CHRONIC LYMPHOCYTIC LEUKEMIA (CLL)

Curs an IV – limba engleza

2012-2013

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CLL - Definition

• CLL is a neoplastic disease characterized by proliferation and accumulation (blood, marrow and lymphoid organs) of morphologically mature but immunologically dysfunctional lymphocytes

CLL - Definition

• Clonal B cell malignancy.

• Progressive accumulation of long lived mature lymphocytes.

• Increase in anti-apoptotic protein bcl-2.

• In most cases, the cells are monoclonal B lymphocytes that are CD5+

• Intermediate stage between pre-B and mature B-cell.

• T cell CLL can occur rarely

Cancer statastics 2000; CA J Clin 2000; 50:7-33

CLL - Epidemiology

• Most common leukemia of Western world.

• Less frequent in Asia and Latin America.

• Male to female ratio is 2:1.

• Median age at diagnosis is 65-70 years.

• Uncommon (10%) in patients under 50 years

• In US population incidence is similar in different races.

CLL - Etiology (1)

• The cause of CLL is unknown

• There is increased incidence in farmers, rubber manufacturing workers, asbestos workers, and tire repair workers

• Genetic factors have been postulated to play a role in high incidence of CLL in some families

CLL - Etiology (2)

• Cytogenetics– clonal chromosomal abnormalities are detected in

approximately 50% of CLL patients – the most common clonal abnormalities are:

• trisomy 12• structural abnormalities of chromosomes 13, 14 and 11

– patients with abnormal karyotypes have a worse prognosis

• Oncogenes– in most cases of CLL is overexpressed the proto-

oncogene c-fgr 9a member of the src gene family of tyrosine kinases

CLL – Initial symptoms

• Approximately 40% are asymptomatic at diagnosis – discovered by a CBC

• In symptomatic cases the most common complaint is fatigue

• Well’s syndrome – increase sensitivity to insects bites

• B symptoms – fever, sweats, weight loss

• Less often the initial complaint are enlarged nodes or the development of an infection (bacterial)

CLL - Clinical findings• Most symptomatic patients have enlarged lymph nodes

(more commonly cervical and supraclavicular) and splenomegaly

• The lymph nodes are usually discrete, freely movable, and nontender

• Hepatomegaly may occure• Less common manifestation are infiltration of tonsils,

mesenteric or retroperitoneal lymphadenopathy, and skin infiltration

• Patients rarely present with features of anemia, and bruising or bleeding

CLL – Lab findings a) Blood test lymphocytosis ≥ 5G/l (4 weeks)

b) Morphology monoconal population of small mature lymphocyte

c) B-cell CLL phenotype clonal CD5+/CD19+ population of lymphocyte

d) Markers of clonality κ/λ light chain restriction; cytogenetical abnormalities

e) Bone marrow infiltrate > 30% of nuceated cells on aspirate

f) Lymph node diffuse infiltrate of small lymphocye

CLL - Laboratory findings (1)

• The blood lymphocyte count above 5.000/mmc

• Leukemic cells have the morphologic appearance of

normal small lymphocytes

• In the blood smears are commonly seen ruptured

lymphocytes (“basket” or “smudge” cells)

• Careful examination of the blood smear can usually

differentiate CLL, and the diagnosis can be confirmed

by immunophenotyping

CLL - Peripheral Blood

• Absolute lymphocytosis• Lymphs = B cells

– Thin cytoplasm– Dense nucleus – Partially aggregated chromatin– No recognizable nucleoli

• Smudge cells

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CLL - Laboratory findings (2)

• Clonal expansion of B (99%) or T(1%) lymphocyte– In B-cell CLL clonality is confirmed by

• the expression of either or light chains on the cell surface membrane

• the presence of unique idiotypic specificities on the immunoglobulins produced by CLL cells

• by immunoglobulin gene rearrangements• typical B-cell CLL are unique in being CD19+ and CD5+

CLL - Immunophenotype

• Detect antigens on surface of cells – Specific antibodies– Use flow cytometry or immunohistochemistry

• CLL = mature B cells– CD5– CD19– CD20 - low– CD22 - low– CD23– Light chains (κ, λ)

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CLL - Immunophenotype scoring system

Scoring system for B-CLL

Membrane marker

Points

1 0

Smlg Weak Moderate/strong

CD5 Positive Negative

CD23 Positive Negative

FMC7 Negative Positive

CD79b (SN8) Negative Positive

1. Matutes E, et al. Leukemia. 1994;8:1640-1645.2. Moreau EJ, et al. Am J Clin Pathol. 1997;108:378-382.

CLL – Other lab tests

• Hypogammaglobulinemia or agamma-globulinemia are often observed

• 10 - 25% of patients with CLL develop autoimmune hemolytic anemia, with a positive direct Coombs’ test

• The marrow aspirates shows greater than 30% of the nucleated cells as being lymphoid

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CLL - Bone Marrow

• Infiltrates of small lymphocytes– Causes neutropenia, anemia,

thrombocytopenia

• Extramedullary expansion– Splenomegaly

– Hepatomegaly

– Lymphadenopathy

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CLL – Lymph node• Loss of nodal architecture• Diffuse infiltration of small

lymphocytes• Lymphadenopathy

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CLL - diagnostic criteria

1) A peripheral blood lymphocyte count of greater than 5 G/L, with less than 55% of the cells being atypical

2) The cell should have the presence of Bcell-specific differentiation antigens (CD19, CD20, and CD24) and be CD5(+)

3) A bone marrow aspirates showing greater than 30% lymphocytes

CLL - Differential diagnosis

• Infectious causes– bacterial (tuberculosis)– viral (mononucleosis)

• Malignant causes– B-cell– T-cell

• leukemic phase of non-Hodgkin lymphomas

• Hairy-cell leukemia

• Waldenstrom macroglobulinemia

• large granular lymphocytic leukemia

Investigations

• Pretreatment studies of patients with CLL should include examination of:– complete blood count– peripheral blood smear– reticulocyte count– Coomb’s test– renal and liver function tests – serum protein electrophoresis– immunoglobulin levels– plasma 2 microglobulin level

• If available immunophenotyping should be carried out to confirm the diagnosis

• Bone marrow biopsy and cytogenetic analysis is not routinely performed in CLL

CLL – Rai staging system

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CLL – Rai stages

CLL – Binet staging system

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CLL – Binet stages

Staging: Rai and Binet staging systems for CLL

Clinical staging systems for CLL

Stage

Value Rai Binet Median survivalLymphocytosis (>15,000/mm3) 0 -

150 months (12.5 years)

Lymphocytosis plus nodal involvement I A <3

node groups101-108 months

(8.5-9 years)

Lymphocytosis plus organomegaly II B >3

node groups60-71 months

(5-6 years)

Anemia (RBCs) IIIHgb <11 g/dL

C

Hgb <10 g/dL 19-24 months

(1.5-2 years)

Lymphocytosis plus thrombocytopenia(platelets)

IVPLT <100,000/mm3 PLT <100,000/mm3

CLL - Markers of poor prognosis

• Advanced Rai or Binet stage

• Peripheral lymphocyte doubling time <12 months

• Diffuse marrow histology

• Increased number of prolymphocytes or cleaved cells

• Poor response to chemotherapy• High 2- microglobulin level

• Abnormal karyotyping

Prognosis: histologic bone marrow patterns

• The different bone marrow patterns probably reflectvariations in amount of lymphoid accumulation duringthe natural course of the disease

Interstitial(low risk)

Diffuse(high risk)

Nodular(low risk)

Randy Gascoyne, MD., Montserrat E, et al. Cancer. 1984;54:447-451.

Prognosis: lymphocyte doubling time

Survival time according to LDT (all stages)

Months

Pro

babili

ty o

f su

rviv

al

1600 20 40 60 80 100 120 140

1.0

0.9

0.8

0.7

0.6

0.5

0.4

0.3

0.2

0.1

0.0

Doubling time ≤12 monthsDoubling time >12 months

1. Montserrat E, et al. Br J Haematol. 1986;62:567-575.

CLL - Genetic abnormalities

Genetic abnormality

Incidence (%)

Median survival (months)

Clinical correlation

13q14 55-62 133-292 Typical morphology Mutated VH genes Stable disease

+ 12 16-30 114-122 Atypical morphology

Progressive disease

del 11q23 18 79-117 Bulky lymphadenopathy Unmutated VH genes

Progressive disease Early relapse post autograft

p53loss/mutation

7 32-47 Atypical morphology

Unmutated VH genes

Advanced disease Drug resistance

1. DÖhner H, et al. N Engl J Med. 2000;343:1910-1916.2. Oscier DG, et al. Blood. 2002;100:1177-1184.

Effect of genetic abnormalities on survival

1. Döhner H, et al. N Engl J Med. 2000;343:1910-1916.

CLL - VH Status

• “Variable” region of immunoglobulin gene

• H=heavy chain

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CLL - VH Mutation

• Naïve B cells have unmutated V regions– Make initial IgM and IgG that bind Ag poorly

• Somatic hypermutation of V region creates memory B cells– Make Ab with higher affinity for Ag

CLL progression is dependent on mutation status of B cells– Unmutated VH = rapid progression, short survival– Mutated VH = slow progression, long survival

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CLL - VH Mutation

Naïve B cellsUnmutated VH

CLLFast progressionShort survival

Memory B cellsMutated VH

CLLSlow ProgressionLong Survival

90

225 3000 50 100 150 200 25025 75 125 175 275

100

80

60

40

0

20

70

50

30

10

Unmutated VH geneMedian = 117 monthsMutated VH geneMedian = 293 months

325

1. Hamblin TJ, et al. Blood. 1999;94:1848-1854.

Perc

ent

surv

ivin

g (

%)

Months

Prognosis: effect of VH gene mutations on survival

Clinical Course of CLL• Disease has a variable course; however, it often progresses

from an indolent lymphocytosis without other evident disease to one of generalized lymphatic enlargement with concomitant pancytopenia

• Progression: bone marrow impairment, susceptibility to infection

• Complications of pancytopenia, including hemorrhage and infection, represent a major cause of death in these patients

CLL - Immunologic Complications • Autoimmune hemolytic anemia

– Coombs’ positive– Clinical hemolysis

• Pure red cell aplasia• Immune-mediated thrombocytopenia• Depressed immunoglobin levels• Granulocytopenia• Hypogammaglobulinemia with advanced disease• Long-term complications: autoimmune phenomena,

Richter’s transformation

CLL - Complications

• Severe systemic infections

• Bleeding

• Richter’s transformation

• Prolymphocytoid transformation

• Secondary malignancies

• Acute myeloid leukemia

Transforming the way we approach CLL therapy

• Traditional treatment goal: Palliation– Age is a factor

– Treat to relieve symptoms

– Continuous/intermittent treatment

– No survival advantage for any tested treatment to date

• New advances may allow treatment for remissionand survival– Treat to complete remission (CR)

– Eliminate minimal residual disease

– Response predicts for survival: More CRs

CLL – treatment (1)

• Watch and wait• Monotherapy

– glucocorticoids– alkylating agents (Chlorambucil, Cyclophosphamide)– purine analogues (Fludarabine, Cladribine, Pentostatin)

• Combination chemotherapy– Chlorambucil/ Cyclophosphamide + Prednisone– Fludarabine + Cyclophosphamide +/- Mitoxantrone– CVP, CHOP

• Monoclonal antibodies (monotherapy and in combination)– Alemtuzumab (anti-CD52)– Rituximab (anti-CD20)

• Splenectomy• Radiotherapy

CLL – treatment (2)

• Hematopoietic stem cell transplantation– allogeneic with reduced intesity conditioning

– autologous

• New and novel agents– Oblimersen – bcl2-directed antisense oligonucleotide

– Lenalidomide

– Flavopiridol

– Anti-CD23

– Anti-CD40

• Vaccine strategies

• Supportive therapy (allopurinol, G-CSF, blood and platelet transfusion, immunoglobulins, antibiotics)

NCI-WG: Indications to Initiate Treatment for CLL

• Constitutional symptoms referable to CLL (B-symptoms)

• Progressive marrow failure• Autoimmune anemia and/or thrombocytopenia

poorlyresponsive to corticosteroids

• Massive or progressive splenomegaly• Massive or progressive lymphadenopathy• Rapid lymphocyte doubling time

MoAbs citotoxic mechanisms

Effector cells/Complement

Apoptosis Radionuclide Toxin/Antibiotic

MoAbs for CLL

Antibody Antigen

Alemtuzumab (Campath-1H) CD52

Rituximab (Rituxan, Mabthera) CD20

Epratuzumab (LymphoCide) CD22

Hu-1D10 (Apolizumab) HLA-DR

IDEC-152 (Lumiliximab) CD23

IDEC-114 CD80

Bevacizumab (Avastin) VEGF

BL-22 CD22( conjugate with Pseudomonas)

MabThera®: a chimeric murine/human MoAb

Variable murine regions bounding CD20 on B cells

Human kappa costant regions

Human domain IgG1 Fc, synergistic with human effector mechanisms

Chimeric IgG1

Rituximab as part of first-line therapy for CLL: Rationale

• Rituximab monotherapy is moderately active in CLL1,2

– Activity is dose dependent (between 500–2250 mg/m2)1

• Rituximab acts synergistically with other cytotoxic agents in vitro– Increases fludarabine activity in NHL cell lines3

– Increases activity of bendamustine, mitoxantrone and other chemotherapeutic agents in CLL cells4

• Rituximab combination therapies (e.g. R-F, R-FC, R-PC, R-FCM,R-Bendamustine, R-Chlorambucil) are now being assessed

Alemtuzumab (anti-CD52) antibody

IgG1 humanised antibody:Low immunogenicity

CD52 antigen: Highly expressed on

all lymphocytes monocytes and macrophages spermatozoa eosinophils

Not expressed on haemopoietic stem cells

Does not modulate/shed Also expressed on the majority of

malignant lymphocytes

Treatment of CLL

CLL – Treatment strategy

Gribben J G Blood 2010;115:187-197