Chronic lymphocytic leukaemia: a short overview

symposium article Annals of Oncology 19 (Supplement 7): vii320 vii325, 2008 doi:10.1093/annonc/mdn460 Chronic lymphocytic leukaemia: a short overview E. Montserrat & C. Moreno Institute of Hematology and Oncology, Department of Hematology, Hospital Clinic, IDIBAPS, University of Barcelona, Barcelona, Spain symposium article introduction Chronic lymphocytic leukaemia (CLL) is characterized by the relentless accumulation of monoclonal CD5+ B lymphocytes in blood, bone marrow and lymphoid tissues. The disease predominates in older individual and its incidence increases with age. The median survival of patients with CLL is around 10 years but the individual prognosis is highly variable. In spite of some advances in its therapy CLL remains incurable. epidemiology The median age of patients at diagnosis is about 70 years [1 3]. The incidence of CLL is about 5 cases per 100 000 people per year, increasing with age [4]. Whereas CLL is the most frequent form of leukaemia in Western countries, where it accounts for 30% of all leukaemias, it only constitutes 10% of all leukaemias in Asian populations [5]. In most series, CLL is more frequent in males than in females. aetiology The aetiology of CLL is unknown. The potential relationship between inflammatory and autoimmune conditions and CLL is conceptually appealing and is the subject of active investigation [6, 7]. Familial cases of CLL strongly suggest a genetic basis for this disease [8]. In about 5% of first-degree relatives of patients with CLL a population of B cells immunophenotypically identical to that of CLL can be detected in peripheral blood. The clinical significance of this fact, which can also be observed in 2 3% of normal subjects, is unclear [8 10]. This situation is known as monclonal B cell lymphocytosis (MBL) and should not be confounded with CLL. biology The CD5+ B cells from which CLL arise constitute a small subpopulation of B lymphocytes with a characteristic immunophenotype, i.e. SmIg (weak), CD5+, CD19+, CD20 (weak) and CD23+. Although the majority of these cells are arrested in the G 0 phase of the cell cycle, between 0.1% and 1% of B CLL lymphocytes are actively duplicating every day [11]. B CLL lymphocytes express anti-apoptotic BCL-2 proteins whereas the pro-apoptotic BCL-X proteins are decreased. This, together with the interaction of neoplastic and stromal cells through a number of chemokines, leads to the accumulation of leukaemic cells [3, 12]. Cytogenetic abnormalities can be detected by interphase fluorescence in situ hybridization (FISH) in up to 90% of cases. The most frequent genetic abnormalities are 13q, 11q, +12, 17p and 6q [13 15]. Although chromosomal translocations have been reported, their real incidence and clinical meaning need further investigation. In cases of disease progression, del (11q), overexpression of c-myc, deletions of the Rb1 gene and mutations of the p53 tumour-suppressor gene have been reported [16, 17]. No genes have been consistently associated with either the cause or the pathogenesis of CLL. However, the 13q14 region involves a number of genes such as mir15a, mir16-1 and Leu2 whose downregulation increases the expression of antiapoptotic BCL-2 protein [18 21]. For many years it was considered that IGVH genes were always unmutated in CLL. One of the most important breakthroughs in the biology of CLL in the last few years has been the demonstration that, contradicting this paradigm, IGVH genes are mutated in 30 40% of CLL cases. Consequently, two forms of CLL are currently accepted (i.e. IGVH-mutated and IGVH-unmutated) [22, 23]. As discussed later, these two forms have different clinical behaviour. Immune disturbances are frequent. Hypogammaglobulinaemia may be found in up to 60% of patients and is considered the major cause of infections in CLL [24]. In addition, a positive direct antiglobulin test (DAT) or clinically overt autoimmune haemolytic anaemia (AHA) can be observed during the course of the disease in 2 35% of patients; immune thrombocytopenia and pure red cell aplasia can also be present, although they are much less frequent than AHA [25, 26]. The mechanisms of immune defects in CLL have not been completely elucidated, although several explanations have been proposed, including abnormalities in T regulatory cells and in non-neoplastic Bcells[27,28]. clinical features About 70% of cases are diagnosed in asymptomatic individuals. In patients with symptoms the most frequent finding is lymphadenopathy. Fever, night sweats or weight loss are not frequent and should raise the possibility of some clinical complication, including disease transformation into a more aggressive disease. The patient may refer with a history of repeated infections or autoimmune haemolytic anaemia in the preceding months or even years. ª The Author 2008. Published by Oxford University Press on behalf of the European Society for Medical Oncology. All rights reserved. For permissions, please email: journals.permissions@oxfordjournals.org

Annals of Oncology In about 10% of patients the disease undergoes transformation into a more aggressive tumour, most commonly diffuse large B-cell lymphoma (DLBCL), a situation also known as Richter s syndrome [29, 30]. In such cases, fever, weight loss, night sweats, enlarged lymphadenopathy, increased lactate dehydrogenase (LDH) serum levels, anaemia, hypercalcaemia, thrombocytopenia and monoclonal gammopathy are the most frequent features [31]. Positron emission tomography/computed tomography is useful to detect disease sites with likely disease transformation [32]. The DLBCL arising in this context may be clonally related or unrelated to the preceding CLL [33]. In some cases disease transformation is driven by Epstein Barr virus (EBV) infection facilitated by immunosuppressive treatments [34]. Infections are frequent. Common bacteria such as Streptococcus pneumoniae, Staphylococcus and Haemophilus influenzae cause most of the infections. Herpes zoster is also frequent. The use of new treatment agents with a highly immunosuppressive effect has led to the observation of infections due to Legionella pneumophila, Pneumocystis carinii, Listeria monocytogenes and cytomegalovirus (CMV). Candida and Aspergillus species are also of concern [35, 36]. Patients with CLL have an increased risk of suffering second cancers [37, 38]. In a study based on data from population-based cancer registries, the observed/expected ratio was 1.20, with an increased risk for Kaposi sarcoma, malignant melanoma, larynx cancers, lung cancer and also bladder and gastric cancer in men. No relationship was found between the characteristics of the disease and its treatment and the incidence of second cancers [39]. diagnosis The diagnosis of CLL requires the sustained detection of more than 5000 monoclonal B lymphocytes in peripheral blood with a characteristic immunophenotype: SmIg (weak), CD5+, CD19+, CD20 (weak) and CD23+. Individuals with fewer than 5000 monoclonal B lymphocytes and lymphadenopathy, splenomegaly, hepatomegaly or cytopenias due to bone marrow infiltration are considered to have CLL, whereas those without those features are categorized as MBL [40]. The natural history of subjects with MBL is not well known. Although some cases of MBL can evolve into CLL, it is worth emphasizing that the diagnosis of MBL is not equivalent to that of CLL. prognosis symposium article The prognosis of patients with CLL is extremely variable. The overall median survival is about 10 years, but besides patients whose disease has an indolent course and who have a survival no different from that of the general population, there are others who have a rapidly evolving and fatal course. Clinical staging systems are the basis for assessing prognosis in patients with CLL [41, 42]. Patients with low-risk disease (Rai 0; Binet A) have a median survival that exceeds 10 years, those with intermediate-risk disease (Rai I, II; Binet B) have a median survival of 5 7 years and patients with high-risk disease have a median life expectancy of usually less than 3 4 years (Table 1). Besides clinical stages, many other prognostic factors have been proposed, the most important of which are shown in Table 2. The correlation of the mutational status of IGVH genes with the clinical outcome has signified important progress in the understanding of the natural history of CLL [43, 44]. About 30 40% of patients with CLL present in their leukaemic cells somatic hypermutations in the rearranged variable regions of the immunoglobulin heavy chains (IGVH). Patients with IGVH-unmutated genes (unmutated-cll) tend to have a more malignant condition, including evidence of advanced, progressive disease, atypical peripheral blood cell morphology, adverse cytogenetic features (i.e. 11q, 17p ), clonal evolution and resistance to therapy than those with mutated IGVH genes (mutated-cll) [45]. The prognostic significance of IGVH mutations is independent from that of clinical stages and cytogenetic abnormalities, particularly in patients with early stage disease [46]. There is, however, an exception to this rule, and that is the expression of the immunoglobulin variable region gene V3-21, which is associated with an inferior outcome independently of the IGVH mutational status [47, 48]. Determining IGVH mutations still requires specific equipment for DNA sequencing, and is time-consuming and relatively expensive. Because of this, many attempts have been made to identify a marker that could be useful as a surrogate or to replace the study of IGVH mutations. CD38 expression on leukaemic lymphocytes was the first marker that was correlated with IGVH mutations [44]. Eventually, however, it was found that the relationship is not absolute and these two markers are currently considered as independent prognostic factors [49, 50]. Recently, it has been found that CD38 and CD49d are consistently co-expressed on CLL cells [51]. CD49d is an Table 1. Rai and Binet staging system for chronic lymphocytic leukaemia Binet stages Rai stages Median survival Low risk A Hb 10, platelets 100 000, 2 sites involved a 0 Lymphocytosis in blood and bone marrow > 10 years Intermediate risk B Hb 10, platelets 100 000, 3 sites involved I Lymphocytosis + lymphadenopathy 5-7 years II Lymphocytocis + splenomegaly and/or hepatomegaly High risk C Hb < 10, or platelets < 100 000 III Lymphocytosis + Hb < 11.0 < 3-4 years IV Lymphocytosis + platelets < 100 000 a Sites involved are liver, spleen and lymph nodes (either unilateral or bilateral) in inguinal, axillary and cervical regions. Volume 19 Supplement 7 September 2008 doi:10.1093/annonc/mdn460 vii321

symposium article Table 2. Chronic lymphocytic leukaemia: prognostic markers Classical prognostic markers Biological prognostic markers Treatment related FISH, fluorescence in situ hybridization. Clinical stages Blood lymphocyte count Lymphocyte morphology in peripheral blood Blood lymphocyte doubling time Degree of bone marrow infiltration (aspirate/biopsy) Serum markers: thymidine kinase b 2 -microglobulin soluble CD23 IGVH mutational status V3-21 gene usage FISH cytogenetics: low risk: normal, 13q ; high risk: +12, 11q, 17p, complex karyotype CD38 expression ZAP-70 expression Response to therapy (minimal residual disease status after therapy) adhesion molecule variably expressed in CLL. Interestingly, a CD49d cut-off of 30% discriminates two groups of patients with different prognosis [52]. This is an interesting marker deserving further investigation. A small number of genes allow us to separate mutated- and unmutated-cll, the most specific of these genes encodes for a 70-kDa zeta-associated protein (ZAP-70). The vast majority of mutated cases are ZAP-70 negative whereas unmutated forms are ZAP-70 positive [53]. In general, there is a good correlation between ZAP-70 expression and IGVH mutations and, not surprisingly, ZAP-70 has a prognostic value by itself [54 57]. The reasons for discrepant cases, which range from 8 25%, may depend on patient selection and methodological variables. Krober et al. [58] have shown that unmutated CLL cases with high ZAP-70 expression frequently present other biological features conveying poor prognosis such as 17p, 11q or V3-21 expression. Rassenti et al. [56] analysed the relative merits of IGVH mutational status and CD38 expression in predicting disease outcome. In that study, ZAP-70 was more useful than IGVH mutations as a predictor of disease progression as measured by the time interval between diagnosis and treatment necessity. Interestingly, among ZAP-70-negative patients IGVH mutational status separated patients with different times to treatment. ZAP-70 and CD38 may provide complementary prognostic information. Patients who express both markers seem to have the poorest prognosis, whereas those in whom neither of these markers is present have good outcome, and discordant cases would fall in a intermediate-risk category [59 61]. Response to therapy is the most important prognostic factor for survival. Predicting subsets of patients who will respond to a given therapy would therefore be useful to avoid unnecessary treatment toxicity, to prevent the emergence of clones resistant to treatment and to select targeted therapies [62 66]. In this regard, abnormalities of 17p reflecting alterations of P53 convey resistance to fludarabine. The power of other markers such as IGVH mutations, ZAP-70 or CD38 expression to predict response to therapy has not been fully investigated in large series of homogeneously treated patients, although there is some indication that the overall reponse rate, and particularly the response duration, might be worse in patients expressing these markers. Recently it has been reported that the presence of unmutated IGVH genes correlates with shorter duration of remission in patients treated with fludarabine, cyclophosphamide and rituximab (FCR)-based therapy, although no differences in survival rates could be observed between unmutated and mutated CLL [67]. treatment Therapy is only indicated when any of the following features is present: (i) Fever without evidence of infection, extreme fatigue, night sweats, weight loss. (ii) Increasing anaemia or thrombocytopenia due to bone marrow failure. (iii) Bulky or progressive lymphadenopathy; massive or progressive splenomegaly. (iv) Autoimmune cytopenias not responsive to corticosteroids. (v) Rapidly rising lymphocyte count in peripheral blood (doubling time <6 months). A marked hypogammaglobulinaemia or increased WBC counts, in the absence of any of the above criteria, are not sufficient to initiate treatment [40]. treatment approaches Annals of Oncology Treatment of patients in early stage (Binet A; Rai 0) has resulted in a delay in the rate of disease progression but no survival benefit [68, 69]. Studies aimed at determining whether patients with early stage but unfavourable markers (e.g. high ZAP-70 expression, increased thymidine-kinase serum levels, rapid doubling time, poor cytogenetics) might benefit from early intervention are under way. A proportion of patients in intermediate stage (Rai I and II; Binet B) have indolent disease; these patients may be followed with no therapy, like those in early stage. However, the majority of patients with intermediate stage and virtually all patients with advanced stage disease (Rai III and IV; Binet C) due to bone marrow infiltration require therapy. Over the last two decades, chlorambucil has been the treatment of choice. The number of complete responses (CR) obtained with chlorambucil is low (10%) and, besides palliation of symptoms, it is doubtful that it has any impact on the natural history of the disease. Because of this, chorambucil is usually given to patients who are not likely to tolerate more intensive therapies. Purine analogues, particularly fludarabine, are the most effective agents for treating CLL. Treatment with fludarabine vii322 Montserrat & Moreno Volume 19 Supplement 7 September 2008

Annals of Oncology results in a much higher CR rate than chlorambucil or alkylating based chemotherapies (20 40% vs 10%) and a longer disease-free interval, although survival is not prolonged [70 72]. The efficacy of fludarabine is improved by combining it with other agents (e.g. cyclophosphamide, mitoxantrone, rituximab) [62, 63, 65, 73, 74]. There are already data indicating that the combination of fludarabine, cyclophosphamide and rituximab (FCR) not only results in a superior response rate than fludarabine or fludarabine plus cyclophosphamide but a longer freedom from progression and, perhaps, a longer survival [62, 63]. Importantly, genetic abnormalities such as 17p and 11q, pointing at defects in P53 and ATM genes, convey poor response to standard therapy. Alemtuzumab (Campath 1H) is a humanized anti-cd52 monoclonal antibody that results in responses of 40 89%, including CR of 2 50%. Responses are better in patients without prior therapy [75, 76]. Toxicity includes rigors, chills, fever, immunosuppression and lymphocytopenia. Opportunistic infections can be observed. Reactivation of CMV is a problem that deserves monitoring and pre-emptive therapy [75]. Responses to alemtuzumab vary for different disease sites, being higher in peripheral blood and bone marrow than in lymph nodes [77]. Interestingly, alemtuzumab may induce responses in patients who are refractory to fludarabine, this indicating that well-known mechanisms of resistance to the drug (e.g. p53 mutations) can be overcome by the monoclonal antibody [78]. Alemtuzumab is being combined with high-dose corticosteroids or fludarabine in some studies [79, 80]. An increasing number of subjects with CLL are being offered stem-cell transplants [81 84]. The necessary condition for the success of an autologous transplant is to achieve CR prior to the procedure, which is extremely unlikely in patients refractory to the newer chemo or chemo-immunotherapy regimens [81]. Moreover, unfavourable prognostic factors for response to conventional chemotherapy (e.g. del(17p), del(11q), unmutated IGVH genes) also predict poor results with autologous transplants [85]. Autologous transplants do not cure CLL but might prolong survival in extremely selected patients (i.e. those sensitive to chemotherapy, without unfavourable prognostic factors and who are transplanted early in the course of the disease) [86]. A problem with autotransplants is the risk of secondary myelodysplasia/acute myeloid leukaemia, particularly in patients conditioned with regimens including total body irradiation. In contrast, allogeneic transplants can cure about 40% of the patients, but at the cost of a high toxicity and mortality [82, 87]. Because of this and the advanced age of most patients with CLL, reduced-intensity conditioning regimens are employed in most patients with CLL requiring a transplant; results are better in patients still responding to therapy [84, 88]. The possibility of performing an allogeneic transplant should be considered in any patient with refractory CLL, an acceptable clinical condition, and willing to accept the risk of the procedure. Stem cell transplants should not be performed outside clinical trials [89]. acknowledgements This work has been performed thanks in part to Redes Temáticas de Cáncer (RETICS 2006, RD06/0020/0051), José Carreras International Foundation (EM/07 and CR/07) and American/European Alliance for CLL (CLL Global Foundation) and Fundación Farreras Valenti. disclosures No significant relationships references symposium article 1. Müller-Hermelink HK, Catovsky D, Montserrat E, Harris NL. Chronic lymphocytic leukaemia/small lymphocytic lymphoma. In: Jaffe ES, Harris NL, Stein H, Vardiman JW (eds): WHO Classification of Tumors Tumors of Haematopoietic and Lymphoid Tissues. Lyon: IARC Press 2001; 127 130. 2. Rozman C, Montserrat E. Chronic lymphocytic leukemia. N Engl J Med 1995; 333: 1052 1057. 3. 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