Diagnosis of CLL  

Dr Samir Agrawal MBChB, Consultant Haematologist, St Bartholomew's Hospital (Barts), London

The following article has been written exclusively for the CLL Support Association. It is aimed at patients and their carers and assumes almost no prior medical knowledge. 

Introduction

A few words about chronic lymphocytic leukaemia (CLL) are necessary in order to explain the diagnostic process. CLL is a malignancy of B lymphocytes, one of several types of cells of the immune system. Disorders of the other lymphocytes also occur – T cells and natural killer (NK) cells – but are much rarer. While CLL is the commonest adult leukaemia in the western world, there are many other types of malignancy of B lymphocytes, globally referred to as B-cell lymphoproliferative disorders (B-LPD).

As suggested by the term ‘leukaemia’, CLL is a disease characterised by an increase in the abnormal B-cells in the blood and bone marrow (BM). Rarely, CLL cells may not be detected in the blood (or BM), but only found in enlarged glands – lymph nodes (see below) – which is a form of CLL localised to the tissues and often referred to as small lymphocytic lymphoma (SLL). SLL is a subset of CLL and should be treated as any other case of CLL.

An increase in B lymphocytes in the blood – a ‘lymphocytosis’ – indicates a B-LPD, however, infections may also lead to a lymphocytosis, usually of T or NK-cells, but also of B-cells. Most commonly, viral infections – which may or may not cause clinical symptoms – are the cause of a lymphocytosis, which resolves spontaneously with time. 

Diagnosis

Clinical features

The clinical presentation of CLL is entirely non-specific, with all the findings also seen in other B-LPD, as well as other malignant and non-malignant conditions. In fact, the commonest presentation of CLL today is as an ‘incidental’ finding on a routine blood test (full blood count) performed for some other reason, for example, prior to an operation or as part of a health check. In other words, the patients had no idea they were unwell! If patients do present because of the CLL itself, this may include: enlarged glands, which are called lymph nodes, in the neck, armpits, abdomen and groin (one or more areas may be affected); recurrent infections (related to the poorly functioning immune system in CLL); weight loss, fevers (not related to infection) and drenching night sweats – globally referred to as ‘B’ symptoms; excessive fatigue; a sensation of fullness in the abdomen (usually due to an enlarged spleen)…

Full Blood Count

The full blood count (FBC) is one of the key tests in the diagnostic process and is the first step. The widespread availability of FBC analysers explains the increasing detection of CLL as an ‘incidental’ finding. The FBC reveals the increase in the lymphocytes in the blood – but cannot tell whether they are B-, T- or NK-cells. The other important information is the level of the haemoglobin (Hb) and platelets, as if they are low, this suggests bone marrow failure (usually due to the CLL) and is an indication for treatment.

From the FBC sample, a drop of blood is spread on a glass slide, stained with dyes and examined under a microscope. This allows the appearance of the cells to be  determined – CLL cells have a characteristic ‘clumped chromatin’ (looks like cracked mud! Figure 1) pattern to the nucleus (the central dark part of the cell) and from this a confident diagnosis of CLL can be made. BUT, some cases of CLL do not have this typical picture and equally other conditions can mimic CLL.

Immunophenotyping

Having identified the lymphocytosis from the FBC, the next step is to determine the nature of the cells – B-, T- or NK-cells – in the blood. This is easily established by immunophenotyping: reagents (antibodies) label the cells only if they are B-cells and emit light, which is then detected by a machine called a flow cytometer. In addition to proving the cells are B-cells, immunophenotyping produces a profile of the different molecules expressed on the surface of the B-cells and establish the typical pattern seen in CLL. This requires a panel of reagents, as no single marker can define a CLL cell, and the British Society of Haematology has published guidelines on this (see website details). It should be noted that different, but similar, panels may also be used to arrive at the same final conclusion. To assist in the diagnostic process with a panel of markers, Table 1 shows the five markers that constitute the so-called ‘CLL score’. A high CLL score of 4 or 5 is almost always proof of CLL. The diagnosis becomes less certain as the CLL score decreases.

 Table 1. Modified CLL scoring system

(A score of 4 or 5/5 is strongly supportive of a diagnosis of CLL. Reference: Moreau EJ, Matutes E, A’Hern RP, et al (1997) Improvement of the chronic lymphocytic leukaemia scoring system with the monoclonal antibody SD8 (CD79b). Am J of Clin Pathol; 108: 378-382)

Bone marrow

A bone marrow test does not have any role in the diagnosis of CLL. In general, a bone marrow biopsy is only performed if: treatment is to be started– as a baseline to then assess response; the platelet count is low and suggestive of idiopathic thrombocytopenia (ITP); anaemia is present and the blood film and Coomb’s test (or DAT) suggest autoimmune haemolytic anaemia (AIHA). Both ITP and AIHA are phenomena where the body destroys its own platelets and red cells, respectively, and such findings are relatively common in CLL.

Radiological imaging

Ultrasound and CT scanning allow more accurate documentation of the enlarged lymph nodes, liver and spleen – all possible features of CLL - than the physical examination. However, these tests do not provide diagnostic information and are usually only necessary as part of the work-up prior to instituting treatment.

Lymph node biopsy

A biopsy of an enlarged lymph node is not routinely performed in CLL, as the diagnosis is usually clear from analysis of the blood. In the majority of cases, a definitive diagnosis of CLL can be made from the FBC and immunophenotyping. However, if the cells in the blood are not typical under the microscope and the CLL score is 3 or less, looking at the cells in a biopsy specimen under a microscope can be useful in definitively making a diagnosis – whether CLL or not.

Other tests at presentation

Other tests that can be performed at diagnosis are usually for prognostic purposes – see ‘Prognostic Factors’ also in this section on the website). Cytogenetic analysis (looking at the chromosomes in the cells) can also help in the diagnostic process in difficult cases. A number of cytogenetic changes are commonly detected in CLL and if present can support the diagnosis of CLL (as well as give prognostic information). Perhaps the most important role of cytogenetics in diagnosis is to specifically exclude mantle cell lymphoma (MCL) - which is generally a more aggressive disease than CLL. There can be significant diagnostic overlap between MCL and CLL, with either disease looking like the other, both under the microscope and by immunophenotyping!

Another test used for prognosis – ZAP70 – can also provide diagnostic information. At Barts and the London, we have been routinely performing ZAP70 measurement for the last 3 years in all cases of B-LPD at diagnosis. If positive, our data indicates that this is highly supportive of a diagnosis of CLL, as opposed to any other B-LPD.

Summary

In general, the diagnosis of CLL is straightforward and can be made confidently by a combination of the FBC, the appearances of blood cells under the microscope and definitively established by immunophenotyping. However, in difficult cases, a lymph node biopsy and cytogenetic analysis of blood or BM cells can help establish a definitive diagnosis. The commonest diagnostic confusion is with MCL and it is important to be aware of the overlap with CLL – with cytogenetics allowing separation of the two conditions. Diagnostic accuracy is essential, as all the prognostic markers available in CLL are only meaningful if the diagnosis really is CLL!

Future directions

For difficult cases, improved diagnostic tools are still required. At Barts and the London, for the last 4 years, the immunophenotyping panel has routinely included a reagent to detect a novel marker on CLL cells, CD160. The addition of this reagent has simplified the diagnosis of CLL from other B-LPD and, perhaps more importantly, particularly aids in the diagnosis of difficult cases, which have a low CLL score. Out of 343 cases of CLL, 339 were positive for CD160. Twenty-eight cases had a CLL score of less than or equal to 3 (ie, not definitely CLL), of which 27 were CD160 positive. This reagent will allow a reduction in the size of the panel for immunophenotyping, making the process simpler, cheaper and faster, as well as increasing diagnostic accuracy.

Ideally, the diagnostic process should be simple, inexpensive, quick and also incorporate the prognostic markers. Building on the established immunophenotypic prognostic markers – CD38 and ZAP70 – will allow a better idea of prognosis at the time of diagnosis. This then needs to be incorporated with the other prognostic markers that do not impact on diagnosis.

December 2006

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