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J. Caers et al.
Serum electrophoresis and immunofixation may not be able to detect light-chain aberrations in patients with oligo-secretory disease, such as light-chain MM. Due to their low molecular weight, these SFLC are rapidly cleared by the kidneys. In such cases, the monoclonal burden should be measured in a 24 h urine collection or in the serum by an automated SFLC immunoassay (Grade 1A), the latter having a higher sensitivity to detect and quanti- fy the involved free light chains.7 In concordance with the International Myeloma Working Group (IMWG), we rec- ommend the performance of serum immunofixation and electrophoresis on serum and urine samples and a SFLC assay for the diagnosis of a monoclonal PC disorder (Grade 1A).
Additional laboratory tests should be performed for the diagnosis and follow-up of MM patients, such as a com- plete blood count to evaluate possible cytopenias, blood smears to look for circulating PC and general biochem- istry tests (renal and liver function tests, calcium, phos- phate, uric acid, albumin, creatinine, lactate dehydroge- nase, C-reactive protein, β2-microglobulin). Quantification of serum immunoglobulins by nephelom- etry enables an indirect measurement of the M-protein or the recognition of a secondary hypogammaglobinemia and is recommended for any patient presenting with a gammopathy (oligo-, poly- or monoclonal) (Grade 1A).
The HevyLite® immunoassay quantifies both the involved and uninvolved intact immunoglobulin chains and quantifies them separately (IgGκ/λ, IgAκ/λ, IgMκ/λ).8 This assay has prognostic value for progression-free and overall survival9 and seems particularly useful when the M- protein is difficult to measure via serum protein elec- trophoresis. This assay is not yet part of the routine workup of MM patients, but may be of value in the follow- up of patients and has been included in clinical trials.10
Urine analysis
Proteinuria should be assessed on urine samples from all patients at diagnosis and during the follow-up. If pro- teinuria is present, it should be quantified in a 24 h urine collection. Total 24 h protein and Bence-Jones proteinuria should be evaluated by densitometry, electrophoresis and immunofixation. To detect low amounts of monoclonal proteins, it is recommended that the urine is concentrated 200-fold.11 These 24 h urine collections are often inconsis- tently performed, resulting in incomplete urine collection. In addition, renal function can influence the accuracy of the results, a fact which should be taken into considera- tion when interpreting laboratory values. The SFLC assay can be used for the follow-up of patients with light-chain MM. A recent French study, focusing on patients with light-chain MM, demonstrated that the SFLC assay is superior to 24 h urine collection for: (i) identifying patients with measurable disease, (ii) following their response to initial therapy, and (iii) giving a prognostic indication of the patients’ response and overall survival.12 This study included 113 patients with light-chain MM, all of whom had an abnormal SFLC ratio and measurable disease parameters in serum, while only 64% patients had measurable M-proteins in the urine, as determined by urine protein electrophoresis. Similar results were found in 576 patients with light-chain MM from the UK Myeloma IX and XI trials. The disease burden of the patients with light-chain MM could be measured and monitored by urine protein electrophoresis in 80% of
cases. Of the remaining patients 113 (97%) had involved free light chains >100 mg/L, which was sufficient to measure response to therapy.13 These two studies con- firmed the importance of SFLC measurements to diag- nose and monitor patients with light-chain or oligosecre- tory MM. The replacement of urine studies by the SFLC assay for all myeloma patients remains controversial, since an Eastern Cooperative Oncology Group study on 399 MM patients (of whom only a minority had light- chain MM disease) found only a weak correlation between results of the SFLC assay and 24 h protein analy- sis.14
In line with the IMWG guidelines,15 we recommend the SFLC assay for the diagnosis and monitoring of patients with oligosecretory disease (Grade 2B). However, for patients with measurable urinary M-proteins, MM should be monitored by 24 h urine collections. When albumin is the dominant protein found in the urine, a glomerulopathy (such as AL-amyloidosis or light-chain deposition disease) should be excluded. The 24 h urine collection remains important when results are discordant.
Bone marrow studies
A BM aspirate enables quantification of infiltrating PC and cytogenetic studies on purified PC. Unfortunately, dilution by peripheral blood during aspiration or the pres- ence of patchy disease (uneven distribution of MM cells throughout the BM) may result in an underestimation of tumor infiltration.16 We therefore recommend an addi- tional BM trephine biopsy, which may generate comple- mentary information (Grade 1B). A BM biopsy correctly identified MM disease in 95% of symptomatic patients with a low PC count on the initial BM smears.16 The cor- rect quantification of BM PC is also important because of the 60% cut-off as a biomarker of malignancy. The IMWG earlier recommended retaining the highest PC infiltration in case of discrepancy. Finally, the monoclon- ality of PC in the diagnostic sample should be confirmed by multiparameter flow cytometry or by immunohisto- chemistry confirming light-chain restriction.
Flow cytometry of bone marrow cells
In cases of monoclonal gammopathies, the most rele- vant information provided by multiparameter flow cytometry is the identification and enumeration of neo- plastic versus polyclonal BM PC. Regardless of the disease category, these neoplastic PC share similar immunophe- notypic features, which are distinct from those of normal PC. Typically, CD38, CD138 and CD45 (together with light scatter characteristics) are the best backbone mark- ers for the discrimination of PC. In addition, expression of CD19, CD56, CD117, CD20, CD28, CD27 and CD81, together with cytoplasmic immunoglobulin light-chain restriction, allows a clear discrimination between nor- mal/reactive versus monoclonal PC17 and was used by the EuroFlow consortium to create a standardized panel allowing the quantification and immunophenotypic char- acterization of neoplastic PC.18
Due to dilution and the sometimes patchy disease dis- tribution, multiparameter flow cytometry often underes- timates the infiltration but remains important for detec- tion of monoclonal PC in the peripheral blood and for the detection of minimal residual disease (MRD) in the BM. The Mayo Clinic group reported on the prognostic importance of circulating neoplastic cells in patients with
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haematologica | 2018; 103(11)