Chronic lymphocytic leukemia: Present status

Annals of Oncology 6: 219-235, 1995. © 1995 Kluwer Academic Publishers. Printed in the Netherlands.

Review Chronic lymphocytic leukemia: Present status E. Montserrat & C. Rozman Postgraduate School of Hematology 'Farreras Valenti', Department of Medicine, University of Barcelona, Hospital Clinic, Barcelona, Spain Summary

Chronic lymphocytic leukemia (CLL) is the form of leukemia which occurs most frequently in Western countries. Its etiology is unknown, and no relationship with viruses or genes has been demonstrated. Epidemiological data suggest that genetic and ambiental factors might be of some significance. Clinical features of CLL are due to the accumulation of leukemic cells in bone marrow and lymphoid organs as well as the immune disturbances that accompany the disease. The prognosis of patients with CLL varies. Treatment is usually indicated by the risk of the individual patient, which is clearly reflected by the stage of the disease. In the early stage (Binet A, Rai O) it is reasonable to defer therapy until disease progression is observed. By contrast, because their median survival is less than five years, patients with

1. Introduction

Chronic lymphocytic leukemia (CLL) is due to the proliferation and accumulation of immune-incompetent B-lymphocytes arrested at an early stage of their differentiation (1, 2]. Clinical features of CLL reflect the accumulation of leukemic cells in bone marrow and lymphoid organs, and the immune disturbances (e.g., hypogammaglobulinemia, autoimmune phenomena) that accompany the disease. The prognosis of patients with CLL varies. Some patients survive for a long time and ultimately succumb to maladies unrelated to CLL, while others have rapidly progressive courses and die soon after diagnosis. In the last two decades, important advances have been made in the biology, natural history, prognosis, and therapy of CLL [3-9]. This progress is summarized in this review. 2. Epidemiology

CLL is rare in people younger than 40, but the incidence increases dramatically with age [10]. The median age of patients at diagnosis is about 65 years [3-5]. The incidence of CLL is particularly high in Western countries (1.5 to 2.5 per 100,000 persons/year), where it accounts for 30% of all leukemias [3-5, 10]. By contrast, in Asian populations CLL constitutes only

more advanced stages require therapy. For almost 50 years, no major advances in the management of CLL, which has revolved around the use of alkylating agents, have been made. In recent years, the therapeutic approach in patients with CLL has changed as a result of the introduction of combination chemotherapy regimens and, in particular, purine analogues. The latter are already the treatment of choice for patients not responding to standard therapies, and their role as front-line therapy is being investigated. Bone marrow transplants are also being increasingly used. It is to be hoped that in years to come the outcome of patients with CLL will be improved by these advances.

Key words: biology, chronic lymphocytic leukemia, natural history, prognosis, treatment

3%-5% of the total number of cases of leukemia; this variation probably reflects not only real differences among populations but also under- and misdiagnoses, as well as variability in access to medical care [10]. Interestingly, in Asians who have emigrated to the U.S. the incidence of CLL is still low [11]. It is higher in Jews originating from Eastern Europe than in Sephardics (proportion: 4:1) [1, 12], and in most series it is also higher in males than in females (average ratio: 2, range: 1.0 to 4.7) [10]. 3. Etiology

The etiology of CLL is unknown; no virus or oncogene has been found to be involved in its pathogenesis. Relationships among various chemicals, drugs, and other environmental agents and the development of the disease are much less well established for CLL than for other forms of leukemia [10]. Ionising radiation does not lead to this type of leukemia [3-5, 10]. An increased risk of CLL in people exposed to a number of chemicals (e.g., benzene, aromatic hydrocarbons) has been reported, although the evidence is weak [10]. In addition, some studies have reported a moderately higher incidence of CLL among farmers, and petroleum and wood workers than among other groups, suggesting an occupational exposure in the etiology of

220 CLL [10]. Familial aggregates of leukemia with other cases of leukemia, lymphoproliferative diseases or autoimmune conditions support the notion of a genetic factor in this disease [13, 14], although studies in twins have indicated an acquired nature [15]. The excessive risk of leukemia in people with one or more firstdegree relatives with CLL has been estimated as 2- to 7-fold greater than the risk in individuals without affected first-degree relatives [16]. HLA information in CLL is contradictory [10].

phocytes were arrested at an early stage of differentiation, studies with anti-idiotypic reagents and mitogenic stimulation have shown that the CLL B lymphocyte can differentiate [3-5, 31]. The CLL cytokine network is complex. A variable pattern has been shown in the response to different cytokines such as IL-2 and B-cell growth factors (e.g., BCGF, IFN alpha and gamma, IL-4). TNF alpha, IL-2 and possibly IL-1 and IL-6, either alone or in synergy, act as growth factors, and IFN-gamma, IFN-alpha, and IL-4 inhibit apoptosis [32-35]. CLL cells also express EL-8 [36].

4. Biology

4.3. Normal cell counterpart of the CLL B cell

4.1. Clonality

CD5+ (Bl-B) B cells constitute a small subpopulan'on of human B lymphocytes in the lymphoid organs and CLL results from an accumulation of clonal B-lympho- peripheral blood (10%-15%) of normal adults. The cytes with a relatively mature phenotype. CLL B-cells normal counterpart of the CD5+ cells which proliferate express a single Ig light chain (kappa or lambda) on the in CLL was initially located at the edge of the germinal cell membrane, a unique immunoglobulin-idiotype center in human lymph nodes [37]. It was subsequently specificity, a single pattern of glucose 6-phosphate observed that a substantial number of B cells from 20dehydrogenase activity, clonal chromosome abnormal- week-old fetal lymph nodes and spleen express the CD 5 marker as well as mu and delta chains on their ities, and unique Ig gene rearrangements [3-5,17]. membrane. These fetal cells also appear to share lectin 4.2. Characteristics of the neoplastic B lymphocyte in non-responsiveness and an inability to cap Smlg of B lymphocytes [8, 38]. CLL The high prevalence of CD 5+ cells during early The neoplastic B cell from CLL is currently known as ontogeny led some authors to assume that the CD5+ the Bl-B cell, in contrast to the normal B cell which is lymphocyte corresponds to an expansion of a B cell designated as the B2-B cell [18]. In the neoplastic clone arrested at an immature stage between pre-B and lymphocyte from CLL, surface membrane immuno- mature-B cell. This hypothesis, however, does not satisglobulin (Smlg) is restricted to a single Ig light chain factorily explain the difficulty that B cell differentiation (kappa or lambda) and frequently expresses IgM or pathways have in integrating CD5 antigen expression, both IgM and IgD. In the latter case, IgM and IgD have the high frequency of hypogammaglobulinemia or the been shown to share idiotypic and antigen specificities. autoimmune phenomena directed against blood cell B cells express the T-cell antigen CD5 as well as several components in CLL. Alternatively, it has been postuother antigens, including DR-related human leukocyte lated that CD 5+ cells could derive from a separate B antigens and B-cell antigens (e.g., CD19, CD20, CD24, cell lineage [4]. CD37, and CD21). CD23 is positive in most cases, whereas membrane positiviry for CD 2 2 is infrequent. 4.4 Cytogenetic abnormalities CLL B-cells also express mouse RBC receptors, but virtually never react with FMC7 and CALLA (CD 10). Approximately 50% of patients with CLL have cytoExpression of some myelomonocytic antigens may be genetic abnormalities; trisomy 12 is found in 11% to observed [3-5,19, 20]. CLL cells also depict the com- 54% of these patients [39-44]. This variation reflects mon CLL antigen (cCLLa) [21]. Mdr genes and p-gly- differences in patient selection and in diagnostic cricoprotein are variously expressed [22-25]. Of the teria as well as in the sensitivity of the techniques used. integrin family, the beta-1 integrin VLA4 is poorly ex- For example, FISH identifies trisomy 12 more frepressed, the VLA3 is constantly present [26, 27]. While quently than does conventional cytogenetics [40, 42, in turn the beta-2 integrin LFA-1 (CD 11 a/CD 18) is 43]. On the other hand, trisomy 12 appears more frenot expressed [28]. quently in 'atypical' CLL, with more prolymphocytes C5+ (Bl-B) lymphocytes from CLL display a re- or lymphoplasmacytoid differentiation than in typical stricted expression of VH and VL genes. Ig VH genes or classic CLL [42, 44]. Recurrent abnormalities inof the relatively small VH subgroups, VH4 and VH5, volving chromosome 13 at band ql4, the site of the and the single copy VH6 gene are rearranged at fre- retinoblastoma (Rbl) suppressor gene, are present in quencies which are disproportionate to the relative one-third to one-fifth of the patients with clonal abnorsizes of these subgroups in the germline DNA. On the malities [39, 40]. Other abnormalities include deletions other hand, the largest VH gene subgroup, the VH3 of the long arm of chromosomes 6 and 11, with variable breakpoints [39, 40]. Deletions of chromosome 6 gene family, seems to be under-represented [29, 30]. Although earlier studies indicated that B-CLL lym- (q21q23) have been associated with a subset of lym-

221 phoid malignancies that exhibit larger prolymphocytoid cells in peripheral blood [45]. 14q+ abnormalities are present in about 15% of the patients. Most of the 14q+ marker chromosomes are due to t(ll;14) translocations involving the bcl-1 gene; however, this abnormality is more specific to other types of lymphoproliferative disorders such as prolymphocytic leukemia and mantle-cell lymphoma than for CLL [46]. About 40% of the patients undergo karyotypic evolution which is usually associated with disease progression [47].

(B2-B) cells and correlates with tumor burden but not with previous treatment. Immune response to vaccines is considered to be hampered by hypogammaglobulinemia [30,64]. Although a marked monoclonal (M) immunoglobulin component, usually of IgM, can be found in about 5% of the patients [65, 66], a small M component can be detected in the serum or urine of 80% of the patients by use of high-resolution techniques [67]. In most cases, the M component is secreted by non-neoplastic (B2-B) cells.

4.5 Oncogenes

4.7. Natural antibody activity

Although initial reports suggested involvement of the bcl-1 and bcl-2 genes in 5% to 15% of the patients [3-5, 48-51], subsequent studies failed to confirm these data. For example, in a recent analysis of 100 patients with well-diagnosed 'classic' CLL, no alteration of the bcl-1 or bcl-2 oncogenes was detected [52], in agreement with other reports [53]. In a few cases, a t(14;19) has been reported with involvement of the bcl-3 gene [54, 55]. Again, this abnormality seems to be extremely rare [55]. Deletions of the Rbl gene have been reported in 13%-31% of the cases [56, 57], but biallelic deletions of the Rbl gene have not been observed, and patients with monoallelic deletions have been shown to have normal Rbl gene mRNA and protein. Characterization of a genomic region localized telomeric to the Rbl gene, identified by the D13S25 probe, might be of great importance. This region has been found to be homozygously deleted in leukemic cells from CLL patients. It is possible that the D13S25 recognized a new tumor suppressor gene implicated in the pathogenesis of CLL [58, 59]. Southern blot analysis identifies bcl-2 expression in only 5% to 10% of CLL. In contrast to follicular lymphomas, the translocation does not involve the 3' region of the bcl-2 gene but rather of the 5' region [50, 60], but bcl-2 is overexpressed even when there is no rearrangement. Thus, in a study using immunoblotting with an antibody specific fo the bcl-2 protein, expression of bcl-2 was detected in more than 70% of the patients [61]. On the other hand, overexpression of the c-myc oncogene, deletions of the Rbl gene, and mutations of the p53 suppressor gene have been reported in a small number of patients with CLL and small lymphocytic lymphomas showing clinical progression and morphologic transformation [62,63].

A high proportion of CLL B cells displays natural antibody activity, with about half of the CLL B cells having rheumatoid-factor activity and about 20% showing multispecific activity against autoantigens such as DNA and cytoskeleton proteins [68]. Autoimmune phenomena are mainly directed against cells of the hematopoietic system, and in most cases they reflect abnormalities in non-neoplastic (B2-B) cells. Nevertheless, some cases have been reported of antibodies produced by the neoplastic CD5+ (Bl-B) cell clone [69].

5. Clinical features As many as 70% of patients with CLL are currently diagnosed in an asymptomatic phase because of the increasing number of medical examinations made because of minor complaints. At the Postgraduate School of Hematology in Barcelona, the proportion of patients diagnosed in an early stage has increased from 40% in the early 1970s to about 70%. A similar trend has been observed in other series [4, 70]. In symptomatic patients, the most frequent findings are generalized lymphadenopathy, fatigue, and weight loss. A history of repeated infections in the months preceding diagnosis is not infrequent. Infiltration of extralymphatic organs and tissues (e.g., pleura, lung, skin, CNS) is extremely rare, although some cases have been reported [71, 72]. In addition, vasculitis [71], hypercalcemia [73], and nephrotic syndrome [74] have also been described. In rare cases (<1% of the patients), spontaneous remissions may be observed at some time during the course of the disease [75]. 6. Complications

4.6 Disglobulinemia 6.1. Autoimmune phenomena Hypogammaglobulinemia occurs in about 60% of the patients and is the major cause of infection in CLL [64]. The pathogenesis of hypogammaglobulinemia is poorly understood and data concerning helper and suppressor lymphocytes, and NK and ADCC activity are contradictory [30, 64]. Hypogammaglobulinemia is considered to be the result of a dysfunction of non-clonal B

A positive direct anti-globulin test has been reported in 7.7% to 35% of patients. The antibodies are typically warm ones of the IgG class. Autoimmune hemolytic anemia occurs in 10% to 25% of the patients and may be triggered by cytotoxic agents used to treat the disease [76, 77]. It has been suggested that the latter

222

might be particularly frequent with fludarabine [78, 79]. Antibody-mediated Coombs' negative hemolysis can occur when the density of the autoantibody on the red-blood cells is below the threshold detectable by the standard antiglobulin test, a possibility that should be borne in mind when dealing with CLL patients with anemia but otherwise low tumor burden. Immune thrombocytopenia is observed in only 2% of the cases. Pure-red cell aplasia and immune neutropenia are even less frequent and, as is the case with immune thrombocytopenia, are difficult to document [80, 81]. Cases of CLL associated with lupus erythematosus [82], Sjogren syndrome [82], and thyroiditis [83] have also been reported. 6.2. Infections

90]. CLL/PL carries a poorer prognosis than classic CLL and there is a strong correlation between the number of prolymphocytes in blood and survival [90, 91]. Although only rarely, the disease sometimes transmutes into acute leukemia and multiple myeloma [92, 93], and cases of Hodgkin's disease [94], myelodysplastic syndrome [95], polycythemia vera [96] and chronic myelocytic leukemia [97] have also been described in patients with CLL. 6.4. Second malignancies The fact that patients with CLL may suffer from second cancers is well known [98-101]. However, risk assessment is not easy because of the variability in the criteria used to evaluate the risk. For example, skin cancers are included in some studies but not in others. In a study based on data from tumor registries, an incidence of 8.9% corresponding to a 28% increased risk of second malignancy was found [102]. In that study, the second cancers most frequently associated with CLL and their relative observed/expected ratios were Hodgkin's disease (7.69), intraocular melanoma (3.79), malignant melanoma (2.79), brain tumors (1.98), and lung cancer (1.90). There is no relationship between the characteristics of the disease and its treatment and the incidence of second cancers, although treatment with continuous chlorambucil was associated with a high number of solid tumors in a large trial.

Infections are primarily due to the hypogammaglobulinemia that up to 60% of the patients present, although other immune defects (e.g., impaired complement activation, T-cells imbalance, NK and ADCC disfunction) may also play a role [64, 84]. Historically, most of the infections were caused by common bacteria such as Streptococcus pneumoniae, Staphylococcus, and Haemophilus influenzae. Herpes zoster is also a common complication. The use of new chemotherapy agents, particularly purine analogues which decrease the absolute number of CD4+ cells in blood, has led to the observation of infections due to opportunistic organisms such as Legionella, Pneumocystits carinni, Listeria, and Cytomegalovirus. Candida and Aspergillus species are also of concern. 7. Diagnostic criteria 6.3. Disease transformation About 3% to 10% of the patients undergo transformation of their disease to a more aggressive type, most frequently large-cell lymphoma (Richter's syndrome) [85]. In these cases, fever, weight loss, night sweats, enlarged lymphadenopathy, increased LDH serum levels, anemia, thrombocytopenia, and monoclonal gammopathy are the most common features [86]. Diagnosis is not always easy since the histologjc transformation may be a phenomenon localized in a given organ (e.g., spleen, lymph nodes of a given territory, extralymphatic tissue) rather than a generalized event. The immunologic, cytogenetic, and molecular characteristics of the lymphoma may be the same [87] or different from those of the original CLL [62]. Stage, prior therapy, and response to therapy are not risk factors, but complex cytogenetic abnormalities are often present [86]. In addition, c-myc overexpression, Rb gene deletion and p53 gene mutation have been reported in CLL and small lymphocytic lymphomas showing progression [62, 63]. The prognosis of the transformation to largecell lymphoma is poor, with a median survival of less than one year [86]. Prolymphocytic leukemia (PL) may present either de novo or as a transformation of CLL (CLL/PL) [88-

The diagnostic workup of patients with CLL is shown in Table 1. The National Cancer Institute/Working Group (NCI/WG) [104] and the International Workshop on CLL (IWCLL) [105] have independently proposed criteria for diagnosing CLL (Table 2). In the past, an absolute lymphocyte count of 15 x 109/L was considered to be the threshold for defining CLL, but the NCI/WG [104] and the IWCLL [105] lowered this threshold to 5 x 109/L and 10 x 109/L, respectively. In fact, CLL can be diagnosed when there is an absolute increase in the number of lymphocytes in blood which are morphologically and immunophenotypically consistent with Table I. Chronic lymphocytic leukemia; Diagnosis and staging. Physical examination Blood cell counts (including reticulocytes) and morphology Cell markers (e.g., Smlg, CD5, CD19, CD23, FMC7; CD4, CD8) Biochemical parameters (including LDH and beta-2 microglobulin serum levels) Coombs test Immunoglobulin levels Immunoelectrophoresis Cytogenetics Bone marrow aspirate and biopsy Chest radiography Chest and abdominal CT scans

223 Table 2. Chronic lymphocytic leukemia: Diagnostic criteria. 1. Absolute lymphocytosis in peripheral blood >5 x 107L (NCI/Working Group) > 1 0 x 107L (IWCLL) 2. The majority of lymphocytes should be small and mature in appearance Morphologic subtypes: 2.1 typical or classic CLL: <10% atypical lymphocytes 2.2 atypical or mixed CLL; CLL/PL: prolymphocytes in blood between 11% and 54% 2.2 atypical or mixed: variable proportion of atypical lymphocytes; <10% prolymphocytes 3. Characteristic immunophenotype Smlg +/-, CD5+, CD19+, CD20+, CD23+, FMC7-/+, CD22+/4. Bone marrow infiltration > 30% lymphocytes in bone marrow aspirate, or consistent pattern in bone marrow biopsy References: see text and refs: [104-107] and [110].

the diagnosis. The most common marker profile in CLL is Smlg +/-, CD5+, CD23+, FMC7-, and membrane CD22 + / - [106]. Assessment of the degree of bone marrow infiltration, either by aspirate or biopsy, is more useful for predicting the outcome of the disease and in evaluating response to therapy than for diagnosis. CLL must be differentiated from other lymphoproliferative disorders, particularly splenic lymphoma with villous lymphocytes, lymphoplasmacytic lymphomas, and mantle-cell lymphoma which may resemble CLL both morphologically and immunophenotypically [107-109]. Likewise, cases of 'CD5-' CLL should always raise the possibility of a disease other than CLL. The French-American-British (FAB) Group has divided CLL into different morphologic variants on the basis of the proportion of atypical lymphoid cells in blood: typical, in which most of the lymphocytes are small and mature in appearance with less than 10% of atypical ones; mixed, CLL/prolymphocytoid (CLL/ PL), when the proportion of prolymphocytes in blood is between 11% and 54%; and, a more vaguely defined mixed form, with a variable proportion of atypical lymphocytes but less than 10% prolymphocytes in blood [110]; whether this latter variety truly represents a form of CLL should be reconsidered in the light of the existence of lymphomas which may be confused with CLL, as mentioned above.

marrow histopathology [114-116], blood lymphocyte levels [117, 118], lymphocyte doubling time [119, 120], lymphocyte morphology [90, 91], and cytogenetic abnormalities [39] are reliable predictors of survival (Tables 3-5). These prognostic factors apply to all patients with CLL regardless of their age [121]. Clinical staging systems are the most useful prognostic parameters. The two schemes most widely used are the ones proposed by Rai et al. and Binet et al. (Tables 1 and 2). Other variables may add discriminant power to clinical stages. For example, among patients in early stage, those with diffuse bone marrow histopathologic pattern and/or rapidly increasing lymphocyte counts are likely to progress whereas those with non-diffuse bone marrow involvement and low, stable blood lymphocyte levels tend to have an indolent and non-progressive disease. This is the so-called smoldering CLL, for which diagnostic criteria have been proposed [103,122,123] (Table 6). There are several karyotype abnormalities related to prognosis [39, 40]. Patients with trisomy 12 as their sole abnormality have a shorter survival than those with other single abnormalities, and patients with 13q- anomalies do as well as those with normal karyotype. The complexity of the karyotype and the proportion of cells with clonal abnormalities have important prognostic significance [39]. Cytogenetic abnormalities correlate with clinical stage [40]. Table 3. Chronic lymphocytic leukemia: Rai staging system. Median survival (years) Low-risk

Stage 0 -

lymphocytosis alone

Intermediaterisk

Stage I -

lymphocytosis lymphadenopathy

Stage II - lymphocytosis spleen and/or liver enlargement

5

High-risk

Stage III - lymphocytosis anemia (Hb < 11 g/dL)

2

Stage IV - lymphocytosis platelets < 100 x 10VL

2

9

Table 4. Chronic lymphocytic leukemia: Binet staging system. Median survival (years)

8. Prognosis The median survival of patients with CLL has increased from about 5 years in the early 1970s to about 8 years in recent series. This is due to the larger proportion of patients currently diagnosed when asymptomatic and in an early stage rather than to treatment progress. Some patients have a survival not different from that of the general population while others have a rapidly fatal course. Clinical stages [111-113], bone

>15

Stage A -• no anemia, no thrombocytopenia less than 3 enlarged lymphoid areas"

12

Stage B -• no anemia, no thrombocytopenia three or more enlarged areas

5

Stage C -• anemia (Hb < 10 g/dl) and/or platelets < 100 x 109/L

2

* Lymphoid areas considered are: cervical, axillary, and inguinal lymphadenopathies (whether uni- or bilateral), spleen, and liver.

224 Table 5. Chronic lymphocytic leukemia; Other prognostic factors. Parameter Bone marrow histopathologic pattern Non-diffuse Diffuse Number of lymphocytes in blood <50 x 109/L >50 x 107L Doubling time > 12 months < 12 months Lymphocyte morphology in peripheral blood <5 percent prolymphocytes >5 percent prolymphocytes Cytogenetic abnormalities Normal karyotype Multiple and complex abnormalities

Median survival (years) 10 3-5 6 3-4 10 5 5-6 3-4

tein detection on neoplastic cells have been correlated with resistance to therapy and poor prognosis in some reports [23, 129]. Finally, response to therapy is associated with a better outcome [120]. 9. Treatment

After diagnosis, patients should be followed without treatment for several weeks to establish the stage of the disease and to assess its pace, which is useful in determining whether or not treatment is necessary. 9.1. When to treat?

A number of factors are generally acknowledged to justify therapy in CLL [131, 132]. Almost none of the conditions listed below are found singly but rather N.B.: number of lymphocytes and prolymphocytes in blood as well occur altogether in the same patient. as doubling time behave as continuous variables: the higher the 1. Disease-related symptoms (i.e., weight loss, fever number of cells or the shorter the doubling time, the poorer the without infection, night sweats, weakness). Since prognosis. In most instances, these poor-risk factors are not found alone but systemic symptoms in uncomplicated CLL are accompanying advanced disease. rare, infection or the transformation of CLL to a References: see text and refs: [39,90,92,114-120]. more aggressive lymphoproliferative disorder (Richter's syndrome, CLL/PL) should be exTable 6. Smouldering chronic lymphocytic leukemia: Diagnostic cluded before assuming that these symptoms are criteria. due to the disease. 2. Development of anemia or thrombocytopenia Montserrat et al. (1988) [103] Stage A due to bone marrow infiltration. Non-diffuse bone marrow histopathology 3. Autoimmune hemolytic anemia or thrombocytoHemoglobin > 13 g/dl penia. In such cases, treatment should be initiatBlood lymphocytes <30 x lO'/l ed with corticosteroids alone and cytotoxic Lymphocyte doubling time >12 months agents added only if no response is observed. French Cooperative Group on CLL (1990) [122, 123] 4. Bulky lymphadenopathy and/or splenomegaly A-l causing compressive problems. Stage A Hemoglobin > 12 g/dl 5. High blood lymphocyte count. The threshold for Blood lymphocytes <30 x 109/l initiation of therapy is difficult to establish; A-2 whereas hyperviscosity and/or leukostasis is Stage A very rare in CLL and has been reported only Hemoglobin > 12 g/dl with lymphocyte counts higher than 500 x 109/L Blood lymphocytes <30 x 1071 [133], blood lymphocyte counts above 30 to 50 Lymphocytes in bone marrow aspirate <80% Number of lymphoid areas" involved <2 x 109/L are associated with shorter survival [117, 118]. In the absence of other unfavorable factors * Lymphoid areas considered are: cervical, axillary and inguinal or symptoms related to hyperleukocytosis, treatlymphadenopathy (whether uni- or bilateral), spleen and liver. ment is not indicated unless the blood lymphocyte count is extremely high (e.g., above 500 x 109/L). The prognostic value of the immunophenotype of neoplastic B lymphocytes is unclear, with many studies 6. Rapidly increasing blood lymphocytosis(i.e., yielding inconsistent or even contradictory results. For doubling time of less than 12 months). example, in a large series of patients a high Smlg inten7. Hypogammaglobulinemia with increased suscepsity or low CD23 membrane expression were associattibility to bacterial infections. Infection susceped with poor prognosis [124], whereas in a similar tibility usually correlates with immunoglobuUn study CD23 expression was found to be associated levels. Nevertheless, even in cases responding to with poor-prognosis indicators, namely, higher WBC therapy hypogammaglobulinemia is usually not count, advanced Rai stage, and lymphadenopathy [51]. corrected. LDH, thymidine-kinase, beta-2 microglobulin, CD23, 8. Massive lymphocytic infiltration of bone marrow and CD25 serum levels have also been found to be of (i.e., diffuse histopathologic pattern). This is usuprognostic value in some studies [125-128]. Overally associated with other poor-prognosis facexpression of mdr-1 and mdr-3 genes and p-glycoprotors, particularly advanced clinical stage. >10 5-6

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9. Complex cytogenetic abnormalities. In most cases, cytogenetic abnormalities are associated with advanced clinical stage and other poorprognosis features. 10. Advanced clinical stage. Clinical stage is the most important parameter for deciding upon therapy. Patients in early clinical stage (Binet A, Rai O) usually have a long survival and should not be treated unless the disease progresses. On the other hand, patients with advanced clinical stage (Binet B, C; Rai HI, IV), have a median survival of less than 5 years and should be treated. To summarize, patients with no symptoms, early clinical stage, non-diffuse bone marrow infiltration, and low and stable blood lymphocyte counts have excellent prognoses and should not be treated unless the disease progresses. By contrast, patients with symptoms due to the disease, advanced clinical stage, diffuse bone marrow involvement or high and rapidly increasing blood lymphocyte counts require therapy. 9.2 Treatment modalities 9.2.1. Radiation therapy P, thymic/mediastinic irradiation, extracorporeal irradiation of blood, phototherapy, and total body irradiation (TBI) [134, 135] have all been used with varying, usually moderate success. In randomized studies, TBI has been found not to be superior to chemotherapy [136-138]. Radiation therapy basically has a palliative role (i.e., treatment of large lymph nodes or spleen causing compressive problems). Low-dose radiation over the spleen may be an alternative to splenectomy [139] and has also been used as part of the systemic treatment of the disease [140]. 32

9.2.2. Splenectomy Patients with autoimmune hemolytic anemia who are resistant to corticosteroids or immunosuppressive and cytotoxic therapy may benefit from splenectomy [141— 144]. 9.2.3. Leukapheresis Leukapheresis has been used to decrease high blood lymphocyte counts in patients with CLL not responding to, or not suitable for chemotherapy. Although significant elevations in hemoglobin levels and platelet counts have been observed in some cases, this effect is only transient [145,146]. 9.2.4. Single-agent chemotherapy The most widely used agent to treat CLL has been chlorambucil at doses of 6 to 8 mg daily, 15 to 20 mg/ m2 every two weeks or 30 to 40 mg/m2 monthly. Response rates are between 40% and 60%, but complete remissions are rare [132,147,148]. The combination of chlorambucil and prednisone does not appear to be superior to chlorambucil alone [140,149]. In one study, daily chlorambucil was associated with an increased

risk of solid neoplasms [103]. In another trial, highdose chlorambucil (15 mg) given daily until remission or the advent of unacceptable toxic effects resulted in a remission rate higher than the one achieved with CHOP (cyclophosphamide, doxorubicin, vincristine, prednisone) [150, 151]. Cyclophosphamide is administered at a dose of 2 to 3 mg/kg day orally or 20 mg/kg every 2 to 3 weeks intravenously [132, 149] and is usually employed in cases of poor tolerance to chlorambucil or clorambucil ineffectiveness. Prednisone (30-60 mg/m2 per day, orally) is the treatment of choice in instances of autoimmune hemolytic anemia or thrombocytopenia associated with CLL. 9.2.5. Combination chemotherapy A wide range of combination chemotherapy regimens (e.g., COP [cyclophosphamide, vincristine, prednisone] [152, 153], the M2 protocol [melphalan, cyclophosphamide, BCNU, prednisone] [154], CAP [cyclophosphamide, doxorubicin, prednisone] [155], CMP [cyclophosphamide, melphalan, prednisone] [156], and CHOP [cyclophosphamide, doxorubicin, vincristine, prednisone] [157-159] has been used to treat CLL. Although these regimens induce a relatively large number of responses, they are no better than chlorambucil with respect to survival. For example, in three randomized trials, results with COP were similar to those with chlorambucil with or without prednisone [140, 160, 161] and in another study CMP was not found to be superior to chlorambucil plus prednisone [156]. The French Cooperative Group on CLL reported that a modified CHOP regimen, with doxorubicin at a low dose (25 mg/m2) was a good therapy for patients in advanced stage (Binet C) on the basis of a randomized trial in which the modified CHOP was compared to COP. Nine months after therapy, 25 of 25 CHOP-treated stage C patients were downstaged to stage A whereas only 8 of 30 COP-treated stage C patients entered stage A. In addition, patients treated with CHOP had a projected survival of 52% at 5 years compared to 14% in those receiving COP [157, 162]. However, in randomized trials from other groups, although the response rate with CHOP (both at modified and standard doses) was usually higher than with chlorambucilbased regimens, no advantage in survival was detected [149, 158, 159]. The fact that in these latter trials the higher remission rate achieved with CHOP did not translate into longer survival may be due to selection bias or to the fact that a proportion of patients failing to respond to clorambucil who were switched to CHOP or other regimens eventually responded. 9.3. Newer treatment approaches 9.3.1. Purine analogues Deoxycoformycin (DCF), fludarabine, and 2-chlorodeoxyadenosine (2-CDA) are purine analogues which have demonstrated high activity in CLL and other chronic lymphoproliferative disorders. All these agents

226 sponse and previous therapy is observed: 17% to 74% (0% to 19% CRs) in patients with previous therapy versus 81% to 100% (25% to 37% CRs) in untreated patients. In addition, the less advanced the stage of the disease, the higher the response rate [184]. Approximately two-thirds of the patients who obtained CR still have residual nodules or interstitial aggregates in the bone marrow biopsy and are classified as nodular CR 9.3.1.1 Deoxycoformycin (DCF). DCF is given at a (nCR). The number of prior treatments, hemoglobin, dose of 4 mg/m2 intravenously every week or every two albumin, and creatinine are reported to predict reweeks. Overall, 152 patients, most of them previously sponse [175, 176]. The combined use of fludarabine treated, received DCF. Their response rate is about with prednisone has provided no better results than 25%, with most responses being partial or minor. Life- fludarabine alone [175]. The survival of patients who threatening infections may be observed in up to one- respond to fludarabine regimens as initial therapy correlates with response and its degree [175, 176]. The third of the patients [163-168]. objectives of current studies are to determine the role 9.3.1.2. Fludarabine. Fludarabine has the highest of fludarabine combined with other agents (e.g., intertherapeutic activity ever reported for a single agent in feron, chlorambucil, doxorubicin, cytosine arabinoside, CLL [169-180]. It is usually given at a daily intra- platinum) [188-190] and to develop an orally active venous dose of 25 mg/m2 for 3 to 5 days, every 28 form of the drug. days, for four to six cycles. As with all of the purine analogues, the most imporAs shown in Table 4, the response rate ranges be- tant side effects of fludarabine are myelosuppression tween 17% and 100%. A high correlation between re- and infections. Thus, there is a decrease in the number of CD4+ cells in blood which may facilitate a greater Table 7. Chronic lymphocytic leukemia; Treatment results with susceptibility to infections, including opportunistic purine analogs. ones (e.g., herpes simplex, herpes zoster, Cytomegalovirus, Pneumocystis carinii, Listeriosis, and Legionella), Agent Patients Prior Responses (%) particularly when fludarabine is given in combination Author Rx PR CI CR with prednisone [175, 191]. Some cases of interstitial pneumonia's have also been described [192, 193]. DeDeoxycoformycin spite the lack of formal proof of its effectiveness, the in16 16 0 NA 19 Riddell et al., 1985 [163| 16 25 25 4 16 Greveretal., 1985 [164] creased risk of infection makes the use of prophylactic 29 NA 3 21 NA O'Dywcr ct al., 1988 [165| antibiotics (e.g., trimetroprim-sulfametoxazol) advis39 26 3 23 28 Dillman et al., 1989 [166| able; likewise, prednisone should be omitted from regi27 26 0 19 Hoetal., 1990 [167] 26 17 35 65 17 0 Dearden et al., 1990 [168] mens containing fludarabine. Other potential complications are fever of unknown origin, tumor lysis Fludarabine 14 32 32 3 68 Greveretal., 1988 |169] syndrome [194], and autoimmune hemolyric anemia 42 42 52 NA Puccioetal., 1991 [170] 0 [78,79]. 20 20 20 35 45 Hiddemann et al., 1991 |171] inhibit DNA synthesis in replicating cells, but their mechanisms of action in lymphoid malignancies are basically unknown. DCF is a potent inhibitor of adenosine deaminase. 2-CDA may induce DNA strand breaks. Both fludarabine and 2-CDA appear to activate in vitro cellular apoptosis. The results reported with these drugs are summarized in Table 7.

Sorcnsen et al., 1992 [172| Zinzani et al, 1992 [173] de Rossi et al., 1993b[174| O'Brien etal.; 1993" [175] Keating et al., 1993 [176] Fenchel et al., 1995 [177] Gillisetal., 1994 [178] Zajaetal., 1994 [179) Spanish Scries, 1995 [180] O'Brien etal., 1993" j 174) Keating et al., 1993 |176| Zajaetal., 1994 [179| 2-Chlorodeoxyadenosine Saven et al., 1991 [181| Tallman et al., 1992 [182] Juliusson et al., 1993 [183] Delannoy et al., 1994 [184] Raietal., 1994 [185] O'Brien etal., 1994 [186] Saven etal., 1993 [187]

637 28 22 169 78 56 10 9 56 95 35 8

637 28 22 169 78 56 10 9 56 0 0 0

90 7 18 22 14 28 20

90 7 18

22C 14" 28" 0

4 0 5

27 46 36 12(26)- 14 19 (19)" 36 5 68 10 30 NA NA 7 23 30 {3iy 19 37 (37)' 6 25 75

0 18 NA NA NA NA NA NA 29 NA NA 0

4 0 39 9 7 0 25

NA NA 16 NA NA 20 NA

40 57 28 23 29 2 60

CR - complete remission; PR - partial remission; CI - clinical improvement; NA - not available. * CR with persistent lymphoid nodules in bone marrow (CRn). b Fludarabine plus prednisone. c Six patients previously treated with fludarabine. d Seven patients previously treated with fludarabine.

9.3.1.3. Chlorodeoxyadenosine (2-CDA). 2-CDA is usually administered at a dose of 0.1 mg/kg per day in a continuous infusion for 7 days. The same dose may be given over 2 hours in a bolus infusion, or subcutaneously, for 5 consecutive days [181-187]. Overall, 199 patients treated with 2-CDA have been reported. Responses are better in patients with no prior therapy (85% with 25% CRs) than in previously treated patients (22% to 83% with 0% to 39% CRs). A rapid initial decrease of the blood lymphocyte count after starting 2-CDA predicts good response [183]. 2-CDA also appears effective when given orally [195]. Bone marrow depression, especially thrombocytopenia, is the most important side-effect and may be particularly marked in patients heavily pretreated, particularly those who have received alkylating agents. As with all of the other purine analogues, 2-CDA selectively depletes CD4positive cells from peripheral blood, which may facilitate infections [196]. In a very small series of patients, 2-CDA was found to be useful in patients resistant to

227

fludarabine [197]. However, this concept was not confirmed in subsequent studies [181, 182, 184-186, 198] (Table 8). In summary, purine analogues, particularly fludarabine, are the treatment of choice for CLL patients not responding to conventional therapy. Nevertheless, heavily pretreated patients do not benefit from these agents and may suffer substantial toxic effects. For this reason, purine analogues should be used earlier in the course of the disease, before a truly resistant disease emerges. Although it is likely that in the near future the purine analogues will be the first choice for CLL treatment, the superiority of these drugs over conventional therapy (e.g., chlorambucil, CHOP) should be confirmed in ongoing randomized trials [199, 200]. Preliminary results from these trials are shown in Table 9. Although a trend to a higher response rate in patients treated with fludarabine may be observed, longer follow-up is necessary to determine whether there are differences in survival. Likewise, there are no studies comparing purine analogues. In clinical practice, bone marrow toxicity, tumor lysis syndrome and opportunistic infections should be of concern when using purine analogues, and measures to prevent these complications should be employed. 9.3.2. Allogeneic bone marrow transplantation Experience with allogeneic bone marrow transplantation in CLL is increasing [201-203] (Table 10). The largest series has been collected by the European and International Bone Marrow Transplant Registries [203]. Fifty-four patients who were reported to these groups received an allogeneic bone marrow transplant because of CLL. Their median age was 41 years (range, 21 to 57 years) and the median interval from diagnosis to transplant was 37 months (range, 5 to 130 months). At the time of transplantation, 3 patients were Rai stage 0; 10, stage I; 10, stage II; 7, stage m, and, 22, stage IV. Twenty-one were considered to have chemotherapy-resistant disease. Most of them received pretransplant conditioning with cyclophosphamide and total body irradiation and graft-versus-host disease prophylaxis with cyclosporine and methotrexate. Thirty-eight patients (70%) achieved hematologic remisTable 8. CLL: Response to 2-CDA of patients previously treated with fludarabine. Author

No. cases

4 Juliusson et al., 1992 [197] Saven et al., 1993 [198] 14 Delannoy et al., 1994 [184] 6 Rai et al, 1994 [185] 7 O'Brien et al., 1994 [186] 28 Total

59

CR 1 _ _ _ 0

PR 3 2

1 5 (2%) (8%)

MR _ 6 _ 20 26 (44%)

sion, and 24 (44%) are alive at a median 27 months (range, 5 to 80) post-transplant. Their three-year survival probability was 46% (95% CI: 32%-60%). Thirty patients died of progressive leukemia (n = 5; 9%) or treatment-related complications ( n - 2 5 ; 46%). This latter result contrasts with the 10% mortality in recent studies from single institutions [201, 202], with the difference probably being due to patient selection criteria. Cytofluorometry and/or molecular biology techniques have shown some of the remissions achieved after transplant to be molecular, with no evidence of residual disease. On the other hand, remissions are more frequent in patients with pre-transplant stable and responding disease than in those with progressive disease. Relapses, sometimes as late as 4 years after transplant, may be observed. Table 9. Fludarabine in CLL therapy: Preliminary results of randomized trials. Study

Treatment

No. pts

27 (46%)

CR - complete response; PR - partial response; MR - minor response (less than PR); NR - no response.

Prior Rx

Responses CR

Hiddemann et al. [199]

Fluda CAP

53 52

Fluda CAP

53 50

French CLL Group [200)

Fluda CAP CHOP

54 66 54

Fluda CAP CHOP

24 24 25

No No

70% 58%

Yes Yes

45% 26%

B B B

No No No

48% 14% 32%

C C C

No No No

58% 54% 67%

B,C B,C

PR

40% 64% 45%

Table 10. Chronic lymphocytic leukemia: Bone marrow transplants. Study

No. pts

Rabinowe et al. 1993 [201] Allogeneic 8 Autologous Khouri et al., 1994 [202] Allogeneicb Autologous

12

11 11

NR _ 8 6 7 6

Stage

IBMTR/EBMT, 1995 [203] Allogeneic 54 EBMT, 1994 [204] Autologous

11

Age

40 (31-54) 45 (27-54) 42 (25-55) 59 (37-66)

CR

CCR'

6

6 (6-18) 10 (6-31)

10

7 6

7 (6-36) 3 (4-29)

BMTrelated deaths

1

1 1

41 (21-57)

38

25 24 (5-80)

50 (41-59)

NR

8 (3-24)

* Continuous complete response: number, follow-up range in months is shown in parentheses. b One syngeneic transplant (alive in CCR at 19 months after transplantation). NR - not reported.

228

9.3.3. Autotransplants Autotransplants have been reported in 34 CLL patients [201, 202, 204] (Table 10), all of whom had advanced disease before transplantation, and all of whom received cyclophosphamide and TBI as the conditioning regimen. In about half of the cases the bone marrow graft was purged with monoclonal antibodies against B-cells (CD19, CD20, CD10, B5), and in two cases, blood progenitor cells were used [204, 205]. With a median follow-up of less than two years, 21 patients remain alive in CR. 9.3.4. Bio therapy New approaches to treating CLL include monoclonal antibodies, interferon, and cytokines. Monoclonal antibodies (MoAb), either alone (e.g., CAMPATH) or conjugated with toxins (B4-blocked ricin), cytotoxic agents or radioisotopes (1131) are being actively investigated; the response is usually partial and transient. MoAb might be useful in eliminating residual disease in patients achieving good response after chemotherapy [206-209]. Interferon (IFN) has no effect in patients with advanced and heavily pretreated disease. In contrast, IFN is more effective in patients with early disease and no prior therapy, although no CR is obtained [210-215]. IFN might further improve responses achieved with chemotherapy [214, 215]. The combined use of IFN and cytotoxic agents [190, 214] deserves investigation in large clinical trials. Interleukins (e.g., IL-2, IL-4, IL-6) are also under study. IL-2 has proven to have limited clinical activity with considerable toxicity [216-218]. 9.4. Therapy for systemic complications Hypogammaglobulinemia is frequent in CLL, particularly in patients with advanced disease and is the major reason why these patients have a high incidence of infections which are the first cause of death. In a placebo-controlled randomized study, 400 mg per kg of immunoglobulin given intravenously at 3-week intervals for one year was found to be effective and well tolerated [219]. Prophylactic immunoglobulin, however, had no impact on survival and cost/benefit considerations render the routine use of immunoglobulin in all patients with hypogammaglobulinemia debatable [220]. Recent studies, however, suggest that lower doses of immunoglobulin (10 g every three weeks; 250 mg/kg every four weeks) might be as effective as higher doses, which has important practical and economic implications [221, 222]. The role of oral antibiotics as prophylaxis has not been formally investigated. As in other settings, recombinant hemopoietic growth factors may overcome treatment-related neutropenia [223]. Erythropoietin may be useful for treating anemia unresponsive to other measures [224].

10. How to treat 10.1. Patients with early and stable disease Treatment of patients in an early stage (Binet A, Rai O) has resulted in a delay in the rate of progression of the disease but no survival benefit [103, 140, 225, 226] and in one study a higher incidence of second neoplasms [103]. These patients should not be treated unless the disease progresses. 10.2. Patients with advanced clinical stage due to high tumor burden and bone marrow failure Chlorambucil (e.g., 0.4-0.8 mg/kg orally every two weeks) has been, and probably still is, the treatment of choice for most of these patients. Although patients treated with CHOP achieve a higher response rate, this does not translate into longer survival. Front-line treatment with CHOP is indicated in the rare rumoral forms of the disease with compressive problems. In such cases, local radiotherapy may also be useful. Patients failing front-line therapy should be treated with combination chemotherapy or fludarabine. At present, frontline therapy with fludarabine or other purine analogues should only be considered within the framework of clinical trials. 10.3 Patients with cytopenias due to immune mechanism (e.g., stage C (111, IV) immune) These patients should be initially treated with corticosteroids (e.g., prednisone 40 mg/m2 daily given orally), with cytotoxic agents (e.g., cyclophosphamide 50 to 100 mg orally per day) being added only in instances of no response after 4 to 6 weeks of treatment. In patients with autoimmune hemolytic anemia not responding to, or difficult to control with, corticosteroids plus cytotoxic agents, high-dose immunoglobulin may be tried. Some of these patients, however, eventually require splenectomy or low-dose spleen radiation. Another form of apparently advanced CLL not due to bone marrow infiltration is pure red-cell aplasia (PRCA) which occasionally may be associated with CLL. Excellent treatment results have been reported with cyclosporine with or without prednisone [227]. 10.4. Patients with hypersplenism There are cases of CLL in which the anemia and/or the thrombocytopenia are entirely or partially due to hypersplenism. In such cases, splenectomy or low-dose radiotherapy over the spleen may be more effective than chemotherapy. 10.5. Younger patients Indication for treatment in younger patients with CLL should be on the basis of prognostic factors. Age alone

229 should not be the criterion for initiating therapy in in these patients, but the goal of treatment, once resolved upon, should be to eradicate the disease. To that end, front-line therapy with either CHOP or fludarabine is more likely to induce CR than chlorambucil. The possibility of performing an allogeneic bone marrow transplant should be considered in any younger patient with CLL for whom an HLA histocompatible donor is available. Under some circumstances (e.g., non-availability of an HLA histocompatible donor, advanced age), and provided CR has been achieved with standard therapy, autografts may be an alternative to allografts. When CR is achieved, bone marrow or peripheral blood cells may be collected to be used either immediately or upon disease progression. Transplants from CD34+ cells also warrant investigation.

11. Evaluation of response to treatment

To standardize response criteria, the IWCLL and the NCI/WG independently defined complete remission (CR), partial response (PR), stable disease (SD), and progressive disease (PD), as shown in Table 11 [104, 105]. CR is difficult to assess with conventional clinical and hematological criteria. More sensitive techniques such as analysis of CD19/CD5-positive cells, kappa/ lambda clonal excess by cytofluorometry or gene rearrangement by PCR are required for detecting residual malignant cells [228].

12. Treatment goals

The survival of some CLL patients is not affected by the disease. Treatment should therefore not be decided upon without taking risk-factors into account. On the other hand, response to treatment is associated with in-

creased survival, although the disease is rarely, if ever, cured. With this background, in selected patients (e.g., younger people needing therapy) every effort should be made to achieve CR. For the majority of the patients, however, the most reasonable therapeutic goals are: a) symptom palliation, b) correction of cytopenias, and c) switching of the disease to a less advanced stage. The benefits of achieving CR should be balanced with the risks associated with therapy, with the most reasonable aim of treatment being to achieve the greatest possible response with acceptable toxicity.

Acknowledgements

This work was supported in part by Grants 93/0303 and 91/213 from 'Fondo de Investigaciones Sanitarias de la Seguridad Social' (FISS) and SAL 89/0963 from 'Comision Interministerial de Ciencia y Tecnologia', Spain.

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Table 11. Chronic lymphocytic leukemia: Response criteria. Response

IWCLL criteria

NCI criteria

CR

No evidence of disease

Absence of lymphadenopathy, hepatomegaly, splenomegaly or constitutional symptoms. Normal blood count: neutrophils > 1.5 x 109/L, platelets > 100 x 109/L, Hb > 11 g/dL, lymphocytes <4.0 x 109/L, BM biopsy with normal cellularity and lymphocytes < 30%.

PR

Change from stage C to stage A or B; from stage B to A

50% reduction in blood lymphocytes and 50% reduction in lymphadenopathy and/or 50% reduction in splenomegaly and/or hepatomegaly. Neutrophils > 1.5 x lO'/L or 50% improvement over baseline; platelets > 100 x 109/L or 50% improvement over baseline; Hb > 11.0 g/dL (not supported by transfusion) or 50% improvement over baseline.

SD

No change in the stage of the disease

No CR, PR, or PD

PD

Change from stage A disease to stage B or C, or from stage B to C

At least one of the following: >50% increase in the size of at least two lymph nodes or new palpable lymph nodes; >50% increase of splenomegaly or hepatomegaly or appearance if there was no transformation to a more aggressive histology, Richter or prolymphocytic leukemia; >50% increase in the absolute number of circulating lymphocytes.

CR - complete response; PR - partial response; SD - stable disease; PD - progressive disease.

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231

52. 53. 54. 55.

56.

57.

58.

59.

60. 61. 62.

63.

64. 65.

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68. 69. 70.

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