Serum-Inert Tandem-Locked Fluorescent Probe for Specific Imaging of Intratumoral Complement System

Area of Science:

• Biomedical Engineering
• Molecular Imaging
• Cancer Biology

Background:

• The complement system plays a role in cancer, but its dynamic behavior in tumors is poorly understood due to limitations of conventional analytical methods.
• Real-time imaging of intratumoral complement activity is challenging due to serum protein interference and lack of specific probes.
• Understanding complement system dynamics is crucial for deciphering the tumor immune microenvironment.

Purpose of the Study:

• To develop a novel molecular optical probe for non-invasive, real-time monitoring of intratumoral complement system activity.
• To overcome challenges associated with serum complement protein interference and nonspecific probe activation.
• To investigate the role of complement activity in different tumor models and its correlation with immune cell infiltration.

Main Methods:

• Synthesis of a tandem-locked fluorescence probe (TECP_CTSB) with high C1r specificity and a cathepsin B (CTSB)-cleavable masking sequence.
• Screening of peptide sequences for enzymatic kinetics toward C1r and CTSB.
• In vivo imaging of intratumoral complement activity in 4T1 and CT26 tumor models.
• Flow cytometry profiling of complement activation-associated fluorescence in tumor tissues.

Main Results:

• The developed probe TECP_CTSB selectively detects intratumoral complement activity without nonspecific activation in circulation.
• Higher intratumoral complement activity was observed in 4T1 tumors compared to CT26 tumors, potentially explaining differences in myeloid-derived suppressor cell infiltration.
• Complement activation-associated fluorescence was primarily detected in cancer-associated fibroblasts and tumor cells, often localized at the tumor periphery.
• Chemotherapy treatment led to enhanced complement activation extending into the tumor interior.

Conclusions:

• The novel fluorescence probe enables unprecedented in vivo imaging and flow cytometry analysis of intratumoral complement activity.
• Findings provide new insights into the differential roles of complement in various tumor types and their immunosuppressive microenvironments.
• The probe holds significant potential for complement system research, cancer diagnostics, and monitoring therapeutic responses.