MALDI-MSI of Immunotherapy: Mapping the EGFR-Targeting Antibody Cetuximab in 3D Colon-Cancer Cell Cultures

Abstract

Immunotherapies are treatments that use a patient's immune system to combat disease. One important type of immunotherapy employed in cancer treatments is the delivery of monoclonal antibodies to block growth receptors. In this manuscript, we develop a methodology that enables accurate and simple evaluation of antibody-type drug delivery using MALDI-MSI. To overcome the mass-range limitation that prevents the detection of large therapeutic antibodies, we used in situ reduction and alkylation to break disulfide bonds to generate smaller fragments. These smaller fragments are more readily ionized and detected by MALDI-MSI without loss of spatial information on the parent drug. As a proof of concept study, we evaluated the distribution of cetuximab in 3D colon cell cultures. Cetuximab is a monoclonal antibody that binds to the extracellular domain of epidermal-growth-factor receptor (EGFR), which is often overexpressed in colorectal cancer (CRC) and mediates cell differentiation, proliferation, migration, and angiogenesis. Cetuximab directly inhibits tumor growth and metastasis and induces apoptosis. By performing on-tissue reduction followed by MALDI-MSI analysis, we successfully mapped the timedependent penetration and distribution of cetuximab in spheroids derived from two different *

Characterization of an Amphiphilic Phosphonated Calixarene Carrier Loaded With Carboplatin and Paclitaxel: A Preliminary Study to Treat Colon Cancer in vitro and in vivo

“…The most common form of colon cancer is adenocarcinoma (Beloqui et al, 2016; Yueh et al, 2016), so we selected HT-29 adenocarcinoma cells as a colon cancer model. These cells are frequently used in tumorigenicity studies (Al-Saffar et al, 2018; Guerrero et al, 2018; Liu et al, 2018) and can form well-differentiated adenocarcinoma consistent with primary colon cancer in nude mice (Handali et al, 2018). We found PTX-CPT-P4C6 nanovesicles to suppress proliferation of HT-29 cells more than Caco-2 cells.…”

3D Culture Systems for Exploring Cancer Immunology

“…Matrix-assisted-laser-desorption-ionization mass-spectrometry imaging (MALDI-MSI) is a widely employed imaging technique that can visualize the spatial arrangement of drugs and metabolites in tissue from the whole-body level to the subcellular level [ 34 ]. In a proof-of-concept study, Liu et al demonstrated that a MALDI-MSI technique can detect and assess the distribution of cetuximab, an anti-EGFR monoclonal antibody, throughout colon cancer spheroids [ 35 ]. Since the MALDI-MSI technique can also be applied to tissue, it is feasible to identify optimal antibodies via a spheroid screening assay, then assess the best antibody candidate in vivo.…”

3D Tumor Models and Their Use for the Testing of Immunotherapies

“…CTOs poorly retain immune cells usually and often require cocultures with immune cells (59). They could show that the diffusion and distribution of Cetuximab in 3D tumor models was similar to those occurring in vivo in previous studies (60). Organoids can be cultured in the presence of immune cells to assess their antitumoral activity as well as to test methods to stimulate them.…”