E3 Ubiquitin Ligase TRIM21 Enhances Macrophage-Mediated Bortezomib Resistance By Inducing M2 Polarization in Multiple Myeloma

Multiple myeloma is a hematologic malignancies characterized by clonal plasma cell abnormalities in the bone marrow. Although new drugs such as proteasome inhibitors(PIs), immunomodulators(IMiD) and autologous hematopoietic stem cell transplantation have significantly prolonged the survival time of MM patients, MM is still incurable, the majority of MM patients relapsed or refractory due to disease progression. It has been shown that the bone marrow microenvironment plays an important role in the proliferation, growth and drug resistance of MM cells. As the major component of the MM bone marrow microenvironment, Macrophages have been shown to mediate the initiation, development and drug-resistant behavior of MM through multiple pathway in existing studies. Thus targeting key molecules that regulate macrophage function in the bone marrow microenvironment and elucidating their specific mechanisms of action; It has important clinical significance to delay the progression of MM.

In the bone marrow microenvironment, there exist M1-type TAMs with anti-tumor effects, and M2-type TAMs that promote tumor progression. These two types can interconvert, showing high plasticity. Here, we found that inducing macrophages to polarize towards M1 or M2 types using GM-CSF and M-CSF respectively, resulted in significantly increased expression of the E3 ubiquitin ligase TRIM21 in M2-type macrophages. The proportion or quantity of M2-type macrophages in the bone marrow microenvironment is related to the treatment outcomes of multiple myeloma (MM). Furthermore, we discovered that in bone marrow biopsy samples from relapsed/refractory MM patients, there is a significant increase in the proportion of M2-type macrophages, accompanied by markedly elevated levels of TRIM21 expression compared to newly diagnosed MM patients.After knocking down TRIM21 in macrophages, there was a significant increase in markers associated with M1 polarization and a noticeable decrease in markers associated with M2 polarization. Moreover, when co-cultured with MM cells, these macrophages exhibited reduced protective capabilities towards MM cells, accompanied by a marked increase in apoptosis levels among the MM cells. Conversely, overexpression of TRIM21 in macrophages led to opposite effects. TRIM21, an E3 ubiquitin ligase, mediates targeted protein degradation via ubiquitin enzymatic activity. Subsequently, we constructed a virus overexpressing TRIM21 with ubiquitin enzymatic activity disabled, confirming that TRIM21 impact on macrophage polarization depends on its ubiquitin enzymatic activity.

Mechanistically, through RNA-seq analysis, we found that overexpression of TRIM21 significantly activates the JAK-STAT pathway in macrophages. Pre-treatment of macrophages with the JAK-STAT pathway inhibitor C188-9 reversed TRIM21-mediated regulation of macrophage polarization and notably reduced its protective effect on MM cells. Further investigation revealed that knocking down TRIM21 led to a significant increase in the protein expression levels of SOCS3, a key negative regulator of the JAK-STAT pathway in macrophages. Immunoprecipitation and ubiquitination experiments confirmed that TRIM21 interacts with SOCS3, and TRIM21 significantly increases its ubiquitination, thereby reducing SOCS3 protein levels.

In summary, our research confirms that TRIM21 induces M2 polarization in macrophages by targeting SOCS3 to activate the JAK-STAT pathway, thereby enhancing its protective effect on MM cells. Targeting TRIM21 could potentially be a strategy to improve MM treatment efficacy.

No relevant conflicts of interest to declare.