Evaluation of a CD206-Targeted Peptide for PET Imaging of Macrophages in Syngeneic Mouse Models of Cancer

Tumor-associated macrophages (TAMs) are large phagocytic cells that play numerous roles in cancer biology and are an important component of the relationship between immune system response and tumor progression. The peptide, RP832c, targets the Mannose Receptor (CD206) expressed on M2-like macrophages and is cross-reactive to both human and murine CD206. Additionally, it exhibits therapeutic properties through its ability to shift the population of TAMs from an M2-like (protumor) toward an M1-like phenotype (antitumor) and has demonstrated promise in inhibiting tumor resistance in PD-L1 unresponsive melanoma murine models. In addition, it has shown inhibition in bleomycin-induced pulmonary fibrosis through interactions with CD206 macrophages.^1,2 Our work aims to develop a novel CD206 positron emission tomography (PET) imaging probe based on RP832c (K_d = 5.64 μM) as a direct, noninvasive method for the assessment of TAMs in mouse models of cancer. We adapted RP832c to incorporate the chelator DOTA to allow for radiolabeling with the PET isotope ⁶⁸Ga (t_1/2 = 68 min; ß⁺ = 89%). In vitro stability studies were conducted in mouse serum up to 3 h. The in vitro binding characteristics of [⁶⁸Ga]RP832c to CD206 were determined by a protein plate binding assay and Surface Plasmon Resonance (SPR). PET imaging and biodistribution studies were conducted in syngeneic tumor models. Stability studies in mouse serum demonstrated that ⁶⁸Ga remained complexed up to 3 h (less than 1% free ⁶⁸Ga). Binding affinity studies demonstrated high binding of [⁶⁸Ga]RP832c to mouse CD206 protein and that the binding of the tracer was able to be blocked significantly when incubated with a blocking solution of native RP832c. PET imaging and biodistribution studies in syngeneic tumor models demonstrated uptake in tumor and CD206 expressing organs of [⁶⁸Ga]RP832c. A significant correlation was found between the percentage of CD206 present in each tumor imaged with [⁶⁸Ga]RP832c and PET imaging mean standardized uptake values in a CT26 mouse model of cancer. The data shows that [⁶⁸Ga]RP832c represents a promising candidate for macrophage imaging in cancer and other diseases.