Secondary hemophagocytic lymphohistiocytosis in children (Review)

Abstract

Hemophagocytic lymphohistiocytosis (HLH) is a rare, life-threatening condition characterized by hyperinflammation in an uncontrolled and ineffective immune response. Despite great improvement in diagnosis and treatment, it still represents a challenge in clinical management, with poor prognosis in the absence of an aggressive therapeutic approach. The present literature review focuses on secondary HLH at pediatric age, which represents a heterogeneous group in terms of etiology and therapeutic approach. It summarizes the most recent evidence on epidemiology, pathophysiology, diagnosis, treatment and prognosis, and provides a detailed description and comparison of the major subtypes of secondary HLH. Finally, it addresses the open questions with a focus on diagnosis and new treatment insights.

1. Introduction

Hemophagocytic lymphohistiocytosis (HLH) is a rare, life-threatening condition characterized by hyperinflammation in an uncontrolled and ineffective immune response (1). It was initially described in early infancy (2), but it may develop at any age, from childhood to adulthood. The spectrum of clinical symptoms of the disease is wide (3,4), ranging from fever, cytopenia and splenomegaly to shock, disseminated intravascular coagulation and multiple organ failure (5). Despite improvement in diagnosis and treatment, HLH represents a challenge in clinical management as it still results in a poor prognosis in the absence of an aggressive therapeutic approach (6,7).

The classical description of the disease distinguishes between primary forms due to a known genetic mutation [familial HLH (FHL), and HLH in the context of congenital immunodeficiency] and secondary forms triggered by an external stimulus [secondary HLH (sHLH)] (7). This classification, though widely accepted and used, is not fully effective in clinical practice (8). Often, due to the severity of the clinical picture, it is necessary to start an aggressive treatment before having clarified the primary or secondary nature of the syndrome (9).

The ongoing advances in genetic diagnosis and a deeper understanding of the molecular pathophysiology suggest new strategies in clinical management (10).

The present study focuses on sHLH at pediatric age and explored its pathogenetic and therapeutic peculiarities. To the best of our knowledge, the current review provided for the first time an extensive and exhaustive synopsis of all the known etiologies with an in-depth discussion of the individual aspects, a detailed overview of the most recent evidence and a summary of the remaining open questions.

2. Search/selection criteria

The present study reviewed the literature available on PubMed (https://pubmed-ncbi-nlm-nih-gov.bvsp.idm.oclc.org/) from 1950 to May 2022 to find all published works on sHLH at pediatric age, including case reports, meta-analyses, randomized controlled trials, reviews or systematic reviews and official guidelines. The PubMed search string was: ‘Secondary HLH OR sHLH’. The PubMed age filter ‘child: birth-18 years’ was applied. Reference lists of all articles were manually searched for cross-references and additional articles were included, if relevant. The present study obtained the full text of each article. Exclusion criteria included: A text language other than English, a focus on adult patients and a focus on primary HLH forms. Each article was selected and analyzed for inclusion by two authors in parallel. If the authors disagreed, a third author was consulted, who was responsible for the final decision.

The website ‘Clinical trial.gov’ was searched for new trends in sHLH therapy. The research string was ‘HLH, hemophagocytic lymphohistiocytoses and hemophagocytic syndrome’. The Clinical trial.gov age filter ‘child: birth-17 years’ was applied. Exclusion criteria included a focus on conditioning regimes for hematopoietic stem cell transplantation (HSCT).

3. Results of literature search

The Pubmed search found 283 articles, of which 199 were considered relevant and included. The manual search for cross references included nine more articles for a total of 204 articles. Clinical trial.gov search found 75 studies, 17 of which were included.

4. Epidemiology

HLH is rare and likely underestimated due to its difficult diagnosis. From the institution of the first international HLH registry in 1989 to the HLH-2004 therapeutic study, >700 patients have been formally evaluated worldwide (6). In Europe and Japan an HLH incidence of 1-2 per million was reported in 2005(11), however, there is the possibility that the diagnosis is under-reported, especially in developing countries (12).

5. Clinical presentation

HLH should be suspected in all patients with prolonged high-grade fever associated with splenomegaly and multiple organ involvement (7). The clinical spectrum of the disease is wide, ranging from mild organ dysfunction to multiorgan failure requiring intensive care. Central nervous system (CNS) involvement is frequent and often severe, even though it is not included in the official diagnostic criteria (7). HLH is rare, and a high grade of suspicion is essential for diagnosis. Differentiation between primary and secondary forms is mandatory since they share the same clinical picture but often require different therapeutic approaches (10).

The typical patient is an infant under 1 year of age, ill-appearing (toxic aspect), occasionally with a critical sepsis-like aspect (8). A younger age at onset suggests an underlying genetic basis, as seen in FHL or in HLH forms arising from genetic primary immunodeficiencies; however, familial HLH can present at any age, including adulthood (13).

Progressive splenomegaly is typically observed in patients, and can be associated with hepatic involvement, neurological signs, respiratory and renal failure (14). Skin rashes, erythroderma, edema or petechiae have been reported (14). Lymphadenopathy is uncommon in patients and indicates a potential underlying lymphoma (15).

Typical laboratory findings include cytopenias (at least bilinear), hypertriglyceridemia, hypofibrinogenemia (suggestive of HLH in the context of general inflammation) and hyperferritinemia. Liver function tests are frequently altered (7). Leukocytosis is not typical of HLH [except in HLH-associated with defined rheumatological conditions/macrophage activation syndrome-HLH (MAS-HLH)] (7). Hemophagocytosis is frequently observed but is neither pathognomonic nor mandatory for diagnosis (7,16).

Atypical forms, usually seen in children older than 1 year, include isolated fever of unknown origin (FUOs), isolated CNS involvement (17) or isolated acute liver failure (18,19).

6. Pathophysiology

The pathogenesis of HLH has mostly been studied in FHL. An inherited defect in the perforin/granzyme pathway or in the fusion of cytotoxic lytic granules with the surface of natural killer (NK) cells causes, in the presence of an external trigger, an over-response by cytotoxic CD8⁺ T lymphocytes. Viral infections are the most common triggers (20).

Cytotoxic CD8⁺ T cells produce large amounts of INFγ, which in turn activates macrophages. Overstimulated macrophages release large amounts of inflammatory cytokines, such as IL-1β, IL-6, IL-12, IL-18 and TNFα. There is also an increased production of IL-10, with inhibitory activity, but not sufficient to limit the phenomenon. IL-12 and IL-18 produced by macrophages in turn stimulate CD8⁺ T cells, amplifying the inflammatory response (1). The resulting tissue damage causes a release of IL-33 and IL1-β, which further activates the macrophages. Activated macrophages engulfblood cells and produce large amounts of ferritin. The ‘cytokine storm’ causes all the clinical manifestations of HLH, from endothelial damage to coagulopathy and multi-organ failure (1,13,21-23). The central role of INFgamma in the pathogenesis of FHL has been demonstrated in a perforin-deficient mouse model (24).

A similar pathogenesis of HLH can also be observed in patients with primary immunodeficiencies involving granule trafficking or exocytosis, such as Hermansky-Pudlak syndrome type 2, Griscelli syndrome type 2 and Chediak-Higashi syndrome II, all with reduced cy-totoxic T lymphocytes (CTL) cytotoxicity (3,4). Advances in genetic diagnosis suggest that cell killing by CTLs and NK cells can be affected from mildly to severely, thus explaining the different HLH phenotypes as a continuum (25). The known mutations in genes related to granule-mediated killing account for FHL and primary forms in general (7). Minor alterations of the CTL and NK activity, together with an external trigger, account for sHLH, severe sepsis, multi-organ failure and HLH in the context of rheumatologic diseases (25).

In all cases, a cytokine storm causing a devastating inflammation is the primal agent of the multiorgan failure, regardless of the underlying defect (24-29) and HLH should be considered a clinical syndrome of hyperinflammation with different phenotypes (10).

Moreover, this finding likely explains the numerous similarities and overlaps between HLH and other systemic inflammatory syndromes, such as septic shock, cytokine release syn-drome (CRS) following viral infections (29) and acute liver failure (30).

In 1994, the Histiocyte Society introduced an etoposide-based treatment protocol for HLH, which is still the gold standard in the therapy of the disease (31). Etoposide, dexamethasone and cyclosporine act on the pathogenetic mechanisms of HLH, inhibiting T cells and the release of inflammatory cytokines. In this respect, etoposide has a very high immunosuppressive activity (32).

Another treatment capable of acting on the pathogenesis of HLH is ATG-based immuno-therapy which has shown efficacy as a first-line therapy for FHL (33). Epstein-Barr virus (EBV)-associated HLH can occur in X-linked lymphoproliferative type (XLP)1 and XLP2, where it is caused by signaling lymphocytic activation molecule-associated protein (SAP) or X-linked inhibitor of apoptosis protein (XIAP) deficiency and by the subsequent uncontrolled EBV infection (34), or in individuals without obvious immune defects. In the latter case, activation of cytotoxic CD8⁺ cells by EBV-infected B cells or a direct activation of CD8⁺ cells or NK cells by the virus has been hypothesized (35,36). The addition of rituximab to an etoposide-based therapy is effective in the treatment of EBV-HLH, reducing the infected B lymphocyte population and EBV load (37).

The case of malignancy-associated HLH includes two completely heterogeneous mechanisms. Firstly, when HLH is a manifestation of the underlying malignancy, particularly in the case of lymphomas, the overproduction of inflammatory cytokines such as INF-γ and TNF-α is likely caused by neoplastic cells (1,38). Secondly, when HLH develops as the consequence of chemotherapy, the process is more likely to be caused by the association of drug-induced CTL suppression and infection (39).

7. Diagnosis

HLH was first described in the 1950s (2), and the first attempt to provide homogeneous diagnostic criteria by the International Histiocyte Society goes back to 1991(40). The HLH 2004 international trial established a clear and small set of diagnostic criteria that is currently the most used (31). Those diagnostic criteria do not include frequent features of the disease such as CNS involvement, and their specificity remains unknown. Furthermore, they are often inconclusive in differentiating primary and secondary forms of HLH in intensive care settings (41).

In order to accelerate diagnosis, rapid screening by flow cytometry has been used to identify some forms of p-HLH through the assessment of expression of intracellular perforin, SAP, and XIAP protein in peripheral blood cells (42) while waiting for the results of the genetic tests. Moreover, some simplified clinical algorithms have been proposed, for example, the ‘H score’ (43,44) scoring system and the ‘Minimal Parameter set’ (45), but none of these scores are conclusive in differentiating secondary HLH forms that might benefit from a different treatment.

Another notable proposal came from the North American Consortium for Histiocytosis (NACHO) in 2019(8): Patients fulfilling the HLH 2004 criteria should be diagnosed as having a ‘HLH syndrome’, while the subsequent diagnosis of ‘HLH disease’ should be restricted to the patients who additionally show distinctive immune dysregulation (FLH, HLH associated with immune compromise, rheumatological HLH/MAS) and are likely to benefit from deep immune suppression. Disorders leading to the HLH clinical symptoms, but not likely to benefit from immune suppression, should be diagnosed as ‘HLH disease mimics’ (such as infection-related HLH and iatrogenic HLH). This proposal clarifies diagnostic and therapeutic priorities, but some of the suggested categories still overlap (such as infection associated-HLH, malignancy-associated HLH and storage disorders) (41).

The most recent consensus guidelines on recognition, diagnosis and management of HLH in critically ill children (46) stress the importance of a high suspicion rate for HLH in all patients admitted to intensive care units who show a disproportionate inflammatory response and/or rapid clinical deterioration. Those patients should be thoroughly investigated for possible triggers of sHLH and treated aggressively.

New diagnostic parameters

Imaging techniques, such as CT and 18F-fluorodeoxyglucose (FDG) positron emission tomography-computed tomography (PET-CT) have been traditionally used in diagnosis to exclude an underlying malignancy. These imaging techniques can also be useful in raising HLH suspicion in the context of FUOs by showing an inflammatory disease involving bone marrow, lymph nodes and the spleen (47,48). Moreover, some 18F-FDG PET/CT metabolic parameters can help identify the etiology of sHLH in children (for example, malignancy and EBV-HLH), and provide directions for further inspection and information about the prognosis (47-49).

A high sCD25/ferritin ratio has been observed in malignancy-associated HLH (M-HLH) secondary to a lymphoma (50) and in HLH secondary to multisystem Langerhans cell histiocytosis (51), and may represent a novel useful marker for the malignancy-associated subset of sHLH (50,51).

A Chinese group suggested that using the T helper (Th)1/Th2 cytokine profile (dosage of sCD25/IFN-γ signature, TNF, IL-10, IL-6, IL-4 and IL-2 on cytometric bead array technique) for early quick diagnosis of HLH. This has demonstrated that a significant increase of IFN-γ and IL-10 and a slight increase of IL-6 is an early, specific and prognostic cytokine pattern for childhood HLH (52). More recently, to discriminate FHL from sHLH, lower IL-4 and IFN-γ levels have been demonstrated to more likely indicate primary HLH (53). This quick cytokine profile seems also useful in discriminating HLH vs. sepsis (54), and to identify EBV-HLH (55), so that a combined approach HLH2004 and Th1/Th2 profile is currently available.

Other new diagnostic tools are being developed, even though they are currently used in preclinical or research settings. Some relevant examples include studies on IFNγ and its ‘signature’ (CXCL9 and CXCL10) as serum biomarkers of disease activity in both primary HLH (pHLH) and sHLH (56), or as predictors of predominant liver involvement in sHLH (57), or studies on soluble TNF-like weak inducer of apoptosis selective elevation in HLH patients (58) and studies on secretory sphingomyelinase upregulation in HLH, with the C16 ceramide:sphingosine ratio determining a poorer prognosis (59).

Genetic analysis of polymorphisms of the TNF-α promoter (60) or of the interferon regulatory factor 5 gene (61) have identified variants that increase susceptibility to sHLH/MAS-HLH.

Determination of the NK-cell dysfunction type (type 1-4) and of the underlying genetic defect has been used to guide HSCT indication (62) and to estimate the risk of progression to acute leukemia (63).

Finally, targeted sequencing of FHL genes appears to be insufficient to identify pathogenic mechanisms in the majority of patients with HLH, and whole exome sequencing (WES) is increasingly used. WES is rapidly expanding the range of causal mutations, with a double sided effect: It accelerates diagnosis in a subset of patients, but has redefined numerous other cases of secondary HLH as primary HLH (54). Furthermore,a proved mutation in HLH-related gene cannot exclude the coexistence of an underlying malignancy (41).

8. Classification

The most recent international classification of HLH is the one provided in 2016 by the Histiocyte Society in the ‘revised classification of histiocytoses and neoplasms of the macrophage-dendritic cell lineages’ (64). In 2019 the NACHO (8) introduced the concept of ‘HLH syndrome’ and proposed to categorize HLH subtypes by specific etiologic associations instead of classifying HLH as ‘primary’ or ‘secondary’. The two classifications substantially overlap, so the present study referred to the 2016 Histiocyte Society classification because it is the most widely accepted and used to date (64).

Infection-associated HLH

Infection-associated HLH represents a challenge in classification. This is because infectious diseases can trigger HLH in both primary and secondary forms, and because septic shock can either be a presentation of HLH syndrome or may mime it, with great implications in therapeutic approach. A thorough infectious screening is thus highly recommended when facing an HLH syndrome (7,8).

Among the case reports the present study reviewed, the most frequent infectious triggers were viruses, such as the herpetic viruses, dengue virus, Crimean Congo hemorrhagic fever virus, Eastern equine encephalitis virus, hepatitis A virus (HAV), hepatitis B virus (HBV), human immunodeficiency virus (HIV), adenovirus, parvovirus B19, measles, influenza virus and severe acute respiratory syndrome coronavirus 2 (Sars-Cov2).

EBV has been frequently reported from all across the world, and especially from the Asian Countries (37,65-73). The reason for this geographical heterogeneity may involve a higher virulence in EBV viral strains circulating in Asia (74), or a different immune predisposition in the Asian patients, but this issue has not been clarified yet. EBV typically targets B cells, but in a subset of patients with EBV-HLH, of prevalent Asian origin again, it infects T or NK cells leading to oligoclonal or monoclonal proliferation and massive cytokine production (69,75). In extreme cases, EBV-HLH clinical picture can be difficult to differentiate from T cell lymphoproliferative disorder (69). EBV-HLH is usually quite aggressive, with a frequent involvement of CNS. Once EBV is demonstrated by serology tests or molecular biology methods, a targeted approach is recommended (74,76). Clinical scores to differentiate low risk vs. high-risk patients have been proposed (76), but have not achieved conclusive results yet. Targeted treatment is discussed later in this review.

Cytomegalovirus (CMV) is typical of newborns and immunocompromised patients (72,73,77-81) and so is the varicella zoster virus (82). Dengue virus plays a significant role in tropical countries (73,83-90). Tick-borne diseases include the Crimean-Congo hemorrhagic fever nairovirus (91,92) and the Eastern equine encephalitis virus (93). Sporadic case reports include HLH related to HAV (90,94), HBV (95), HIV (89,96), Adenovirus (72,89,97,98), Parvovirus B19(72), Measles (88) and Influenza virus (99-101).

Sars-Cov2 infection, causative of the recent COVID19 pandemic, has been reported to also be a HLH trigger, mostly in adults (102,103). In 2021, a Mexican group reported the unusual case of a 7-years old boy that developed a severe inflammatory syndrome triggered by Sars-Cov-2 and Dengue virus coinfection (104). The authors diagnosed both multisystemic inflammatory syndrome in children (MIS-C) for the presence of coronary involvement and HLH for fulfilling the HLH 2004 criteria. The patient required mechanical ventilation and maximal support treatment for 2 weeks, but completely recovered. The authors agree that it is a very unusual presentation of HLH. In 2022, an Iranian group described the case of an 8-year-old boy who developed MIS-C with heart failure requiring heart transplantation (105). Liver and bone marrow biopsy, together with typical laboratory findings, fulfilled HLH04 diagnostic criteria. He was initially treated with intravenous immunoglobulins (IVIGs) and steroids, followed by heart transplantation and the consequent heavy immunosuppression. Unfortunately, the patient died 20 days after surgery. A third case, regarding a female newborn birthed from a SARS-CoV-2-positive mother via cesarean section at 35 weeks of gestation, was reported from Jordan in 2022(106). The newborn tested positive for SARS-CoV-2 on the first day after birth and progressively developed typical HLH syndrome from the 6th day of life. The patient died on day 51 from severe respiratory failure.

Bacterial infections associated to HLH have frequently been reported in tropical countries and include Brucella (72,107-109), Salmonella enteritidis (73,90,110), Tuberculosis (88,90,111), sepsis by streptococcal infection [group B streptococcus (112) and Streptococcus suis (113)], by Listeria (72) or without identified pathogen (90). Orientia tsutsugamushi, the causing agent of scrub typhus, has been reported in India (73,89,90) and Korea (114-117). Ehrlichia chaffeensis, the agent of human monocytic ehrlichiosis, has been reported from the US (118-122). A case of S. pneumoniae 23A, serotype that is not included in the pneumococcal 13-valent conjugated vaccine (PCV-13) has been reported from Japan (123).

HLH in kidney transplant recipients has been associated with Ehrlichiosis in one case (121), and with Bartonella henselae in one other case (124). A case associated with Serratia marcescens has been reported in a preterm newborn (125).

Fungal infection-associated HLH has been reported in immunocompromised hosts (being treated for aplastic anemia and preB-acute lymphoblastic leukemia, respectively), caused by Trichosporon asahii (126).

Leishmania (71,72,127-135) and Plasmodium (90) [falciparum (136) and vivax (137)] have been reported as parasitic triggers of HLH.

M-HLH

M-HLH represents another challenging category, because it can be the initial manifestation of an underlying malignancy, it may develop during therapy as the consequence of chemotherapy or of treatment-induced mutations [treatment-induced HLH (Ch-HLH)], or it can develop as a consequence of an intercurrent infection (138-141).

HLH as a manifestation of underlying malignancy. The excess of proinflammatory cytokines produced by activated T cells infiltrating or surrounding the tumor or by the neoplastic T-cells in T/NK cell lymphoma is likely to cause M-HLH (1,38). In rare cases, the diagnosis of HLH anticipates that of malignancy by several weeks (138,142).

M-HLH is common in adults, where it accounts for ~50% of HLH cases (15), while it is significantly rarer in children, with a prevalence of 8-11% (138,143).

Lymphomas, though relatively uncommon at pediatric age, are frequently reported in association with HLH (138). Among the cases we reviewed in the present study, HLH has been associated with Hodgkin lymphoma (73,121,140,144), anaplastic large cell lymphoma (73,145,146), peripheral T-cell lymphoma (145,147), post-transplant lymphoproliferative disorder-lymphoma (148), subcutaneous panniculitis-like T-cell lymphoma (71) extranodal NK/T cell lymphoma, hepatosplenic T-cell lymphoma, systemic EBV-positive T-cell lymphoma of childhood (145). In addition, HLH and lymphoma can be alternative diagnoses as well (149), and extreme caution is required in differentiating the two conditions before starting steroid therapy.

Acute leukemia (both myeloblastic and lymphoblastic) presenting as HLH syndrome has also been reported (72,121,138-141,145).

Ch-HLH

Among the cases we reviewed in the present study, HLH developed on therapy or after treatment for juvenile myelomonocytic leukemia (150), acute monocytic leukemia (151), Langherhans cell histiocytosis (152) and solid tumors [Wilms tumor (153), neuroblastoma (72,154), rhabdomyosarcoma (140)]. Langerhans cell histiocytosis has been associated to HLH at presentation, during therapy or as a consequence of viral infection (140,142,155).

The immunosuppression induced by treatment frequently causes viral infections or reactivations, that in turn can trigger HLH. Among the cases reviewed, viral reactivation included EBV, CMV, respiratory syncytial virus, BK virus, human herpes virus 6, adenovirus and parvovirus B19 (141,156).

MAS-HLH

HLH syndrome occurring in the context of rheumatological disorders has been commonly referred to as MAS (157). While traditionally considered separate entities that share common features, it is now clear that they should be viewed as the same disease, regardless of differences in presentation and treatment (64). However, the detailed description of MAS-HLH is beyond the scope of the present work.

HLH associated with iatrogenic immune activation

Various emerging therapies, such as chimeric antigen receptor T cells (CAR-T cells) and Blinatumomab, have been associated with CRS (29). CRS shows major overlaps with HLH, so numerous authors classify it as a form of sHLH (158-160); some other authors, conversely, restrict the diagnosis of HLH following CAR-T infusion (carHLH) to the cases in which a severe CRS is associated to the typical HLH laboratory findings, often including hemophagocitosis (161,162).

CAR-T cells were officially licensed for use in children and young adults by the U.S. Food and Drug Administration in 2017 for the treatment of refractory B-cell acute lymphoblastic leukemia (163). CRS is a common severe adverse reaction following such therapies, especially in the acute phase (29). It probably results from the high amount of IFN-γ and IL-6 produced by the constitutionally activated CAR T cells (164) or by the subsequently activated macrophages (165-167) and by the targeted tumor cell lysis (168). Development of carHLH seems associated with pre-infusion NK lymphopenia (although it is not associated with impaired NK function) that is further amplified by treatment (161).

The St. Jude Children's Research Hospital group recently described a cohort of 27 pediatric patients treated with CD-19 CAR-T cells (162): 12/27 patients developed CRS alone, while four progressed to carHLH despite appropriate therapy. The carHLH subgroup showed higher and more sustained inflammation parameters and a poorer antileukemic response and survival. Blinatumomab, as well, has been associated with CRS and/or HLH in adults (29) and children (169).

Patients treated with T-activating therapies should be closely monitored for HLH, in order to optimize diagnosis and therapeutic approach.

Additional iatrogenic causes of cytokine storm include rituximab (170), gene therapies, immune checkpoint inhibitors, allogenic HSCT and heart transplantation (29).

Transplant-related HLH

HLH has been described in the context o HSCT (171,172) and, more rarely, in kidney (124,165) and liver (173-175) transplant recipients. Post-transplant HLH (PT-HLH) might be triggered by a combination of tissue damage, immunosuppressive therapy (176), alloimmune response, residual malignancies or by infections (171). Viral infections or reactivations, such as EBV and CMV but also gastrointestinal viruses, represent the most frequent trigger, but bacterial and fungal infections have also occasionally been described (124,165,172,177-179).

HLH post-HSCT (post-HSCT HLH) can occur within the first 30 days after transplantation (early onset) or later (late onset) (171). Late onset is usually related to infectious events, while for early onset the causes are not fully understood. In some cases the triggering factor may be the residual disease (171). PT-HLH diagnosis is extremely challenging because of the complex clinical and laboratory picture of the affected patients, for whom specific diagnostic criteria do not exist yet, and a high suspicion rate is essential (180).

In early onset post-HSCT HLH, aspects of differential diagnosis may occur with engraftment syndrome and acute graft-vs.-host disease (171). The treatment of PT-HLH is also particularly complex, since we are dealing with immunosuppressed patients, often with transplant-related toxicity (171). In EBV-related forms, the use of rituximab plays an important role (37,181).

HLH of unknown/uncertain origin

HLH has recently been described in the context of febrile infection related epilepsy syndrome (FIRES) (182), an epileptic encephalopathy characterized by refractory status epilepticus following unspecific febrile illnesses. The pathophysiology of FIRES has not been completely understood, but it likely to depend, once more, on dysfunctional activation of the innate immune system (182). Moreover, though rare, this association stresses the need of screening for patients with HLH presenting with severe CNS symptoms.

A curious case of HLH following spider bite (Loxosceles reclusa) has been reported from the US (183).

Drug reaction with eosinophilia and systemic symptoms induced by vancomycin, carbamazepine and levetiracetam has been described as an occasional trigger of HLH (72).

HLH and metabolic disorders

Among the cases reviewed in the present study, several patients were diagnosed with sHLH/HLH syndrome in the context of an inherited condition associated with metabolic diseases, such as Wolman's disease (184), galactosemia, Gaucher disease (185), Niemann-Pick disease, methylmalonic acidemia and propionic acidemia (186). Long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency (187), type 1 glycogen storage disease (188), lysosomal acid lipase deficiency (189), mucopolysaccharidosis-plus syndrome (190).

These patients fulfilled the HLH 2004 criteria, but it is possible that some HLH features, such as splenomegaly and cytopenias, were caused by the metabolic disease itself rather than by immune hyperactivation (184-190). On the other hand, it is possible that metabolites accumulation activated macrophages and triggered a proper sHLH. The link between metabolic disorders and HLH needs to be clarified, but an extensive screening for underlying metabolic diseases should be performed in patients presenting with HLH (184,185).

9. Treatment

The aim of HLH treatment is to suppress the life-threatening inflammation that leads to organ damage (23). The first goal invariably is to induce disease remission by controlling the hyper-activated T cells and the cytokine storm they generate (7,191). The second step, instead, may differ according to the underlying condition: in FHL and HLH cases, due to primary immunodeficiency, allogeneic HSCT is the only curative treatment that is currently known, while there is no consensus on sHLH best treatment. The present study agrees that, whenever possible, sHLH should be addressed according to the specific etiology, while the debate on whether treatment of the triggering disease may alone control sHLH or not is still open (7,191,192).

‘Old but gold’-The HLH 94/04

Since a neat distinction between FHL and sHLH is often impossible in the clinical setting, the HLH 94/04 protocol (7,193) suggests a pragmatic approach to HSCT indication, which remains a mainstay of therapy.

HLH 94 induction chemotherapy is based on steroids and etoposide administration for 8 weeks with or without intrathecal methotrexate, followed by maintenance with cyclosporine A (CSA), etoposide and dexamethasone pulses while waiting for HSCT (193). Chemotherapy might be suspended after 8 weeks of treatment once clinical remission is obtained and a clear genetic base is excluded (193). Otherwise, if a clear genetic base is proved, or in case of reactivation, HSCT is recommended (193).

Treatment intensification was proposed in 2004, but failed to improve the outcome (6). The Histiocyte Society thus recommends HLH-94 as the standard of care (7).

New therapeutic frontiers

The heavy mortality rate before HSCT (7) and the significant toxicities of chemotherapy urge the clinicians to find new treatments (6). This is especially true in patients with sHLH, whose prognosis widely varies from a 55% survival at 3 years with the standard protocols (193) to 100% survival in specific conditions (such as Leishmania-triggered HLH when treated appropriately) (135).

The ongoing recognition of the central role played by cytokines suggests a treatment shift towards targeted therapies. Whether these drugs can eradicate HLH alone or require the addition of etoposide remains to be clarified, as well as how they are best used [including doses, drug combinations and use in adults/children (6).

Emapalumab (NI-0501; Novimmune SA), an anti-IFN-γ monoclonal antibody has been used successfully in pediatric patients with refractory or previously untreated pHLH (194,195).

The most recent clinical trials focus on hybrid immunotherapy (ATG + etoposide + dexamethasone; NCT01104025), monoclonal antibodies such as Alemtuzumab (196) (NCT02472054) and tyrosine-kinase inhibitors such as ruxolitinib (197) (NCT04999878; NCT04120090; NCT04551131; NCT03795909; ChiCTR2000029977) and zanubrutinib (NCT05320575). Studies on selectively cytokine-targeting molecules include Tocilizumab (198) (NCT02007239), Anakinra (NCT02780583) and Emapalumab/NI-0501(195) (NCT05001737; NCT03311854; NCT03312751; NCT01818492). In the subset of EBV-HLH patients, the ongoing clinical trials focus on Rituximab (NCT05384743), anti CD20 monoclonal antibody Sintilimab and Lenalidomide (NCT05258136), Tabelecleucel (NCT04554914), programmed cell death protein 1 antibody alone (NCT05039580) or in association with Lenalidomide (NCT04084626).

In intensive care settings, nonspecific physical therapies showed interesting results, at least in the acute phase (199,200). If a massive release of cytokines/a cytokine storm is responsible for organ damage, blood purification techniques can blunt the process with a rapid non-selective effect, potentially translating into survival benefit (199). These techniques include plasma exchange, which has been successfully applied in medium resource contexts (such as Turkey and India) (201,202), and hemofiltration combined with highly immune adsorbent cartridges (119,191,200). HLH recommendations have recognized blood purification as a potential salvage treatment in adults (203). Moreover, these non-specific therapies might be combined with targeted therapies (204), but do not replace appropriate treatment of the underlying trigger of HLH, especially in patients with refractory disease (205).

The most recent consensus guidelines on recognition, diagnosis and management of HLH in critically ill children (46) stress the importance of aggressive treatment. Steroids, in particular, should not be delayed as part of first-line treatment in different protocols (HLH-94 and ATG-based protocols) and are effective in both primary and secondary HLH.

A Canadian group reported nine cases of pediatric liver transplantation in acute liver failure secondary to sHLH, suggesting that HLH might not be an absolute contraindication to transplantation and instead play a role in a very restricted subgroup of patients (19).

Specific treatment considerations for infection-related HLH

Treatment of infection-associated HLH deserves specific considerations according to the underlying agent, since some forms respond to anti-infectious treatment [namely Leishmania (131-134), Ehrlichiosis (122), scrub typhus (89,90) and other tropical diseases-associated forms], while reaction to some other pathogens displays intermediate features and benefits of standard aggressive immunosuppression.

Several studies (131,132,134,206) have reported complete response of Leishmania-associated HLH to amphotericin B therapy +/- steroids, with an excellent prognosis [3 year overall survival rate 24% in pHLH vs. 100% in visceral leishmaniasis-associated HLH; P<0.001; 2021 Iranian cohort of 60 children (133)]. A Chinese retrospective study (135) reported good results with antiparasitic therapy associated with chemotherapy according to the HLH94 protocol in patients with visceral leishmaniosis-HLH; however, the cohort includes children and adults and it is impossible to discriminate whether treatment intensification depends on age.

Human monocytic ehrlichiosis, can also trigger an HLH form that has proved to respond to antimicrobial therapy alone (doxycycline) (122).

Scrub typhus, dengue fever and tropical infections in general respond to steroids alone or to regimens without cytotoxic drugs in cases with mild to moderate disease activity (89,90).

The hepatitis-associated HLH cases reviewed in the present study have experienced complete recovery on antiviral therapy and ruxolitinib in the case of a patient with HBV (95) and after IVIG-infusion in the cases of patients with HAV (94).

A patient developing HLH after HIV diagnosis has required both antiretroviral therapy and chemotherapy according to HLH94 protocol (96).

The Crimean Congo hemorrhagic fever-associated HLH has been resolved using antiviral therapy combined with IVIG therapy and plasma exchange (91).

The case of EBV-HLH is more complex. EBV-HLH standard treatment is by the HLH94 protocol, and early initiation of etoposide-based regimens improves survival in severe EBV-HLH (8,74,207). Chemotherapy can be combined with targeted therapies such as Rituximab (37,181) or Ruxolitinib (208). Treatment intensification with the L-DEP regimen (PEG-asparaginase in combination with liposomal doxorubicin, etoposide and high-dose methylprednisolone) has been reported as salvage therapy and bridge to alloHSCT for children with refractory disease (209). Blood purification techniques (plasma exchange and continuous renal replacement therapy) have proved to be safe and effective in addition to standard chemotherapy in severe cases (199). On the other hand, patients presenting with mild-to-moderate disease activity may only require immunomodulating therapies, such as IVIGs, CSA and steroids (69). A Japanese group (210) described a cohort of 22 patients aged 6 months to 41 years in which >60% recovered after immunotherapy (IVIG, CSA 6 mg/kg/day, prednisone); they suggest that early immunotherapy may modulate T-cell activation and reduce the chance of unnecessary chemotherapy. Sporadic cases of spontaneous resolution have been reported: An American group (68) reported the cases of two adolescents who fulfilled the HLH2004 diagnostic criteria in the context of an acute EBV infection. In both cases, treatment was planned according to HLH94 protocol, but the patients spontaneously improved before the initiation of therapy. The authors recommend caution in starting aggressive treatment, especially when patients experience mild-to-moderate symptoms. These differences in treatment approaches and outcomes underline the need for evidence-based risk stratification criteria in patients with EBV-HLH.

Specific treatment considerations for M-HLH

Malignancy-triggered forms should be distinguished from the forms that occur after chemotherapy (Ch-HLH) to optimize the therapeutic approach.

Ch-HLH. Ch-HLH is often caused by infections related to immunosuppression, and benefits of anti-infectious therapy, steroids and IVIGs.

HLH as a manifestation of underlying malignancy. M-HLH is associated with increased mortality and the early detection of malignancy can favorably influence the prognosis. Treatment is not homogeneous, and can address the underlying malignancy first, be focused on HLH or a combination of the two (41,138,145,211-213). The published series refer to small numbers and suggest an individualized therapeutic approach. Aggressive HLH-directed therapy at the onset of malignancy-associated HLH can delay or complicate anticancer treatment (39,41).

Specific treatment considerations for Rheumatologic HLH (MAS-HLH)

MAS-HLH usually responds to steroid therapy or to anakinra (recombinant human IL-1 receptor antagonist), especially when given in the early course of therapy (214,215). The present study refers to specific international guidelines for more detailed considerations on MAS-HLH (157,216,217).

Specific treatment considerations for HLH associated with iatrogenic immune activation

The cytokine release syndrome following CAR-T or Blinatumomab administration usually responds to Tocilizumab, steroids, Anakinra or to a combination of such molecules (29,165-167,169). Whether carHLH should be differentiated from a severe form of CRS is still an open question, and a more precise definition of both conditions is essential in order to guide therapeutic choices in the most complex cases (162,218).

Specific treatment considerations for transplant-related HLH

Post-transplant HLH requires prompt and aggressive treatment because of the high risk of graft failure, but no international consensus exists on treatment protocol. Reported therapeutic approaches range from corticosteroids, IVIGs and low-dose etoposide (219-221) to standard HLH94 protocol with eventual rescue HSCT (172). Ruxolitinib was recently reported as salvage therapy in 2 children, with alternating results (222). Outcome is unclear, usually compromised by graft failure.

10. Prognosis

The prognosis is related to the cause that induced sHLH, with the worst outcomes in M-HLH. In the 1980s, long-term survival in HLH was <5% (223). The HLH-94 protocol improved life expectancy up to 54% of 5-year survival, with similar results in FHL and sHLH patients. Unfortunately, the subsequent 2004 revision failed to further improve the outcome (6).

An alternative therapeutic approach consisting of steroids, antithymocyte globulin and cyclosporin has obtained satisfactory results, with a 70% remission rate (33). In general, in HLH the highest mortality is in the first weeks of therapy. In patients with pHLH who underwent hematopoietic stem cell transplantation, a 5-year probability of overall survival of ~70% has been observed (224). Overall, HLH still carries a heavy burden of mortality up to the present day.

11. Conclusions

HLH is a rare disease that may develop from childhood to adult life, but HLH subtypes spread with different frequency according to the age of the patient. In early infancy, FLH forms are most typical. They are linked to genetic defects affecting the cytotoxic activity of T lymphocytes and NK cells, and invariably require allogeneic HSCT.

Secondary forms become more frequent with increasing age. They represent a heterogeneous group in terms of etiology and therapeutic approach. The severity of the clinical picture often requires heavy immunosuppression and the use of etoposide while trying to define the etiology. Some significant exceptions exist: i) Less aggressive therapeutic approaches proved to have efficacy in MAS-HLH and in the majority of infection-triggered HLH forms; and ii) anti-tumor therapy, alone or in combination with corticosteroids, can usually control M-HLH when hyperinflammation is triggered by the tumor itself.

Various viruses and EBV can trigger FHL or cause a HLH syndrome in the absence of known genetic alterations. The improvement of genetic investigations will probably clarify the genetic background of several patients with ‘secondary’ forms of HLH.

The future goal in addressing this rare and potentially fatal condition will be to provide a tailored therapy for each patient, based on its genetic and biological characteristics.

Funding Statement

Funding: No funding was received.

Availability of data and materials

Not applicable.

Authors' contributions

All authors substantially contributed to the present work, both in literature review and in the elaboration of the present article. CB, IM and CO acquired, analyzed and interpreted the literature data. CB, IM and FST wrote and edited the manuscript. FST supervised the study. All authors have read and approved the final manuscript, and agree to be accountable for all aspects of the work. Data authentication is not applicable.

Ethics approval and consent to participate

Not applicable.

Patient consent for publication

Not applicable.

Authors' information

Dr Fabio S. Timeus, ORCID 0000-0003-0095-283X.

Competing interests

The authors declare that they have no competing interests.