Hematopathology LECTURE OUTLINE

Hematopathology LECTURE OUTLINE This document contains the text shown on the lecture slides. The purpose is the aid students taking notes during lect...
Author: Milo Logan
7 downloads 2 Views 958KB Size
Hematopathology LECTURE OUTLINE

This document contains the text shown on the lecture slides. The purpose is the aid students taking notes during lectures. The following is not what you have to learn for the exam, many things are mentioned only to present complexity or to spark interest.

2014/15

Hematopathology SLIDES 1. 2. 3. 4. 5. 6.

PREPARATIONS 1. 2. 3. 4.

Toxoplasma lymphadenitis Follicular lymphoma CLL/SLL in lymph node Diffuse large B-cell lymphoma Hodgkin’s lymphoma CML, CP, smear

Burkitt’s lymphoma Lymphomatous polyposis Multiple myeloma CML - splenomegalia

EXAM TOPICS 1. 2. 3. 4. 5. 6. 7. 8.

Reactive lymph node changes Indolent B cell lymphomas (FL, CLL, MCL, MZL) High grade B cell lymphomas (BL, DLBCL). Plasma cell neoplasms Hodgkin’s lymphoma T/NK cell lymphomas. Non-neoplastic bone marrow diseases AML and MDS Myeloproliferative neoplasms

Hematopathology

Hematopoiesis Lymphoid cell populations Reactive lymph node alterations

Hematopoiesis

Normal ranges in blood Neutrophil 1.8 – 5.4 ×109 Eosinophil 0.0 – 0.5 × 109

Production

Basophil

0.0 – 0.08 × 109

Leukocyte

60 ×109 /day

Monocyte

0.3 – 0.8 × 109

Rbc

210×109 /day

Lymphocyte

1.3 – 3.6 × 109 Thrombocyte

140×106 /day

Red blood cell Thrombocyte

×1012

4.5 – 6 140 – 440 ×109

Hematopoiesis thrombocyte

>16 weeks

erythrocyte

LT-HSC 6-8 weeks

mast cell

ST-HSC MEP Stem cell

BMCP

CMP

eosinophil

MPP self renewal supporting 2-6 weeks hematopoiesis active LT-HSC quiescent LT-HSC

basophil

GMP

neutrophil

CLP

monocyte/ macrophage

BCP „myeloid” LT-HSC „lymphoid” LT-HSC

TNKP B-cell T-cell

NK-cell

dendritic cell

Hematopoiesis thrombocyte

MYELOID

erythrocyte

LT-HSC

mast cell

ST-HSC MEP

basophil

BMCP

CMP

eosinophil

MMP GMP

neutrophil

CLP

monocyte/ macrophage

BCP TNKP B-cell

LYMPHOID T-cell

NK-cell

dendritic cell

HISTIOCYTIC

B-lymphoid cell populations CLP

5’

V genes

D genes

C genes

J genes

3’

µ δ γ DHJH

3’

5’

pro-B (BCP)

µ δ γ CDR3

VHDHJH V

pre-B

D N1

TdT VHDHJH VLJL

(terminal dezoxyribonucleotidil transferase)

immature B-cell

5’

IgM+ mature IgD+

Bone marrow

(naive) B-cell

µ δ γ

IgM

VDJ

J N2

C genes

3’

µ δ γ δ γ

IgD

B-lymphoid cell populations CLP

SOMATIC HYPERMUTATION

DHJH V

pro-B (BCP)

D

J

VHDHJH

Centrocyte

pre-B VHDHJH VLJL

FDC

SHM

immature B-cell

T-cell

IgM+ mature IgD+

Bone marrow

(naive) B-cell

activated B-cell

Centroblast

B-lymphoid cell populations CLP

ISOTYPE SWITCH (CLASS SWITCH RECOMBINATION) VDJ

DHJH

5’

µ δ γ εγ α

IgM

pro-B (BCP)

C gének

5’

VHDHJH

CSR Centro-

pre-B VHDHJH VLJL

cyte FDC

SHM

immature B-cell

T-cell

IgM+ mature IgD+

Bone marrow

3’

εγ α

IgE

3’

(naive) B-cell

activated B-cell

Centroblast

B-lymphoid cell populations plasma cell CLP DHJH

pro-B (BCP)

B1 B-cell?

plasma cell memory B-cell marginal zone B-cell?

VHDHJH

pre-B VHDHJH VLJL

plasmablast

extrafollicular B-cell

Centrocyte CSR

FDC

SHM

immature B-cell

T-cell

IgM+ mature IgD+

Bone marrow

(naive) B-cell

activated B-cell

Centroblast marginal zone

T-lymphoid cell populations 90% DβJβ

pro-T

γ/δ T-cell TCRβ V

VβD β J β pre-T VβD β J β VαJ α

CD4+/CD8T-cell

TCR αβ TCR γδ IgHL

NKT-cell (CD4-, CD8+/-)

J

TCRα V

CD4-/CD8+ T-cell

medulla

D

TCRγ

CD4+/CD8+ T-cell

Thymus-cortex

TCRδ

J

diversity 1016 1018 1011

T-lymphoid cell populations 90% DβJβ

pro-T

γ/δ T-cell

VβD β J β pre-T VβD β J β VαJ α

TH1-cell TH2-cell

CD4+/CD8T-cell

CD4+/CD8+ T-cell

TH17-cell CD4-/CD8+ T-cell

Treg-cell

CD8+ memory T-cell

TFH-cell cytotoxic T-cell

Thymus-cortex

medulla

NKT-cell (CD4-, CD8+/-)

CD4+ memory T-cell

Lymphocyte differentiation and maturation TCR/BCR rearrangement IgH/IgL TCRαβ TCRγδ

no costimulation: apoptosis anergy

costimulation: immune response

TCR/BCR signal

activation

Positive selection eliminating useless cells

Negative selection eliminating autoreactive cells

apoptosis

other survival signal

Lymphoid cell populations – B cells B1 B-cell in mice: lymphocyte in body cavities, innate immune cells debatable in humans, present in blood? spontaneous IgM production, T-cell independent activation

Marginal zone B-cell T-cell independent activation low affinity, IgM antibodies, isotype switching possible heterogenous population in humans: B1? + memory B-cells? + MZ B-cells?

Follicular B-cell T-cell dependent activation high affinity antibodies, isotype switching (IgG)

Lymphoid cell populations – T- and NK-cells CD4+ T-cell modulating limmune/inflammatory reactions „helper” and „regulatory” cells

CD8+ T-cell cytotoxic T-cells granzyme B, perforin, etc. enzymes inducing membrane damage

NKT-cell similar to NK-cells, rare T-cell population, usually suppressing function

γδ T-cell has a role in mucosal immunity, analogue of B1 B-cells

NK-cell innate immunity, not a T-cell cell lysis/induction of apoptosis without antibody recognition

Lymphatic tissue abnormalities General definitions lymphadenopathy

lymph node enlargement

lymphadenomegaly

lymph node enlargement

lymphadenitis

lymph node inflammation

Clinical characteristics localized vs. generalized painful vs. painless soft vs. firm/hard rapid growth vs. persitant, stable

General differential diagnosis reactive changes/inflammation hematological malignancy metastasis

Reactive lymph node abnormalities Non-specific lymph node changes follicular hyperplasia paracortical hyperplasia sinus histiocytosis

Specific lymphadenitis Toxoplasma lymphadenitis cat scratch disease

Infectious mononucleosis

Non-specific lymph node abnormalities Follicular hyperplasia germinal centrum reaction B-cell expansion due to humoral immune response Examples: characteristic in children lymph node/tonsil enlargement may be due to autoimmune disease may be due to HIV+

>1 cm usually is abnormal soft, painful– inflammation firm, painless – chr. inflammation or neoplasm

Non-specific lymph node abnormalities Paracortical hyperplasia hyperplasia of T-cell compartment

Examples: lymph node near tumors dermatopathic lymph node (eg. dermatitis) viral infection (herpes, mononucleosis, etc.)

Non-specific lymph node abnormalities Sinus histiocytosis expansion of sinuses due to macrophages

Examples: anthracosis Rosai-Dorfmann disease (sinus histiocytosis with hemophagocytosis)

Specific lymph node abnormalities Toxoplasma lymphadenitis Toxoplasma gondii – protozoon, human is not a definite host toxoplasma lymphadenitis diffuse toxoplasmosis with immunosuppression – encephalitis, myocarditis, etc. intrauterine toxoplasma infection – CNS, eye, liver changes, growth retardation

Piringer-Kuchinka lymphadenitis follicular hyperplasia epithelioid cell microgranulomas monocytoid B-cell hyperplasia

Specific lymph node abnormalities Cat scratch disease Bartonella henselae (G-negative) asbcedating reticulohistiocytic lymphadenitis suppurating granulomas

Others Kikuchi-Fujimoto lymphadenitis (necrotizing reticulohistiocytic lymphadenitis) granulomatous lymphadenitis – Myocbacterium, Yersinia, Pastorella, Chlamydia, etc. drug induced lymphadenopathy sarcoidosis Castleman disease others

Infectious mononucleosis Microbiology Epstein-Barr virus gamma-herpesvirus, DNA virus

Pathogenesis lytic infection of epithelium latent infection of B-cells T-cell immune response - atypical mononuclear cells humaral immune response

Clinicopathology general signs of inflammation lymph node and spleen enlargement leukocytosis atypical mononuclear cells

B-lymphoid populations plasma cell CLP

EBV

DHJH

pro-B (BCP)

VHDHJH

pre-B VHDHJH VLJL

plasma cell

lytic infection

latency I (survival) memory B-cell

plasmablast

extrafollicular B-cell

Centrocyte CSR

FDC

SHM

immature B-cell

activated B-cell IgM+ mature latency III IgD+ (naive) (proliferation) B-cell Bone marrow

T-sejt latency II Centro(differentiation) blast marginal zone

Infectious mononucleosis

EBER: EBV encoded small RNAs intranuclear RNA

Lytic infection: EBNA1 Latency III: EBER, EBNA1, EBNA2, LMP1 Latency II: EBER, EBNA1, LMP1 Latency I: EBER, EBNA1, LMP2

EBNA1: EB nuclear antigen1 EBV genome replication LMP1: Latent membrane protein 1 EBV associated diseases: infectious mononucleosis Burkitt lymphoma Hodgkin lymphoma immunosuppression related DLBCL extranodal NK/T-cell lymphoma primary effusional lymphoma nasopharyngeal carcinoma lymphoepithelial carcinoma

Hematopathology

foundations of the WHO classification morphology and immunophenotype genetic alterations clinical characteristics

Hematopoietic and lymphoid neoplasms lymphoid neoplasms myeloid neoplasms histiocytic and dendritic cell neoplasms

Lymphoid neoplasms – basic concepts

Hodgkin lymphoma vs non-Hodgkin lymphoma leukemia vs lymphoma nodal vs extranodal lymphoma aggressive vs indolent lymphoma epidemiology clinical signs staging

Lymphoid neoplasms – basic concepts Hodgkin lymphoma

non-Hodgkin lymphoma

described by Thomas Hodgkin

every other lymphoma

mature B-cell lymphoma characteristic morphology characteristic clinical picture excellent prognosis

Lymphoid neoplasms – basic concepts Leukamia

Lymphoma

neoplastic cells in blood

neoplasm of lymphatic tissue

manifestation, not disease

lymph node enlargement

many lymphomas may be leukemic

may be extranodal

leukemias may be aleukemic

may appear as leukemia

Lymphoid neoplasms – basic concepts Nodal lymphoma

Extranodal lymphoma

lymphatic tissue involvement:

often with disseminated lymphoma

lymph node, spleen, thymus,

primary extranodal lymphoma:

(Waldeyer’s ring )

GI tract, skin, CNS, head and neck, testis, etc.

Lymphoid neoplasms – basic concepts Aggressive lymphoma

Indolent lymphoma

rapid proliferation/progression

slow progression, little proliferation

higher chance of being treatable

usually uncurable

„blastic”:

„cytic”

lymphoblast, centroblast, immunoblast pediatric lymphomas

older age

due to immunosuppresion:

chronic lymphcytic activation

EBV, HHV8

alterations of the memory B cells H.pylori, B.burgdorferi, C.psitacci HCV

Lymphoid neoplasms – basic concepts Epidemiology

Prognosis

4-5% of all adult cancer (7-10th)

very heterogeneous

incidence: 19.7/100,000 (USA, 2013.)

summary: 5 year survival - approx. 69%

most frequent cancer in children (ALL) 1.5× more frequent in males approx. 85% B-cell NHL approx. 5% T-cell NHL approx. 10% HL

Lymphoid neoplasms – basic concepts Clinical picture Lymph node enlargement painless, firm Rapid tumor growth – extranodal lymphoma Splenomegaly Hepatomegaly Bone marrow failure anemia, thrombocytopenia, neutropenia Systemic signs autoimmune hemolytic anemia, infections B signs weight loss, fever, nigh sweats

Lymphoid neoplasms – basic concepts Staging Ann-Arbor staging I

one lymph node region

II

more lymph node regions on one side of diaphragm

III

lymph nodes on both sides of diaphragm

IV

bone marrow, liver, lung, pleura, etc.

E:

extranodal involvement, or spread from lymph node

B:

B signs

X:

>10 cm „bulky” involvement

Lymphoid neoplasms – morphology CD (cluster of differentiation) markers (cell surface epitopes) B-cell markers

T-cell markers

CD19: general B-cell

CD2, CD3, CD5, CD7: general T-cell

CD20: mature B-cell

CD1a ,CD34, TdT: precursor T-cell

CD21: C3d receptor (EBV), mature B-cell

CD4, CD8: T-cell subpopulations

CD23: FCεRII (IgE receptor), mature B-cell

CD56: NK-cell

CD34, TdT, CD10: precursor cells

granzyme B: cytotoxic T-cell

CD10, Bcl6: germinal center B-cell CD138, Mum1: plasma cell CD5: small fraction of mature B-cells

Other markers CD34: precursor cells CD45: general leukocyte marker

Lymphoid neoplasms – genetics IgH/TCRγ gene rearrangement – clonality test monoclonal

single PCR product is dominant

polyclonal

many PCR products in the sample

oligoclonal

few PCR products in the sample

Alternative testing of clonality detecting light chain restriction in case of B cells

Lymphoid neoplasms – genetics Translocations

IgH/IgL translocations

Gene rearrangement (VDJ/VJ) V

D

IgH/Partner

IgH region

C

J

5’

3’

µ δ γ

E

E

3’

5’

µ δ γ CDR3 D

V TdT

N1

J N2

Class switch recombination Centrocyte FDC T-cell Centroblast

Somatic hypermutation (SHM)

Partner gene region

Examples t(8;14) IgH/C-Myc

BL

t(11;14) IgH/CyclinD1

MCL

t(14;18) IgH/Bcl2

FL

Point mutations eg. Bcl6, Myc, Bcl2, p53 Translocations eg. C-Myc, Bcl6

Lymphoid neoplasms – WHO classification Precursor B-cell lymphomas B-lymphoblastic leukemia/lymphoma, with recurrent genetic aberrations BCR-ABL1 translocation MLL translocation ETV6-RUNX1 translocation Hyperdiploidity Hypodiploidity IL3-IGH translocation E2A-PBX1 translocation B-lymphoblastic leukemia/lymphoma, NOS Mature B-cell lymphomas Chronic lymphocytic leukemia/Small lymphocytic lymphoma B-cell prolymphocytic leukemia Splenic marginal zone lyphoma Hairy cell leukemia Splenic B-cell lymphoma/leukemia, NOS Splenic diffuse red pulp small B-cell lymphoma Hairy cell leukemia variant Lymphoplasmacytic lymphoma Heavy chain diseases Gamma heavy chain disease Mu heavy cahin disease Alpha heavy chain disease Plasma cell neoplasms Monoclonal gammopathy of unknown significance Plasma cell myeloma Solitary bone plasmacytoma Extraosseal plasmacytoma Extranodal marginal zone lymphoma in MALT (MALT lymphoma) Nodal marginal zone lymphoma Follicular lymphoma Primary cutaneous follicular lymphoma Mantle cell lymphoma

Precursor T-and NK-cell lymphomas T-lymphoblastic leukemia/lymphoma Hodgkin lymphomas Nodular lymphocyta predominant Hodgkin lymphoma Classic Hodgkin lymphoma Nodular sclerosis Lymphocyte rich Mixed cellularity Lymphocyte depletion

Diffuse large B-cell lymphoma, (DLBCL) NOS T-cell/histiocyte rich large B-cell lymphoma Primary CNS DLBCL Primary cutaneous DLBCL, leg type EBV-positive DLBCL of the elderly Chronic inflammation associated DLBCL Lymphomatoid granulomatosis Primary mediastinal (thymicus) large B-cell lymphoma Intravascular large B-cell lymphoma ALK-positive large B-cell lymphoma Plasmablastic lymphoma Large B-cell lymphoma arising in HHV8-associated multicentric Castleman disease Primary effusional lymphoma Burkitt lymphoma B-cell lymphoma, unclassifiable, intermediate between DLBCL Burkitt lymphoma B-cell lymphoma, unclassifiable, intermediate between DLBCL and Hodgkin lymphoma

Mature T- and NK-cell lymphomas T-cell prolymphocytic leukemia T-cell large granular cell leukemia Chronic NK-cell lymphoproliferative disorders Aggressive NK-cell leukemia Pediatric EBV-positive T-cell lymphoproliferative disease Pediatric systemic EBV-positive T-cell lymphoproliferative disease Hydroa vacciniforme-like lymphoma Adult T-cell leukemia/lymphoma Extranodal NK/T-cell lymphoma, nasal type Enteropathy associated T-cell lymphoma Hepatosplenic T-cell lymphoma Subcutaneous panniculitis-like T-cell lymphoma Mycosis fungoides Sézary syndrome Primary cutaneous CD30+ T-cell lymphoproliferative diseases Primary cutaneous anaplastic large-cell lymphoma Lymphomatoid papulosis Primary cutaneous peripheral T-cell lymphoma, rare variants Primary cutaneous gamma-delta T-cell lymphoma Primary cutaneous CD8+ aggressive epidermotropic cytotoxic T-cell lymphoma Primary cutaneous CD4+ small/medium sized T-cell lymphoma Peripheral T-cell lymphoma, NOS Angioimmunoblastic T-cell lymphoma Anaplastic large-cell lymphoma, ALK positive Anaplastic large-cell lymphoma, ALK negative

B-cell lymphomas (that will be discussed) Precursor B-cell lymphomas Acute lymphoblastic leukemia/lymphoma (ALL/LBL)

Aggressive, mature B-cell lymphomas Burkitt lymphoma (BL) Diffuse large B-cell lymphoma (DLBCL)

Indolent, mature B-cell lymphomas Chronic lymphocytic leukemia/Small lymphocytic lymphoma (CLL/SLL) Follicular lymphoma (FL) Marginal zone lymphomas (MZL)

Other, mature B-cell lymphomas Mantle cell lymphoma (MCL) Plasma cell neoplasms (PCN)

B-cell lymphomas ALL

plasma cell CLP PCN

pro-B (BCP)

memory B-cell extrafollicular B-cell

VHDHJH

pre-B VHDHJH VLJL

DLBCL

plasma cell

DHJH

plasmablast

CLL/SLL MZL

Centrocyte CSR

DLBCL

FDC

SHM

immature B-cell

T-cell MCL IgM+ mature IgD+

Bone marrow

(naive) B-cell

activated B-cell

BL CentroDLBCL blast marginal zone

FL

Precursor B-cell lymphoma/leukemia Acute lymphoblastic leukemia (lymphoblastic lymphoma) Epidemiology 80-85% of precursor cell leukemias, 10% of such lymphomas are B-cell 75% of patients are under 6 years

Symptoms anemia, thrombocytopenia usually lymph node, spleen and liver enlargement bone pains

Morphology lymphoblastic infiltration in bone marrow usually leukocytosis in peripheral blood CD34+, CD19+, CD10+

Genetics hyperdiploidity – favorable prognosis translocation producing gene fusion eg. t(9;22) – BCR-ABL1 – unfavorable eg. t(12;21) – ETV6-RUNX1 – favorable

Mature B-cell lymphoma - aggressive Burkitt lymphoma (BL) History 1958: Denis Parsons Burkitt: recognized BL in Uganda tried chemotherapy with success 1964: Michael Anthony Epstein/Yvonne Barr: recognition of virus 1967: Werner és Getrude Henle: EBV in mononucleosis

Epidemiology third of pediatric NHL more frequent in males endemic: Africa (near Equator): malaria and EBV sporadic: anywhere, 30% EBV+ with immunsuppression: HIV+, 25-40% EBV+

Symptoms endemic: tumor of head and neck area sporadic: abdominal and retroperitoneal rarely leukemic manifestation

Mature B-cell lymphoma - aggressive Burkitt lymphoma (BL) Morphology diffuse, medium sized blastic infiltrate high mitotic and apoptotic activity („starry sky pattern”) CD20+, CD10+, Mib1 > 95%

Genetics MYC-translocation: t(8;14) – IGH/C-MYC t(2;8) – IGL/C-MYC t(8;22) – IGK/C-MYC

Mature B-cell lymphoma - aggressive Diffuse large B-cell lymphoma (DLBCL) Epidemiology 20-30% of adult NHL (most frequent) 60-70 year is average primary (de novo) secondary (transformation): CLL, FL, MZL, NLPHL immunsuppression increases risk, mostly EBV+

Symptoms rapidly growing tumor, approx. 60% extranodal 5 year survival approx. 60%

Classification DLBCL, NOS – GC-type, non-GC-type T-cell/histiocyte rich large B-cell lymphoma (TCRBCL), Primary CNS DLBCL, Primary cutaneous DLBCL, leg type, Elderly EBV+ DLBCL Other large cell lymphomas: Primary mediastinal large B-cell lymphoma (PMBCL), Lymphomatoid granulomatosis, Intravascular large B-cell lymphoma, Primary effusional lymphoma, etc.

Mature B-cell lymphoma - aggressive Diffuse large B-cell lymphoma (DLBCL) Morphology diffuse, „blastic” infiltrate centroblastic/immunoblastic/anaplastic CD20+, everything else is variable

Genetics Bcl6 translocation (30-40%) Bcl6 mutation (70%) IGH/BCL2 (15%) IGH/C-MYC (10%)

Mature B-cell lymphoma - indolent Chronic lymphocytic leukemia (CLL) Epidemiology most frequent adult leukemia most frequent lymphoma in Hungary incidence: 3-5/100,000, 90% > 50 years rarely familial

Symptoms often symptomless lymphocytosis splenic and lymph node enlargement B-signs anemia, thrombocytopenia autoimmune manifestations: AIHA, ITP different staging system, since it is often disseminated

Mature B-cell lymphoma - indolent Chronic lymphocytic leukemia (CLL) Morphology Blood: monoclonal B-cell lymphocytosis in blood CD5+, CD19+, CD20+, CD23+

Bone marrow: nodular or diffuse infiltrate

Lymph node: diffuse, small lymphocytic infiltrate, sometimes pseudofollicles if restricted to lymph node: small lymphocytic lymphoma (SLL)

Mature B-cell lymphoma - indolent Chronic lymphocytic leukemia (CLL) Pathogenesis Mutated IgH gene

Non-mutated IgH gen

(„postfollicular”)

(„prefollicular”)

approx. 40%

approx. 60%

rarely IgG/IgA

characteristic VDJ gene usage

poly/autoreactive BCR

poly/autoreactive BCR

slow progression

faster progression

usually no treatment is needed

usually requires treatment

long survival similar to an autoimmune disease

Mature B-cell lymphoma - indolent Chronic lymphocytic leukemia (CLL) Pathogenesis - theory Mutated IgH gene

Non-mutated IgH gen

(„postfollicular”)

(„prefollicular”)

polyreactive, anergic IgM+ memory cells

IgM+ memory cells

(marginal zone B-cells)

(marginal zone B-cells)

clonal expansion due to restriction

ongoing aberrant SHM

of the memory B cell compartment

(AID expression)

Mature B-cell lymphoma - indolent Chronic lymphocytic leukemia (CLL) Treatment only observation cytostatic drugs chemotherapy anti-CD20 (Mabthera), anti-CD52 (MabCampath) treatment

Progression advanced stage (anemia, thrombocytopenia develops) prolymphocytic transformation Richter-transformation: DLBCL develops

Mature B-cell lymphoma - indolent Follicular lymphoma (FL) Epidemiology 2nd most frequent NHL in Western world (approx. 20-30% of lymphomas) rare in Asia mean age: 60 years

Symptoms slow, painless lymph node enlargement 50-60% bone marrow involvement may be extranodal

Morphology follicles without mantle zones no apoptotic activity centrocytoid/centroblastoid cells CD19+, CD20+, CD10+, Bcl6+, Mib1 low

Mature B-cell lymphoma - indolent Follicular lymphoma (FL) Genetics 50-90% IgH/BCL2 translocation occurs in bone marrow at DJ rearrangement, detectable in some memory cells in adults may develop due to SHM in germinal centers increased Bcl2 expression – apoptosis resistance clone „gets stuck” in germinal center accumulating genetic damage

Prognosis usually treatment is not needed approx. 30-50% transforms to DLBCL in 10 years

Mature B-cell lymphoma - indolent Marginal zone lymphoma (MZL)

Extranodal marginal zone lymphoma (MALToma)

Nodal marginal zone lymphoma

Splenic marginal zone lymphoma

Mature B-cell lymphoma - indolent Extranodal marginal zone lymphoma (MALToma)

H.pylori gastritis

Epidemiology 7-8% of lymphomas, more frequent in women, around 60 years stomach – 30% eye, salivary gland, skin, lung, etc. acquired MALT

Symptoms depends on localization often chronic inflammation: stomach – H.pylori, salivary gland – Sjögren thyroid gland – Hashimoto, conjunctiva – C. trachomatis may be large, bulky, often disseminated (extranodal!)

high grade transformation

clonal B-cells

MALT lymphoma

Mature B-cell lymphoma - indolent Extranodal marginal zone lymphoma (MALToma) Morphology centrocytoid cells with wide cytoplasm infiltrate originating in lymphoid follicles lymphoepithelial lesion

Genetics stomach: approx. 25% t(11;18)(q21;q21) – AIP2/MALT1 not stomach: apporx. 20-25% (14;18)(q32;q21) – IGH/MALT1

Mature B-cell lymphoma - indolent Splenic marginal zone lymphoma (SMZL) Epidemiology rare (3 g/l 55 years mild male dominance first curable lymphoma; now 80-90% is cured

Symptoms cervical, axillary, mediastinal lymph node enlargement rarely B signs

Mature B-cell lymphoma – Hodgkin lymphoma Classic Hodgkin lymphoma Morphology rare neoplastic cells – large, bizarre cells Reed-Sternberg cells: binucleated/multinucleated Hodgkin cells: mononuclear „mummy” cells: apoptotic, shrunken cells lacunar cells: clear halo many inflammatory/reactive cells T-cells, eosinophils, macrophages, plasma cells

Mature B-cell lymphoma – Hodgkin lymphoma Classic Hodgkin lymphoma Immunophenotype CD45-, CD20-/+ CD30+, CD15+, fascin+ Bcl6+, Mum1+ EBV+ (LMP1, EBER; latency type II) – 30-70%

Mature B-cell lymphoma – Hodgkin lymphoma Classic Hodgkin lymphoma Nodular sclerosis cHL 50-70% of cHL mean age: 15-35 years most frequently mediastinal EBV+/-

Mixed cellularity cHL 25-30% of cHL mean age: 40 years rarely mediastinal, often EBV+

Lymphocyte rich cHL rare, more frequent in males mean age: 30-50 years usually localized

Lymphocyte depletion cHL very rare, more frequent in males mean age: 30-40 years aggressive, disseminated

Mature B-cell lymphoma – Hodgkin lymphoma Nodular lymphocyte predominant Hodgkin lymphoma Epidemiology approx. 5% Hodgkin lymphomas mean age: pediatric and between 30-40 years tends to recur, but good prognosis

Symptoms cervical and axillary lymph node enlargement

Morphology L&H cells/popcorn cells

Phenotype CD45+, CD20+, CD30-, CD15-, EBVBcl6+, Mum1+

Genetics 50% 3q27 (Bcl6) translocation

Mature B-cell lymphoma – Hodgkin lymphoma L&H-cell

Pathogenesis HRS-cells are monoclonal SHM+ approx. 25% destroyed IgH mutation

PB CC

abnormal transcription autocrine programs function LMP1 apoptosis (EBV) resistance increased proliferation

FDC T-cell CB

EBV? preapoptotic CG-cell destroyed IgH gene lacking transcription factors

NFκB activation

HRS-cell cytokine production

T-cell lymphomas

Precursor T-cell lymphomas

Extranodal T-cell lymphomas

Acute lymphoblastic leukemia/ lymphoma (ALL/LBL)

NK/T-cell lymphoma, nasal type

Cutaneous T-cell lymphomas

Enteropathy associated T-cell lymphoma Hepatosplenic T-cell lymphoma

mycosis fungoides (MF) Sézary syndrome (SS)

Nodal T-cell lymphomas Peripheral T-cell lymphoma, NOS Anaplastic large-cell lymphoma Angioimmunoblastic T-cell lymphoma

Leukaemic T-cell lymphomas Adult T-cell leukemia Large granular lymphocyte (LGL) leukemia

Precursor T-cell lymphoma/leukemia Acute lymphoblastic leukemia (lymphoblastic lymphoma) Epidemiology 80-85% of precursor cell leukemias, 10% of such lymphomas are B-cell 75% of patients are under 6 years

Symptoms leukemia – like B cell lymphoma – usually mediastinal tumor

Morphology diffuse lymphoblastic infiltration CD3+, TdT+, CD34+

Genetics TCRα (14q11), TCRβ (7q35) translocation usually with homeobox genes

Cutaneous T-cell lymphomas Mycosis fungoides Definition epidermotropic cutaneous mature T-cell lymphoma

Epidemiology 50% of cutaneous lymphomas mean age: 55-60 years, may appear anytime 2× more frequent in males

Symptoms indolent, long course, localized to the skin for a long time rarely, late: lymph node, liver, spleen, bone marrow

Cutaneous T-cell lymphomas Mycosis fungoides Macroscopy

patch (premycotic) stage

Microscopy Pautrier microabscesses Sézary cells

plaque stage

tumor stage

Cutaneous T-cell lymphomas Mycosis fungoides Immunophenotype cutaneous CD4+ memory T-cell phenotype: TCRβ+, CD3+, CD2+, CD5+, CD4+ often CD7 loss rarely CD8+

Genotype monoclonal TCR gene rearrangmenet

Pathogenesis - theories chronic antigen-stimulation? – no evidence chronic exposition (eg. like contact dermatitis), bacterium? chr. inflammation? viral infection? – HTLV, CMV?

Cutaneous T-cell lymphomas Sézary syndrome Definition erythroderma, generalized lymph node enlargement neoplastic cells in peripheral blood (leukemia)

Epidemiology 5% of cutaneous lymphomas mean age: 60 years 2× more frequent in males

Morphology similar to mycosis fungoides

Nodal T-cell lymphomas Peripheral T-cell lymphoma, NOS (not otherwise specified) Definition heterogenous group

Epidemiology 30% of T-cell lymphomas adult, 2× frequent in males

Symptoms disseminated lymph node enlargement, often B signs often bone marrow and spleen involvement often dermal and GI tract involvement eosinophilia frequent 5 year survival: approx. 25%

Nodal T-cell lymphomas Peripheral T-cell lymphoma, NOS (not otherwise specified) Morphology heterogeneous usually diffuse infiltrate in lymph nodes medium sized, clear cells

Immunophenotype CD3+, CD2+, CD5+/-, CD7+/CD4 more frequently, than CD8 may be CD30+

Nodal T-cell lymphomas Anaplastic large-cell lymphoma Definition CD30+ T-cell lymphoma ALK+ systemic ALCL ALK- systemic ALCL cutaneous ALCL

Epidemiology ALK+ ALCL: 10-20% of lymphomas in children 3% in adults

Symptoms B-signs, disseminated lymph node enlargement frequent extranodal involvement

Nodal T-cell lymphomas Anaplastic large-cell lymphoma Morphology/immunophenotype anaplastic lymphoid cells CD30+ frequently CD3-, but T-cells! cytotoxic markers

Genetics t(2;5)(p23;q35): NPM-ALK1 variants

Prognosis ALK+ ALCL: 5 year survival: 75-90% ALK- ALCL: 5 year survival: 20-30%

Nodal T-cell lymphomas Angioimmunoblastic T-cell lymphoma Definition CD4+ follicular helper T-cell neoplasm (CD4+, PD1+)

Epidemiology 15-20% of T-cell lymphomas adulthood, male = female frequency

Symptoms disseminated lymph node enlargement, often B signs spleen, liver, bone marrow involvement, hypergammaglobulinemia skin rashes, pruritus, arthritis frequently EBV+ B-cell expansion

Extranodal T-cell lymphomas NK/T-cell lymphoma, nasal type Asia, Middle/South America nasal cavity, nasopharynx, oral cavity EBV+, NK, or NKT-cell phenotype angiocentric growth, aggressive (lethal midline granuloma)

Enteropathy associated T-cell lymphoma rare, intraepithelial T-cell neoplasm often associated with coeliac diseas; poor prognosis ulcerative jejunitis, intestinal perforation CD4-, CD8+/-, cytotoxic phenotype, TCRβ+

Hepatosplenic T-cell lymphoma γδ-T-cell neoplasm, cytotoxic phenotype mean age is 35 years, hepatosplenomegaly, bone marrow involvement poor prognosis: 20% of blood volume is lost exsanguination: bleeding out (hypovolemic shock)

Chronic blood loss compensation – increased iron utilization iron deficiency anemia tissue hypoxia – increased ventillation, reduced cardiac output weekness, fatigue, pallor

Anemia – reduced production Reduced proliferation aplastic anemia reduced proliferation/survival of stem cells and progenitor cells unknown cause in 50% of cases drugs, viral hepatitis, thymoma, irradiation, etc. reduced proliferation due to genetic damage of progenitors T-cell activation induced myelosuppression

anemia due to bone marrow infiltration (myelophthisis) metastasis, fibrosis, granulomatous inflammation, storage disease, etc. teardrop red blood cells + leukoerythroblastosis

anemia due to chronic disease chr. renal disease, RA, IBD, carcinomas chr. inflammation – hepcidin↑, erythropoietin↓, thrombopoietin↓

Anemia – reduced maturation Iron deficiency anemia (cytoplasmic maturation is inhibited) metabolism: 3.5/2.5 g iron, 15-20% in iron storage pools; 1-2 mg/day excretion 10% of adults are iron deficient (in developed countries) causes:

chronic bleeding – GI, female genital tract insufficient diet malabsorption syndromes

microcytic/ hypochromic anemia anisocytosis

Anemia – reduced maturation Megaloblastic anemia (nucelar maturation is inhibited) folate (B9): casues of def.: cobalamin (B12): causes of def.:

DNS synthesis, repair, methylation pregnency, obesity, malabsorption, inadequate diet DNS synthesis, fat metablosm, etc. intrinsic factor def. (autoimmune gastritis) – anaemia perniciosa malabsorption

megaloblasts in bone marrow macrocytosis, anisocytosis

Anemia – hemolytic Intrinsic abnormalities spherocytosis sickle cell anemia thalassemia

Extrinsic abnormalities autoimmune hemolytic anemia microangiopathy hemolytic anemia

Anemia – hemolytic Spherocytosis usually AD cytoskeleton-cell membrane interaction failure (spectrin, ankyrin, etc. mutation) spherocytes –sequestered in spleen splenomegaly, cholelithiasis aplastic crisis – parvovirus B19 infection

Anemia – hemolytic Structure of hemoglobin

Haemoglobin A (α2β2)

Haemoglobin A2 (α2δ2) 1-3% normally Haemoglobin F (α2γ2) 51/48%, vvt > 6.1 / 5.4 × 1012 /l polyglobulia: > 185/165 g/l hemoglobin concentration polycythemia: increased red blood cell mass

polycythemia vera neoplastic disease

secondary polycythemia

Secondary polycythemia Secondary polycythemia

Hypoxia heart/lung disease smoking sleep apnoe renal artery stenosis

Congenital Acquired EPO not↓

No hypoxia

EPO↓

Hgb-O2 affinity

P50 ↓

Hemoglobinopathy 2,3BPG-deficiency

drugs paraneoplastic (liver/kidney cc, hemangioblastoma) EPOR mutation

P50 normal

VHL mutation HIF2A mutation based on Tefferi A. et al. Nat Rev Clin Oncol 2009

Granulocytic disorders

Leukopenia

Leukocytosis

Neutropenia Basic concepts leukopenia: < 4000 / µl WBC neutropenia: < 1500 / µl absolute neutrophil count agranulocytosis: < 100 / µl absolute neutrophil count

Casues Reduced production

Increased destruction

suppression of granulopoesis

immunological damage

damage of precursors

splenomegaly

ineffective hematopoesis

peripheral consumption

congenital disease

Neutropenia Consequences bacterial/fungal infection frequent oral, necrotizing, gangrenous inflammation frequent airway infections

Morphology hypocellular or hypercellular bone marrow

Leukocytosis Basic concepts leukocytosis: > 10000 / µl EBC leukemoid reaction: severe leukocytosis, with immature precursors

Causes bacterial infection – granulocytosis viral infection – lymphocytosis allergic reactions – eosinophilia paraneoplastic: Hodgkin lymphoma, plasma cell myeloma

Thrombocytosis Basic concepts normal thrombocyte count: 150 - 400,000 / mm3

Causes inflammation (IL6)

IBD, RA, tuberculosis

paraneoplasia

carcinoma

iron deficiency

Thrombocytopenia Basic concepts normal thrombocyte count: 150 – 400.000 / mm3 bleeding: 10-20.000 / mm3

Casues: A. Reduced production: bone marrow insufficiency, infiltration B. Reduced survival: ITP, TTP, HUS, DIC C. Sequestration: hypersplenism

Hematopoiesis thrombocyte

MYELOID

erythrocyte

LT-HSC

mast cell

ST-HSC MEP

basophil

BMCP

CMP

eosinophil

MMP GMP

neutrophil

CLP

monocyte/ macrophage

BCP TNKP B-cell

LYMPHOID T-cell

NK-cell

dendritic cell

HISTIOCYTIC

Histiocyte neoplasms Langerhans cell neoplasms (histiocytosis X) Epidemiology 0.5-0.9/100,000 children, rarely adults high mortality before age of 2 (30-50%)

Clinical characteristics eosinophil granuloma: 5-15 years, solitary lesion: bone, lung, soft tissue Hand-Schüller-Christian disease: 2-10 years, multiple bone lesions, diabetes insipidus Letterer-Siwe disease: < 2 years, disseminated; bone and soft tissue involvement

Histiocyte neoplasms Langerhans cell neoplasms (histiocytosis X) Morphology Langerhans cell: dendritic cell in skin/mucosa morphologo: oval nucleaus, nuclear groove immunohistochemistry: CD1a, langerin, CD68 elektronmicroscopí: Birbeck-granulum frequent BRAF mutation

Histiocyte neoplasms Dendritic cell sarcomas Epidemiology very rare

Clinical characteristics lymph node involvement soft tissue/organ manifestation lung/liver metastasis may occur

Types interdigitating dendritic cell sarcoma follicular dendritic cell sarcoma others

Myeloid neoplasms myeloproliferative neoplasm increased 6-10/100,000 incidence proliferation 50-70 years most frequent (some exceptions)

myelodysplastic syndromes patients 80% are above 60 years 60-69 years:

7.1/100,000 incidence

over 80 years:

35.5/100,000 incidence

acute myeloid leukemia mean age: 60 years approx. 3/100,000 incidence

maturation defect

maturation defect and increased proliferation

Myeloproliferative neoplasms

Epidemiology 6-10/100,000 incidence 50-70 years most frequent (some exceptions)

Classification chronic myeloid leukemia polycythemia vera essential thrombocythemia primary myelofibrosis chronic neutrophilic leukemia mastocytosis MPN with eosinophilia

Chronic myeloid leukemia

1845 – first clinicopathological description of CML 1960 – description of Ph-chromosome (first cytogenetic change in malignancy) 1973 – description of Ph-translocation 1984 – BCR-ABL1 fusion protein altered adhesion in bone marrow increased proliferation increased survival more frequent commitment 1996 – first tyrosine kinase inhibitor (imatinib)

Chronic myeloid leukemia Epidemiology 1-2/100,000 incidence 50-60 years most frequent PTE Path: mean age: 52.9 years (7-84 years)

Clinical characteristics 20-45% symptomless loww of weight, fatigue, night sweats, abdominal dyscomfort leukocytosis (usually >100,000 /µl) thrombocytosis (usually >450,000 / µl) splenomegaly

Ph-negative MPNs Polycythemia vera

Essential thrombocythemia

Primary myelofibrosis

1-3

1-2

0.5-1.5

60 years

60 years

70-80 years

symptoms

hyperviscosity, thrombosis, spleen↑

thrombosis, bleeding, erythromelalgia

B-signs, anemia, liver/spleen↑

peripheral blood

erythrocytosis (Hgb>185, Htc>51%), mild neutrophilia, basophilia

thrombocytosis (>450), tct anisocytosis

thrombocytosis/CMLlike, leukoerythroblastosis, anisopoikilocytosis

bone marrow

trilinear proliferation mature, heterogenous megakaryocytes

normocellular hyperlobulated megakaryocytes

myeloid proliferation atypical megakaryocytes

cytogenetics

20%, not specific

< 5% , not specific

35-40%, not specific

mol. biology

JAK2

JAK2, MPL, other

JAK, MPL, other

transformation

2-3%

15 years

>15 years

3-10 years

incidence age

survival

Mastocytosis Epidemiology heterogenous diseases cutaneous mastocytosis: pediatric, usually < 6 months systemic mastocytosis: 50-80 years Classification cutaneous mastocytosis urticaria pigmentosa diffuse cutaneous mastocytosis solitary mastocytoma systemic mastocytosis bone marrow, skin, GI tract

MPNs are tyrosine kinase diseases

JAK2exon12 ≈4%

BCR-ABL1 100%

CML

JAK2V617F ≈95%

PRV

MPLW515 ≈5%

MPLW515 ≈10%

JAK2V617F ≈50%

JAK2V617F ≈50%

KITD816V ≈85%

PMF

Mastocytosis

ET

PDGFRα PDGFRβ FGFR1 abnorm. 100%

PDGFR/ FGFR1 neoplasms

Myeloproliferative neoplasms Summary • diseases of old age • increased proliferation – tyrosine kinases are often involved • complications due to high blood counts • risk of acute leukemia (blastic crisis) • risk of bone marrow fibrosis/insufficiency

Myelodysplastic syndromes

heterogenous diseases clonal HSC disease ineffective hematopoiesis

risk of acute leukemia diseases of old age

Myelodysplastic syndromes

Epidemiology 80% of patients are over 60 years 60-69 years:

7.1/100,000 incidence

over 80 years:

35.5/100,000 incidence

below 14 years: 0.1/100,000 incidence males are involved more frequently (2×) Etiology congenital bone marrow insufficiency, previous chemotherapy ionising radiation, chemical exposure (agriculture, chemical industry) Clinical characteristics neutropenia, anemia, thrombocytopenia

Myelodysplastic syndromes Morphology blood and bone marrow cytology/histology morphological signs of dysplasia dyserythropoesis dysgranulopoesis dysmegakaryopoesis blast ratio ring sideroblasts monocytes Genetics clonal cytogenetic/molecular genetic abnormalities Clinical picture exluding potential causes (eg. alcoholism, chronic liver disease, vitamin deficiency, etc.)

MDS – ineffective hematopoiesis

proaptotic bone marrow microenvironment inflammatory cytokines? (eg. TNFα, IFNγ) increased proliferation

T-cell mediated myelosuppression chronic, autoimmune-like activity?

restricted stem cell repertoir restricted differentiation potential myeloid as default pathway

Acute myeloid leukemia

Epidemiology 60 years mean age approx. 3/100,000 incidence

Etiology ionizing radiation chemotherapy: alkilating agents, topoisomerase inhibitors chemicals: pesticides, benzene, etc. genetic diseases: Down, Fanconi anemia, Bloom, Wiscott-Aldrich, etc.

Clinical picture complications of cytopenias: weakness, infections, bleeding extramedullary manifestation

Acute myeloid leukemia

Definition >20% blast in blood/bone marrow presence of characteristic genetic change + increased blasts „blast” may be: myeloblast promyelocyte monoblast promonocyte megakaryoblast erythroblast

Acute myeloid leukemia

Peripheral blood

Bone marrow

anemia, thrombocytopenia

hyper/hypocellular

may be aleukemic

may be fibrotic

Flow cytometry blast ratio determining lineage leukemia-associated immunophenotype (LAIP)

Molecular genetics RT-PCR based detection of translocations demonstrating mutations

Cytogenetics karyotyping and FISH

Acute myeloid leukemia Molecular pathogenesis type I mutation signal transduction pathway – increased proliferation eg.: NRAS/KRAS, FLT3, KIT, JAK2, PTPN11 type II mutation transcription factors – abnormal differentiation eg.: CBF, MLL, NPM1, CEPBA, RUNX1, RARA type III mutation? epigenetic regulation – abnormal differentiation eg.: TET2, IDH1/2, DNMT3A, ASXL2, EZH2

Pedersen-Bjergaard J et al. Leukemia. 2006;20(11): 1943–1949.

AML – WHO classification t(8;21)(q22;q22) – RUNX1-RUNX1T1

AML with recurrent genetic change

inv(16)(p13q22) – CBFB-MYH11 t(15;17)(q24;q21) – PML-RARA

AML with myelodysplasia associated changes

t(9;11)(p22;q23) – MLLT3-MLL

AML, therapy related

inv(3)(q21q26) – RPN1-EVI1

t(6;9)(p23;q34) – DEK-NUP214

t(1;22)(p13;q13) – RBM15-MKL1

AML, not otherwise specified

NPM1 mutation CEBPA mutation

Myeloid sarcoma

AML – t(15;17)

Characteristics 5-10% AML young adults DIC frequent favorable prognosis ATRA treatment induced differentiation myeloblasts around 10% variant translocations eg. t(11;17)(q23;q11) are not favorable hypergranular (classical) hypogranular (microgranular): bilobed, folded nucleus – monocyte-like strong CD33, CD177, however CD34-, HLADRPML-RARA fusion, abnormal PML localisation