Table 4.
Recommendations for the work-up of suspected newly diagnosed Waldenström’s macroglobulinemia patients.
| Diagnostic studies |
| (1) BM biopsy is mandatory for a correct diagnosis to distinguish between the different forms of IgM monoclonal gammopathies, and it is particularly informative in distinguishing between WM and MYD88L265P MZL cases with IgM monoclonal protein |
| (2) BM aspiration samples have to be sufficient, both in terms of quantity and quality, to perform MFC, cytogenetic and molecular studies in both unselected and CD19+ enriched samples. This implies sampling up to 12–20 ml of BM distributed in 3 EDTA tubes (flow cytometry, molecular and cell enrichment) and 1 sodium heparin tube (FISH studies) |
| (3) MFC studies have to be done in the non-fractionated BM aspiration to distinguish and assess the size of the tumor clone and exclude other B-LPDs and plasma cell dyscrasias. To do so, both the lymphoplasmacytic and plasma cell compartments should be characterized and counted |
| (4) MYD88L265P assessment is mandatory during the diagnostic work-up of WM and related disorders, and must be done in BM samples, using ASqPCR or dPCR |
| (5) MYD88L265P molecular analysis on CD19+ enriched samples is not mandatory but is required if using low sensitivity techniques (i.e., Sanger sequencing) or for samples with low infiltration |
| (6) MYD88L265P assessment in DNA from PB alone is not sufficient for a correct diagnosis due to frequent false negative results, even if the sample has been enriched in CD19+ cells, or ctDNA was used, and it is therefore not recommended |
| (7) In patients with a high probability of WM diagnosis but no detection of MYD88L265P in the BM with reliable sensitive methods, it is highly recommended to do complete MYD88 gene sequencing by Sanger or NGS, especially if BTKi are planned to be used. Sample must be enriched in CD19+ cells by immunomagnetic methods or flow cytometry sortinga |
| (8) CXCR4WHIM analysis is not currently considered mandatory in all WM patients outside clinical trials. However, if the patient is going to be treated with BTKi, it is highly recommended to perform this analysis to unveil treatment resistance mechanisms and anticipate slow responses. Analysis should be done by Sanger or NGS, using genomic DNA from BM cells after CD19+ enrichment. If CD19+ cell enrichment is not feasible, more sensitive approaches such as ASqPCR or dPCR on unsorted samples must be done, considering that these methodologies do not cover all mutationsa |
| (9) FISH studies with suitable probes for 6q21–25 and TP53/17p should be carried out by local or reference laboratories in BM samples enriched in CD19+ cellsa |
| (10) Alternative samples from other tissues (PB, lymph nodes, CSF, pleural or peritoneal effusions) can be used for additional diagnostic characterization if indicated. In this case, the presence of sufficient tumor cells in the cellular suspensions must be warranted by flow cytometry |
| Research studies |
| (11) New promising approaches with good sensitivity (Cast-PCR, high resolution melting analysis, dPCR, etc.) could be used as alternatives for detecting the MYD88L265P mutation and CXCR4 mutations. However, they still have to be considered as investigational and require validation before they could be considered reliable for diagnostic purposes |
| (12) MYD88L265P assessment in DNA from PB could be of help in the diagnosis of WM, especially when the sample has been enriched in CD19+ cells, or when ctDNA is used for the study. However, the real feasibility of this approach remains to be assessed and should be investigated, especially in the context of clinical trials |
| (13) Although PB studies are a good addition to the diagnostic work-up, their exclusive use, without BM evaluation, could be of value in low-risk IgM-MGUS (monoclonal protein <15 g/l, normal free light-chain ratio). However, they should still be considered under the umbrella of research studiesa |
| (14) Although TP53 mutation analysis is of high interest in WM due to prognostic and potential therapeutic implications, especially when immunochemotherapy is planned, it should be considered investigational at this moment. This analysis can be performed by reference laboratories using Sanger sequencing or NGS, using genomic DNA from BM cells after CD19+ cell enrichmenta |
ASqPCR allele-specific quantitative PCR, BM bone marrow, BTKi Bruton’s tyrosine kinase inhibitors, B-LPDs B-cell lymphoproliferative disorders, Cast-PCR Competitive Allele-Specific TaqMan® PCR, CSF cerebrospinal fluid, ctDNA circulating tumor DNA, dPCR digital PCR, FISH fluorescence in situ hybridization, IgM-MGUS IgM monoclonal gammopathy of uncertain significance, MFC multiparametric flow cytometry, MZL marginal zone lymphoma, NGS next-generation sequencing, PB peripheral blood, WM Waldenström’s macroglobulinemia.
aStatements that required a second round.