Monoclonal protein polymerisation is the cause of IgM multiple isotype-matched monoclonal proteins

Letter to the Editor,

The recently published study by Valentine and Dawney ¹ contained interesting insights about multiple isotype-matched monoclonal proteins (MIMMP) and the effect of treatment with a reducing agent. Their findings suggested that, in most cases, reducing agent treatment caused IgA but not IgG MIMMP to resolve into a single band. IgA contains an additional polypeptide chain compared to IgG: the J chain. This allows disulphide bond mediated binding between the heavy chains to form the natural IgA dimers, and in some cases larger polymers. ² The hypothesis is that formation of polymers by some IgA monoclonal proteins gives rise to additional bands seen on electrophoresis. Treatment of the sample with reducing agent breaks the disulphide bonds linking the heavy chains, releasing IgA monomers. This provides the rationale for why IgA MIMMP often resolve to a single band. Given that IgM also contains the J chain and naturally forms pentamers, it is reasonable to suggest that IgM MIMMP might be due to polymerisation in the same way as for IgA. We decided to verify this in our centre.

Samples containing either IgA or IgM MIMMP were prospectively identified from routine requests for serum electrophoresis. A Sebia Capillarys 2 capillary electrophoresis instrument (Sebia, France) was used. The selected samples were re-analysed, pre- and post-treatment with a reducing agent: 4 μL of 0.5 mol/L dithiothreitol (DTT, in Sebia IF/IT diluent) was added to 240 μL serum, mixed and incubated at room temperature for 10 min before analysis. Monoclonal protein identification and quantitation pre- and post-treatment was compared.

15/16 IgA and 12/12 IgM MIMMP resolved into a single monoclonal protein band after DTT treatment. Monoclonal protein quantitation was statistically significantly lower post-DTT compared to pre-DTT: p = 0.015 for IgA and p = 0.002 for IgM MIMMP. The median (range) difference in quantitation was ─0.6 g/L (─5.2 to +0.7) for IgA and ─1.6 g/L (─4.4 to ─0.1) for IgM MIMMP.

These data confirm the findings of Valentine and Dawney for IgA MIMMP. In addition, we demonstrate that the majority of apparent IgM MIMMP are due to polymerisation of a single IgM monoclone. This finding is worth noting, but only of practical interest if DTT treatment causes a clinically significant difference in monoclonal protein quantitation. The International Myeloma Working Group response criteria define a 25% decrease in quantitation as minimal response and 25% increase as progressive disease. ³ Similarly the UK Myeloma Forum MGUS guidelines define 25% increase in quantitation (and absolute difference ≥5 g/L) as significant, warranting re-referral. ⁴ We observed a difference in quantitation ≥25% in only 3/28 samples (2 IgM, 1 IgA), though the absolute difference was non-significant in each case (2, 2, 3 g/L). There was no distinguishing feature of these three samples. Given the generally small difference in quantitation, and noting the importance of consistent monoclonal protein quantitation over time, routine DTT treatment of all MIMMP is not indicated. In our centre, we DTT treat new IgA or IgM MIMMP, but only report post-treatment quantitation if the difference is significant. Subsequent samples from the same patient are treated the same, ensuring consistency of quantitation.