Exploring EGFR, Nectin-4, and TROP-2 as Therapeutic Targets for Bladder Cancer Photoimmunotherapy
Isis Wolf1,2, Nora Giess1,2, Céline Roider1,2
1Department of Urology, Medical Center-University of Freiburg, Hugstetter Str. 55, 79106 Freiburg, Germany.
Background:
Non-muscle invasive bladder cancer (NMIBC) has limited therapeutic options and high recurrence rates. Photoimmunotherapy (PIT) enables targeted tumor ablation using antibody-photosensitizer conjugates and light activation. We evaluated EGFR, Nectin-4, and TROP-2 as PIT targets using cysteine-modified antibodies conjugated to the photosensitizer WB692-CB2.
Methods:
Antibodies derived from Cetuximab (Cmb, anti-EGFR), Enfortumab (Enf, anti-Nectin-4), and Sacituzumab (Sac, anti-TROP-2) were engineered with T120C and D265C mutations in the heavy chains for site-specific dye conjugation. Binding of the conjugates to BC cells was tested by flow cytometry and light-induced cytotoxicity of the conjugates, alone or in combination, was assessed by viability assays following irradiation.
Results:
Cysteine-modified antibodies were produced as intact IgG molecules and were efficiently conjugated with WB692-CB2 without loss of antigen specificity. SacT120C/D265C-WB692-CB2 showed the highest target binding and achieved near-complete cell killing at a red-light dose of 32 J/cm2. CmbT120C/D265C-WB692-CB2 required a fourfold higher light dose for comparable efficacy, while EnfT120C/D265C-WB692-CB2 demonstrated lower potency. No cytotoxicity was observed in antigen-negative cells. Combined treatment enhanced cytotoxicity, indicating additive phototherapeutic effects.
Conclusions:
Our findings suggest that PIT targeting EGFR, Nectin-4, or TROP-2 merits further preclinical development as a targeted therapeutic approach for NMIBC, including potential combinatorial or personalized strategies.
Insights
Photoimmunotherapy (PIT) targeting TROP-2 shows promise for non-muscle invasive bladder cancer (NMIBC). This approach uses antibody-photosensitizer conjugates for precise tumor ablation, offering new therapeutic avenues for NMIBC patients.
Area of Science:
- Oncology
- Immunotherapy
- Bioconjugation
Background:
- Non-muscle invasive bladder cancer (NMIBC) presents limited treatment options and high recurrence rates.
- Photoimmunotherapy (PIT) offers targeted tumor ablation via antibody-photosensitizer conjugates activated by light.
- Evaluating epidermal growth factor receptor (EGFR), Nectin-4, and TROP-2 as potential PIT targets.
Purpose of the Study:
- To engineer cysteine-modified antibodies targeting EGFR, Nectin-4, and TROP-2 for photoimmunotherapy.
- To assess the binding affinity and light-induced cytotoxicity of antibody-photosensitizer conjugates in bladder cancer cells.
- To determine the most effective target for PIT in NMIBC.
Main Methods:
- Site-specific conjugation of photosensitizer WB692-CB2 to engineered cysteine residues (T120C, D265C) on antibodies targeting EGFR, Nectin-4, and TROP-2.
- Flow cytometry to confirm antigen binding of the antibody conjugates to bladder cancer cells.
- Cell viability assays to measure light-induced cytotoxicity following irradiation.
Main Results:
- Cysteine-modified antibodies were successfully conjugated with WB692-CB2 while retaining antigen specificity.
- Sacituzumab (anti-TROP-2) conjugate demonstrated superior target binding and near-complete cell killing at 32 J/cm² red light.
- Cetuximab (anti-EGFR) and Enfortumab (anti-Nectin-4) conjugates showed lower potency, with no observed cytotoxicity in antigen-negative cells.
- Combined treatments exhibited enhanced cytotoxicity, suggesting additive phototherapeutic effects.
Conclusions:
- Targeting TROP-2 via PIT with Sacituzumab conjugate shows significant potential for NMIBC treatment.
- EGFR and Nectin-4 are also viable targets for PIT, though less potent in this study.
- Further preclinical development of PIT targeting these antigens is warranted for NMIBC, including combination and personalized strategies.
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