Pathologie Stephan Dirnhofer

Myeloproliferative Neoplasms (MPN)

PathoBasic: Bone Marrow 20. Oktober 2015

• Cytology

• Cytochemistry

• Histology

• Immunohistochemistry

• FISH

• Flow cytometry

• Genetics

• Cytogenetics

• FISH

• Molecular genetics

• In vitro Culture

• Clinical information

BM investigation: diagnostic tools

&

BM aspirate BM trephine biopsy

- Cytology - Flow cytometry - Karyotype, FISH - Molecular biology

- Histology - Immunohistochemistry - FISH - Molecular biology

COMPLEMENTARY INFORMATION

Courtesy of Prof. L. De Leval

Strength and limits of BM cytology

Strength

Morphology – dysplastic changes

Auer rods, Chediak Higashi granules

Blast counts

Ring sideroblasts

Cytochemistry

Limits

Hodgkin lymphoma/FL

Fibrosis

Cellularity

Topography of the cells

Carcinoma infiltration

Granulomas

PAS Iron

Strength and limits of BM histology

Strength

Cellularity

Topography of the hematopoiesis

Immunohistochemistry

CD34+ counts and localization

Hodgkin and Reed Sternberg cells

Evaluation of fibrosis

Limits

Dysplastic changes

Blast cells

Auer rods and other changes

CD34

Gö

CD30

Clinical Information is essential

• Blood count (CBC) with differential

• Aspirate, FCM, Genetics

• Drug history

• Previous diagnosis of (hemtological) disease

• (Previous) Therapies

• Serology

• ………

• i.e. complete

N.S.

MPN

• CML, BCR-ABL1 +

• CNL

• ET

• PV

• PMF

• CEL, NOS

• SM

• MPN,u

• MPN/MDS

• aCML

• CMML

• JMML

• RARS-T

• MDS

Kralovics et al (2005) NEJM, 352: 1779-90

JAK2 Mutation in MPN

MPN mutations overview

MPN “phenotypic driver” mutations

mainly

initiation

MPN progression

“non-phenotypic driver” mutations

“important passenger mutations”

PV

ET

PMF

sAML

AML

MDS

ALL

CML

RARS-T

MPL

JAK2

exon 12 exon 16V617F TET2 DNMT3a CBL

IDH1

IDH2 EZH2CALR ASXL1

Courtesy of Prof. R. Skoda

ET: Diagnostic Criteria (WHO 2008)

1. Sustained platelet count ≥450x109/L

2. Bone marrow biopsy specimen showing proliferation mainly of the megakaryocytic lineage with increased numbers of enlarged, mature megakaryocytes. No significant increase or left-shift of neutrophil granulopoiesis or erythropoiesis

3. Not meeting WHO criteria for PV, PMF, BCR-ALB1 positive CML or MDS or other myeloid neoplasm

4. Demonstration of JAK2 V617F or other clonal marker, or in the absence of JAK2 V617F, no evidence for reactive thrombocytosis

ET: all 4 criteria

ET: Morphology

H&E

PV: Diagnostic Criteria (WHO 2008)

Major criteria

1. Hb >18.5 g/dl in men, >16.5 g/dl in women or > RCV

2. JAK2 V617F or similar mutation

Minor criteria

1. BM biopsy: panmyelosis

2. EPO below reference range

3. Endogenous EC formation in vitro

PV: 2+1 or 1.+2

PV: Morphology

Post-PV/ET myelofibrosis (spent phase)

~ 30 %

Blast phase

~ 10 - 15 %

Manifestation

(t)

10 - 15 yrs.

definite increase in red cell mass

Evolution

Initial stage Polycythemic stage Terminal stage

Transformation

Dynamics of the disease process in PV (ET)

~10%-15% mimicking “ET”

JAK2 +/- JAK2 +/+ EPO

Thiele & Kvasnicka, 2008

post-PV/ET MF: Diagnostic Criteria

Required criteria:

- Previous dx of WHO-defined PV/ET

- BM-fibrosis grade 2-3

Additional criteria (2 required):

- Anemia

- Leukoerythroblastic PB picture

- Increasing splenomegaly

- Increased LDH

- Development of constitutional symptoms

(WHO 2008)

PMF: Diagnostic Criteria (WHO 2008)

Major criteria

1. Presence of megakaryocyte proliferation/atypia, accompanied by reticulin and/or collagen fibrosis, or

in the absence of fibrosis, the megakoryocyte changes accompanied by an increased bm cellularity characterized by granulocytic proliferation (i.e. prefibrotic cellular-phase disease).

2. Not meeting criteria for PV, CML, MDS, or other clonal marker

3. JAK2 V617F or other clonal marker (e.g. MPL W515K/L),or

in the absence of a clonal marker, no evidence that the bm fibrosis or other changes are secondary

PMF: Diagnostic Criteria (WHO 2008)

Minor criteria

1. Leukoerythroblastosis

2. Increase in serum LDH

3. Aneamia

4. Splenomegaly

PMF: all 3 major plus 2 minor criteria

Key features of prePMF according to WHO

increased cellularity (age-matched) with predominant neutrophil

granulopoiesis

increased megakaryopoiesis, small to large

atypical histotopography of megakaryocytes

endosteal translocation

formation of dense clusters*

distinctive nuclear features of megakaryocytes

hypolobulation (bulbous/cloudlike)

maturation defects

* WHO definition of a megakaryocyte cluster: 3 or more megakaryocytes lying strictly adjacent

- without other hematopoietic cells lying in between

Courtesy of Prof. Kvasnicka

PMF: Morphology

PMF: Morphology

Göm

PMF: Morphology

Disease evolution in Primary Myelofibrosis

Courtesy of Prof. Kvasnicka

Myelofibrosis - Definition

• Increase of reticulin fibers (+/- collagenous fibers) in

the bone marrow

• +/- Osteosclerosis

• Focal/diffuse

• Grade 0-3

• Not a specific diagnosis/entity

• Symptom for underlying disease

• “Clinical thermometer of the bone marrow”

• Biopsy: “sine qua non”

A biopsy is essential whenever there is myelofibrosis,

and the classification of some entities, particularly MPN,

relies heavily on trephine sections.

J.W. Vardiman, WHO 2008

• MPN: BM-Biopsy essential

• Major and/or minor diagnostic criteria (WHO 2008)

MF-0

MF-2

MF-1

MF-3

Grading of BM (Reticulin-)fibrosis

Grading of Collagen and Osteosclerosis

Coll-0

Coll-1

Coll-2

Coll-3

Os-0

Os-1

Os-2

Os-3

According to upcoming 2015 consensus criteria

Kvasnicka et al, Histopathology 2015

Summary

• BM investigation: interdisciplinary diagnostic approach

• MPN: histology essential diagnostic step

• Myelofibrosis:

• symptom – not a specific entity

• only by histology, grade 0-3

• Gentics: JAK2, CALR, MPL

Well, ...it looks like

you broke the

Jamishidi-needle

...

• Causes of discrepancies

• Incorrect interpretation

• Sampling disparity

• Different perspective of the

same disease/situation

• Different methods reveal

diverse aspects

• What to do?

• Mutual information

• Review of the case

• Discussion and consensus

• Consider the worse result

• Comprehensive result to the

treating physician

How to deal with contradictive findings?

Dirnhofer S, Went Ph and Tichelli A. 2007

?

Myelofibrosis - causes

• MPN

• MDSf

• MPN/MDS

• AML (APMF, M7)

• Lymphoma-Infiltration (HCL, FL, HL)

• Metastasis

• Autoimmune-Myelofibrosis

• Drugs, RT

• Chronic inflammatory disorders

• Metabolic disorders (Vit-D deficiency, renal, …)

• Idiopathic