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Hydrogen Peroxide-Inducible PROTACs for Targeted Protein Degradation in Cancer Cells
Dehao Yu1, Heli Fan1, Zhili Zhou2
1Tianjin Key Laboratory on Technologies Enabling Development of Clinical Therapeutics and Diagnostics, School of Pharmacy, The Province and Ministry Co-sponsored Collaborative Innovation Center, for Medical Epigenetics, Tianjin Medical University, Tianjin, 300070, China.
Abstract:
Proteolysis-targeting chimeras (PROTACs) provide a powerful technique to degrade targeted proteins utilizing the cellular ubiquitin-proteasome system. The major concern is the host toxicity resulting from their poor selectivity. Inducible PROTACs responding to exogenous stimulus, such as light, improve their specificity, but it is difficult for photo-activation in deep tissues. Herein, we develop H2 O2 -inducible PROTAC precursors 2/5, which can be activated by endogenous H2 O2 in cancer cells to release the active PROTACs 1/4 to effectively degrade targeted proteins. This results in the intended cytotoxicity towards cancer cells while targeted protein in normal cells remains almost unaffected. The higher Bromodomain-containing protein 4 (BRD4) degradation activity and cytotoxicity of 2 towards cancer cells is mainly due to the higher endogenous concentration of H2 O2 in cancer cells (A549 and H1299), characterized by H2 O2 -responsive fluorescence probe 3. Western blot assays and cytotoxicity experiments demonstrate that 2 degrades BRD4 more effectively and is more cytotoxic in H2 O2 -rich cancer cells than in H2 O2 -deficient normal cells. This method is also extended to estrogen receptor (ER)-PROTAC precursor 5, showing H2 O2 -dependent ER degradation ability. Thus, we establish a novel strategy to induce targeted protein degradation in a H2 O2 -dependent way, which has the potential to improve the selectivity of PROTACs.
Insights
New Proteolysis-targeting chimeras (PROTACs) are activated by hydrogen peroxide (H₂O₂) in cancer cells. This targeted protein degradation strategy improves selectivity and reduces toxicity in normal cells.
Area of Science:
- Chemical Biology
- Drug Discovery
- Molecular Biology
Background:
- Proteolysis-targeting chimeras (PROTACs) offer a method for targeted protein degradation via the ubiquitin-proteasome system.
- Current PROTACs face challenges with host toxicity due to poor selectivity.
- Inducible PROTACs enhance specificity but face limitations in deep tissue activation.
Purpose of the Study:
- To develop novel hydrogen peroxide (H₂O₂)-inducible PROTAC precursors for targeted protein degradation.
- To enhance the selectivity of PROTACs by leveraging the endogenous H₂O₂ levels in cancer cells.
- To investigate the potential of H₂O₂-dependent PROTACs for cancer therapy.
Main Methods:
- Design and synthesis of H₂O₂-inducible PROTAC precursors (2/5).
- Utilized H₂O₂-responsive fluorescence probe (3) to characterize H₂O₂ levels in cancer cells.
- Employed Western blot assays and cytotoxicity experiments to evaluate PROTAC efficacy and selectivity.
Main Results:
- H₂O₂-inducible PROTAC precursors (2/5) effectively degraded targeted proteins (BRD4, ER) in cancer cells.
- PROTAC precursor 2 demonstrated enhanced degradation of Bromodomain-containing protein 4 (BRD4) and cytotoxicity in H₂O₂-rich cancer cells (A549, H1299) compared to normal cells.
- The H₂O₂-dependent degradation mechanism was validated for both BRD4 and estrogen receptor (ER) targets.
Conclusions:
- A novel strategy for H₂O₂-dependent targeted protein degradation was established.
- This approach significantly improves PROTAC selectivity by exploiting differential H₂O₂ concentrations between cancer and normal cells.
- H₂O₂-inducible PROTACs hold promise for developing safer and more effective cancer therapeutics.
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