Poster Presentation 35th Lorne Cancer Conference 2023

Bacopaside II increases doxorubicin accumulation and overcomes ABCC3-associated chemotherapy resistance in triple negative breast cancer (#327)

Kenny Yeo 1 , Bimala Dhakal 1 , Saifei Liu 1 , Runhao Li 1 , Jennifer Hardingham 1 , Bimala Dhakal 2 , Celine Li 2 , Kevin Aaron Fenix 2 , Joanne Young 1 , Yoko Tomita 3 , Timothy Price 3 , Amanda Townsend 3 , Eric Smith 1
  1. Solid Tumour Group, Basil Hetzel Institute for Translational Health Research, The Queen Elizabeth Hospital, Woodville South, SA, Australia
  2. Surgical Science Research Group, Basil Hetzel Institute for Translational Health Research, The Queen Elizabeth Hospital, Woodville South, SA, Australia
  3. Medical Oncology, The Queen Elizabeth Hospital, Woodville South, South Australia, Australia

Introduction: Neoadjuvant chemotherapy is the standard of care for many triple negative breast cancer (TNBC) patients. Unfortunately, approximately half will develop chemotherapy resistance, leading to poor overall survival. Chemo-resistance is driven by multiple factors, including over-expression of ATP-binding cassette (ABC) drug transporter gene ABCC3. There is a need for therapies that overcome chemo-resistance. Bacopaside II (BII) is a major active constituent of the medicinal herb Bacopa monnieri, that has been used for centuries in traditional Ayurvedic medicine. We previously showed that BII inhibits proliferation, migration and invasion of breast cancer cells. However, it is not known if BII can overcome ABCC3-mediated chemotherapy resistance in TNBC.

Methods: In TNBC cell lines (DU4475, HCC1143, MDA-MB-231 and MDA-MB-453) ABCB1, ABCC3 and ABCG2 transcript expression was measured by qRT-PCR. Growth inhibition (IC50) was determined using crystal violet staining. Apoptosis and loss of cell membrane integrity (necrosis) was determined by kinetic live-cell imaging measuring activated caspase-3/7 and propidium iodide (PI) staining. Doxorubicin intracellular accumulation following treatment with BII or ABC transporter inhibitor cyclosporine A (CsA) was measured by flow cytometry. Chemo-resistance was induced in MDA-MB-231 by long-term scaffold-less 3D-culture.

Results: ABCC3 expression was relatively high in HCC1143, intermediate in MDA-MB-231 and MDA-MB-453, and low in DU4475, and correlated with doxorubicin resistance. BII IC50 for HCC1143, MDA-MB-231, MDA-MB-453 and DU4475 was 18.1µM (95%CI:17.6-18.6), 13.4µM (13.2-13.5µM), 18.5µM (18.0-18.9), and 20.5µM (19.3-22.0). Kinetic live-cell imaging of caspase-3/7 and PI staining demonstrated that ≥15µM BII induced apoptosis in HCC1143. In contrast, BII induced rapid loss of membrane integrity (≥15µM) and apoptosis (≥20µM) in MDA-MB-231. Non-cytotoxic concentrations of BII (10µM) and CsA (5µM) significantly increased doxorubicin accumulation in HCC1143 and MDA-MB-231. MDA-MB-231 3D-culture of increased expression of ABCC3 (8.6-fold), but not ABCB1 and ABCG3, increased resistance to doxorubicin (10-fold), paclitaxel (2.1-fold), and 5-fluorouracil (1.4-fold), and decreased doxorubicin accumulation, whilst BII sensitivity was unaltered.

 Conclusions: These results suggest that bacopaside II may increase doxorubicin accumulation and overcome ABCC3-associated chemo-resistance in TNBC.