Eliciting effective antitumor immune responses in patients who fail checkpoint 
inhibitor therapy is a critical challenge in cancer immunotherapy, and in such 
patients, tumor-associated myeloid cells and macrophages (TAMs) are promising 
therapeutic targets. We demonstrate in an autochthonous, poorly immunogenic 
mouse model of melanoma that combination therapy with an agonistic anti-CD40 mAb 
and CSF-1R inhibitor potently suppressed tumor growth. Microwell assays to 
measure multiplex protein secretion by single cells identified that untreated 
tumors have distinct TAM subpopulations secreting MMP9 or cosecreting CCL17/22, 
characteristic of an M2-like state. Combination therapy reduced the frequency of 
these subsets, while simultaneously inducing a separate polyfunctional 
inflammatory TAM subset cosecreting TNF-α, IL-6, and IL-12. Tumor suppression by 
this combined therapy was partially dependent on T cells, and on TNF-α and 
IFN-γ. Together, this study demonstrates the potential for targeting TAMs to 
convert a "cold" into an "inflamed" tumor microenvironment capable of eliciting 
protective T cell responses.