PLASMA CELL

NAIVE -BLYMPHOCYTE

MCLt(11;14)CCND1

CLL

FLt(14;18)

bcl-2

MEMORY CELL

BLt(8;14)c-myc

MALTt(11;18)API2-MLT

Patogénesis Linfomas de células B

ALL/LBLLCLt(3q)bcl-6

CLL

Linfomas Indolentes y AgresivosCaracterísticas histopatológicas

• Células pequeñas

• Baja actividad proliferativa

• Crecimiento no-

destructivo ( “Homing” )

• Respuesta a influencias

reguladoras

• Células grandes

• Actividad proliferativa

alta

• Crecimiento destructivo

• Crecimiento Autonomo

Indolente (Bajo grado) Agresivos (Alto grado)

• Curso clinical indolente

• Supervivencia larga

• No curable con quimioterapia

• Ausencia de “plateau” en las

curvas de supervivencia

• Curso clinical agresivo

• Supervivencia corta sin

tratamiento

• Posible larga superviencia

(curación)

• “Plateau” en las curvas de

supervivencia

Linfomas Indolentes y AgresivosCaracterísticas Clinicas

Indolente (Bajo grado) Agresivos (Alto grado)

PLASMA CELL

NAIVE -BLYMPHOCYTE

MCLt(11;14)CCND1

CLL

FLt(14;18)

bcl-2

MEMORY CELL

MZLt(11;18)API2-MLT

Linfomas de células B pequeñas

CLL

Clasificación REAL/OMS

Histological distribution

2%

8%

6%

36%1%2%

7%

7%

7%

24%

LymphocyticLymphoplasmocytoidMALTFollicularMantle-cellDiffuse large-cellBurkittAnaplasicPeripheral T-cellOther

NHL Classification Project, 1997

N=1.403

Chronic Lymphocytic Leukemia/ Small Lymphocytic Lymphoma

Definition

Neoplasm of monomorphic small, roud B-lymphocytes in peripheral blood, bone marrow and lymph nodes, admixedwith prolymphocytes and paraimmunoblasts(pseudofollicles), usually expressing CD5 abd CD23.

SLL is the same disease but restricted to tissues withoutevidence of leukemic involvement

WHO 20001

CLL / SLL : Clinical Features

• Most frequent leukemia in Western

countries (15 cases /100,000 /year)

• Median age at diagnosis: 70 yrs

• 80% of patients are asymptomatic

at diagnosis

• Lymphadenopathy,splenomegaly,

hepatomegaly, extranodal

infiltrates (20-30%)

• Median survival 10 years

• No effective therapy

Age at diagnosis

n=553

0.3% 2%

10%

21%

33%

26%

8%

25

50

75

100

125

150

175

200

< 3031-40

41-5051-60

61-7071-80

>80

Years

% p

atie

nts

aliv

e

0,00,10,20,30,40,50,60,70,80,91,0

0 4 8 12 16 20 24

n=553

HCP, Barcelona

CD20 CD79a

CD 3 CD 5 CD 23

CLL/SLL: Perifollicular pattern

CD23 CD5

Morphological Spectrum in CLL Transformation

• Large Cell Lymphoma­ Variable number of large lymphoid cells­ Expansion of proliferative growth centers­ Proliferative activity

• Hodgkin’s Disease­ Isolated RS cells in a background of CLL

Variable expresion of B-cell markers, CD30 and CD15Generally EBV positive

­ Typical HD associated with CLL­ Typical HD anatomically separated from CLL

Áreas típicas de LLC, con centros de proliferación

Áreas con agregados de histiocitos epitelioides

CD15

CD30

VEB(LMP-1)

CLL Chromosomal Abnormalities and Prognosis

Aberration No. Patients (%)

Median SRV (months)

17p del 3-7 32

11q del 11-25 79

Trisomy 12 10-16 114

Normal 18-22 111

13q del 31-44 133

Döhner et al. NEJM 2000

p53

ATM

CLL: From Naive to an AntigenExperienced Cell

Antigen SelectionIg Somatic Mutations

Class Switch

Naïve

V D J cµ c?

Memory

Plasma Cell

T Cell-Independent Response

Mutated-CLL

UnMutated-CLL

Clinical and PathologicalCharacteristics of U-CLL and M-CLL

M>F

Intermediate/high

< 12 mths

Atypical

17p-, 11q-

CD38/CD69

Uniformly short

Yes

Usual (3-4 years)

Gender

Rai Stage

LDT

Morphology

Genetic Alterations

Activation Markers

Telomers

BCR Signaling

Treatment Requirement

(Time from diagnosis)

M=F

Low

>12 mths

Typical

13-

CD71/CD62L

Diverse lengths

No

Unusual (8-11 years)

Unmutated-CLL Mutated-CLL

ZAP-70 FITC->

IgVH

CD

3 P

E +

CD

56

PE-

>

LLC-48Un Mutated

100 101 102 103 104

74%

LLC-31Mutated

12%100 101 102 103 104

Zap70 Expression and Ig Gene Mutations in CLL

Crespo et al. N Engl J Med 2003

ZAP=0 ZAP=1 ZAP=2

Carreras et al J Pathol 2005

Bosch et al, 2003

0 2 4 6 8 10 12 14 16

Years

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

Pro

babi

lity

ZAP-70 < 20% (n=98)

ZAP-70 > 20% (n=80)

Binet A

Months

Pro

bab

ility

of

Pro

gre

ssio

n0 48 96 144 192 240

100

90

80

70

60

50

40

30

20

10

0

ZAP-70 < 20

ZAP-70 > 20

Binet A

ZAP-70 Expression Predicts Survival and Progression in CLL

Flow Cytometry

CLL/SLL: Summary

• MORPHOLOGY:– Small round cells, prolymphocytes, paraimmunoblasts– Diffuse pattern with pseudofollciles

• PHENOTYPE– B-Cells, CD5 +, CD23 +, CD10-, Cyclin D1 -

• GENETIC ALTERATIONS– Unmutated Ig genes/ Hypermutated Ig genes– Tri 12, Del 13q, Del 11q, Del 17p

• CLINICAL– Mean age 70 – Peripheral blood and bone marrow, lymphadenopathy,

splenomegaly, hepatomegaly– SLL: No peripheral blood involvement (rare)– Median survial 7-10 years

Mantle Cell Lymphoma

• Morphological Spectrum

• Clinical Manifestations

• Prognostic parameters

• Molecular pathogenesis

• Therapeutic strategies

t(11;14)(q13;q32) Translocation

Breakpoints in BCL-1 RegionChr 14

JH

Chr 11

MTCTEL

p94CEN

CCND1

MCL: Clinical Characteristics

• Male to Female 2-8:1

• Median age 54-68 (range 29-85)

• Stage IV > 60%

• B Symptoms 35%

• Generalyzed lymphadenopathy

• Bulky Disease 18% (5-25%)

• Extranodal involvement 30-50%

• Splenomegaly 30-60%

• Leukemic phase

Mantle Cell Lymphoma: Architectural Spectrum

Mantle Zone Nodular Diffuse

Mantle Cell Lymphoma: Cytological Variants

Typical Small cell

Mantle Cell Lymphoma: Cytological Variants

PleomorphicBlastic

Blastoid MCL

• Typical 7 %

• Blastic 20 %

• Pleomorphic 87 %

(Ott G et al Blood 1997)

Proliferation Tetraploidy

(Bosch et al Cancer 1998)

Marginal Zone Pattern in MCL

Cyclin D1 Cyclin D1

MCL PhenotypeCD20

Cyclin D1 CD5

CD3

Gastrointestinal Involvement in MCL

• Clinico-pathological Presentation

• Lymphomatous Polyposis

• Microscopical Infiltration

• Clinical manifestations in 10-25 % of patients

• Asymptomatic more common (80%)

Extranodal Involvement in MCL

• Gastrointestinal 9-30 %

• Waldeyer’s Ring 2-18 % • CNS 5-20 %

• Liver 5-20 %

• Lung & Pleura 2-17 %• Peripheral Blood 24-90 %

Peripheral Blood Involvement in MCL

• 20-60% of patients, Defining criteria for involvement?• Controversial prognostic significance

Rosenwald A et LLMPP, Cancer Cell 2003

0 2 4 6 8 10 12 14 16Years

0

0,25

0,5

0,75

1

ProliferationIndex >40

ProliferationIndex <40

p<0.0001

Scharader et EMCL, Br J Haematol 2005

Cyclin D1 Negative MCL Variant

Cyclin D1

Cyclin D3 Cyclin D2

Fu et al, Blood 2005; in press

Mantle Cell Lymphoma:Summary• MORPHOLOGY:

– Mantle zone pattern, nodular Diffuse– Classical (small irregular cells) and blastoid variants

• PHENOTYPE– B-Cells, CD5 +, CD23 -, CD10-, Cyclin D1 +

• GENETIC ALTERATIONS– t(11;14); bcl-1 rearrangements; Cyclin D1 overxpression– Gains 3q, Deletions 11q; 13q; 9p; 17p; Complex karyotypes

• CLINICAL– Mean age 60, Male predominance– Generalyzed lymphadenopathy, Extranodal involvement– Median survial 3-5 years

Pro

babi

lity

Years

0.0

0.2

0.4

0.6

0.8

1.0

0 2 4 6 8 10

N=175

Progression-free survival

Overall survival

Follicular lymphoma

?Median age: 60 years

? Advanced stage: 80%

?Nodal involvement; B.M.+

?CR rate: 10-80%

?OS: 6 - 10 years

?No plateau in OS curve

Linfoma Folicular vs Hiperplasia Folicular

• Irregularidad de los foliculos• Escasa zona del manto• Monotonía celular• Ausencia de macrofagos• Escasa proliferacion

• Folículos de tamaño similar• Polarización del foliculo• Heterogeneidad celular• Macrófagos • Proliferación marcada

Hiperplasia FolicularLinfoma Folicular

Grados en Linfoma FolicularGrados DefiniciónGrado 1 0-5 cb per hpfGrado 2 6-15 cb per hpfGrado 3 > 15 cb per hpf

3a Centrocitos presentes3b Centroblastos

Hans Blood 2003; 101:2363-7Martin AR Blood 1995; 85:2671

Grado 1

Grado 3a Grado 3b

Grado 2

Follicular Lymphoma Grade 3b

? FL with a pure population of centroblasts

?More commonly CD10 negative (50%)

? Frequent plasmacytoid differentiation (40%)

? Frequent 3q27 chromosomal breaks and uncommont(14;18)

? DLBCL component

? Possible related to DLBCL rather than to FL grade 1,2, or 3a

Ott et al Blood 2002; 99;3806Bosga-Bouwer et al; Blood 2003; 101: 1149

Patrón Arquitectural

Folicular > 75%

Folicular y difuso 25-75%

Focalmente folicular < 25%

CD10CD20

CD20 CD3

CD10bcl-2

Modulación de la expresión de CD10

Reordenamiento y Expresión de Bcl-2

MBR (60%)

>20 kB

mcr(20%)

ExonI

ExonII

ExonIII

5’ 3’icr

Bcl-2

Bcl-2 Protein Expression

Normal quiescent lymphocytesNegative in normal germinal centersNegative in reactive Monocytoid B-cells

PositiveFollicular lymphomasCLL, MCL, MZLDiffuse large B-cell Lymphomas

NegativeBurkitt’s lymphomaALCL ALK +

Bcl-2 staining Pitfalls

• Increased number of T-cells in reactive follicles

• Primary lymphoid follicles (CD10, bcl-6, IgD)

• Nodular mantle cell lymphoma (CD10, bcl-6, Cyclin D1)

• FL negative for bcl-2 expression

IgD

MostlyT cell genes?

Mostlymacrophage

genes?

Prognostic Model in Follicular Lymphoma

p= 9.8 x 10-15p= 9.8 x 10-15

“In situ” Follicular Lymphoma

CD10

Bcl-2

Follicular Lymphoma: Summary• MORPHOLOGY:

– Nodular, Diffuse– Grade 1, Grade 2, Grade 3a and 3b

• PHENOTYPE– B-Cells, CD5 -, CD23 -, CD10+, Cyclin D1 -, bcl-2+, bcl-6 +

• GENETIC ALTERATIONS– t(14;18); bcl-2 rearrangements– t(3q27); bcl-6 (FL 3b)

• CLINICAL– Mean age 59, Occasional cases in young adults and children– Generalyzed lymphadenopathy, BM: 40%– Asymptomatic– Median survial 8-10 years

Marginal Zone Lymphomas in the WHO Classification

• Marginal Zone B-cell Lymphoma of MALT

• Nodal Marginal Zone B-cell Lymphoma

• Splenic Marginal Zone B-cell Lymphoma

Extra Nodal MZL: Morphology

MZL: Phenotype

IgD

CD20 CD 5

CD 43

CD10CD10

MZL: Follicular Colonization

Bcl-2Bcl-2

MZL: Follicular Colonization

CD23 Bcl-6 CD10

MZL: Follicular Colonization

Extra Nodal MZL: Etiological Factors

Chronic Inflammatory ResponseStomach H pyloriOcular Chlamydia PsittaciSalivary Gland Sjogren’sThyroid Hashimoto’sSkin BorreliaOther ?

HCV?

Genetic and enviromental backgroundPolymorphismsThymic MZL Asians

Tumor Site, Etiology, and Tumor Progression in MALT Lymphomas

Normal Cell

Oligoclonal Expansion

MALT LymphomaHP dependent

MALT LymphomaHP independent

Transformation

Traslocaciones en linfomas MALT

t(1;14)Wild type

Ye et al Am J Pathol 2000Streubel et al Leukemia 2005

t(11;18)

• t(11;18) API2-MALT1

• t(1;14) bcl-10

• t(14;18) MALT1

• t(3;14) FOXP1

Bcl-10 Bcl-10Bcl-10

FOXP1

Nodal Marginal Zone Lymphomas

1986-87 Sheibani et al : Monocytoid B-cell LymphomaCousar et al: Parafollicular lymphomaNodal and extranodal localizationMonocytoid differentiation

• 41% patients had evidence of extranodal lymphoma

• Evidence of primary nodal MZL– Long follow-up without evidence of extranodal site

• Nodal MZL may be heterogeneous

(Am J Surg Pathol 1999; 23: 59 – 68)

Nodal MZL (MALT Type) :Morphology

Nodal Marginal Zone Lymphoma

Nodal Marginal Zone Lymphoma

Nodal Marginal Zone Lymphoma

Splenic Marginal Zone Lymphoma(Splenic lymphoma with villous lymhocytes)

• MORPHOLOGY:– Small round lymphocyteswith pale cytoplasms and ocasional

larger cells– Lymphoid proliferation replaces germinal cetners and merges

with marginal zone

• PHENOTYPE– B-Cells, IgD+, CD5 -, CD23 -, CD10-, Cyclin D1 -, bcl-2+, bcl-6 -

• GENETIC ALTERATIONS– Losses of 7q21-32

• CLINICAL– Adults (>50y) with spelnomegaly– Leukemic involvement with villous lymphocytes– Bone marrow involvement– Autoimmune thrombocytopenia or anemia– Indolent clincal course and long survival– Occasional cases transform to DLBCL

Linfoma de Zona Marginal Esplénica

CD20

IgD

kappa Ki-67

Linfoma de Zona Marginal Esplénica

Bcl-2

Linfoma Linfoplasmacitoide / Linfoplasmacítico

• MORFOLOGIA: Espectro de diferenciación plasmacelularPlasmacitoide: Leucemia ? , Paraproteína ?Plasmacítico: Leucemia ? , Paraproteína ?

• FENOTIPO: IgM, IgD citoplásmicaCD5 ± , CD23 ±CD38 ± , EMA ±

• GENETICA t(9;14)(p13;q32); PAX5 ?

• CONTROVERSIA: L Linfoplasmacitoide una variante de CLLSignificado pronóstico

Kappa Lambda

CD5 CD23

Diferenciación Plasmocelular en LNH

• CLL (Linfoma linfoplasmocitoide)

• Linfoma FolicularAreas Interfoliculares

• Linfoma de Zona Marginal Zonación topográficaEn ocasiones masiva: Plasmacitoma

• Linfoma B de Células GrandesLinfoma Plasmablástico (?)

Linfomas de Células PequeñasProblemas diagnósticos

• Similitudes citologicas

– Nucleos redondeados

– Nucleos hendidos

– Diferenciacion Plasmocelular

• Patrón Nodular

– FL vs MCL nodular

– Colonización centro germinal

• Patrón Zona Marginal

– MZL, CLL, MCL, FL

CLL/SLL MCL

MCL

FL

CD10

Cyclin D1

MARGINAL ZONE PATTERN

FL

CLL MZL

CD23

Cyclin D1

CD5

p27

Diagnosis

Ig

CD20

CD5

CD43

CD23

CD10

Bcl-6

Cyclin D1

p27

MCL MD/? ++ + + - - - + -

CLL M/D + + + + - - - +

FL M/G ++ - - - + + - +

LLP M ++ - + - - - - +

MALT M ++ - +/- - - - - +

SMZL M/D ++ - - - - - - +

Immunophenotype in Malignant Lymphomas