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EpCAM-Binding DARPins for Targeted Photodynamic Therapy of Ovarian Cancer
Dirk van den Brand1,2, Sanne A M van Lith3, Jelske M de Jong1
1Department of Biochemistry, Radboud Institute for Molecular Life Sciences (RIMLS), Radboud University Medical Center, Geert Grooteplein 28, 6525 GA Nijmegen, The Netherlands.
Abstract:
Ovarian cancer is the most lethal gynecological malignancy due to late detection associated with dissemination throughout the abdominal cavity. Targeted photodynamic therapy (tPDT) aimed at epithelial cell adhesion molecule (EpCAM), overexpressed in over 90% of ovarian cancer metastatic lesions, is a promising novel therapeutic modality. Here, we tested the specificity and activity of conjugates of EpCAM-directed designed ankyrin repeat proteins (DARPins) with the photosensitizer IRDye 700DX in in vitro and in vivo ovarian cancer models. EpCAM-binding DARPins (Ec1: Kd = 68 pM; Ac2: Kd = 130 nM) and a control DARPin were site-specifically functionalized with fluorophores or IRDye 700DX. Conjugation of anti-EpCAM DARPins with fluorophores maintained EpCAM-specific binding in cell lines and patient-derived ovarian cancer explants. Penetration of DARPin Ec1 into tumor spheroids was slower than that of Ac2, indicative of a binding site barrier effect for Ec1. DARPin-IRDye 700DX conjugates killed EpCAM-expressing cells in a highly specific and illumination-dependent fashion in 2D and 3D cultures. Furthermore, they effectively homed to EpCAM-expressing subcutaneous OV90 xenografts in mice. In conclusion, the high activity and specificity observed in preclinical ovarian cancer models, combined with a high specificity in patient material, warrant a further investigation of EpCAM-targeted PDT for ovarian cancer.
Insights
Targeted photodynamic therapy (tPDT) using EpCAM-binding DARPins conjugated to IRDye 700DX shows high specificity and activity against ovarian cancer. This approach effectively targets EpCAM-expressing cells in preclinical models, warranting further investigation for ovarian cancer treatment.
Area of Science:
- Oncology
- Biotechnology
- Photochemistry
Background:
- Ovarian cancer is a lethal gynecological malignancy often detected late, with metastasis throughout the abdominal cavity.
- Epithelial cell adhesion molecule (EpCAM) is overexpressed in over 90% of ovarian cancer metastatic lesions, making it a key therapeutic target.
- Targeted photodynamic therapy (tPDT) offers a promising approach for localized cancer treatment.
Purpose of the Study:
- To evaluate the specificity and efficacy of designed ankyrin repeat protein (DARPin) conjugates with the photosensitizer IRDye 700DX for EpCAM-targeted tPDT in ovarian cancer.
- To assess the binding characteristics and tumor penetration of EpCAM-specific DARPins.
- To determine the in vitro and in vivo anti-cancer activity of EpCAM-targeted tPDT.
Main Methods:
- Site-specific functionalization of EpCAM-binding DARPins (Ec1, Ac2) and a control DARPin with fluorophores or IRDye 700DX.
- Assessment of EpCAM-specific binding in ovarian cancer cell lines and patient-derived explants.
- Evaluation of DARPin penetration into tumor spheroids and cell killing efficacy in 2D and 3D cultures.
- In vivo studies using subcutaneous OV90 xenografts in mice to assess tumor homing.
Main Results:
- Conjugation of anti-EpCAM DARPins with fluorophores preserved EpCAM-specific binding.
- DARPin Ec1 exhibited slower tumor spheroid penetration compared to Ac2, suggesting a binding site barrier effect.
- DARPin-IRDye 700DX conjugates demonstrated potent, illumination-dependent killing of EpCAM-expressing cells in vitro.
- Effective homing of conjugates to EpCAM-expressing xenografts was observed in vivo.
Conclusions:
- EpCAM-targeted tPDT using DARPin-IRDye 700DX conjugates displays high specificity and activity in preclinical ovarian cancer models.
- The observed specificity in patient-derived material further supports the potential of this therapeutic strategy.
- Further investigation of EpCAM-targeted tPDT is warranted for the treatment of ovarian cancer.
