p53 – “Guardian of the Genome”
Cancer is a genetic disease caused by changes in DNA, particularly mutations caused by DNA damage. Failure to repair DNA damage can lead to cancer-prone mutations. It is therefore critical to repair DNA damage or, when the damage is too extensive, eliminate cancerous cells entirely. The key cellular component charged with this awesome responsibility – to be the “guardian of the genome” – is the tumor suppressor protein, p53.
Since its discovery in 1979, p53 has been extensively studied due to its central role as the body’s first line of defense against cancer. p53 links together multiple pathways that patrol the cell for DNA damage and cellular stresses. When DNA damage is detected, and when oxidative or other cellular stresses exceed thresholds for normal cellular function, p53 activates critical genes that prevent propagation of cancerous cells. Some of these apply the brakes on cell division in a process call cell cycle arrest; others attempt to fix the DNA damage or reduce stress on the cell. When the damage is too extensive to allow comprehensive and accurate repair, p53 activates other genes that proactively destroy the cell in a process called apoptosis. Apoptosis is a form of programmed cell death that can eliminate cancer cells and prevent tumor formation.
p53 is the Most Commonly Mutated Protein in Human Cancers
DNA damage and mutations can occur anywhere in the genome, including the gene that encodes the p53 protein. In fact, mutations in p53 are the most common mutations in cancer, occurring in half of all tumors. When mutations occur in the TP53 gene they can introduce corresponding mutations in the p53 protein that not only diminish its tumor suppressive function but also lead to the acquisition of new properties that promote cancer cell growth. When p53 no longer functions normally, propagation of cancerous cells can proceed unchecked and lead to tumor formation. The incidence of TP53 mutation has been shown to increase following chemotherapy or radiation, resulting in resistance to these treatment modalities and limiting future therapeutic options.
To date, more that 25,000 unique TP53 mutations have been reported and thus a key challenge in the development of p53-targeted anti-cancer therapies is the vast number of mutations that can compromise its tumor suppression activity. The most common of these are missense mutations, involving the site-specific exchange of one amino acid for another, and account for 75% of all p53 mutations. Approximately 95% of p53 mutations occur in the DNA-binding domain, the crucial central portion of the protein responsible for binding DNA and activating the critical genes needed for cell cycle arrest and apoptosis. Given the very large diversity of possible p53 mutations, a therapeutic agent that targets one or even a small subset of TP53 mutations would likely be of limited benefit. In contrast, a more effective approach would be a general, direct reactivation of mutant p53 and restoration of normal p53 function.
Our Approach to Reactivating p53
Mutations in p53 destabilize the structure of the protein, rendering mutant p53 unable to safeguard genome integrity and destroy cancerous cells via apoptosis. Some mutations not only turn off normal p53 function but also endow mutant p53 with new properties that facilitate cancer cell replication, tumor formation and even metastasis. The sheer number of possible mutations, each one a unique protein target, complicates the design of a therapeutic agent that is capable of reactivating mutant p53. Yet within this complexity lies an opportunity to exploit the unifying property of p53 mutations: structural alteration or misfolding that is due to mutation-induced protein destabilization. A therapeutic agent that can broadly stabilize normal structure and restore protective functions to diverse p53 mutants has the potential to revolutionize the treatment of cancer.
This hypothesis guided our founders in their search for molecules that could reactivate multiple different p53 mutants in cancer cells and selectively destroy those cells through apoptosis. From a collection of thousands of molecules they identified one, PRIMA-1, with the desired activity. Further study of PRIMA-1 not only provided a deeper understanding of the mechanism of action but also directly led to eprenetapopt, our lead drug candidate. Laboratory studies of eprenetapopt, also known as APR-246, have produced promising results regarding its potential activity in many different p53 mutations. Eprenetapopt/APR-246 is currently being studied in two Phase 1b/2 studies and we are seeking enrollment in a pivotal Phase 3 study.
Mechanism of Action
When APR-246 is administered to patients it is converted into the active drug, methylene quinuclidinone (MQ). MQ binds to p53 at two cysteine residues in the DNA-binding domain and stabilizes mutant p53 in the normal, functional structure. Research conducted by our founders and our collaborators have confirmed not only the sites where MQ binds to mutant p53 but also the restoration of the correct protein structure. Stabilization of normal protein structure – the key step in the APR-246 mechanism of action – reactivates mutant p53, which can then trigger responses such as cell cycle arrest and apoptosis.
In vitro and in vivo experiments have demonstrated that APR-246, via MQ, also impairs the ability of tumor cells to respond to oxidative stress. In parallel to the binding mutant p53, MQ depletes intracellular glutathione and inhibits enzymes involved in the control of reactive oxygen species (ROS) that can lead to oxidative DNA damage. As a result, ROS levels, oxidative stress and DNA damage are augmented. These effects may contribute to the anticancer activity of APR-246 as well as the selectivity for cancer cells versus normal healthy cells. Severe and sustained oxidative stress, induced by MQ, is therefore an important secondary feature of the APR-246 mechanism of action.
MQ not only restores the ability of a cell to response to oxidative stress and DNA damage via reactivation of mutant p53 but also produces severe and sustained oxidative stress to which reactivated p53 can respond. The overlap of these features, linked together by MQ, may in turn provide more efficient induction of apoptosis and elimination of cancer cells.
Recent Scientific Publications & Presentations
Eprenetapopt (APR-246) and Azacitidine in TP53-Mutant Myelodysplastic Syndromes
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J Clin Oncol. 2021 Jan 15:JCO2002341. Online ahead of print. / D.A. Sallman, et al
Publication of data from an investigator-initiated trial for which financial support was provided by Aprea Therapeutics, Inc.
Eprenetapopt Plus Azacitidine in TP53-Mutated Myelodysplastic Syndromes and Acute Myeloid Leukemia: A Phase II Study by the Groupe Francophone des Myélodysplasies (GFM)
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J Clin Oncol. 2021 Feb 18:JCO2002342. Online ahead of print. / T. Cluzeau, et al
Publication of data from an investigator-initiated trial for which financial support was provided by Aprea Therapeutics, Inc.
APR-246 Combined with Azacitidine (AZA) in TP53 Mutated Myelodysplastic Syndrome (MDS) and Acute Myeloid Leukemia (AML). A Phase 2 Study By the Groupe Francophone Des Myélodysplasies (GFM)
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25th Congress of the European Hematology Association / T. Cluzeau, et al
June 2020
Presentation of data from an investigator-initiated trial for which financial support was provided by Aprea Therapeutics, Inc.
Phase 1b/2 Combination Study of APR-246 and Azacitidine (AZA) in Patients with TP53 Mutant Myelodysplastic Syndromes (MDS) and Acute Myeloid Leukemia (AML)
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61st American Society of Hematology Annual Meeting / D.A. Sallman, et al
December 2019
Presentation of data from an investigator-initiated trial for which financial support was provided by Aprea Therapeutics, Inc.