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A21 Development of an Nrf2-KEAP1 protein-protein disruptor for proof of concept
  1. Alberto Bresciani1,
  2. Mauro Cerretani1,
  3. Melania D’Amico1,
  4. Paola Fezzardi1,
  5. Mariana Gallo1,
  6. Antonino Missineo1,
  7. Daniele Moretti1,
  8. Alessia Santoprete1,
  9. Sara Tambone1,
  10. Licia Tomei1,
  11. Celia Dominguez2,
  12. Ignacio Munoz Sanjuan2,
  13. Leticia Toledo Sherman2,
  14. Larry Park2
  1. 1IRBM Science Park S.p.A., Roma, Italy
  2. 2CHDI Foundation, Los Angeles, CA, USA


Mechanisms that activate innate antioxidant responses, as a way to mitigate oxidative stress at the site of action, hold much therapeutic potential in diseases, such as the Huntington’s disease, where the use of frank antioxidants has not yielded positive results. The nuclear factor (erythroid-derived 2)-like 2 (NRF2) is a transcription factor whose activity upregulates the expression of antioxidant and cell detoxifying enzymes such as oxidoreductase, catalases and oxygenases in response to oxidative stress. NRF2 activation has a strong interplay with other cellular pathways such as apoptosis, autophagy and cell death, thus regulating cell fate when an oxidative insult cannot be resolved. NRF2 levels are tightly regulated by KEAP1, a sensor of oxidative stress or reactive electrophilic compounds. KEAP1 binds NRF2 and facilitates its ubiquitination and subsequent degradation. Under ROS or reactive electrophiles, modification of KEAP1 cysteines leads to conformational changes that release NRF2 for nuclear translocation and transcription of the antioxidant enzyme cascade.

To date, most NRF2 activating compounds published modulate NRF2 levels via covalent binding to KEAP1 cysteines. As a consequence, these present several issues in terms of off-target effects, toxicity and inappropriate pharmacokinetics at the desired site of action. Recently, compounds that reversibly disrupt the NRF2-KEAP1 interaction have been described, opening the field to a new era of safer NRF2 activators.

Here we describe the design and early phases of a drug discovery program aimed at identifying new chemical classes able to induce the NRF2 response via reversible binding of KEAP1. With this regards, new and more informative biochemical assays were developed that enabled a better prioritisation and characterisation of tool compounds and hit molecules. In addition, the monitoring of cellular events, starting from target engagement down to protection from reactive oxygen species insults, led to the validation of non-covalent KEAP1 binders on primary neurons and astrocytes derived from a mouse model of the Huntington’s disease.

  • oxidative stress response
  • NRF2 KEAP1 displacement
  • neuroprotection

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