The antigen causing IgG hexamerization in patients with Chronic lymphocytic leukemia (CLL) is Alpha 2 macroglobulin (A2M)

Chronic lymphocytic leukemia (CLL) is the most common leukemia in adults
in the western world. One of the treatments offered for CLL is
immunotherapy. These treatments activate various cellular and biochemical
mechanisms, using the complement system. Recently it was shown that the
complement system in CLL patients is persistently activated at a low level
through the classical pathway (CP). The mechanism of chronic CP activation
involves the formation of IgG-hexamers. According to recent studies,
formation of ordered IgG-hexamers occurs on cell surfaces via specific
interactions between Fc regions of the IgG monomers, which occur after
antigen binding. The present study investigated the formation of
IgG-hexamers in CLL patients and normal (non-malignant) controls (NC),
their ability to activate complement, their incidence as cell-free and
cell-bound forms and the identity of the antigen causing their formation.
Sera from 30 patients and 12 NC were used for separation of IgG-hexamers.
The obtained IgG-hexamers were measured and used for assessment of CP
activation. For evaluation of the presence of IgG-hexamers on blood cells,
whole blood samples were stained and assessed by flow cytometry. Serum
levels of IgG-hexamers were higher in CLL and they activated the
complement system to a higher extent than in NC. Alpha 2 macroglobulin
(A2M) was identified as the antigen causing the hexamerization of IgG, and
was found to be part of the hexamer structure by mass spectrometry,
Western blot and flow cytometry analysis. The presence of A2M-IgG-hexamers
on B-cells suggests that it may be formed on B cells surface and then be
detached to become cell-free. Alternatively, it may form in the plasma and
then attach to the cell surface. The exact timecourse of A2M-IgG-hexamers
formation in CLL should be further studied. The results in this study may
be useful for improvement of current immunotherapy regimens.