Evaluation of In Vitro Phototoxicity of a Minibody-IR700 Conjugate Using Cell Monolayer and Multicellular Tumor Spheroid Models

Abstract

Photodynamic therapy (PDT) is a treatment strategy that utilizes photosensitizers (PSs) and light of a specific wavelength to kill cancer cells. However, limited tumor specificity is still a drawback for the clinical application of PDT. To increase the therapeutic efficacy and specificity of PDT, a novel human minibody (MS5) that recognizes a cell surface receptor expressed on various cancer cells was labeled with the hydrophilic phthalocyanine PS IR700 to generate an MS5-IR700 conjugate that is activated by near-infrared (NIR) light. The phototoxicity of the conjugate was mainly tested against the PC3 prostate cancer cell line. The MS5-IR700 conjugate killed PC3 cells after NIR light irradiation as compared to untreated cells or cells treated with IR700 alone. Time-course analysis of cell viability revealed a high percentage of cell death during the first hour in PC3 cells exposed to the MS5-IR700 conjugate and NIR light irradiation. After irradiation, the MS5-IR700 conjugate-treated PC3 cells displayed cellular swelling, round shape, and rupture of the cell and nuclear membranes. In a co-culture model, the MS5-IR700 conjugate killed MS5-positive Ramos lymphoma cells specifically, while leaving MS5-negative cells unaffected. In line with the data obtained with the monolayer cultures, the MS5-IR700 conjugate also killed PC3 cancer cell spheroids. The treatment induced relocation of heat shock protein 70 and calreticulin to the cell surface, implying the induction of immunogenic cell death. Overall, the data suggest that the developed MS5-IR700 conjugate is a promising therapeutic agent that warrants further preclinical studies.

Precision Killing of M2 Macrophages with Phage-Displayed Peptide-Photosensitizer Conjugates

“…The selection of M2 macrophage-specific phage clones should pave the way for multiple targeted therapies. Recently, we have developed an improved version of photoimmunotherapy that uses single chain Fv antibodies to direct the IR700 photosensitizer to cancer cells [ 26 ]. Given its high hydrophilicity, the IR700 photosensitizer, developed by Kobayashi’s lab, requires a ligand to bind and kill target cells after near infrared (NIR) light exposure [ 26 , 27 , 28 ].…”

“…Recently, we have developed an improved version of photoimmunotherapy that uses single chain Fv antibodies to direct the IR700 photosensitizer to cancer cells [ 26 ]. Given its high hydrophilicity, the IR700 photosensitizer, developed by Kobayashi’s lab, requires a ligand to bind and kill target cells after near infrared (NIR) light exposure [ 26 , 27 , 28 ]. Hence, we hypothesized that the combination of IR700 with the phage displaying the KML peptide (M2-specific) may kill M2, but not M1 macrophages.…”

“…Light microscopy also revealed severe alterations in the cell morphology post-NIR irradiation (Upper panels). We and others have shown that once the monoclonal antibody-IR700 conjugate binds to the target cell and is exposed to NIR light, it can result in rapid and irreversible damage to the cell membrane [ 26 , 28 , 30 ].…”

“…Some of the selected phage-displayed peptides were converted into phage-IR700 conjugates and shown to kill M1 and/or M2 macrophages. One of the unique characteristics of the developed phage-IR700 conjugate is that it gains photocytotoxicity only when it is bound to the target cell membrane and is activated by NIR light [ 26 , 27 ]. Only the phage-displayed peptides conjugated to the IR700 photosensitizer demonstrated cell killing when irradiated with NIR light (690 nm).…”

Correction: Sioud et al. Evaluation of In Vitro Phototoxicity of a Minibody-IR700 Conjugate Using Monolayer and Multicellular Tumor Spheroid Models. Cancers 2021, 13, 3356

“…The authors wish to make the following corrections to this paper [ 1 ]: In the original article, there was an error in Figure 4. The control panel (BF, FDA, and PI) was erroneously duplicated during the final figure assembly, resulting in two identical panels (control and IR700).…”