In vivo Assessment of the Impact of Molecular Weight on Constructs of 68Ga-DOTA-Manocept in a Syngeneic Mouse Tumor Model

Abstract

Purpose: Manocept™ constructs are mannosylated amine dextrans (MADs) that bind with high affinity to the mannose receptor, CD206. Tumor-associated macrophages (TAMs) are the most numerous immune cells in the tumor microenvironment and a recognized target for tumor imaging and cancer immunotherapies. Most TAMs express CD206, suggesting utility of MADs to deliver imaging moieties or therapeutics to TAMs. The liver Kupffer cells also express CD206, making them an off-target localization site when targeting CD206 on TAMs. We evaluated TAM targeting strategies using two novel MADs differing in molecular weight in a syngeneic mouse tumor model to determine how varying MAD molecular weights would impact tumor localization. Increased mass dose of the non-labeled construct or a higher molecular weight (HMW) construct were also used to block liver localization and enhance tumor to liver ratios.

Procedures: Two MADs, 8.7 kDa and 22.6 kDa modified with DOTA chelators, were synthesized and radiolabeled with ⁶⁸Ga. A HMW MAD (300 kDa) was also synthesized as a competitive blocking agent for Kupffer cell localization. Balb/c mice, with and without CT26 tumors, underwent dynamic PET imaging for 90 min followed by biodistribution analyses in selected tissues.

Results: The new constructs were readily synthesized and labeled with ⁶⁸Ga with ≥ 95% radiochemical purity in 15 min at 65 °C. When injected at doses of 0.57 nmol, the 8.7 kDa MAD provided 7-fold higher ⁶⁸Ga tumor uptake compared to the 22.6 kDa MAD (2.87 ± 0.73%ID/g vs. 0.41 ± 0.02%ID/g). Studies with increased mass of unlabeled competitors showed reduced liver localization of the [⁶⁸Ga]MAD-8.7 to varying degrees without significant reductions in tumor localization, resulting in enhanced tumor to liver signal ratios.

Conclusion: Novel [⁶⁸Ga]Manocept constructs were synthesized and studied in in vivo applications, showing that the smaller MAD localized to CT26 tumors more effectively than the larger MAD and that the unlabeled HMW construct could selectively block liver binding of [⁶⁸Ga]MAD-8.7 without diminishing the localization to tumors. Promising results using the [⁶⁸Ga]MAD-8.7 show a potential path to clinical applications.

Keywords: CD206 PET imaging; DOTA-tilmanocept; Molecular weight comparison; Syngeneic mouse tumor model.