endpoint, therefore confirming the positive benefit of emapalumab in patient's refractory or intolerant to conventional HLH therapies. Taken together, these findings also suggest that the clinically objective ORR, utilized in the pivotal emapalumab trial, may be used as a primary endpoint in primary HLH. Disclosures Locatelli: Jazz Pharmaceeutical: Speakers Bureau; Medac: Speakers Bureau; Miltenyi: Speakers Bureau; Bellicum Pharmaceutical: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Amgen: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Jordan:Sobi: Consultancy. Allen:Sobi: Other: Scientific Steering Committee, Data And Safety Monitoring. Rizzari:Sobi: Consultancy, Other: Advisory Board. Rao:Sobi: Consultancy, Other: Advisory Board. Sevilla:Amgen: Other: Advisory Board; Rocket Pharma: Consultancy; Sobi: Other: Advisory Board; Novartis: Other: Advisory Board. Henry:Sobi: Consultancy. De Benedetti:Abbvie: Research Funding; F Hoffmann-La Roche AG: Research Funding; Novartis Pharma: Research Funding; Pfizer: Research Funding; Sanofi-Aventis: Research Funding; Sobi: Consultancy, Research Funding. Grom:Sobi: Consultancy; Novartis Pharma: Consultancy; AB2Bio: Consultancy. Stoltenberg:Sobi: Current Employment. Vågerö:Sobi: Consultancy. de Min:Sobi: Consultancy.

Abstract: Introduction Primary HLH (pHLH) is a rare immune regulatory disorder invariably lethal if untreated. It is driven by pathologic immune activation, leading to the development of fever, splenomegaly, cytopenias and coagulopathy, which ultimately may cause multi-organ failure (MOF) and death. Based on data from murine models of primary and secondary HLH (sHLH), and observational studies in patients with HLH, the high production of IFNγ is thought to be a critical factor driving development of the disease. Immune-chemotherapy, primarily etoposide-based regimens, is at present the only pharmacological approach able to control HLH and bring patients to curative allogeneic hematopoietic stem cell transplant (allo-HSCT). In spite of recent attempts to further intensify treatment regimens, mortality and morbidity remain high, in part due to drug-related toxicities. NI-0501 is a fully human, high affinity, anti-IFNγ mAb that binds to and neutralizes human IFNγ, offering a novel and targeted approach for the control of HLH. Methods An open-label Phase 2 study has been conducted in United States and Europe to evaluate safety and efficacy of NI-0501 in children with either confirmed or suspected pHLH. NI-0501 was administered at initial dose of 1 mg/kg every 3 days, with possible dose increase guided by PK data and/or clinical response in each patient, on initial background dexamethasone dose of 5-10 mg/m2. Treatment duration ranged from 4 to 8 weeks. Ability to move to allo-HSCT, relevant HLH disease parameters, and 8-week survival were assessed. Study Population A total of 13 patients were enrolled: 8F/5M, median age 1.0y (range 2.5mo-13y). Twelve patients received NI-0501 as a second line treatment after having received conventional therapy and either reactivating, obtaining unsatisfactory response, or being intolerant to therapy. One patient was treated with NI-0501 in first line. Nine patients carried a known HLH genetic defect (3 FHL2, 2 FHL3, 2 GS-2, 1 XLP1, 1 XLP2). The majority of patients were at the severe end of HLH spectrum, in compromised general condition (2 patients in ICU) and carrying significant toxicities from previous HLH treatments. At baseline ferritin was elevated in 12/13 patients and sCD25 in 8, cytopenias were present in 10 patients, splenomegaly in 8, hypofibrinogenemia and hypertriglyceridemia in 9. Liver and CNS involvement were present in 7 and 3 patients, respectively. Results Overall, NI-0501 treatment significantly improved parameters of HLH disease activity (Fig. 1), and 9 of 13 patients achieved a satisfactory response. Seven patients have proceeded to allo-HSCT. Allo-HSCT is planned for two patients with good HLH control upon identification of an appropriate donor. In one patient (who achieved disease control with first line NI-0501) allo-HSCT is not yet planned given the absence of a causative HLH gene mutation. Eleven of 13 patients were alive at 8 weeks. Two patients died of HLH/MOF at day 53 and 36, respectively. CNS signs and symptoms resolved in the 2 evaluable patients. Greater than 50% reduction of dexamethasone dose was possible in 50% of patients during the first 4 weeks of NI-0501 treatment. IFNγ neutralization was demonstrated by a sharp decrease in CXCL9, a chemokine exquisitely IFNγ-induced. CXCL9 levels tightly correlated with IFNγ production, estimated by measuring IFNγ bound to NI-0501 (Fig. 2), suggesting CXCL9 as a potential new biomarker for HLH. NI-0501 was well tolerated and no safety concern was identified. None of the infections known to be favored by IFNγ neutralization were reported, and no infections occurred in patients who had not previously received chemotherapy. Seven patients reported at least one SAE, all assessed by the DMC as not related to NI-0501 administration. No unexpected events attributable to “off target” effects of NI-0501 (myelotoxicity, hemodynamic alterations) were observed. Conclusions Neutralization of IFNγ by NI-0501 offers an innovative targeted and potentially less toxic approach to HLH management. The results of this study show that NI-0501 is a safe and effective therapeutic option in patients with pHLH who have demonstrated unsatisfactory response, or are intolerant, to conventional therapy. Furthermore, therapy with NI-0501 was not associated with any of the typical short- or long-term toxicities reported for etoposide-based regimens. Assessment of NI-0501 as first line treatment for pHLH is ongoing. Disclosures: Jordan: Novimmune: Consultancy. Allen:Roche: Consultancy , Other: unpaid ; NovImmune: Consultancy , Other: unpaid. De Benedetti:Novimmune: Consultancy. Grom:Novartis: Consultancy , Research Funding ; Novimmune: Consultancy. Ballabio:Novimmune SA: Employment. Ferlin:Novimmune SA: Employment. De Min:Novimmune SA: Employment.

Abstract: Introduction: Primary hemophagocytic lymphohistiocytosis (pHLH) is a rare, genetic life-threatening syndrome characterized by hyper-inflammation that is mainly driven by high production of interferon (IFN)- 𝛾 , leading to the development of fever, splenomegaly, cytopenias and coagulopathy. There are currently no approved treatments for HLH, and recent attempts to improve the dexamethasone/etoposide-based regimen (HLH-94) did not show a significant improvement in overall probability of survival. Emapalumab (NI-0501) is a fully human, anti-IFN- 𝛾 monoclonal antibody that binds to and neutralizes IFN- 𝛾 and is in development for treatment of HLH. Methods: This open-label pivotal study (NCT01818492) includes patients ≤18 years with a diagnosis of pHLH based on genetic confirmation, family history, or the presence of ≥5 of the 8 HLH-2004 diagnostic criteria. Patients were either treatment-naïve or had failed previous conventional HLH therapy prior to study entry. The emapalumab initial dose was 1 mg/kg given intravenously every 3-4 days. Subsequent doses could be increased up to 10 mg/kg based on the evolution of clinical and laboratory response parameters. Emapalumab was administered concomitantly with 5 to10 mg/m2/day of dexamethasone which could be tapered during the study. Treatment duration was 8 weeks (with possible shortening to a minimum of 4 weeks). Treatment could be extended up to allogeneic hematopoietic stem cell transplantation (HSCT) whenever needed. The primary efficacy endpoint of the study was overall response at end of treatment assessed by pre-defined objective parameters. Overall Response Rate (ORR) was assessed as normalization or at least 50% improvement from baseline of fever, splenomegaly, cytopenias, hyperferritinemia, fibrogen and/or D-Dimer levels, central nervous system (CNS) abnormalities, with no sustained worsening of sCD25 serum levels. The primary analysis used an exact binomial test to evaluate the null hypothesis that ORR be at most 40% at a one-sided 0.025 significance level. Data presented are from 34 patients of whom 27 entered the study after failing conventional HLH therapy. Following completion of the main study patients entered into an extension phase (NCT02069899). The data cut-off applied is July 20 2017. Results: Patient characteristics are summarized in Table 1. Disease presentation at study entry was consistent with the broad spectrum of pHLH abnormalities, both in terms of HLH-2004 diagnostic criteria and other known HLH features; over 30% of patients had signs and/or symptoms of CNS disease. Efficacy results are summarized in Table 2. ORR was significantly higher than the pre-specified null hypothesis of 40%; thus the primary endpoint was met. The response rate based on investigator's clinical judgement was 70.6% and 70.4% in the two groups. Emapalumab infusions were in general well tolerated, with mild to moderate infusion-related reactions reported in 27% of patients. The observed safety events pre-HSCT conditioning mostly included HLH manifestations, infections or toxicities due to other administered drugs. Infections caused by pathogens potentially favored by IFN- 𝛾 neutralization occurred in 1 patient during emapalumab treatment (Disseminated histoplasmosis), and resolved with appropriate treatment. No off-target effects were observed. Conclusions: This is the first prospective HLH study that reports response rates based on pre-defined objective criteria. Our results indicate that emapalumab should be considered as a new therapeutic option in pHLH thanks to its targeted mode of action. Treatment with emapalumab was able to control HLH activity with a favorable safety and tolerability profile in a very fragile population. The majority of patients proceeded to HSCT with favorable outcome. Disclosures Jordan: Novimmune: Consultancy, Membership on an entity's Board of Directors or advisory committees. Allen:Novimmune: Membership on an entity's Board of Directors or advisory committees. Sevilla:Rocket Pharmaceuticals Inc: Honoraria, Patents & Royalties; Novimmune: Other: currently participating in and have participated in Novimmune-sponsored clinical trials within the past two years . Grom:Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; AB2Bio: Consultancy, Membership on an entity's Board of Directors or advisory committees; NovImmune: Consultancy, Membership on an entity's Board of Directors or advisory committees. De Benedetti:Novartis: Consultancy, Research Funding; SOBI: Consultancy, Research Funding; Roche: Consultancy, Research Funding; Sanofi: Consultancy, Research Funding; UCB: Consultancy; Eli-Lylli: Consultancy; Abbvie: Research Funding; Novimmune: Research Funding; Pfizer: Research Funding. Ferlin:Novimmune: Employment, Equity Ownership, Patents & Royalties. Ballabio:Novimmune: Employment, Equity Ownership. De Min:Novimmune: Employment, Equity Ownership.

Abstract: Autoinflammatory diseases can cause irreversible tissue damage due to systemic inflammation. Recently, the Autoinflammatory Disease Damage Index (ADDI) was developed. The ADDI is the first instrument to quantify damage in familial Mediterranean fever, cryopyrin-associated periodic syndromes, mevalonate kinase deficiency and tumour necrosis factor receptor-associated periodic syndrome. The aim of this study was to validate this tool for its intended use in a clinical/research setting. Methods The ADDI was scored on paper clinical cases by at least three physicians per case, independently of each other. Face and content validity were assessed by requesting comments on the ADDI. Reliability was tested by calculating the intraclass correlation coefficient (ICC) using an ‘observer-nested-within-subject’ design. Construct validity was determined by correlating the ADDI score to the Physician Global Assessment (PGA) of damage and disease activity. Redundancy of individual items was determined with Cronbach’s alpha. Results The ADDI was validated on a total of 110 paper clinical cases by 37 experts in autoinflammatory diseases. This yielded an ICC of 0.84 (95% CI 0.78 to 0.89). The ADDI score correlated strongly with PGA-damage (r=0.92, 95% CI 0.88 to 0.95) and was not strongly influenced by disease activity (r=0.395, 95% CI 0.21 to 0.55). After comments from disease experts, some item definitions were refined. The interitem correlation in all different categories was lower than 0.7, indicating that there was no redundancy between individual damage items. Conclusion The ADDI is a reliable and valid instrument to quantify damage in individual patients and can be used to compare disease outcomes in clinical studies.