The tumour microenvironment predominantly consists of macrophages with phenotypes ranging from pro-inflammatory (M1-like) to anti-inflammatory (M2-like). Trehalose-6,6'-dibehenate (TDB) displays moderate anti-tumour activity and stimulates M1-like macrophages via the macrophage inducible C-type lectin (Mincle) resulting in IL-1β production. In this study, we examined if monosodium urate (MSU), a known vaccine adjuvant, can boost IL-1β production by TDB-stimulated macrophages. We investigated the effect of MSU/TDB co-treatment on IL-1β production by GM-CSF (M1-like) and M-CSF/IL-4 (M2-like) differentiated mouse bone marrow macrophages (BMMs) and found that MSU/TDB co-treatment of GM-CSF BMMs significantly enhanced IL-1β production in a Mincle-dependent manner. Western blot analysis showed that increased IL-1β production by GM-CSF BMMs was associated with the induction of pro-IL-1β expression by TDB rather than MSU. Flow cytometry analysis showed that MSU/TDB co-stimulation of GM-CSF BMMs led to greater expansion of CD86/MHC II and CD86/MHC II subpopulations; however, only the latter showed increased production of IL-1β. Together, these findings provide evidence of the potential to use MSU/TDB co-treatment to boost IL-1β-mediated anti-tumour activity in M1-like tumour-associated macrophages.