Professor Yehui Shi: The Epigenetic Regulation of HR+ Metastatic Breast Cancer—A Review of the Development Path of HDACi Entinostat

Editor's Note: Histone deacetylases (HDACs) are overexpressed in various tumors and are closely related to tumor progression and drug resistance. In hormone receptor-positive (HR+) breast cancer, HDAC overexpression leads to resistance to endocrine therapy and CDK4/6 inhibitors. The targeted application of HDAC inhibitors (HDACi) can effectively block this process, bringing better prognosis to patients. Entinostat, introduced in 1997, has undergone over two decades of meticulous research and development, and has become the first HDACi to achieve overall survival (OS) benefits in patients with HR+ metastatic breast cancer in China, offering new treatment options for Chinese patients. Professor Yehui Shi from the Tianjin Medical University Cancer Hospital will review the milestone events in the clinical exploration and application of HDACi entinostat in breast cancer, allowing the light of epigenetic regulation to shine on patients with HR+ metastatic breast cancer.

• HDAC Regulation Mechanism and the Discovery of Entinostat

Histone deacetylases (HDACs) are histone-modifying enzymes that regulate epigenetic changes. They control the acetylation process of histone post-translational modifications by removing the N-acetyl group from acetylated lysine residues in histones and non-histone proteins, thereby inhibiting DNA transcription and causing gene silencing. HDAC overexpression is closely related to the development and drug resistance of breast cancer and has become an important target for the development of anti-cancer drugs. In 1964, Vincent G. Allfrey et al. first observed that lysine acetylation in histones can significantly alter the biological properties of proteins, including protein binding, stability, and enzyme catalytic activity. Shortly thereafter, histone acetyltransferases (HATs), histone deacetylases (HDACs), and acetylated lysine-binding proteins were identified as transcriptional regulators. To date, 18 types of HDACs have been discovered in mammals. The term HDAC generally refers to the 11 zinc-dependent enzymes, which are further classified into four categories based on their sequence homology and cellular localization.

Under normal conditions, histone acetylation facilitates the dissociation of DNA from the histone octamer, relaxing the nucleosome structure and allowing transcription factors and co-transcription factors to bind to specific DNA targets, thereby activating gene transcription. HDACs, on the other hand, deacetylate lysine residues, exerting the opposite effect. In breast cancer, overexpressed HDACs reduce the acetylation levels of ERα promoters, inhibit MEF2 activity, and cause resistance to endocrine therapy. Additionally, HDACs can suppress TP53, leading to the loss of p21 and resistance to CDK4/6 inhibitors. HDAC overexpression-mediated reduction in histone acetylation levels is a common epigenetic abnormality, and targeting HDACs can reverse this acetylation imbalance, exerting anti-tumor effects.

In the 1970s, researchers discovered that butyrate salts have strong histone acetylation effects, marking the first global identification of compounds with HDAC inhibitory activity. However, due to their low anti-proliferative activity and short half-life, butyrate salts were not suitable for drug development. As understanding of HDAC structure deepened, researchers found that HDAC inhibitors (HDACi) generally have three pharmacophore groups: the cap structure (cap region) that interacts with amino acid residues at the edge of the HDAC active site entrance, the zinc-binding group (ZBP) that chelates the catalytic zinc ion at the bottom of the active site pocket, and the linker region that connects the cap and ZBG regions and interacts with the hydrophobic channel of the active site. In 1997, during the search for compounds that could inhibit HDAC, researchers discovered a series of benzamide derivatives with inhibitory activity and successfully identified one of the most active compounds—entinostat (MS-27-275).

Entinostat is an orally administered HDACi that can specifically inhibit Class I (mainly HDAC1 and HDAC3) and Class IV HDACs. Its unique mechanism of action has shown significant advantages in breast cancer treatment. It not only corrects abnormal epigenetic states but also overcomes CDK4/6 inhibitor resistance through multiple pathways, such as activating p21 expression and upregulating PTEN levels to inhibit the PI3K/AKT signaling pathway. It can also overcome endocrine therapy resistance by functionally activating ER and reshaping chromatin-associated proteins to restore ER promoter silencing, and it exerts immunomodulatory effects, thereby achieving a comprehensive attack on breast cancer cells.

• Key Clinical Studies of Entinostat

In the Phase I clinical study of entinostat, its pharmacokinetic characteristics showed linear changes within the dose range of 2-12 mg/m². The time to peak plasma concentration of entinostat under fasting conditions was 0.25-2 hours. Early clinical studies explored dosing based on body surface area, and subsequent studies supported a fixed dose of 5 mg/week through covariate analysis of body surface area, BMI, and weight, which was applied in later clinical trials. Entinostat has certain inhibitory effects on P450 liver enzymes, undergoes enterohepatic circulation, has an average half-life of 33-150 hours, and can penetrate the blood-brain barrier to some extent.

The Phase II ENCORE301 study in the United States showed that for 130 patients with advanced HR-positive breast cancer who had progressed after non-steroidal aromatase inhibitor (AI) therapy, entinostat combined with exemestane significantly improved median progression-free survival (mPFS) compared to placebo combined with exemestane, with results of 4.28 months vs. 2.27 months (HR = 0.73, P = 0.060). High acetylation expression levels in patients were associated with longer mPFS. Additionally, median overall survival (mOS) was extended to 28.13 months vs. 19.84 months (HR = 0.59, P = 0.036). Entinostat achieved clinical benefits regardless of whether patients had primary or secondary resistance.

In China, the Phase III randomized, double-blind, controlled clinical trial (EOC103A3101) of entinostat was led by Academician Binghe Xu from the Cancer Hospital of the Chinese Academy of Medical Sciences. The study included patients aged 18-75 years with HR+/HER2- locally advanced or metastatic breast cancer that had relapsed or progressed after previous endocrine therapy, including both pre- and post-menopausal patients. Pre- and peri-menopausal patients required concurrent medical castration. Patients were randomly assigned in a 2:1 ratio to receive entinostat + exemestane (n = 235) or placebo + exemestane (n = 119). With a median follow-up of 15.7 months, the mPFS in the entinostat group was significantly improved compared to the placebo group, at 6.32 months vs. 3.72 months, reducing the risk of disease progression or death by 24% (HR = 0.76, 95% CI: 0.58-0.98, P = 0.046). The mOS in the entinostat group reached 38.39 months, extending survival by over 9 months compared to the placebo group and reducing the risk of death by 17% (HR = 0.83, 95% CI: 0.62-1.10, P = 0.184), demonstrating more significant survival benefits. Patients with CDK4/6 inhibitor resistance and those who had previously received salvage chemotherapy also showed good efficacy.

Subgroup analysis showed that entinostat provided consistent benefits across all subgroups, with more pronounced effects in patients without visceral metastasis, those with primary endocrine resistance, and those who had not previously used fulvestrant. In terms of safety, most adverse reactions in the entinostat group were mild to moderate. The main adverse events leading to discontinuation of treatment included neutropenia, elevated γ-glutamyl transferase, and anemia, but overall safety was manageable.

This study is the first Phase III clinical trial of an HDACi to achieve OS benefits in HR+/HER2- breast cancer, with a patient population that included both pre- and post-menopausal individuals, closely resembling clinical practice. It confirmed the efficacy and safety of entinostat in patients with HR+/HER2- advanced breast cancer who had previously received endocrine therapy. Chinese patients with HR+/HER2- breast cancer have higher mutation rates in several breast cancer-specific genes compared to Caucasian populations, and Asian individuals have higher HDAC1 expression, making them more likely to benefit from HDACi treatment.

Based on these results, on April 24, 2024, the National Medical Products Administration (NMPA) of China approved the combination of entinostat and an aromatase inhibitor (AI) for the treatment of HR+/HER2- locally advanced or metastatic breast cancer that has relapsed or progressed after endocrine therapy. This approval offers a new treatment option for Chinese patients.