The following material was provided by The Binding Site, LTD, which is the company who developed and markets the FreeLite Test.
Multiple myeloma and related B Cell lineage tumours pose many diagnostic problems both for clinicians and laboratory staff. Failure to diagnose MM at first presentation can have severe consequences in terms of renal failure, amyloidosis, fractures or paralysis from spinal collapse.
A new diagnostic serum test (Freelite™) has recently been described1 that enables the rapid identification and subsequent monitoring of MM patients.
Serum Samples: Laboratories currently perform serum protein electrophoresis (SPE) tests on serum samples from suspected MM patients and immunofixation electrophoresis (IFE) if the SPE result is suspicious. These assays are known to be inadequate2, lack sensitivity and are prone to visual misinterpretation.
Approximately 15% of all cases of MM are Bence Jones (light chain) myeloma. These are not readily detected on SPE and most will not be identified.
Between 1 and 3% of MM are non-secretory myelomas these patients are negative by SPE and IFE on both serum and urine.
Consequently, if serum alone is tested, up to 18% of MM patients can be missed.
24-Hour Urine Samples: In order to exclude Bence Jones myeloma, 24-hour urine specimens should be tested. These samples must be sent to the laboratory, where they are concentrated x100, before being tested by electrophoresis and examined for the presence of free light chains. Unfortunately, problems of patient compliance with urine collection, and the transportation of large volumes of urine to the laboratory, means that urine samples are not always tested.
Even when urine samples are received many laboratories are not able to identify the presence of Bence Jones protein (BJP). A recent distribution of urine on the UK National External Quality Assessment Scheme (NEQAS), indicated that 35% of laboratories were unable to detect BJP at approximately 60mg/L.3
Consequently, Bence Jones myeloma patients are frequently not diagnosed on first presentation.WHAT ARE FREE LIGHT CHAINS?
When immunoglobulin molecules are produced by the plasma cells in the bone marrow, the heavy chains (G, A, M, D, or E) and the light chains (kappa or lambda) are produced separately. The kappa and lambda molecules are bound to the heavy chains and intact immunoglobulins are assembled, then transported to the surface of the plasma cell.
Free kappa and lambda light chains are produced in excess and small amounts are found in the serum of healthy individuals.
The fact that bound light chains and free light chains are structurally identical, has, for many years, frustrated attempts to accurately measure serum levels of free light chains in the presence of an overwhelming excess of bound light chains.WHAT IS FREELITE?
Freelite™ is a new immunodiagnostic assay system permitting, for the first time, the accurate and rapid quantitation of free kappa and free lambda light chain concentrations in serum, urine and cerebrospinal fluid.
Affinity purified antibodies, reacting specifically with the free forms of kappa and lambda light chains, are pre-coated onto latex particles. When reacted with patient test samples containing free light chains, the latex particles aggregate causing an increase in turbidity within the reaction vessel. The increase in turbidity is measured by the laboratory analyser, compared to known reference standards and expressed as a concentration in mg/L. The ratio of free kappa/lambda concentrations can then be calculated.FREE LIGHT CHAINS IN DISEASE
Certain diseases can affect the production of free light chains by the plasma cells in the bone marrow, resulting in abnormal levels in the serum, urine or cerebrospinal fluid. Freelite may be used as a screening test to measure the free kappa and free lambda levels, as a tool for monitoring the response to therapy and as a serum marker of progression/relapse.
Multiple Myeloma (MM) is a malignant clonal proliferative plasma cell disorder. It is characterised by >30% plasma cells in the bone marrow, anaemia, lytic bone lesions and a serum and/or urine M component. In a recent trial1, serum samples from patients with multiple myeloma or Waldenström’s macroglobulinaemia, who had intact monoclonal immunoglobulins, were tested with Freelite. All samples contained increased concentrations of the clonal free light chain.
Light Chain Myeloma (Bence Jones). Approximately 15% of MM patients have no detectable whole immunoglobulin M component in the serum only free light chains in the serum (and usually in the urine - see graph, opposite). These patients frequently show no serum abnormality when tested on SPE. Similarly, urine dipsticks used to detect proteinuria are insensitive to Bence Jones protein and invariably give false negative results. Consequently, these patients are difficult to diagnose and are in danger of being missed at presentation.
When a clonal proliferation of plasma cells start to develop, free light chain serum concentrations increase. These small molecules are rapidly cleared by the renal tubules, so, in early disease, the urine is negative for Bence Jones protein. As the tumour mass expands, free light chain serum concentrations increase until the resorptive capacity of the renal tubules is finally exceeded. At this point, free light chain flows into the urine and the patient will test positive for Bence Jones protein.
In a recent study1, serum samples from patients with Bence Jones MM were tested with Freelite and all patients showed elevated levels of the clonal free light chain and normal levels of the alternate free light chain. Interestingly, two of these patients were in clinical remission, with no Bence Jones protein detectable in the urine but both had abnormal serum Freelite results. It is suggested that serum Freelite testing may provide a suitable alternative to 24-hour urine testing in the diagnosis and monitoring of Bence Jones MM.
Smouldering Myeloma (SM) is an asymptomatic condition where patients have a monoclonal band in the serum (>30g/L), >30% plasma cells but no anaemia or bone lesions. Like MGUS, these patients should not be treated but it is essential to monitor them on a regular basis for signs of progression to MM.
Non-Secretory Multiple Myeloma (NSM)
Between 1 and 3% of all patients with MM have an absence of detectable monoclonal proteins in both serum and urine using conventional laboratory tests. Thus, the diagnosis and monitoring of patients with NSM is dependent upon clinical assessment and bone marrow biopsy.
In a recent study5, serum samples from 28 patients with NSM were tested with Freelite. Of these, 19 patients were clearly clonal with abnormal free kappa/lambda ratios. A further 8 patients gave abnormally elevated or suppressed results for one or both free light chains. Only 1 patient was found to have normal results.
Follow-up studies were performed on 6 of the patients and in all cases, free light chain serum levels could be used to monitor progress.
It is therefore proposed that Freelite will be a useful tool in the diagnosis and monitoring of NSM.
Monoclonal Gammopathy of Undetermined Significance (MGUS) is an asymptomatic condition which occurs in approximately 3% of the population >70 years of age. It is associated with the presence of a monoclonal band (<20g/L) in the serum, no Bence Jones protein in the urine and <5% plasma cells in the bone marrow.
Although MGUS patients should not be treated it is essential that they be monitored on a regular basis, as a significant percentage progress to develop MM or a related malignant condition.
The finding of Bence Jones protein in the urine of MGUS patients is associated with an increased risk of malignant evolution to MM4, therefore the serum free light chain levels in these patients may provide useful prognostic information.
Primary Amyloidosis and Light Chain Deposition Disease
Primary amyloidosis (AL) is characterised by fibrillar deposition of circulating free light chains in a wide range of organs. The diagnosis of AL is characteristically made by biopsy of rectal mucosa or by subcutaneous fat pad aspiration. In Light Chain Deposition Disease (LCDD) the light chain is deposited largely in the kidneys. In a recent study the rate of detection of serum or urine free light chains in a group of AL and LCDD patients was significantly improved using the Freelite assays over conventional IFE techniques6.
In the USA Freelite is FDA cleared as an aid in the diagnosis and monitoring of multipe myeloma, Lymphocytic neoplasms, Waldenströms macroglobulinaemia and connective tissue diseases such as systemic lupus erythematosus. Clearance for AL and LCDD is pending.
Freelite is a new and exciting breakthrough in the detection and management of B Cell tumours. Laboratories worldwide are beginning to offer these tests, so if you would like to know more about the use of free light chain measurements in the clinical environment please contact The Binding Site.View FREELITE abstracts
- Bradwell et al, Highly sensitive automated immunoassay for immunoglobulin free light chains in serum and urine. (In Press) Clin Chem 2001;47:4,p673-680
- Sheehan et al. Demonstration of serum monoclonal immunoglobulin in a case of non-secretory myeloma by immunoisoelectric focusing. J Clin Pathol. 1985;38:806-809.
- Ward AM, White P, Beetham R. Monoclonal proteins in urine: preliminary results of a UK NEQAS survey, UK NEQAS for monoclonal proteins. Sheffield: UK NEQAS for Immunology and Immunochemistry 1998.
- Baldini et al. Role of different haematological variables in defining the risk of malignant transformation in monoclonal gammopathy. Blood 1996;87:912-18.
- Drayson et al. Serum free light chain measurements for identifying and monitoring patients with non-secretory multiple myeloma. Blood 2001;97:9,p2900-2902
- Abraham et al. Detection in serum of immunoglobulin free light chains in primary amyloidosis and light chain deposition disease by nephelometry. Clin Chem 2001;47:6,pA32, No.107