1. TREATMENT OF HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS G.E. Janka Hamburg
Iran November 2018

2. HAEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Diagnostic criteria1
Familial disease/known genetic defect
Clinical and laboratory criteria (5 out of 8)
Fever
Splenomegaly
Cytopenia of => 2/3 cell lines
Hemoglobin <90g/l, platelets <100x109/l, ANC <1x109/l
Hypertriglyceridemia and/or hypofibrinogenemia
Fasting triglycerides =>3mmol/l, fibrinogen <1.5g/l
Haemophagocytosis in bone marrow, CSF, lymphnodes
Ferritin =>500µg/l
sCD25 =>2400U/ml
Decreased or absent natural killer cell activity
Strong supportive evidence are cerebral symptoms with moderate pleocytosis and
or elevated protein, elevated transaminases, bilirubin and LDH
1HLH Study Group of the Histiocyte Society, Henter et al. PBC 2007

3. New diagnostic criteria
MAS - HLH
New diagnostic criteria
Multistep process:
Delphi survey to identify MAS features potentially suitable als criteria
Data collection on patients with sJIA-associated MAS (362), patients with sJIA not complicated by MAS (404), and patients with systemic infections (345)
Selection of candidate criteria from patients with consensus (391/428) in 28
experts (cut-off values calculated by ROC)
Selection of final criteria by consensus conference
Final criteria
Ravelli 2016
MAS is diagnosed in a febrile patient with sJIA if the following criteria are met:
Ferritin > 684ng/mL and any 2 of the following
Platelet count ≤ /µL
Aspartate aminotransferase (GOT) > 48 U/L
Triglycerides > 156 mg/dL
Fibrinogen ≤ 360 mg/dL

4. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
MAS as first presentation of soJIA versus FHL/VA-HLH
Retrospective analysis from German pediatric HLH database (Lehmberg J. Pediat 2013)
pHLH = 90, VAHS = 42, MAS in soJIA= 27
Variables in favour of MAS ROC-AUC Sensitivity Specificity
ANC ≥ 1.8 x 109/L
CRP ≥ 90mg/L
sCD25 ≤ 7900 U/L
pHLH: n = 258, MAS in soJIA: n = (Minoia F, J Pediatr 2017)
Development of „MH Score“ with 6 variables most closely associated with pHLH:
Age ≤ years Platelets ≤ /nL
ANC ≤ /nL Hemoglobin ≤ g/L
Fibrinogen ≤ g/L Splenomegaly present
Score assigned depending on statistical weight of each variable in multivariate analysis
(median score: genetic HLH 97, MAS 12; best cut-off 59: sensitivity 91%, specificity 93%)

5. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
How to differentiate between genetic and acquired HLH?
Identification of an infectious agent not helpful
severity of disease not helpful
Age helpful
~90% of children below 1 year will have
genetic HLH
Do not forget: mutations in HLH-related genes are
being increasingly identified in adolescents or adult patients
Functional tests (degranulation, expression of perforin, SAP and
XIAP) or mutation analysis allow differentiation between genetic and
acquired forms, but are not widely available or still too expensive.
The distinction between genetic and acquired HLH is not important for initial treatment which is guided mainly by the severity of the disease!

6. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
General statement
It is important to realize that HLH is a syndrome where
often an underlying trigger/condition can be identified.
In children infections (mostly viral), autoinflammatory/ autoimmune diseases, malignant diseases and (rarely) metabolic diseases have to be considered.
The search for a trigger is of utmost importance! Especially in adults and older children lymphomas (ultrasound, x-ray, CT, MRI) must be ruled out.
Usually HLH-directed therapy is needed in addition to the specific treatment of the triggering condition
(exceptions: leishmaniasis, tuberculosis).

7. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Underlying conditions and classification in children
Genetic HLH Acquired HLH
● Familial HLH (FHL)
(„Farquhar`s disease“) ● Infectious agents
● Endogenous products
- tissue damage
- radical stress
- metabolic products
● Autoimmune/autoinflammatory
● Immune deficiency diseases (Macrophage activation
syndromes syndrome; MAS)
Chédiak-Higashi syndrome 1
Griscelli syndrome  Malignant diseases
X-linked proliferative syndrome
Other inborn immune deficiencies ● Acquired immune defects
Frequency ~ 1 : 5
bi-allelic mutations
Perforin
UNC13D
Syntaxin11
UNC18B
HLH

8. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Principles of treatment
Treatment for HLH depends on the underlying disease
and severity of symptoms.
Children with MAS are usually treated successfully with
high-dose corticosteroids ± ciclosporin A.
In non-responders anakinra is effective.
HLH cases with less severe symptoms may only need
corticosteroids ± immunoglobulins –
but be aware of the dynamics of the disease!

9. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Principles of treatment
HLH therapy is a two-sided sword
On one hand hyperinflammation has to be suppressed
to prevent the dangerous consequences of
hyperinflammation; on the other hand immune-
suppressive and cytostatic treatment may be
counterproductive for control of infectious triggers
and recovery of the bone marrow.
In contrast to malignant diseases the aim is not to destroy as many
cells as possible by intensive therapy but just to down-regulate the
hyperactive immune response and to reduce the number of activated
(infected) cells in the hope to achieve a new balance.

10. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Principles of treatment
Suppression of hyperinflammation
corticosteroids
immunoglobulins
cyclosporin A
anti-cytokine treatments
antibodies
Elimination of (infected) cells: APCs, lymphocytes
etoposide, rituximab, corticosteroids
antithymocyte globulin/alemtuzumab
Treatment of the trigger
Replacement of the defective immune system
by stem cell transplantation in genetic cases

11. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Treatment
Immunosuppressive/immunomodulatory agents
- Corticosteroids
- Intravenous immunoglobulins
- Cyclosporin A
- Cytokine removal (plasmapheresis, cytokine-absorbers)
- Antagonists of single cytokines: IL-1, IL-6, INFγ
- Inhibition of Janus kinase 1/2: ruxolitinib
- Antibody against IL-2 receptor: basiliximab
Cytotoxic drugs
- Corticosteroids
- Etoposide (Ambruso 1980)
- T-and B-cell antibodies (ATG, alemtuzumab, rituximab)

12. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Founding of the Histiocyte Society in 1985
Founding of the HLH Study Group in 1989
Founding members
Maurizio Aricò
Jan-Inge Henter
Blaise Favara
Diane Komp
Christian Nezelof
Göran Elinder
Gritta Janka
Jon Pritchard

13. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Two international studies: HLH-1994 and HLH-2004
Follow-up publication by Trottestam H, Blood 2011

14. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
International treatment protocols for HLH
A HLH-1994
B HLH-2004
CSA starts upfront
Prednisolone is added to i.th. therapy

15. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Treatment results with protocol HLH-1994 and HLH-2004
Study HLH-1994
n = 232
33 (14%) died < 2 months
23 (10%) died >2 <12 months
7 ( 3%) died >12 months
40 (17%) died after HSCT
Mortality after
HSCT
40/118 (34%)
Study HLH-2004
n = 369
50 (14%) died < 2 months
17 ( 6%) died >2 <12 months
8 ( 2%) died > 12 months
64 (17%) died after HSCT
Mortality after
HSCT
64/185 (35%)
Trottestam H, Blood 2011; Bergsten E, Blood 2017

16. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
n = 201
n = 369
n = 201
Results of
HLH-2004
n = 369
n = 168
n = 240
n = 240
n = 168
Bergsten et al.
Blood 2017
n = 232
n = 232

17. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
HLH Steering Committee of the
Histiocyte Society
Recommendations for the Use of Etoposide-Based Therapy and Bone Marrow Transplantation for the Treatment of HLH
Ehl S. et al. Journal of Allergy Clinical Immunology Practice 2018
Recommendation of HLH Study Group to use the HLH-94
protocol with HLH-2004 diagnostic criteria

18. Authors of „Recommendations for use of protocol HLH-1994
AnnaCarin Horne
Elena Sieni
Michael Jordan
Jan-Inge Henter
Paul La Rosée
Itziar Astigarraga
Eiichi Ishii
Stephan Ehl
Melissa Hines
Kim Nichols
Tatiana Greenwood
Kai Lehmberg
Zhao Wang
Gritta Janka
Rafal Machowicz

19. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Treatment results with T-cell antibodies
Antithymocyte globulin (Mahloui N 2007)
Results: 28 patients with FHL; 1-2 courses; CR 23/28; PR 5/28
22/28 HSCT; survival 17/22 (77%)
HIT-HLH USA and EURO-HIT-HLH
ATG instead of the first three doses of etoposide
Results not better and not worse than HLH-1994; no publication yet

20. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Treatment results with T-and B-cell antibodies
Alemtuzumab
Pilot study ( ) (Moushous, HS meeting 2016)
23 patients with FHL, 1-3 courses
Results: CR: 87%, PR 9%, 1 nonresponder; 22/23 patients HSCT
Ongoing study (NCT ) (Moushous, HS meeting 2018)
so far 20 patients with FHL; planned 30;
Campath d1 0.5mg/kg; d 2+3 1mg/kg + Pred d mg/kg, then taper, + CSA
same response rate; ~50% received haplo-BMT
Rituximab (Chellapandian, Rituximab Study Group 2013)
Retrospective survey; 42 patients with EBV-HLH; median 3 doses
(2/3 within 1 mos after diagnosis; combination with other HLH drugs)
Results: significant decrease in EBV load and ferritin; significant increase in platelets and SGOT.

21. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Salvage therapy
Patients with refractory/relapsed HLH, previously treated with steroids + etoposide or steroids + ATG.
At least 2 patients treated.
Literature survey by Working Group on Second-line Treatment in HLH
(R. Marsh, PBC 2017)
Anakinra: 3 patients with MAS (3 CR) (Behrens E, Miettunen PM )
ATG: 2 patients with FHL (2 CR) (Mahlaoui N)
Alemtuzumab: 24 patients with HLH (16 PR, 8 NR) (Marsh R, Strout MP, Gerard LM)
Liposomal doxorubin, etoposide and methyl-prednisolone
Thirty-four adult patients without lymphoma: 12 CR, 14 PR, 8 NR (Wang Z)

22. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
New drugs
Interferon-γ antibody Emapalumab (NI-0501)
NCT : phase II/III study, 20 countries, still recruiting
Inclusion: children with primary HLH; reactivated disease, or
unsatisfactory response, or intolerant to standard therapy
Exclusion: secondary HLH, HLH in rheumatic or malignant diseases
Treatment: antibody 1mg/kg every 3 days + dexamethasone
Results:
Eleven of 13 patients alive after 8 weeks (1 first-line treatment)
Satisfactory response in 9/13 patients (improvement in HLH parameters)
- Seven of 13 patients proceeded to HSCT
- Resolution of CNS symptoms in 2 evaluable patients
- Reduction of DEX in 50% of patients in first 4 weeks.
Update HS meeting 2016: 31 patients recruited,
25 with mutations; 5 first line-treatment
Jordan M, Blood LBA 2015

23. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
New drugs
Ruxolitinib
Ruxolitinib is an oral anti-inflammatory and immunosuppressive
Janus kinase (JAK) inhibitor, approved for myelofibrosis.
Activity has been shown in rheumatoid arthritis, GvHD, psoriasis, ulcerative colitis, and other diseases.
JAK1 and JAK2 control signaling of many cytokines by transmitting cytokine-induced signals, notably from INF-γ, IL-2 and IL-6.
Ruxolitinib is being evaluated in 2 ongoing clinical trial for HLH in adults).There are several recent case reports.

24. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Ruxolitinib
Das et al. (Kim Nichols` group) Blood 2016
Two mouse models for HLH:
PRF1 -/- mouse and CpG induced model for secondary HLH
Preemptive treatment with ruxolitinib starting day 4 (to day 9)
Prevention of HLH symptoms, hypercytokinemia and
tissue inflammation, enhancement of survival
Maschalidi et al. (Alain Fischer`s group) Blood 2016
Two mouse models for HLH:
PRF1 -/- mouse and RAB27a -/-
Treatment of manifest HLH with ruxolitinib for 14 days
starting day 7 (PRF1), or day 10 (RAB27A) post infection
Enhancement of survival (comparable with INF-γ antibody treated mice)
Correction of cytopenias, reduction of tissue inflammation, reversal of
CNS symptoms (RAB27A-/- mouse)

25. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Possible therapeutic algorithm
Less severe cases:
no prolonged neutropenia
no organ failure
no CNS symptoms
no severe coagulopathy
Consider to wait for effect of antiinfectious therapy in mild disease
Functional studies
Severe cases
Steroids, IVIG
EBV-HLH: + Rituximab
good response
24-48 hrs.
stop when CR
and acquired HLH
no response poor PR
stop when CR and acquired HLH
HLH-94
good response
progressive
Unsatisfactory response
CSA
Consider to add
basiliximab/
daclizumab
repeat IVIG
alemtuzumab
(INFγ antibody, ruxolitinib)
cytokine absorption
other cytostatics (e.g. DEP)
salvage
continuous poor response
Consider early transplant

26. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Stem cell transplantation: Reduced intensity conditioning
Author number conditioning regime survival
Horne A, BJH BU, CYT, VP16 ± ATG %
Cooper N BMT FU, MEL, alemtuzumab %
(2-105 mos, med 36)
Marsh R Blood FU, MEL, alemtuzumab pOS 92%
(at 3 years)
Lehmberg K Haemat FU, TREO, THIO (14), alemtu 100% (7-31,med 16mos)
manuscript in preparation %
Allen C, Blood FU, MEL, alemtuzumab % at 18 mos
RIC conditioning is associated with less toxicity and better survival,
but a with a higher frequency of mixed chimerism, necessitating
intervention with DLIs or second transplants

27. HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS
Stem cell transplantation with RIC: mixed donor chimerisms (DC)
(Hartz B HS Blood 2016)
103 patients with FHL from 7 countries, DC < 75%; overall survival 82%
17 (16%) reactivations (10 before d180); overall survival 9/17
18 pts. 2nd transplants (9 with previous reactivations); mortality 33%
Key messages:
● Patients with reactivations before d180 have variable DC and
frequently viral triggers; underlying immunosuppression plays a role.
● A DC of 20-30% protects from reactivation after d180
● DC below 30% does not always result in reactivation of HLH
► 5 pts. line-specific DC (CD3) <10% yrs (med. 5.1) without relapse
● In patients with low DC, risk of 2nd HSCT must be weighed against
risk of reactivation

28. Thank you for your attention!
Our HLH Team
Hamburg: Kai Lehmberg
Udo zur Stadt
Ingo Müller
Anke Clodius
Manuela Adao
Gritta Janka
München: Karin Beutel
Freiburg: Stephan Ehl
Miriam Heizmann
Carsten Speckmann
Sandra Amman
Ilka Fuchs