pediatric cns-isolated hemophagocytic lymphohistiocytosis · lymphohistiocytosis (hlh). 1,2...

ARTICLE OPEN ACCESS CLASS OF EVIDENCE

Pediatric CNS-isolated hemophagocyticlymphohistiocytosisLeslie A Benson MD Hojun Li MD PhD Lauren A Henderson MD MMSc Isaac H Solomon MD PhD

Ariane Soldatos MD MPH Jennifer Murphy MS BSN Bibiana Bielekova MD Alyssa L Kennedy MD PhD

Michael J RivkinMD Kimberly J DaviesMD AmyPHsu BA StevenMHollandMDWilliamAGahlMD PhD

Robert P Sundel MD Leslie E Lehmann MD Michelle A Lee MD PhD Sanda Alexandrescu MD

Barbara A Degar MD Christine N Duncan MDdagger and Mark P Gorman MDdagger

Neurol Neuroimmunol Neuroinflamm 20196e560 doi101212NXI0000000000000560

Correspondence

Dr Gorman

markgorman

childrensharvardedu

or Dr Duncan

christine_duncandfciharvardedu

AbstractObjectiveTo highlight a novel treatable syndrome we report 4 patients with CNS-isolated inflammationassociated with familial hemophagocytic lymphohistiocytosis (FHL) gene mutations (CNS-FHL)

MethodsRetrospective chart review

ResultsPatients with CNS-FHL are characterized by chronic inflammation restricted to the CNS that isnot attributable to any previously described neuroinflammatory etiology and have germlinemutations in known FHL-associated genes with no signs of systemic inflammation Hemato-poietic stem cell transplantation (HCT) can be well tolerated and effective in achieving ormaintaining disease remission in patients with CNS-FHL

ConclusionsEarly and accurate diagnosis followed by treatment with HCT can reduce morbidity andmortality in CNS-FHL a novel treatable syndrome

Classification of evidenceThis study provides Class IV evidence that HCT is well tolerated and effective in treating CNS-FHL

MORE ONLINE

Class of EvidenceCriteria for ratingtherapeutic and diagnosticstudies

NPuborgcoe

These authors contributed equally to the manuscript

daggerThese authors contributed equally to the manuscript and are co-corresponding authors

From the Department of Neurology (LAB MJR MPG) Boston Childrenrsquos Hospital Dana-FarberBoston Childrenrsquos Cancer and Blood Disorders Center (HL ALK LEL BADCND) Department of Rheumatology (LAH RPS) Boston Childrenrsquos Hospital Department of Pathology (IHS SA) Boston Childrenrsquos Hospital Boston MA UndiagnosedDiseasesProgram (AS JM WAG) National Human Genome Research Institute Neuroimmunological Diseases Section (BB) Laboratory of Clinical Immunology andMicrobiology NationalInstitute of Allergy and Infectious Diseases Laboratory of Clinical Immunology and Microbiology (APH SMH) National Institute of Allergy and Infectious Diseases Bethesda MDand Division of Hematology Oncology (MLA) and Marrow and Blood Cell Transplantation Childrenrsquos Hospital at Montefiore Bronx NY

Funding information and disclosures are provided at the end of the article Full disclosure form information provided by the authors is available with the full text of this article atNeurologyorgNN

The Article Processing Charge was funded by the authors

This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives License 40 (CC BY-NC-ND) which permits downloadingand sharing the work provided it is properly cited The work cannot be changed in any way or used commercially without permission from the journal

Copyright copy 2019 The Author(s) Published by Wolters Kluwer Health Inc on behalf of the American Academy of Neurology 1

Despite advances in diagnostic capabilities in neuro-immunology highlighted most dramatically by antineuronalantibody biomarkers some patients continue to have idiopathicor poorly defined chronic andor treatment-refractory neuro-inflammatory disorders Neuroinflammatory findings arepresent in more than half of the patients with hemophagocyticlymphohistiocytosis (HLH)12 However systemic in-flammation is a fundamental diagnostic component of HLHwhich is traditionally characterized by cytokine storm multi-organ dysfunction and ultimately death if untreated3 HLHpathogenesis has been elucidated through the discovery of fa-milial HLH (FHL)-associated genes which encode proteinsrequired for CD8+ lymphocyte and natural killer (NK) cellcytotoxicity Deficiency in cytotoxic killing results in antigenpersistence leading to the hyperinflammatory state associatedwith FHL4 The genes most commonly mutated in FHL arePRF1 (FHL2) and UNC13D (FHL3)5 Hematopoietic celltransplantation (HCT) is the only definitive cure for FHL67

Although common at diagnosis of systemic HLH CNS mani-festations rarely precede systemic findings12 Hemophagocy-tosis has been found in a brain biopsy of a patient not fulfillingthe systemic HLH criteria8 Patients with PRF1mutations havebeen reported with isolated CNS inflammation for weeks tomonths before therapy initiation19ndash12 2 such patients un-derwent HCT911 These reports raise but do not answer thequestion as to whether isolated chronic CNS inflammation canoccur in the absence of systemic HLH findings

Here we describe 4 patients with progressive CNS-isolated in-flammation and mutations in FHL-associated genes who neverfulfilled the systemic diagnostic criteria for HLH except fordefective NK cell activity reflective of their underlying geneticmutations The neuropathologic details of 3 of the patients13 andHCT outcome details of all 4 patients14 have been previouslyreported HCT was well tolerated in these patients and resultedin arrest of disease progression highlighted by the lack of de-velopment of morbidity in an asymptomatic patient with pro-gressive radiologic neuroinflammatory findings

Case reportsBaseline characteristics at diagnosis for all patients are describedin the table

Patient 1The index case presented as a 5-year-old girl with headachevomiting mild ataxia multifocal contrast-enhancing cerebral

and cerebellar T2-hypertintense lesions (figure 1 figure 2) andCSF pleocytosis meeting the diagnostic criteria for a de-myelinating clinically isolated syndrome (CIS)16 Her symp-toms and imaging findings responded to high-doseglucocorticoids (methylprednisolone 30 mgkg IV for 5 days)but returned with steroid tapers despite monthly IV immuno-globulin (IVIg) (1 gkgdose) Serial brain MRI identifiedenhancement patterns reminiscent of chronic lymphocytic in-flammation with pontine perivascular enhancement responsiveto steroids (CLIPPERS)17 Biopsy of a cerebellar lesion iden-tified a mixed chronic inflammatory infiltrate with perivascularinflammation diffusely involving the gray and white matter andleptomeninges (figure 1) Induction treatment with gluco-corticoids and cyclophosphamide followed by mycophenolatemofetil per small vessel CNS vasculitis protocol18 slowed butdid not halt disease progression She experienced progressivetremor ataxia dysarthria dysconjugate gaze and slowed speechand response time Extensive evaluation including meta-genomic next-generation sequencing of CSF and brain tissuewas unrevealing for infection Given her severe symptomatol-ogy worsening course and prominent T cells on repeat biopsy(left thalamic lesion data not shown) natalizumab whichinhibits T-cell entry into the CNS was given under a compas-sionate use investigational new drug protocol (IND 114796150 mg for 4 mgkg every 4ndash5 weeks) While on natalizumabher condition clinically and radiographically improved and thenstabilized for 3 years before eventual clinical and radiographicprogression The patient was referred to the UndiagnosedDiseases Program at the NIH1920 but during the evaluationshe developed rotary nystagmus and worsened ataxia with lossof independent ambulation partly responsive to high-dosemethylprednisolone Ultimately CSF immunophenotypingand soluble biomarker analysis21 revealed activation of the in-nate immune system PRF1 gene sequencing identified biallelicpreviously reported pathogenic mutations 5 years after initialpresentation CSF neopterin was significantly elevated (table)Throughout her course patient 1met no systemicHLH criteriaexcept for decreased NK cell activity initially attributed tochronic immunosuppression Following diagnosis she receivedCNS-directed HLH therapy with intrathecal methotrexate andhydrocortisone and systemic dexamethasone with clinical im-provement and normalization of the neopterin level beforeundergoing unrelated donor HCT at age 10 years

Patient 2A 6-year-old girl developed vomiting headache psychiatricsymptoms and cognitive decline over 3 months and then hadfocal status epilepticus prompting hospitalization Additionalsymptoms noted during the hospitalization included left

GlossaryADEM = acute disseminated encephalomyelitis CIS = clinically isolated syndrome CLIPPERS = chronic lymphocyticinflammation with pontine perivascular enhancement responsive to steroids FHL = familial hemophagocytic lymphohistio-cytosis GVHD = graft-versus-host disease HCT = hematopoietic cell transplantation HLH = hemophagocyticlymphohistiocytosis IVIg = IV immunoglobulinMDEM = multiphasic ADEM NK = natural killerWBC = white blood cell

2 Neurology Neuroimmunology amp Neuroinflammation | Volume 6 Number 3 | May 2019 NeurologyorgNN

Table Characteristics of patients with CNS-FHL

Patient 1 2 (at diagnosis) 3 4 2 (At relapse)

Age at symptom onset (y) 5 6 7 NA 9 (Symptom onset 13 moposttransplant)

Sex F F F F

Age at FHL diagnosis (y) 10 8 13 11 9

Neurologic symptoms

Ataxia Y Y Y N N

Weakness Y Y Y N N

Seizure Y Y Y N Y

Headache Y Y N N Y

Vomiting Y Y N N N

Cognitive decline Y Y N N N

Other symptoms Esotropia nystagmus tremorand dysarthria

Dystonia and dysphagia Diplopia and mood lability N Dystonia

Immunotherapy before HLHdiagnosis

Oral steroids Y Y Y N

IV steroids Y Y Y N

IVIg Y Y Y N

Cyclophosphamide Y Y Y N

Mycophenolate mofetil Y N Y N

Other immunotherapy Natalizumab Plasmapheresis Rituximab and azathioprine N

Mutation PRF1 c452AgtT (pH151L) andc666CgtA (H222Q)

PRF1 c443CgtG (pA148G) andc666CgtA (H222Q)

UNC13D c2346_2349delGGAG(pR782fs) c2588GgtA (pG863D)


PRF1 c666CgtA (H222Q)heterozygous mutation in the firstHCT donor

Systemic HLH evaluationa No fever or splenomegaly WBC13 Hgb 160 Plt 359 ferritin 365fibrinogen 407 and triglycerides56

No fever or splenomegaly WBC22 Hgb 136 Plt 423 ferritin 208fibrinogen 443 and triglycerides99

No fever or splenomegaly WBC442 Hgb 142 Plt 346 ferritin719 fibrinogen 365 andtriglycerides 52



Soluble IL-2 receptor (unitmL normal lt1100)

265 209 710 647 519

Continued

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Table Characteristics of patients with CNS-FHL (continued)


NK cell function (lytic unitsnormal gt26)

17 00 Lymphopenia too severe 00 01

NK cell perforin positivity(normal 73ndash91)

0 0 Lymphopenia too severe Not performed 52

NK cell CD107a Not performed Expression 10 (normal 11ndash35) Lymphopenia too severe Expression 3 (normal 11ndash35) Not performed

Mean fluorescence 164 (normal207ndash678 MCF)


ESR (mmhr)CRP (mgdL) 7017 5lt003 1205 7lt003 8011

CSF WBC (per microliter) 10 2 10 7 0

CSF neopterin (nmolLnormal lt33)

48 150 24 22 14

Pre-HCT therapy

Enteral dexamethasone Y Y Y Y Y

Intrathecal methotrexatehydrocortisone

Y Y N N Y

Duration of follow-up post-HCT (mo)

36 13 mo to relapse 28 25 11 mo

Post-HCT imaging Nonew lesions no enhancementand diffuse encephalomalacia

Nonew lesions no enhancementand diffuse encephalomalaciauntil 13 months posttransplant

No new lesions and noenhancement


No new lesions no enhancementand diffuse encephalomalacia

Pre- and post-HCT ModifiedRankin Scale15

4 4 2 0 4

Abbreviations ESR = erythrocyte sedimentation rate FHL = familial hemophagocytic lymphohistiocytosis HCT = hematopoietic cell transplantation HLH = hemophagocytic lymphohistiocytosis IVIg = IV immunoglobulinNA = not applicable NK = natural killer N = no Y = yesa Fever gt380degC WBC = white blood cell count (times103 per μL) Hgb = hemoglobin (gdL) Plt = platelet count (times103 per μL) ferritin (ngmL) fibrinogen (mgdL) and triglycerides (mgdL)

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hemiparesis and dystonia Brain MRI identified contrast-enhancing cerebral and cerebellar white matter T2 hyperin-tense lesions (figures 1 and 3) Shemet the criteria for and wasdiagnosed with acute disseminated encephalomyelitis(ADEM)16 and received pulse methylprednisolone IV for 3days and IVIg followed by tapering daily oral prednisolonewith gradual partial improvement However 4 months afterthe seizure while off immunotherapy she re-presented with

headache vomiting lethargy worsened motor and languagefunction and new MRI lesions prompting additional IVmethylprednisolone and 5 sessions of plasmapheresis Thepresentation now met the criteria for multiphasic ADEM(MDEM) She was then treated with 3 doses of monthly IVcyclophosphamide Over the next 8 months she had in-terspersed periods of improvement and worsening including2 episodes of status epilepticus Treatments during this time

Figure 1 Imaging and histopathologic characteristics of patients with CNS-FHL

FLAIR (row A) and post-contrast T1 (row B) MRI findings for patients before initial diagnosis or at relapse Note the varied lesion appearance with smallmultifocal confluent and tumefactive lesions Punctate and curvilinear enhancement resembling CLIPPERS is common to 3 of the 4 patients early in diseasePatient 4 had multifocal small lesions with a single area of enhancement Avid enhancement was a common feature of new or active lesions Over timelesions became confluent and diffuse atrophy evolved in patients 1 and 2 Representative histopathology (row C) images from prediagnosis or relapsebiopsies demonstrating a diffuse mixed chronic inflammatory infiltrate in patient 1 foci of perivascular lymphocytes (arrow) extending into the parenchymafor patient 2 prediagnosis perivascular (arrow) and parenchymal mixed chronic inflammatory infiltrate with small vessel vasculitis in patient 3 anddestructive perivascular (arrow) and parenchymal lymphohistiocytic infiltrate in patient 2 at relapse (magnification times40 objective) Post-HCT FLAIR (row D)and post-contrast T1 (row E)MRI images demonstrate resolved enhancement and no development of new lesions for patients 1 3 and 4 after HCT 1 and forpatient 2 after HCT2 CLIPPERS = chronic lymphocytic inflammationwith pontine perivascular enhancement responsive to steroids FLAIR = fluid-attenuatedinversion recovery HCT = hematopoietic cell transplantation

NeurologyorgNN Neurology Neuroimmunology amp Neuroinflammation | Volume 6 Number 3 | May 2019 5

included IV and oral corticosteroids and IVIg Serial brainMRI showed accumulation of multifocal T2 hyperintenselesions becoming confluent in the bilateral periventricularwhite matter as well as progressive supra- and infra-tentorialatrophy necrosis and scattered hemorrhages (figure 3)Nineteen months after her first symptoms she developeddysphagia and worsened dystonia and was referred to BostonChildrenrsquos Hospital Biopsy of a frontal lobe lesion showedperivascular lymphocytic foci with parenchymal extension

(figure 1) CSF neopterin was elevated Based on imagingsimilarities to patient 1 reanalysis of previously performedwhole-exome sequencing with attention to FHL genes iden-tified biallelic PRF1 mutations (1 previously reported inFHL2 1 novel) She met no systemic HLH criteria except fordecreased NK cell activity She received intrathecal metho-trexate and hydrocortisone and systemic dexamethasone be-fore undergoing HLA-matched sibling donor HCT at age 9years

Figure 2 Patient 1 MRI evolution over time

(A) Confluent cerebellar FLAIR lesions with punctate folia and curvilinearenhancement without SWI change evolved to cerebellar atrophy with con-fluent nodules of enhancement and associated accumulation of micro-hemorrhages Pattern persists posttransplant but without enhancement(B) Supratentorial multifocal asymmetric small well-circumscribed lesionsall with avid enhancement without initial SWI change evolving to confluentFLAIR lesions with punctate and confluent areas of enhancement and SWIchange over time leading up to transplant Imaging remains stable post-transplant FLAIR = fluid-attenuated inversion recovery SWI = susceptibility-weighted imaging

Figure 3 Patient 2 MRI evolution over time before relapse

(A) Cerebellar FLAIR lesions with punctate and curvilinear enhancementwithout SWI change evolved to cerebellar atrophy with confluent nodules ofenhancement and associated accumulation of microhemorrhages Patternpersists posttransplant but enhancement responded to treatment (B)Supratentorial multifocal asymmetric lesions with white matter corticaldeep gray and insular predilection with initial punctate enhancementevolving to curvilinear cortical enhancement No initial SWI change evolvingto punctate and confluent areas of SWI change consistent with hemorrhageover time leading up to transplant Posttransplant FLAIR lesions becomesmaller with further global brain atrophy FLAIR = fluid-attenuated inversionrecovery SWI = susceptibility-weighted imaging


Patients 3 and 4Patient 3 presented as a 7-year-old girl with diplopia righthemiparesis ataxia CSF pleocytosis (white blood cell [WBC]count 10μL with 85 lymphocytes) and multifocal brainlesions including a large brainstem lesion (figure 1) leadingto diagnosis of a demyelinating CIS Symptoms and MRIfindings transiently improved with methylprednisolone IV 30mgkgd for 5 days followed by prednisolone starting at 1mgkgd and tapered over 3 months but she developedseizures and new contrast-enhancing lesions similar toCLIPPERS (figures 1 and 4) 7 months after initial pre-sentation Biopsy of a frontal lobe lesion showed extensive

vascular perivascular and parenchymal mixed inflammatoryinfiltrates with occasional small vessel vasculitis (figure 1)Despite treatment with a small vessel CNS vasculitis pro-tocol18 symptoms including altered mental status and seizuresrecurred 6 months after glucocorticoid discontinuation whilestill on mycophenolate mofetil 600 mgm2dose twice dailyOver the next 4 years subsequent treatments (in addition tooral steroids) included infliximab (5 mgkgdose at 0 2 and 6weeks and every 4 weeks thereafter for 9 months) cyclo-phosphamide (10 mgkgdose every 2 weeks for 3 months)rituximab (3 cycles of 1000 mg times 2 doses) azathioprine(100 mg twice daily) and IVIg (05gkg every 4 weeks) withperiods of improvement followed by breakthrough clinical andMRI disease activity Based on our experience with patients 1and 2 FHL genetic testing 7 years after symptom onset iden-tified biallelic pathogenic mutations inUNC13D Patient 3 metno systemic HLH criteria although she was too lymphopenicfrom previous therapy to assess NK cell function and CSFneopterin was normal During her course she had recurrentsinusitis mildly low IgG and IgA and elevated gamma deltaT cells (up to 19) while on immunosuppression thesefindings were attributed to medication toxicity but may havebeen related to herUNC13Dmutations She received systemicdexamethasone therapy before undergoing unrelated donorHCT at age 14 years

Patient 4 the asymptomatic sister of patient 3 was found at age12 years to have the same biallelic UNC13D mutations duringevaluation as a potential bone marrow donor for her sisterBrain MRI demonstrated multifocal white matter lesions oneof which was contrast enhancing (figure 1) Two of the lesionsincreased in size over 6 weeks Neurologic examination wasnormal CSF demonstrated pleocytosis (WBC count 7μL92 lymphocytes) and oligoclonal bands but normal CSFneopterin Laboratory evaluation demonstrated absent NK cellfunction and decreased CD107a expression Similar to hersister she was hypogammaglobulinemic (IgG 482) Shemet noother systemic HLH criteria She received oral dexamethasonetherapy before undergoing unrelated donor HCT

MethodsPatients and medical record reviewThis study was approved by the Boston Childrenrsquos Hospitaland National Human Genome Research Institute InstitutionalReview Boards Clinical information was obtained from themedical record

HLH diagnostic testing and pre-HCT therapyPRF1 and UNC13D gene sequencing was performed at theNIH or Cincinnati Childrenrsquos Hospital NK cell functionaltesting perforin staining and CD107a mobilization wereperformed at Cincinnati Childrenrsquos Hospital Intrathecalmethotrexate and hydrocortisone and systemic dexametha-sone were dosed as previously described3 with dexametha-sone weaned after HCT


(A) Initial attack and follow-up FLAIR and T1 post-contrast images showingtumefactive brainstem lesions with heterogeneous enhancement patternSignificant improvement with steroids at 4-month follow-up (B) Attack 7months from onset with multifocal FLAIR lesions throughout the brain andspinal cord with punctate speckled enhancement throughout (C) Pre- andpost-HCT images of the cerebellar involvement with minimal atrophy overtime in this patient despite 7 years of disease FLAIR = fluid-attenuated in-version recovery HCT = hematopoietic cell transplantation


HCT procedureReduced-intensity bone marrow conditioning with alemtu-zumab fludarabine and melphalan7 was dosed as previouslydescribed Hematopoietic stem cells were harvested from thedonor bone marrow Graft-versus-host disease (GVHD)prophylaxis included cyclosporine and methylprednisoloneWhole blood and T-cell donor chimerism studies were per-formed by the American Red Cross using the short tandemrepeats-PCR method Further details of the transplantationprocess and transplant outcomes are published separately14

Data availabilityPatient data were obtained from their electronic medicalrecords for this study All abstracted data are included withinthe manuscript table and figures If additional details areneeded they can be requested from the correspondingauthor

ResultsHematopoietic stem cell transplantation andCNS-FHL disease responseFollowing HCT patients 1 3 and 4 demonstrated normali-zation of NK cell activity and resolved MRI contrast en-hancement did not develop new brain lesions (figures 1ndash4) orfurther neurologic deficits and had immunosuppressive glu-cocorticoids discontinued by 3 months post-HCT Patient 1had improved cranial nerve function strength coordinationand tremor but continued to have significant physical deficitsrequiring a wheelchair or 11 assistance for ambulation andcognitive deficits with a full-scale IQ of 70 Patient 2 hadimmunosuppressive glucocorticoids discontinued by 2months post-HCT Her dystonia mental status and strengthimproved at 1 year following HCT but remains wheelchairdependent Patient 3 had improved diplopia gait strengthand seizures with resolution on levetiracetam Patient 4remained neurologically asymptomatic post-HCT None ofthe 4 patients experienced graft failure or acute or chronicGVHD Length of follow-up for all 4 patients ranged from 2 to3 years posttransplant

Disease recurrence and second HCTThirteen months following HCT patient 2 experiencedseizures headache and worsening dystonia Biopsy of a newfrontal lobe white matter enhancing lesion (figure 1) showeda dense destructive perivascular and parenchymal lympho-histiocytic infiltrate (figure 1) Pathologic evaluation excludedposttransplant lymphoproliferative disorder and extensiveinfectious testing was negative She was diagnosed with re-lapsed CNS-FHL Clinical and laboratory findings at relapseare described in the table She met no systemic HLH criteriaexcept decreased NK cell function Her NK cell function wasdecreased despite donor chimerism greater than 46 in wholeblood T cells and NK cells which is typically consideredadequate Her sibling donor possessed a monoallelic PRF1mutation Upon relapse NK cell testing in the donor revealed

absent function although decreased NK cell function haspreviously been noted in patients with heterozygous PRF1mutations and the significance is uncertain22 Patient 2 re-peated remission induction with systemic dexamethasone andhad a stable MRI (not shown) without new enhancementbefore undergoing second HCT from an unrelated donorusing an alternative reduced-intensity bone marrow condi-tioning regimen of fludarabine alemtuzumab and busulfan

Twelve months posttransplant patient 2 had normal NK cellfunction and no contrast-enhancing lesions on MRI(figure 1)

DiscussionWe describe 4 patients with isolated CNS-FHL who lackedany systemic HLH manifestations or typical laboratory ab-normalities (other than decreased NK cell function) butcarried FHL-associated mutations in PRF1 or UNC13D It islikely that additional genetic modifying factors exist thatpredispose patients to CNS-isolated disease or that specificneuroinflammatory events lead to CNS involvement withouttriggering systemic inflammation We did not identify an in-fectious trigger in our cohort including negative testing forEpstein-Barr virus but we cannot exclude an undetected or-ganism or virus Further studies will be required to identify theinherited and environmental risk factors for CNS-isolatedinflammation

Our findings of CNS-FHL as a cause of treatment-refractoryisolated neuroinflammation are an important advance in thedifferential diagnosis of neuroinflammatory disorders Somenotable features may facilitate diagnosis Patients 1ndash3 hadoverlapping symptoms including seizures (33) weakness(33) ataxia (23) cognitive decline (23) eye movementabnormalities (23) headache (23) and vomiting (23)Symptoms typically improved with immunotherapy but re-curred both during weans and active treatment Relapses wereespecially common when corticosteroids were weaned ordiscontinued The treatment-refractory and steroid-dependent nature of the symptoms was striking Someshared imaging characteristics include multifocal T2fluid-attenuated inversion recovery hyperintense lesions with avidenhancement and accumulation of microhemorrhages evi-dent on susceptibility-weighted imaging Punctate and curvi-linear lesions evolved in the 3 symptomatic patients at times intheir illnesses

In these patients CNS-FHL mimicked several CNS in-flammatory disorders including CLIPPERS MDEM andsmall vessel CNS vasculitis These diagnoses were made bytreating clinicians and presentations met the diagnostic cri-teria The clinical radiographic and pathologic features forpatient 1 meet proposed CLIPPERS diagnostic criteria23 Inaddition patients 2 and 3 had radiographic features includinghomogeneous enhancing nodules and curvilinear and


punctate lesions as seen in CLIPPERS Patient 2 met thecriteria for MDEM16 early in her course Some patients withMDEM harbor anti-myelin oligodendrocyte glycoprotein anti-bodies which were not clinically available when patient 2 pre-sented but should now be obtained in patients with MDEMPatient 3 met the diagnostic criteria for small vessel CNS vas-culitis including pathologic confirmation

As above there may be clinical and radiographic features thatare suggestive of CNS HLH however many of these featuresoverlap with other neuroinflammatory disorders As seen inpatients 1 and 2 CNS HLH can be a destructive process withsignificant morbidity As such and in the interest of earlierdiagnosis and definitive treatment we propose that pediatricpatients with treatment-refractory or recurrent CNS in-flammation of uncertain etiology including presentationsconsistent with CLIPPERS MDEM (especially if anti-myelinoligodendrocyte glycoprotein antibodies are negative) andsmall vessel CNS vasculitis undergo genetic testing and NKcell functional testing for FHL Patients with refractory orrecurrent neuroinflammation often require long-term immu-nomodulatory therapy which can reduce the yield of a brainbiopsy and alter functional testing for FHL therefore obtain-ing functional diagnostic studies before treatment is preferredNK cell function and FHL genetic testing may eliminate theneed for brain biopsy in some patients Furthermore ourresults illustrate the importance of screening siblings of patientswith CNS-FHL as early detection and treatment can limit thedevelopment of clinical symptomatology and permanent po-tentially devastating neurologic sequelae

CSF neopterin is associated with CNS involvement of HLHamong a broad group of neuroinflammatory disorders24

Levels were elevated in patients 1 and 2 at diagnosis and werenormal following apparent disease control CSF neopterinmay provide a useful biomarker of CNS-FHL disease activityand treatment efficacy

In addition to the importance of diagnostic clarity and geneticcounseling the diagnosis of CNS-FHL has profound treat-ment implications Compared with current primary neuro-inflammatory standard-of-care therapies HCT representsa fundamentally different treatment paradigm correcting anunderlying genetic etiology as opposed to controlling in-flammatory disease with chronic immunosuppression Wefound that HCT using reduced-intensity conditioning regi-mens was tolerated by patients with CNS-FHL without en-graftment failure or evidence of acute GVHD Three of the 4patients achieved CNS-FHL disease control following the firstHCT One patient (patient 2) relapsed in the setting ofa heterozygous PRF1 mutated donor marrow with persis-tently decreased NK cell function despite adequate donorchimerism but achieved normal NK cell function and diseasecontrol after a second HCT using a different bone marrowdonor These findings underscore the potential importance ofscreening prospective sibling stem cell donors with geneticand functional testing

Although all patients in this cohort achieved positive out-comes and are off chronic immunosuppressive therapydelayed diagnoses resulted in significant disability especiallyfor patients 1 and 2 Longer-term follow-up is required toassess response durability given reports of CNS-isolatedrelapses following HCT for systemic HLH2526 Follow-upwill also help clarify optimal post-HCTmonitoring protocolsincluding MRI CSF donor chimerism and NK cell functionThese initial results support efforts to pursue early and ac-curate diagnosis and early treatment with HCT to reducemorbidity and mortality in CNS-FHL a novel curablesyndrome

AcknowledgmentThe authors gratefully acknowledge Biogen for providingnatalizumab through compassionate use This work wassupported in part by the Intramural Research Programs of theNational Human Genome Research Institute the NationalInstitute of Neurologic Disorders and Stroke the NationalInstitute of Allergy and Infectious Diseases and the CommonFund Office of the Director NIH Bethesda MD

Study fundingThis work was supported by Intramural Research Programs ofthe NHGRI NINDS NIAID and the NIH Common FundOffice of the Director NIH Bethesda MD (Z1D-HG200352)

DisclosureLA Benson received research support from Biogen BostonChildrenrsquos Hospital Office of Faculty Development and theNMSS and served as an expert consultant for Vaccine InjuryCompensation Program H Li received research support fromthe American Society of Hematology their spouse isemployed by Novartis LA Henderson received speakerhonoraria for systemic juvenile idiopathic arthritis with Sobifor a talk and received research support from the NIAMS andRheumatology Research Foundation IH Solomon receivedpublishing royalties from Elsevier and consulted for S2NHealth A Soldatos received research support from theNINDS J Murphy reports no disclosures B Bielekova re-ceived royalties from the NIH for patents related to daclizu-mab received research support from the NINDS BiogenAbbVie and Santhera Pharmaceuticals and is an employee ofthe Intramural Research Program of the NIHNIAID ALKennedy received research support from the NIHNCI MJRivkin received research support from the NINDS KJDavies reports no disclosures AP Hsu is a federal govern-ment employee of the NIH SM Holland reports no dis-closures W Gahl received travel funding from the CystinosisResearch Network served as associate editor of MolecularGenetics and Metabolism receives licensing royalties froma patent on ManNAc and received research support from theNIH RP Sundel received payment from Medical EducationResources received speaker honoraria from Boston IBDconference and for a talk on CRMO served as section editorof Pediatric Rheumatology and UpToDate received publishing


royalties from UpToDate consulted for American College ofRheumatology Vasculitis Committee on Pediatric Vasculitisand Growthpoint is a member of the Lyme GuidelinesCommittee received research support from SimulConsultand NIH and served as an expert for Sanofi LE LehmannMA Lee S Alexandrescu and BA Degar report no dis-closures CN Duncan served on the advisory board of Pfizerand Bluebird Bio received publishing royalties from Upto-Date consulted for Bluebird Bio Magenta and AbGenomicsdid review document preparation for Peer View Instituteprepared course content for Open CME and received re-search support from the St Baldrickrsquos Foundation MPGorman received research support from Pfizer NovartisBiogen and the NMSS Go to NeurologyorgNN for fulldisclosure

Publication historyReceived byNeurology Neuroimmunology amp Neuroinflammation January8 2019 Accepted in final form February 15 2019

Appendix Author contributions

Name Location Role Contribution

Leslie ABenson MD

Department ofNeurology BostonChildrenrsquosHospital BostonMA

Author Designed andconceptualized thestudy acquisition ofdata drafted themanuscript andrevised themanuscript forintellectual content

Hojun Li MDPhD

Dana-FarberBoston ChildrenrsquosCancer and BloodDisorders CenterBoston MA


Lauren AHendersonMD

Department ofRheumatologyBoston ChildrenrsquosHospital BostonMA

Author Acquisition of dataand revised themanuscript forintellectual content

Isaac HSolomon MDPhD

Department ofPathology BostonChildrenrsquosHospital BostonMA


ArianeSoldatos MDMPH

UndiagnosedDisease Programof the NationalInstitute ofNeurologicDisorders andStroke BethesdaMD


JenniferMurphy NP


Author Revised themanuscript forintellectual content

Appendix (continued)


BibianaBielekova MD


Author Acquisition andinterpretation ofimmunophenotypingdata participation indiagnostic workupand revised themanuscript forintellectual content

Alyssa LKennedy MDPhD



Michael JRivkin MD

Department ofNeurology BostonChildrenrsquos HospitalBoston MA


Kimberly JDavies MD



Amy P Hsu BA Laboratory ofClinicalImmunology andMicrobiologyNational Instituteof Allergy andInfectiousDiseasesBethesda MD

Author Diagnosed andperformed genetictesting for patient andrevised themanuscript forintellectual content

Steven MHolland MD

Laboratory ofClinicalImmunology andMicrobiologyNational Institute ofAllergy andInfectious DiseasesBethesda MD

Author Care for Patient 1 andrevised themanuscript forintellectual content

William AGahl MD PhD

National HumanGenome ResearchInstituteBethesda MD


Robert PSundel MD



Leslie ELehmann MD



Michelle ALee MD PhD

Division ofHematologyOncology andMarrowampBloodCellTransplantationChildrenrsquos Hospitalat MontefioreBronx NY


SandaAlexandrescuMD




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hemophagocytic lymphohistiocytosis Neurology 2012781150ndash11562 Horne A Trottestam H Arico M et al Frequency and spectrum of central nervous

system involvement in 193 children with haemophagocytic lymphohistiocytosis Br JHaematol 2008140327ndash335

3 Henter JI Horne A Arico M et al HLH-2004 diagnostic and therapeutic guidelinesfor hemophagocytic lymphohistiocytosis Pediatr Blood Cancer 200748124ndash131

4 Degar B Familial hemophagocytic lymphohistiocytosis Hematol Oncol Clin NorthAm 201529903ndash913

5 Zhang K Filipovich AH Johnson J Marsh RA Villanueva J Hemophagocytic lym-phohistiocytosis familial In Pagon RA Adam MP Ardinger HH et al editorsGeneReviewsreg Seattle University of Washington 1993

6 Fischer A Cerf-Bensussan N Blanche S et al Allogeneic bone marrow trans-plantation for erythrophagocytic lymphohistiocytosis J Pediatr 1986108267ndash270

7 Marsh RA KimMO Liu C et al An intermediate alemtuzumab schedule reduces theincidence of mixed chimerism following reduced-intensity conditioning hematopoi-etic cell transplantation for hemophagocytic lymphohistiocytosis Biol Blood MarrowTranspl 2013191625ndash1631

8 Shinoda J Murase S Takenaka K Sakai N Isolated central nervous system hemo-phagocytic lymphohistiocytosis case report Neurosurgery 200556187

9 Moshous D Feyen O Lankisch P et al Primary necrotizing lymphocytic centralnervous system vasculitis due to perforin deficiency in a four-year-old girl ArthritisRheum 200756995ndash999

10 Tesi B Chiang SC El-Ghoneimy D et al Spectrum of atypical clinical presentationsin patients with Biallelic PRF1 Missense mutations Pediatr Blood Cancer 2015622094ndash2100

11 Khazal S Polishchuk V Soffer G Prinzing S Gill J Mahadeo KM Allogeneic he-matopoietic stem cell transplantation is associated with cure and durable remission oflate-onset primary isolated central nervous system hemophagocytic lymphohistiocy-tosis Pediatr Transpl 201722e13101

12 Algahtani H Absi A Bassuni W Shirah B Adult-onset hemophagocytic lymphohis-tiocytosis type 2 presenting as a demyelinating disease Mult Scler Relat Disord 20182577ndash82

13 Solomon IH Li H Benson LA et al Histopathologic correlates of familial hemo-phagocytic lymphohistiocytosis isolated to the central nervous system J NeuropatholExp Neurol 2018771079ndash1084

14 Li H Benson LA Henderson LA et al Central nervous system-restricted familialhemophagocytic lymphohistiocytosis responds to hematopoietic cell transplantationBlood Adv 20193503ndash507

15 Bigi S Fischer U Wehrli E et al Acute ischemic stroke in children versus youngadults Ann Neurol 201170245ndash254

16 Krupp LB Tardieu M Amato MP et al International Pediatric Multiple SclerosisStudy Group criteria for pediatric multiple sclerosis and immune-mediated centralnervous system demyelinating disorders revisions to the 2007 definitions Mult Scler2013191261ndash1267

17 Pittock SJ Debruyne J Krecke KN et al Chronic lymphocytic inflammation withpontine perivascular enhancement responsive to steroids (CLIPPERS) Brain 20101332626ndash2634

18 Hutchinson C Elbers J Halliday W et al Treatment of small vessel primary CNSvasculitis in children an open-label cohort study Lancet Neurol 201091078ndash1084

19 Gahl WAMarkello TC Toro C et al The National Institutes of Health UndiagnosedDiseases Program insights into rare diseases Genet Med 20121451ndash59

20 Gahl WA Mulvihill JJ Toro C et al The NIH Undiagnosed Diseases Program andNetwork applications to modern medicine Mol Genet Metab 2016117393ndash400

21 Han S Lin YC Wu T et al Comprehensive immunophenotyping of cerebrospinalfluid cells in patients with neuroimmunological diseases J Immunol 20141922551ndash2563

22 Rubin TS Zhang K Gifford C et al Perforin and CD107a testing is superior to NKcell function testing for screening patients for genetic HLH Blood 20171292993ndash2999

23 Tobin WO Guo Y Krecke KN et al Diagnostic criteria for chronic lymphocyticinflammation with pontine perivascular enhancement responsive to steroids (CLIP-PERS) Brain 20171402415ndash2425

24 Howells DW Strobel S Smith I Levinsky RJ Hyland K Central nervous systeminvolvement in the erythrophagocytic disorders of infancy the role of cerebrospinalfluid neopterins in their differential diagnosis and clinical management Pediatr Res199028116ndash119

25 Bock AM LeVeque M Camitta B Talano JA Successful treatment of recurrent CNSdisease post-bone marrow transplant in children with familial hemophagocytic lym-phohistiocytosis Pediatr Blood Cancer 2016632154ndash2158

26 Lounder DT Khandelwal P Chandra S et al Incidence and outcomes of centralnervous system hemophagocytic lymphohistiocytosis relapse after reduced-intensityconditioning hematopoietic stem cell transplantation Biol Blood Marrow Transpl201723857ndash860



Barbara ADegar MD



Christine NDuncan MD


Author Designed andconceptualized thestudy and revised themanuscript forintellectual content

Mark PGorman MD




DOI 101212NXI000000000000056020196 Neurol Neuroimmunol Neuroinflamm

Leslie A Benson Hojun Li Lauren A Henderson et al Pediatric CNS-isolated hemophagocytic lymphohistiocytosis

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is an official journal of the American Academy of NeurologyNeurol Neuroimmunol Neuroinflamm

Despite advances in diagnostic capabilities in neuro-immunology highlighted most dramatically by antineuronalantibody biomarkers some patients continue to have idiopathicor poorly defined chronic andor treatment-refractory neuro-inflammatory disorders Neuroinflammatory findings arepresent in more than half of the patients with hemophagocyticlymphohistiocytosis (HLH)12 However systemic in-flammation is a fundamental diagnostic component of HLHwhich is traditionally characterized by cytokine storm multi-organ dysfunction and ultimately death if untreated3 HLHpathogenesis has been elucidated through the discovery of fa-milial HLH (FHL)-associated genes which encode proteinsrequired for CD8+ lymphocyte and natural killer (NK) cellcytotoxicity Deficiency in cytotoxic killing results in antigenpersistence leading to the hyperinflammatory state associatedwith FHL4 The genes most commonly mutated in FHL arePRF1 (FHL2) and UNC13D (FHL3)5 Hematopoietic celltransplantation (HCT) is the only definitive cure for FHL67

Although common at diagnosis of systemic HLH CNS mani-festations rarely precede systemic findings12 Hemophagocy-tosis has been found in a brain biopsy of a patient not fulfillingthe systemic HLH criteria8 Patients with PRF1mutations havebeen reported with isolated CNS inflammation for weeks tomonths before therapy initiation19ndash12 2 such patients un-derwent HCT911 These reports raise but do not answer thequestion as to whether isolated chronic CNS inflammation canoccur in the absence of systemic HLH findings

Here we describe 4 patients with progressive CNS-isolated in-flammation and mutations in FHL-associated genes who neverfulfilled the systemic diagnostic criteria for HLH except fordefective NK cell activity reflective of their underlying geneticmutations The neuropathologic details of 3 of the patients13 andHCT outcome details of all 4 patients14 have been previouslyreported HCT was well tolerated in these patients and resultedin arrest of disease progression highlighted by the lack of de-velopment of morbidity in an asymptomatic patient with pro-gressive radiologic neuroinflammatory findings

Case reportsBaseline characteristics at diagnosis for all patients are describedin the table

Patient 1The index case presented as a 5-year-old girl with headachevomiting mild ataxia multifocal contrast-enhancing cerebral

and cerebellar T2-hypertintense lesions (figure 1 figure 2) andCSF pleocytosis meeting the diagnostic criteria for a de-myelinating clinically isolated syndrome (CIS)16 Her symp-toms and imaging findings responded to high-doseglucocorticoids (methylprednisolone 30 mgkg IV for 5 days)but returned with steroid tapers despite monthly IV immuno-globulin (IVIg) (1 gkgdose) Serial brain MRI identifiedenhancement patterns reminiscent of chronic lymphocytic in-flammation with pontine perivascular enhancement responsiveto steroids (CLIPPERS)17 Biopsy of a cerebellar lesion iden-tified a mixed chronic inflammatory infiltrate with perivascularinflammation diffusely involving the gray and white matter andleptomeninges (figure 1) Induction treatment with gluco-corticoids and cyclophosphamide followed by mycophenolatemofetil per small vessel CNS vasculitis protocol18 slowed butdid not halt disease progression She experienced progressivetremor ataxia dysarthria dysconjugate gaze and slowed speechand response time Extensive evaluation including meta-genomic next-generation sequencing of CSF and brain tissuewas unrevealing for infection Given her severe symptomatol-ogy worsening course and prominent T cells on repeat biopsy(left thalamic lesion data not shown) natalizumab whichinhibits T-cell entry into the CNS was given under a compas-sionate use investigational new drug protocol (IND 114796150 mg for 4 mgkg every 4ndash5 weeks) While on natalizumabher condition clinically and radiographically improved and thenstabilized for 3 years before eventual clinical and radiographicprogression The patient was referred to the UndiagnosedDiseases Program at the NIH1920 but during the evaluationshe developed rotary nystagmus and worsened ataxia with lossof independent ambulation partly responsive to high-dosemethylprednisolone Ultimately CSF immunophenotypingand soluble biomarker analysis21 revealed activation of the in-nate immune system PRF1 gene sequencing identified biallelicpreviously reported pathogenic mutations 5 years after initialpresentation CSF neopterin was significantly elevated (table)Throughout her course patient 1met no systemicHLH criteriaexcept for decreased NK cell activity initially attributed tochronic immunosuppression Following diagnosis she receivedCNS-directed HLH therapy with intrathecal methotrexate andhydrocortisone and systemic dexamethasone with clinical im-provement and normalization of the neopterin level beforeundergoing unrelated donor HCT at age 10 years

Patient 2A 6-year-old girl developed vomiting headache psychiatricsymptoms and cognitive decline over 3 months and then hadfocal status epilepticus prompting hospitalization Additionalsymptoms noted during the hospitalization included left

GlossaryADEM = acute disseminated encephalomyelitis CIS = clinically isolated syndrome CLIPPERS = chronic lymphocyticinflammation with pontine perivascular enhancement responsive to steroids FHL = familial hemophagocytic lymphohistio-cytosis GVHD = graft-versus-host disease HCT = hematopoietic cell transplantation HLH = hemophagocyticlymphohistiocytosis IVIg = IV immunoglobulinMDEM = multiphasic ADEM NK = natural killerWBC = white blood cell





Sex F F F F


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Ataxia Y Y Y N N

Weakness Y Y Y N N

Seizure Y Y Y N Y

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Lauren AHendersonMD









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Michael JRivkin MD



Kimberly JDavies MD





Steven MHolland MD






Robert PSundel MD



Leslie ELehmann MD






SandaAlexandrescuMD

































Barbara ADegar MD






Mark PGorman MD



















Reprints
















































Leslie ABenson MD



Hojun Li MDPhD



Lauren AHendersonMD









JenniferMurphy NP





BibianaBielekova MD






Michael JRivkin MD



Kimberly JDavies MD





Steven MHolland MD






Robert PSundel MD



Leslie ELehmann MD






SandaAlexandrescuMD

































Barbara ADegar MD






Mark PGorman MD



















Reprints

































Barbara ADegar MD






Mark PGorman MD



















Reprints



















Reprints




pediatric cns-isolated hemophagocytic lymphohistiocytosis · lymphohistiocytosis (hlh). 1,2...

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