Laboratory parameters such as level of fibrinogen, ferritin, C-reactive protein and lymphocyte counts may help to distinguish Hemophagocytic lymphohistiocytosis (HLH) from COVID-19-related multisystem inflammatory syndrome in children (MIS-C).

Most children with a SARS-CoV-2 infection are asymptomatic or exhibit mild symptoms (1, 2). However, in April 2020, a rare but serious clinical entity has been described in children, encompassing symptoms of hyperinflammation, resembling toxic shock syndrome (TSS) or Kawasaki disease (KD) (3–5), now generally referred to as either multisystem inflammatory syndrome in children (MIS-C) or pediatric inflammatory multisystem syndrome—temporally associated with SARS-CoV-2 (PIMS-TS). In May 2020, the World Health Organization (WHO) has published a MIS-C case definition for patients under the age of 19 years with fever ≥3 days and fulfilling 4 out of the 4 following criteria: (i) at least two clinical signs of multisystem inflammatory involvement (ii) elevated markers of inflammation (iii) no other microbial cause of inflammation—including bacterial sepsis and TSS—and (iv) evidence of current or past SARS-CoV-2 infection (https://www.who.int/publications/i/item/multisystem-inflammatory-syndrome-in-children-and-adolescents-with-covid-19). Some of the typical laboratory findings of MIS-C include elevated C-reactive protein (CRP), ferritin, D-dimers and fibrinogen, troponin and N-terminal pro brain natriuretic peptide (NT-pro BNP), high neutrophil to lymphocyte ratio and low platelets (6).

Hemophagocytic lymphohistiocytosis (HLH) resembles MIS-C. Diagnosis of HLH is defined by the presence of 5 out of the following 8 criteria: (i) fever, (ii) hemophagocytosis in bone marrow or other organs, (iii) bicytopenia, (iv) enlarged spleen, (v) ferritin > 500 ug/L, (vi) soluble IL-2 receptor >2,400 U/mL, (vii) hypofibrinogenemia or hypertriglyceridemia, and (viii) decreased NK-cell cytotoxicity (7). HLH may be secondary to other diseases, but in every patient fulfilling the diagnostic criteria of HLH, inborn defects in lymphocyte cytotoxic activity should be considered, also known as primary HLH (8). This is paramount given that early diagnosis is crucial for a successful outcome. Primary HLH includes X-linked lymphoproliferative disease (XLP1)—a rare immunodeficiency characterized by immune dysregulation—caused by mutations in the SH2 domain–containing protein 1A (SH2D1A) gene, which encodes signaling lymphocytic activation molecule (SLAM)–associated protein (SAP). Clinical manifestations of XLP1 include lymphoma, dysgammaglobulinemia and HLH, the latter often triggered by Epstein-Barr virus (EBV) infection (9).

Here we report a case of fatal HLH with liver failure, at first considered to be MIS-C, in a patient tested positive for COVID-19, later diagnosed with EBV infection and underlying XLP1.

The previously healthy 6-year-old boy presented at our emergency department after a history of 10 days of fever. Five days earlier, the diagnosis of a SARS-CoV-2 infection had been made through a positive reverse transcription polymerase chain reaction (RT-PCR) from a nasopharyngeal swab. On admission, physical exam was remarkable for fever (39.8°C), tachycardia (150 bpm), hypotension (75/38 mmHg), tachypnea (50/min), and abdominal pain (Table 1). Chest radiography was compatible with right lower lobe pneumonia and moderate pleural effusion (Figure 1A). Abdominal ultrasound revealed hepatosplenomegaly with ascites. Echocardiographic investigation showed normal cardiac function and absence of coronary abnormalities. Blood tests revealed anemia (hemoglobin 108 g/L), thrombocytopenia (103 G/L), and normal neutrophil and lymphocyte counts (Table 2). While CRP was normal, ESR (36 mm/h) and ferritin (3,995 ug/L) were elevated. Liver parameters were abnormal (ALT 511 U/L, bilirubin 75 umol/L, GGT (gamma-glutamyltransferase) 396 U/L, and albumin 22 g/L) and there was profound hyponatremia (124 mmol/L). Troponin and NT-pro BNP were normal. Fibrinogen was decreased (1.09 g/L) with elevated D-dimers (6.4 mg/L). Multiplex PCR was negative for respiratory viruses including SARS-CoV-2 on admission and SARS-CoV-2 antibodies were later found to be absent.

The patient fulfilled the diagnostic criteria of MIS-C at time of admission and was transferred to intensive care. Empiric antibiotic treatment was started and, as he developed acute respiratory distress in the context of pulmonary edema (Figure 1B) and assumed MIS-C, treatment with intravenous immunoglobulins (IVIG; 2 g/kg) and methylprednisolone (30 mg/kg/day) was initiated. Twenty-four hours later, in the absence of clinical improvement, therapy was escalated with interleukin (IL)-1 receptor antagonist anakinra (8–9 mg/kg/day s.c.). The patient's clinical condition continued to worsen. He developed liver failure, lactacidemia and profound coagulopathy, highly suggestive of HLH. Therefore, anakinra was stopped and treatment with anti-human-T-lymphocyte-immunoglobulin (ATG) and ciclosporin initiated. Suspicion of primary HLH was something later strengthened by the detection of very high EBV copy numbers in the blood (20 × 10⁶ copies/mL). Rituximab was added once EBV results were available. Despite added support using molecular adsorbent recirculation system (MARS), the patient's condition further deteriorated and he succumbed on day 8 after admission. Immunological investigations on day 6 had shown normal CD107a expression in NK cell degranulation assay and normal intracellular perforin expression by flow cytometry. However, an X-linked mutation (c.163C > T; p.R55X; position on chrX:124365786; reference genome: GRCh38.p12) in the SAP/ SH2 domain–containing protein 1A (SH2D1A) gene was found, which has previously been described to be disease-causing. Thus, the diagnosis of an X-linked lymphoproliferative syndrome type 1 (XLP1) was made post-mortem. The mother is carrier of a de novo heterozygous mutation. A detailed timetable of the patient's clinical history is shown in Table 3.

Our patient initially presented with fever, pneumonia and rapidly evolving ARDS. Due to previous infection with SARS-CoV-2 the suspected initial diagnosis was MIS-C. He ultimately succumbed secondary to liver failure as a consequence of EBV-triggered HLH. These two disease entities, MIS-C and HLH, have an overlapping clinical spectrum but when comparing our patient's laboratory findings with the ones of a total of 58 previously described MIS-C patients (6), there were some key distinguishing findings: (i) Fibrinogen is usually elevated in MIS-C patients. This is in contrast to hypofibrinogenaemia found in our patient. (ii) MIS-C patients usually have low lymphocytes, while total lymphocyte counts were normal in our patient. (iii) Elevated levels of ferritin are common to both entities but, excessively high concentrations, with a 10-fold increase compared to MIS-C patients, were observed in our patient (6). (iv) All MIS-C patients reported by Whittaker et al. had CRP levels > 100 mg/L whereas our patient's CRP was normal, possibly secondary to liver failure.

It is important to evaluate clinical and laboratory findings in a patient with suspected MIS-C very carefully, as PCR might be already negative and you may have to wait for serological results.

Whether SARS-CoV-2 infection may have triggered this fulminant EBV infection is speculative: viral clearance could have occurred during these days. Serology might have remained negative, as dysgammaglobulinemia is a known feature in about one third of XLP1-patients.

Initial treatment for both diseases consists of corticosteroids, in MIS-C patients also of high-dose IVIG. Prior HLH treatment protocols have included IVIG too, so initial anti-inflammatory and immunomodulatory treatment regimens are similar. In patients with MIS-C not responding to initial treatment, escalation includes the use of biologicals, such as IL-1 or IL-6 blocking agents. For primary HLH, there are several treatment schemes available, including VP-16, alemtuzumab, anti- thymoglobulin and ciclosporin. If there is evidence of EBV-driven disease, treatment with a monoclonal anti-CD20 antibody (rituximab) has been used successfully. Recently, treatment with Janus kinase inhibitors (e.g., ruxolitinib) has also shown promising results (10). In primary HLH, the aim of treatment is to induce remission from the hyperinflammatory state, but only hematopoietic stem cell transplantation is curative.

It remains unclear whether our patient's SARS-CoV-2 infection had an impact on the course of disease. We believe that it was a simple coincidence, but it may be possible that the negative serology for SARS-CoV-2 (despite positive anti-EBV IgM) may be partly explained by the underlying disease, as antibody deficiency is a possible feature in XLP1 (9).

XLP1 may manifest in male patients at the same age as typical MIS-C and shares some clinical and laboratory features, especially at presentation. Clinical course and some laboratory parameters such as level of fibrinogen, ferritin, C-reactive protein and lymphocyte counts may help to distinguish HLH from MIS-C. Elevated aminotransferases in patients with MIS-C have been shown to be associated with more severe disease and higher inflammatory parameters, but acute liver failure in both, MIS-C and HLH, is very rare. Primary HLH and XLP1 in particular, should be part of the differential diagnosis of MIS-C. This can be rapidly evaluated by measuring SAP-expression by flow cytometry. It is absolutely necessary to distinguish MIS-C from primary HLH as prognosis and management differ. Curative treatment by hematopoietic stem cell transplantation should be considered in primary HLH and genetic counseling of affected families is needed.

The parents have given consent and the patient was enrolled in an ongoing, institutional review board-approved study (PB_2016_02280) at the Division of Immunology at the University Children's Hospital Zurich, Switzerland; ClinicalTrials.gov. NCT02735824.

Whole exome sequencing and analysis of sequences have been performed as described previously (11).

The original contributions presented in the study are included in the article/supplementary materials, further inquiries can be directed to the corresponding author/s.

Written informed consent was obtained from the minor(s)' legal guardian/next of kin for the publication of any potentially identifiable images or data included in this article.

SP wrote the manuscript. NR, FB, FG, GB-R, MA, AW, PM, VM, and SG contributed to patients' care, data collection, and interpretation. MK contributed to writing of the manuscript. SV and TM carried out functional analysis of lymphocytes. LO performed bioinformatics analyses of exome sequencing data. JP coordinated the study, was involved in the care of the family, and wrote the manuscript. All authors provided critical feedback and contributed to the final version of the manuscript.

JP is member of a data monitoring committee for Leniolisib for Novartis and advisory board for Emapalumab for SOBI. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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