The Chronic Lymphocytic Leukemia (CLL)

The Chronic Lymphocytic Leukemias (1) The group of clonal diseases characterized by proliferation and accumulation of small, mature lymphocytes in blood, bone marrow and lymphoid tissues (lymph nodes, spleen) Neoplastic lymphocytes belong most often to B-cell lines and they have the special for B-cell antigenes on their surface; exceptionally neoplastic lymphocytes belong to T-cell lines or NK-cell According to REAL (Revised European-American Lymphoma)/ /WHO classification CLLs belong to the group of: lymphoproliferative diseases lymphomas

Lymphoproliferative diseases Primary lymphatic system (central) bone marrow thymus Secondary lymphatic system (peripheral ) spleen lymph nodes MALT (The mucosa-associated lymphoid tissue = also called mucosa-associated lymphatic tissue)

Clinical stages of lymphomas according to Ann Arbor’s classification II III IV A: no general symptoms B: general symptoms such as fever, night sweats, weight loss Lister T, et al. J Clin Oncol 1989; 7:1630

Ann Arbor’s classification is specific for all lymphomas CLL is classified according to Rai and Binet classification

WHO classification B-Cell neoplasms - Precursor B-cell neoplasm: B-lymphoblastic leukemia/lymphoma - Mature (peripheral) B-cell neoplasms: B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma B-cell prolymphocytic leukemia Lympfioplasmacytic lymphoma Splenic marginal zone B-cell lymphoma (+ /- villous lymphocytes) Hairy cell leukemia Plasma cell myeloma/plasmacytoma Extranodal marginal zone B-cell lymphoma of MALT type Nodal marginal zone B-cell lymphoma (+/— monocytoid B cells) Follicular lymphoma Mantle-cell lymphoma Diffuse large B-cell lymphoma Mediastinal large B-cell lymphoma Primary effusion lymphoma Burkitts lymphoma/Burkitt cell leukemia T-cell and NK-cell neoplasms - Precursor T-cell neoplasm: T-lymphoblastic lymphoma/leukemia - Mature (peripheral) T-cell neoplasms: T-cell prolymphocytic leukemia T-cell granular lymphocytic leukemia Aggressive NK-cell leukemia Adult T-cell lymphoma/leukemia (HTLV1 +) Extranodal NK/T-cell lymphoma, nasal type Enteropathy-type T-cell lymphoma Hepatosplenic gamma-delta T-cell lymphoma Subcutaneous panniculitis-like T-cell lymphoma Mycosis fungoides/Sezary syndrome Anaplastic large-cell lymphoma, T/null cell, primary cutaneous type Peripheral T-cell lymphoma, not otherwise characterized Angioimmunoblastic T-cell lymphoma Anaplastic large-cell lymphoma, T/null cell, primary systemic type

The Chronic Lymphocytic Leukemia (1) Chronic lymphocytic leukemias are derived from: B-cell line B-cell chronic lymphocytic leukemia B-cell chronic prolymphocytic leukemia Hairy cell leukemia Splenic marginal zone B-cell lymphoma ( + /- villous lymphocytes) T-cell line T-cell chronic lymphocytic leukemia T-cell chronic prolymphocytic leukemia T-cell granular lymphocytic leukemia Chronic lymphocytic leukemias differ form each other in biology, morphology, antigen structure of the cell and in clinical course

The B-CLL - definition B-CLL is a neoplastic disease characterized by proliferation and accumulation of small, mature, long-living lymphocytes in blood, marrow and lymphoid tissues (lymph nodes, spleen) This lymphocytosis leads to specific clinical and laboratory symptoms of B-CLL The neoplastic lymphocytes have on their surface the special for B-cell line antigens – CD19, CD20 and also CD5, CD23, and a very weak expression of surface immunoglobulin

B-CLL epidemiology Most common adult leukemia in Europe and North America (in USA incidence of about 3/100.000 population) predominantly, CLL is a disease of elderly (50-55 years) 40% of leukemias in patients over 60 years old Morbidity: Men 2,2-3,69 / 100 000 / year Women 0,9-1,59 / 100 000 / year /men affect twice as often as women; 2:1 ratio of male to female / CLL morbidity rapidly increases with age (especially between 50 and 60 years of age) in 98% of patients the leukemic cells are a monoclonal population of mature B lymphocytes with low-density surface immunoglobulin death from infections, BM failure, high-grade transformation (Richter's syndrome), kachexia

B-CLL etiology & pathogenesis (1) the cause of CLL is unknown there is increased incidence in farmers, rubber manufacturing workers and tire repair workers genetics factors have been postulated to play a role in high incidence of CLL in some families

B-CLL etiology & pathogenesis (2) Cytogenetics - clonal chromosomal abnormalities are detected in approximately 50% of CLL patients Immunoglobulin genes - monoclonal surface immunoglobulin is expressed by over 90% of patients (60% kappa and 40% lambda light chains) nearly half of all cases have leukemia cells that express mutated immunoglobulin variable region genes (Ig VH genes) - associated with more indolent disease Immunologic abnormalities autoimmune disease (hemolytic anemia and thrombocytopenia, pure red cell aplasia) hypogammaglobulinemia cellular immune defects

B-CLL clinical symptoms (1) often none! - 25% of patients are asymptomatic and the diagnosis is typically accidental unspecific: night sweats, fever, weakness (many patients have fatigue, reduced exercises tolerance or malaise, weight loss) recurrent infections (bacterial, viral – Herpes Zoster, fungal) – they are the most common cause of death bleeding and symptoms of anemia and thrombocytopenia Lymphadenopathy (lymph node enlargement) at diagnosis - nontender in 80% of patients later - may become very large splenomegaly - mild to moderate in 50% of patients hepatomegaly some organs infiltration (lungs, pleura, skin and soft tissue) Blood lymphocytosis does not cause symptoms!

B-CLL clinical symptoms (2) Cervical and axillary limfadenopathy in 60-years old patient with B-CLL

B-CLL clinical symptoms (2) Cervical and axillary limfadenopathy in 70-years old patient with B-CLL

B-CLL clinical symptoms (3) Cervical limfadenopathy in patient with B-CLL

B-CLL clinical symptoms (3) The CLL patient can have splenomegaly

B-CLL clinical symptoms (3) The CLL patient has splenomegaly, which is visble

B-CLL laboratory features (1) Morphology: Leucocytosis with monoclonal lymphocytosis of greater than 5.000/ul. anemia Because of „displacement ” and/or autoimmunohemolic (10-20% of patients have a positive direct antiglobulin test; AIHA is commonly connected with the presence of warm auto- antibodies IgG class – rapidly increasing fatigue, skin getting yellow, anemia with enlarged reticulocytosis, higher level of bilirubin) pure red cell aplasia is very rare (selective aplasia of red cell line in bone marrow) thrombocytopenia and/or immunologic (about 5% of B-CLL patients have anty-platelet antibodies) protein electrophoresis – Hipogammaglobulinemia, monoclonal protein in 5% of patients

B-CLL laboratory features (2) Peripheral blood smear: Lymphocytosis small, mature, morphologically normal Smudge cells Neutropenia Because of „displacement ” and/or autoimmunologic

B-CLL laboratory features (3) Bone Marrow smear (cytological examination) extensive replacement of marrow element by mature lymphocytes (more than 30%)

B-CLL laboratory features (4) Bone Marrow Biopsy (histological examination): Lymphocyte infiltration nodular infiltration, interstitial infiltration, difussed infiltration mixed infiltration Difussed infiltration (unfavourable prognostic factor)

B-CLL laboratory features (5) Bone Marrow Biopsy Interstitial infiltration

B-CLL laboratory features (6) lymph node finding (histopathological examination) - diffuse infiltration of small lymphocytes identical to low-grade, small lymphocytic lymphoma

B-CLL laboratory features (7) Immunophenotype: CD5+/CD19+/CD23+/CD20+, sometimes also CD38+, low expression of CD22; lack expression of CD 10-, CD 103-, 90% of the patient have a very weak expression of surface immunoglobulin (kappa or lambda light chain, IgM, IgD)

B-CLL features (8) Radiological examinations (X-ray, ultrasonography, CT, ...) Serological examinations (direct and indirect antiglobulin tests) Biochemical examinations (lactate dehydrogenase, 2-microglobulin)

B-CLL laboratory features (9) Cytogenetic examinations - clonal chromosomal abnormalities are detected in approximately 50% of CLL patients deletion 13 (13q14.3) trisomy 12 structural abnormalities of chromosomes 11 (11q-), 14, 17 Genomic aberrations found in approximately 50% of CLL

Diagnosis of B-CLL Blood test lymphocytosis ≥ 5G/l (6 weeks) Morphology monoclonal population of small mature lymphocyte B-cell CLL phenotype clonal CD5+/CD19+ population of lymphocyte Markers of clonality κ/λ light chain restriction; cytogenetical abnormalities Bone marrow infiltration > 30% of nucleated cells on aspirate Lymph node diffuse infiltrate of small lymphocytes

RAI’s CLINICAL STAGING SYSTEM Stage Clinical Features at Diagnosis Median Survival (years) Low risk Blood lymphocytosis>5G/l, Bone marrow lymphocytosis>30% >12,5 I Intermediate risk and enlarged lymph nodes 8 II and enlarged spleen and/or liver 6 III High risk and anemia (Hb < 11g/dl) 1,5-2 IV and thrombocytopenia(< 100 000 /ul)

CLL – Rai stages

BINET’s CLINICAL STAGING SYSTEM Stage Clinical Features at Diagnosis Median Survival (month) A Blood lymphocytosis>5G/l, Bone marrow lymphocytosis>30% and less than 3 areas of palpable lymphoid-tissue enlargement Without anemia (Hb >= 6,21 mmol/l, 10 g/dl) and thrombocytopenia > 120 month B and 3 and more areas of palpable lymphoid-tissue enlargement 60 month C with anemia (Hgb <10g/dL) or thrombocytopenia (Plt <100.000/uL) 24 month An area: cervical, axillary left, axillary right, inquinofemoral left, inquinofemoral right lymph nodes, spleen, liver

CLL – Binet’s stages

New prognostic indicators in B-CLL (1) Prognostic factor Good prognosis Bad prognosis Clinical stage according to Binet Rai A B, C I, II, III, IV Bone marrow infiltration in - bone marrow biopsy - cytological examination Leucocytosis Prolymphocytes in peripheral blood Leukemia cell doubling time Non-Difussed infiltration <=80% lymphocytes <= 50 x 109/l <= 10% > 12 months Difussed infiltration > 80% lymphocytes > 50 x 109/l >10% <= 12 months

New prognostic indicators in B-CLL (2) Prognostic factor Good prognosis Bad prognosis Serum markers: lactate dehydrogenase activity (LDH) ß2-mikroglobulin activity lymphocyte’s thymidine kinase activity CD23 expression Normal range Elevated Clonal chromosomal abnormalities Normal karyotype isolated del (13q) Del (11q) Del (17p) CD 38 expression <= 30 % > 30%

New prognostic indicators in B-CLL (3) Prognostic factor Good prognosis Bad prognosis The mutational status of immunoglobulin variable region of heavy chain genes (IgvH) mutated unmutated ZAP–70 expression low (< 20%) high ( > 20 %) Survivin absence presence

New prognostic indicators in B-CLL (4) - summary clinical stage bone marrow histology (diffuse replacement carries worst prognosis) leukemia cell doubling time (less than 1 year - worse prognosis) percentage of prolymphocyte high cell-surface expression of CD38 ZAP-70 expression serum level of: B2-microglobulin; thymidine kinaze, LDH, sCD23 IgVH mutational status genetic features - FISH cytogenetic low-risk: normal kariotype; isolated del(13q) high-risk: del(17p0, del(11q), trisomy 12

CLL : ZAP-70 ZAP70 is an intracellular protein which is strongly correlated with the VH status in CLL

CLL – treatment (1) We have to remember: B-CLL – indolent lymphoma, but incurable Elderly patients – risk of additional diseases Course of the disease can be very long, indolent for many years, patient can die because of another reason which is not connected to B-CLL. Decision about treatment depends on clinical stage, prognostic factors and patient’s condition Indications to treatment: III/IV stage according to Rai’s classification Progressive disease (rapidly increasing lymphadenopathy, infections, general symptoms) leukemia cell doubling time <6 (12) months rapidly increasing organomegaly Secondary anemia, neutropenia, thrombocytopenia because of bone marrow infiltration Richter’s syndrome

CLL – treatment (2) Watch and wait Monotherapy Glucocorticoids (autoimmunological complications) alkylating agents (Chlorambucil, Cyclophosphamide) purine analogues (Fludarabine, Cladribine, Pentostatin) Combination chemotherapy Chlorambucil/Cyclophosphamide + Prednisone purine analog (Fludarabine) + Cyclophosphamide +/- Mitoxantrone CVP, CHOP (Cyclophosphamide, Doxorubicin, Vincristin, Prednisone) Monoclonal antibodies (monotherapy and in combination) Alemtuzumab (anti-CD52) = CAMPATH Rituximab (anti-CD20) = Mabthera antiCD23 etc. monoclonal antibodies conjugated with radionuclides = Ibritumomab tiuxetan = Zevalin Splenectomy (hypersplenism) Radiotherapy (massive lymphadenopathy)

CLL – treatment (3) Hematopoietic stem cell transplantation autologous - still no cure with auto-SCT allogenic with reduced intensity conditioning Even RIC-SCT is still a risky procedure - indicated only in high-risk disease Can allo-SCT cure CLL? - YES 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)

Response criteria (NCI working group 1996) Complete response (for at least 2 months) clinical features – normal morphology – normal (Hb>11 g/dl; Pt>100 000 /ul, lymphocytes <4000 G/l; neutrofiles >1500 G/l)) bone marrow - lymphocytosis less than 30% Partial response Stable Disease Progressive Disease

Richter’s Syndrome is always the transformation of CLL into an aggressive Lymphoma – diffuse large cell lymphoma (DLCL) or Hodgkin‘s lymphoma usually evolves after a long indolent course - can occur as 1st manifestation of CLL: Primary Richter‘s - but still CLL has a poor prognosis