Breast cancer is the most prevalent malignancy in women and the second leading cause of cancer-related mortality worldwide. Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer and is characterized by high metastatic potential, severe side effects, and lack of targeted therapies. Therefore, novel and targeted TNBC treatment strategies are warranted. DNA polymerase 1 (POLA1) and histone deacetylase 11 (HDAC11) are critical regulators of cell proliferation, DNA replication, and epigenetic mechanisms, making them promising therapeutic targets in TNBC. We have previously shown that the adamantyl retinoid ST1926 is a POLA1 inhibitor with potent anti-tumor activities in breast cancer cells. We have synthesized two ST1926 analogues, MIR002 and GEM144, which exhibit dual inhibitory activity against POLA1 and HDAC11. We investigated the anti-tumor properties of these adamantyl compounds in human TNBC in vitro and in vivo models, focusing on GEM144 due to its superior pharmacokinetic properties.

In silico analysis was conducted to evaluate the expression of POLA1 and HDAC11 in human TNBC versus normal breast tissues. POLA1 and HDAC11 levels were determined by immunoblotting techniques in different subtypes of human TNBC cell lines (basal-like HCC1806, luminal androgen receptor (apocrine) MDA-MD-453, and the mesenchymal stem-like MDA-MB-231) as well as the normal-like breast cell line, MCF10A. The mechanism of action of GEM144 was studied by assessing TNBC cell viability, cell death, DNA damage, and apoptosis induction. For in vivo studies, the mouse intraductal breast cancer model using female NSG mice was explored as a true orthotopic model for breast cancer studies. Tumor progression and treatment response were monitored using the in vivo imaging system (IVIS).

In silico analysis did not indicate any statistically significant differences in the transcript levels of POLA1 and HDAC11 between human TNBC tumors and normal breast tissues. However, basal protein levels of POLA1 and HDAC11 were elevated in several TNBC cell lines compared to normal-like breast cells. ST1926, MIR002, and GEM144 reduced viability and increased cell death in the different tested TNBC cell lines at sub-µM concentrations while sparing normal-like breast cells. GEM144 induced DNA damage in treated TNBC cells, as shown by increased H2AX levels, and resulted in apoptosis, confirmed by TUNEL positivity and PARP cleavage. In vivo administration of GEM144 at 20 mg/kg reduced tumor burden in HCC1806 intraductal xenografts, without any major signs of toxicity.

Our findings highlight the therapeutic potential of DNA polymerase inhibitors in TNBC. Next, we will evaluate the effectiveness of GEM144 in combination with standard chemotherapies in TNBC cell lines and patient-derived xenografts.