The results presented here indicate the potential differential role of TGF-β1 in pathological cell proliferation in individuals with monoclonal gammopathies. The elevated level of TGF-β1 was confirmed in individuals positive for a paraprotein of IgA isotype compared to IgG and IgM individuals. Also, higher TGF-β1 level was detected in monoclonal IgM with λ light chain. Although patients with IgA paraprotein exhibit higher levels of TGF-β (Fig. 2A, 2B), creating a positive feedback loop, the relationship is not linear. Specifically, the level of TGF-β1 did not correlate with the level of monoclonal IgA. This lack of correlation could be attributed to differences in the sensitivity of individual patients/clones to TGF-β1. Furthermore, in IgG monoclonal gammopathies, TGF-β1 functions by supporting residual humoral immunity, specifically the levels of IgM (Fig. 2B). This underscores the importance of treatment stratification to identify and address the unique characteristics of patients more effectively.
Knowing which type of monoclonal immunoglobulin (paraprotein) class is prevailing may help clinicians estimate the course of disease if detected monoclonal gammopathy is determined as multiple myeloma, such that people with IgA myeloma are seen to have worse prognosis than those with IgG myeloma [16]. IgA myeloma is associated with a poorer survival outcome (higher median survival risk) and a higher risk of multiple meyeloma recurrence. Patients with IgA paraprotein often have specific genetic changes, like chromosome 13 deletion, leading to worse prognosis and was even found to be resistant to new therapeutic strategies with thalidomide and bortezomib [17]. Deviations in kappa to lambda light chains ratio can occur due to different factors such as age and diseases, especially in the context of monoclonal gammopathies, where an overproduction of one type of light chain can occur. Abnormalities in this ratio can play a role in the diagnosis and monitoring of diseases involving plasma cell disorders [18].
IgA, though relatively unexplored in its role in serum [19] is the predominant type found in the gut, functioning effectively in mucosal regions where antigens are prevalent [20]. Decades of studies have illuminated the elaborate processes behind immunoglobulin production, such as class switching and intraclonal class switching [21]. In 1980, Gearhart and co-researchers discovered that B lymphocytes in mice, stimulated by antigens, can produce IgA through two pathways: a direct switch from IgM to IgA or via two successive switches from IgM to IgG and then to IgA during clonal expansion, the process by which a single immune cell or a group of genetically identical immune cells replicate and increase in number [22].
In the context of multiple myeloma, cells in the bone marrow environment secrete cytokines like TNF (tumor necrosis factor), TGF-β (transforming growth factor beta), and VEGF (vascular endothelial growth factor) [23]. TGF-β plays a vital role, with its active form regulating cell growth. However, in multiple myeloma, cells evade TGF-β's growth-regulating effects, contributing to disease development/progression. TGF-β, a protein that is found circulating in the blood plasma, comes in latent form, needing activation for binding to type II (TβRII) and type I (TβRI) receptors on cell membranes [24]. The active form of TGF-β1 has a brief half-life in contrast to latent TGF-β1 [12]. In simple terms, when cells lose their ability to respond to TGF-β, they produce more TGF-β. In the case of multiple myeloma, the cells do not have mutations in specific (TβRI or TβRII) genes related to TGF-β, and they do produce the necessary proteins inside the cell. However, these proteins do not reach the cell surface because they form receptor complexes inside the cell [25]. This process allows multiple myeloma cells to escape the normal growth-regulating effects of TGF-β, which contributes to the development of the disease [25]. Research on MM.1S B lymphoblasts demonstrated that TGF-β could suppress cancer cell growth by increasing levels of E2F1 (DNA-binding family of transcription factors) a protein essential in controlling the cell cycle and proliferation [26].
TGF-β is known to influence IgA production. Experiments with mice and human B cells further reinforced the connection between TGF-β and IgA production. In fact, one of the pillars of immunology articles, Sonoda et al (1989), were the first to bring to daylight that IgA switching is induced by TGF-β. Back in 1989 they found that adding TGF-β to LPS stimulated murine B cells significantly increased IgA production while decreasing IgM, IgG1, IgG3, and total Ig production [27, 28] and suggested IgA switching rather than selectively stimulating post-switch. Another study by Ehrhardt and colleagues in 1992 showed that TGF-β impacts B cell survival and that IgA secretion depends on the B cell activation method used [29].
TGF-β is produced not only by specific parenchymal cells but also by hematopoietic cells and it functions where it is made. Thus, a process called IgA class switch recombination (CSR) is not limited to specific body regions, such as mucosal area [30]. Van Vlasselaer found that when TGF-β 1 was added to cultures of pure human splenic B cells along with mitogens (used to stimulate cell division and proliferation) and activated cloned CD4 + T cells, it significantly increased IgA production [31]. This effect was specific to IgA, as there was no impact on IgM or IgG production. They believed this increase in IgA synthesis indicated IgA class switching, a process observed in the early stages of cultures and previously studied by Coffman et al. (1989). This suggests that interaction between CD4 and class II MHC molecules is necessary for productive T-B cell interactions leading to IgA production [31].
Deleting TGF-β receptors in B cells resulted in reduced IgA antibodies, highlighting the significance of TGF-β signaling in IgA synthesis. Cazac and Roes, in their experiments with mice, confirmed that deleting the ligand-binding chain of the TGF-β receptor (TβRII) in CD19 + B cells resulted in reduced production of IgA antibodies in both Peyer’s patches and lamina propria along with low level of detection in the serum [32]. When these modified mice were immunized, they could not produce specific IgA antibodies either in serum or in nasal and bronchoalveolar lavages [32].
When examining IgA, IgG, and IgM concentrations in blood samples tested for presence of monoclonal immunoglobulins we found moderate to strong positive correlation between serum concentrations of non-paraprotein related immunoglobulin isotypes such as IgM and IgA isotypes in IgG monoclonal gammopathies, IgG and IgM isotypes in IgA monoclonal gammopathies, and IgG and IgA in IgM monoclonal gammopathies (Fig. 1B and Table S2; online supplementary material). This result could mean that proliferation of a malignant clone similarly affects the residual antibody mediated immunity. It is known that infectious pathogens are associated with diverse B-cell malignancies, either through direct cell infection leading to transformation or via antigen (Ag)-induced stimulation causing indirect cell transformation, or a combination of both mechanisms. Chronic cancer-associated inflammation is established in hematological malignancies, especially in myeloma [33] and chronic myeloproliferative neoplasms (MPNs) [34, 35]. In the case of monoclonal gammopathy of undetermined significance (MGUS), where the paraprotein level is < 30 g/L and the monoclonal immunoglobulin may constitute 20–70% of total immunoglobulin, the production of polyclonal IgG is sustained [36]. Multiple myeloma is associated with increased risk of infection, but we have little knowledge regarding antibody levels against specific bacteria. Even though polyclonal immunoglobulin isotypes may initially be below the reference range or recover in myeloma patients on antibiotic prophylaxis at the onset of anti-myeloma treatment, there is a potential decline in the proportion of protective antibody (IgG) levels against specific bacteria compared to the reference range in patients at the time of diagnosis. This emphasizes the necessity for developing strategies to protect patients in remission from bacterial infections during therapy administration and vaccination programs [37].
Beta-2-microglobulin is a valuable serum marker of tumor load in hematologic malignancies, and in multiple myeloma for staging disease in multiple myeloma patients [38]. It has been assessed that correlation of serum β2-microglobulin levels to tumor mass determined by whole-body MRI can correctly show the extension of myeloma infiltrate in patients with preserved residual renal function, consequently the stage of the disease [39]. In our study, in IgA monoclonal gammopathy, including IgA monoclonal gammopathy with λ light chain (mIgAλ), the β2 microglobulin levels positively correlated with the levels of total serum IgA and negatively correlated with residual polyclonal IgM and IgG isotypes (Table S2). Although the data with reference to this topic are limited, a group of authors, in in vitro study, have shown a significant correlation between beta2 microglobulin secretion and the immunoglobulin type of paraprotein in multiple myeloma: the highest level of secretion was noted in IgG and IgA multiple meyeloma. They suggested the hypothesis of a direct secretion of beta2 microglobulin by myeloma cells and emphasized the importance of beta2 microglobulin as a determination marker in most of the patients with multiple myeloma [40].
Studies have shown that serum levels of TGF-β may change with age. The relationship between TGF-β levels and age, however, is complex and may vary depending on the context, which is why we are facing inconsistencies regarding the levels, either increase or decrease, of TGF-β reported. It is possible that TGF-β levels may change differently depending on the type and microenvironment of tissues or in response to different stimuli. In recent years, numerous studies have revealed the various mechanisms of context-dependent effects of TGF-β signaling [21]. Our results show negative correlation between age and TGF-β1 levels in IgA monoclonal gammopathies.