Clinical Diagnosis of ITP · T cell B cell Megakaryocyte Treatment Who: - Platelet count, symptoms,...

ISTH Advanced Training Course

Dubai, UAE


Clinical Diagnosis of ITP

Nichola Cooper

September 2016


Dubai, UAE

Disclosures for

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In compliance with COI policy, ISTH requires the following

disclosures to the session audience:

Research Support/P.I. Pfizer

Employee No relevant conflicts of interest to declare

Consultant Novartis, Amgen

Major Stockholder No relevant conflicts of interest to declare

Speakers Bureau No relevant conflicts of interest to declare

Honoraria Amgen, Novartis

Scientific Advisory

BoardAmgen, Novartis

Presentation includes discussion of the following off-label use of a drug or medical device:

<N/A>


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Immune Thrombocytopenia (ITP)

Isolated thrombocytopenia (platelets <100)

Exclusion of other causes:

Inherited platelet disorders

Increased destruction: (other autoimmunity, viruses)

Decreased production: (viruses, infiltration, marrow failure)

1 in 40,000 F>M, approximately 1-2% familial

Definitions:

Newly diagnosed (0-3 months)

Persistent (3-12 months)

Chronic (>12 months)

Refractory


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ITP terminology: 3 ITP phases

0–3 months 3–12 months >12 months

Newly diagnosed ITP

Persistent ITP Chronic ITP

Possibility of spontaneous remission:

0 to 12 months

Rodeghiero F, et al. Blood 2009; 113: 2386–93.


Dubai, UAE

Bone marrow

APAs

T cell

B cell

Megakaryocyte

Treatment

Who:

- Platelet count, symptoms,

psychology

When:

- Newly diagnosed,

persistent, chronic

What:

- Steroids, IVIg, rituximab,

TPO-RAs, MMF, other…

Challenges in managing ITP

Diagnosis

Pathogenesis

Treatment

APA, against phospholipid antigens; IVIg, intravenous immunoglobulin;

TPO-RA, thrombopoietin receptor agonist; MMF, mycophenolate mofetil


Dubai, UAE

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Overview

There is no diagnostic test for ITP, it is a

diagnosis of exclusion

The pathology of ITP is unclear and may

have multiple causes

This is relevant when considering

treatment, which should be tailored to

individual patient details


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Pathogenesis (known and presumed)

Genetic susceptibility vs environment

Serum derived effects: antiplatelet antibodies

Unregulated T helper cells and activated

cytotoxic T cells

Inhibition of megakaryopoiesis (?)


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Genetic susceptibility vs infection

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Cines DB et al. Blood 2009;113:6511-6521


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Platelet count after infusion with patient plasma

Antiplatelet antibodies: 19511 Inhibition of megakaryocytes byplasma from ITP patients: 20042

What is the evidence for immune

pathology in ITP? B-cell disease

1. Harrington WJ et al. J Lab Clin Med 1951;38:1–10;

2. McMillan R et al. Blood 2004;103:1364–1369

Time

Pla

tele

ts (

x10

9/L

)

0

400

600

800

1000

200

1 2 3 1 2 3 4 5 6 7 8 9Hours Days

Co

ntr

ol m

egak

aryo

cyte

s(%

)

100

75

50

25

0


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T-cell proliferation in response to own platelets and Th1-mediated disease: 1990s1

Cytotoxic T-cell attack against platelets: 20032

What is the evidence for immune

pathology in ITP? T-cell-mediated

1. Semple JW & Provan D. Curr Opin Hematol 2012;19:357–362;

2. Olsson B et al. Nat Med 2003;9:1123–1124

200

24

20

16

12

3H

-th

ymid

ine

inco

rpo

rati

on

(x

10–3

)

8

4

00 50 100 150

Platelets (x109/L)

40

30

20

10

0

Control

Active ITP

ITP in remission

Pla

tele

t ly

sis

CD4

IL-2IFN-γIL-15

IL-4IL-10TGF-β

Active disease(Th1)

Remission(Th2/3)

CD, cluster of differentiation; IFN-γ, interferon-γ; IL, interleukin;

TGF-β, transforming growth factor-β; Th, helper T cell


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Activated T cells,

Th1,

Th17 profile1–4

Pro-inflammatory

cytokines1–6

Impaired Treg

function7

Immunological profile in ITP

Figure reproduced with permission from Chong BH. Blood 2010;116:4388–4390 (professional

illustration by Marie Dauenheimer); 1. Gernsheimer T. Oncologist 2009;14:12–21; 2. Semple JW &

Freedman J. Blood 1991;78:2619–2625; 3. Semple JW et al. Blood 1996;87:4245–4254; 4. Wang T et

al. Haematologica 2005;90:914–923; 5. Ogawara H et al. Eur J Hematol 2003;71:283–288;

6. Olsson B et al. Nat Med 2003;9:1123–1124; 7. Yu J et al. Blood 2008;112:1325–1328 Treg, regulatory T cell


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Complex ITP pathophysiology underlies

platelet destruction

MHC, major histocompatibility complex;

Tc, cytotoxic T cell; TCR, T-cell receptor

Th cell

IL-2IFN-γ

B cell

Impaired megakaryocyte

maturation

Reduced platelet production

Megakaryocyte

Tc cell

Tc-cell-mediated

platelet

destruction

Platelets

Epitope

spreading

Macrophage

Platelet

phagocytosis

CD80

CD80

CD28

CD28

CD40

CD40

MHC IITCR

CD154

CD154

Platelet autoantibody production

Adapted from Stasi R et al. Thromb Haemost 2008;99:4–13


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MK development:GATA1, RUNX1, FLI1,

ANKRD26, NBEAL2, GFI1N

TPO

Stem cell niche Vascular nicheMigration: SD1a

Pro-platelet development: Cytoskeleton: Rock, myosin

(inhibition Inc)

Collagen, VWF, fibrinogen (GpIb-IX-V, aIIbB3)

Sheerstress

Blood vessel

Bo

ne

Many factors are involved in making

platelets

Adapted from Pecci A & Balduini CL. Br J Haematol 2014;165:179–192 MK, megakaryocyte; TPO, thrombopoietin


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Ashwell–Morell receptor in hepatocytes:

A possible method of platelet regulation

Kile BT. Nat Med 2015;21:11–12

Old platelets are recognized by the Ashwell–Morell receptor (AMR) in hepatocytes causing thrombopoietin production


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Thrombopoietin levels are lower than

expected in ITP

1. Kosugi S et al. Br J Haematol 2003;93:704–706;

2. Aledort LM et al. Am J Hematol 2004;76:205–213

14

Thro

mb

op

oie

tin

leve

l

Normal

12

10

8

6

4

2

0ITP Aplastic

anaemia

No correlation between platelet count and TPO levels in patients with ITP1

No significant difference in TPO levels between patients with ITP and controls2


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Thymus

Bone Marrow

Liver

Spleen

Lymph node

Blood vessel


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Causes of Thrombocytopenia

Platelet production and regulation is not

fully understood

Thrombocytopenia can be the end result of

many events

Gene defects in megakaryocyte development

Bone marrow infiltration

Infection

Increased destruction (APAs and T cells)

? Abnormalities of thrombopoietin regulation


Dubai, UAE

Clinical diagnosis of ITP: medical history

• Presenting Complaint

– Extent of bleeding (inherited platelet disorders)

• Past Medical History

– Infections (immunodeficiency)

– Previous bleeding (tonsils, teeth)

• Review Of Systems

– Joints, rashes, hair loss, mouth ulcers, night sweats, weight loss (lupus, lymphoproliferative disorders)

• Family History

– Bleeding disorders, thrombocytopenia, autoimmune diseases


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Examination

Extent of bleeding, area of bleeding

Bleeding scores

Lymphadenopathy (not consistent with ITP)

Splenomegaly (not consistent with ITP)

Hepatomegaly (not consistent with ITP)

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pseudothrombocythaemia

ErythrocytesFragmentation: TTP/HUS/DIC/HELLPMacrocytes: megaloblastosis, haemolysisSpherocytes: Evans

PlateletsLarge: hereditary thrombocytopeniasSmall: Wiscott-Aldrich syndromeLarge and small: ITP

LeukocytesToxic granulesPMN InclusionsAtypical lymphocytes (infection, ITP)Blasts: leukaemia

? Bone marrow examination

Blood film examination


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.

Drachman J G Blood 2004;103:390-398

©2004 by American Society of Hematology

Other causes of thrombocytopenia

based on platelet size


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Small platelets MPV <7fL Normal platelets MPV 7-11 fL

Large/Giant platelets, MPV >11 fL

Wiskott-Aldrich syndrome Familial platelet disorder/AML

MHY9-related diseasesMay-Hegglin anomalySebastian syndromeFechtner syndromeEpstein syndrome

X-linked thrombocytopenia Chromosome 10/THC2 Bernard-Soulier syndrome

CongenitalAmegakaryocyticthrombocytopenia

Paris-Trousseauthromboctopenia/Jacobsen syndrome

Thrombocytopenia and absent radii

Velcardiofacial/DiGeorgesyndrome

GATA1 mutation

Grey platelet syndrome


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Platelet size for distinguishing between inherited

thrombocytopenias and immune thrombocytopenia: a

multicentric, real life study

British Journal of HaematologyVolume 162, Issue 1, pages 112-119, 25 APR 2013 DOI: 10.1111/bjh.12349http://onlinelibrary.wiley.com/doi/10.1111/bjh.12349/full#bjh12349-fig-0001

ITPControl

http://onlinelibrary.wiley.com/doi/10.1111/bjh.2013.162.issue-1/issuetoc

http://onlinelibrary.wiley.com/doi/10.1111/bjh.12349/full#bjh12349-fig-0001


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Investigations in persistent ITP

Immunodeficiency:

Immunoglobulins

Lymphocyte subsets

Platelet disorders:

vWAg (children)

Autoimmunity:

DRVVT

ACL

Autoimmune profile

TFTs

Infections:

Hep B, Hep C, HIV,

CMV

H pylori


Dubai, UAE

Controversial Investigations in

persistent ITP

Bone marrow examination: Guidelines recommend only in those with abnormal features,

those over 60 and pre-splenectomy

Anti-platelet antibodies: not recommended (not sensitive or

specific enough to direct treatment)


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Bone marrow in ITP

Useful to confirm no other features causing

thrombocytopenia

Useful to describe megakaryocyte features

Useful for a baseline test pre long term

treatment (3 of our cohort have developed

other bone marrow conditions – OS, MF, MDS)

May be useful in the future to direct treatment

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We analysed the interaction of CD4 cells

CD8 cells with megakaryocytes in bone

marrow

29Thomas Mayo Unpublished work Thomas Mayo and Anwar Sayed


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ITP patients have higher MGK density in

the bone marrow but no difference in size

Mann Whitney U

Thomas Mayo


Dubai, UAE

ITP patients have similar T-cell numbers

to disease controls

Mann Whitney U

Thomas Mayo


Dubai, UAE

But more CD4/MK interactions and

clustering (more than 1 CD4 per MK)

Mann Whitney U

Thomas Mayo


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Increased CD4/MK interactions in

patients with chronic disease

Mann Whitney U

Thomas Mayo


Dubai, UAE

No difference in CD8/MK interactions or

CD8/MK clustering

Mann Whitney U

Thomas Mayo


Dubai, UAE

Co

ntro

l

I TP

0

2 0

4 0

6 0

8 0

1 0 0

C D 8 I F N y+

CD

8%

Co

ntro

l

I TP

0

1 0

2 0

3 0

4 0

C D 8 I L - 2+

CD

8%

Peripheral blood inflammatory CD8

is significantly higher in ITP patients

** **

** P value <0.01

CD8 INF g CD8 IL2

Control Patient Control Patient

Anwar Sayed


Dubai, UAE

Expression of proinflammatory markers

is not increased in CD4 cells of ITP

patients

Co

ntro

l

I TP

0

2 0

4 0

6 0

8 0

C D 4 T N F a+

CD

4%

Co

ntro

l

I TP

0

2 0

4 0

6 0

8 0

C D 4 I L - 2+

CD

4%

*

* P value <0.05

CD8 TNF a CD8 IL2

Control Patient Control Patient

Anwar Sayed


Dubai, UAE

C o n t r o l I T P

0

2

4

6

8

1 0

R e g u l a t o r y T c e l l

%

of

R

eg

ul

at

or

y

T

ce

ll

s i

n

CD

4+

po

pu

la

ti

on

C o n t r o l

I T P

Regulatory T cells are significantly

reduced in the periphery in ITP patients

*

* P value <0.05

Control Patient

Anwar Sayed


Dubai, UAE

Bone marrow changes

May reflect disease activity

Correlation with clinical findings and

peripheral blood abnormalities is in

progress

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Antiplatelet antibodies are difficult to

measure

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Published in: Linlin Shao; Yang Wu; Hai Zhou; Ping Qin; Heyu Ni; Jun Peng; Ming Hou; Platelets 2015, 26, 495-497.DOI: 10.3109/09537104.2014.948838Copyright © 2014 Informa UK Ltd.

Successful treatment with oseltamivir

phosphate (tamiflu) in a patient with

chronic immune thrombocytopenia

positive for anti-GPIb/IX autoantibody


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Potential use for APAs

Identification of specific APAs may allow:

Identification of types of ITP

May be able to predict bleeding and

May predict treatment responses

New methods are needed

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Thrombopoietin levels may distinguish

between consumptive or hypoproliferative

causes of thrombocytopenia

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TPO level

☐Consumptive Hypoproliferative

L

Maker et al Haematology 2013: Thrombopoietin levels in patients with disorders of platelet production: Diagnostic potential and utility in predicting response to TPO Receptor agonists


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Reduced absolute immature platelet

fraction (IPF), but increased % IPF in

ITP

Barsam, et al Blood: Platelet production and platelet destruction: assessing mechanisms of treatment effect in immune thrombocytopenia

IPF %

Absolute IPF

ITP patients

Controls


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IPF may allow a better understanding of

treatment effects

Barsam, et al Blood: Platelet production and platelet destruction: assessing mechanisms of treatment effect in immune thrombocytopenia


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Relevance to ITP patients

Variable findings in the pathology

Only 60% have identifiable antibodies

T cell and bone marrow changes variable

Variable responses to many treatments

with no biomarkers

60% respond to splenectomy

50% respond to rituximab

30-50% respond to immunosuppression

Wide variability in responses shows a

diverse disease

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Treatment recommendations based on

platelet count

Images courtesy of N Cooper

Treat adults

Only treat if bleeding, requiring surgery or requiring anticoagulation or antiplatelet agents

Treatment depends on individual patient factors: Age, other

comorbidity, success of treatment

0 10x109/L 30x109/L 150x109/L

Platelet count

Only treat children with symptoms


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Risk of fatal/severe bleeding in ITP

Presence of other conditions such as

hypertension or cerebrovascular disease

Increased rates of fatal haemorrhage rates

with age

0.004 per patient-year <40 yrs

0.012 per patient-year 40 to 60 yrs

0.130 per patient-year >60 yrs

Increased risk of VTE

George JN. J Thromb Haemost. 2006;4(8):1664-72.; Cohen YC, et al. Arch Intern Med. 2000;160:1630–38; Sarpatwari A, et al. Haematologica. 2010 Jul;95(7):1167-75.


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Management of immune thrombocytopenia (ITP)

Newly diagnosed ITP (0-3 months)Steroids or IVIG

Persistent ITP (3 to 12 months)MMF or thrombopoietin receptor

agonists or Rituximab

Chronic ITP (>12 months)Continuous TPO-RAs

Repeated rituximab (+/- dexamthasone)Continuous MMF

Other: (danazol, dapsone, hydroxychloroquine)

Splenectomy

ThrombocytopeniaDiagnosis: infection, drugs, inherited, acquired


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What factors are involved in treatment

decision making?

Emergency therapy

HRQoL Efficacy

CostPatient

type Ease of use

SafetyClinic

attendance

Rituximab: Increased response in women within first 2 years of diagnosis

MMF: Less effective in patients with virus-associated ITP

TPO-RAs: Potential risk of thrombosis

Consider other factors


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Refractory ITP (approx 5%)

Consider the diagnoses, is it ITP?

Role at this stage for bone marrow

examination

Role for antiplatelet antibody testing

? Role for NGS

There is a real need for better

understanding of ITP and better diagnostic

markers

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Summary

ITP is a diagnosis of exclusion of other

causes of thrombocytopenia

Better understanding of the underlying

pathology is needed

Anti-platelet antibodies may be helpful

TPO levels, bone marrow biopsies and

NGS have potential

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Plan

Establish better antiplatelet antibody testing

Establish a B and T cell

immunophenotyping panel to differentiate

patients in to subtypes

Analyse more bone marrow samples

Correlate TPO levels with outcome

Prospective studies to establish biomarkers

Better phenotyping will help to guide

genomic studies…….- 53 -

Clinical Diagnosis of ITP · T cell B cell Megakaryocyte Treatment Who: - Platelet count, symptoms,...

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Transcript of Clinical Diagnosis of ITP · T cell B cell Megakaryocyte Treatment Who: - Platelet count, symptoms,...