Poster Presentation 32nd Lorne Cancer 2020

Synergistic inhibition of cancer stem cells (CSC) by combined Lysine Specific Demethylase-1 (LSD1) and Poly (ADP-Ribose) polymerase-1 (PARP1) inhibition. (#333)

Thiru PRASANNA 1 2 3 , Fan Wu 1 , Christopher Sutton 1 , Desmond Yip 2 3 , Laeeq Malik 2 3 , Jane E Dahlstrom 3 4 , Kum Kum Khanna 5 , Sudha Rao 1 5
  1. Science, University of Canberra, Bruce, ACT, Australia
  2. Medical Oncology, The Canberra Hospital, Canberra, ACT, Australia
  3. ANU Medical School, Australian National University, Acton, ACT, Australia
  4. Department of Anatomical Pathology, , ACT Pathology, Canberra health Services, Canberra, ACT, Australia
  5. 5QIMR Berghofer Medical Research Institute, Brisbane, Queensland, Australia

Introduction: Despite impressive responses and progression-free survival benefit, resistance to PARP inhibitor (PARPi) is universal. We explored PARP1/LSD1 interaction and the effect of combined inhibition, especially their effect on CSC.

Methods: Synergistic effect of LSD1 and PARP inhibition was tested in-vitro and in-vivo by siRNA knockdown and drug inhibition in breast cancer (BC) cell lines.

Results: PARP1 was mostly cytoplasmic biased in MCF7 cells, however, stimulation with PMA resulted in nuclear translocalization of PARP1, suggesting potential role of nuclear PARP1 in epithelial mesenchymal transition. PARP1 and LSD1 colocalized strongly in human metastatic brain tissue from BC (PCC 0.6) and in MDA-MB-231 (PCC 0.9). PARP1 or LSD1 knockdown did not reduce CD44hi/CD24lo CSC population in MDA-MB-231. However, combined LSD1 and PARP1 knockdown reduced the number of CSCs significantly. Similarly, combination phenelzine (an irreversible LSD1 inhibitor) and olaparib (PARPi) at low dose significantly reduced CSC (>20%), though no effect was seen with either agent alone. This effect was not seen in cells treated with GSK2879552 (irreversible LSD1 inhibitor) or veliparib.  Supporting these findings, PARP1 depletion upregulated mesenchymal genes or proteins (MYC, ALDH1A1, CD44) or transcription factor SNAI2 and histone demethylase, LSD1. PARP1, LSD1 and POL II were enriched in uPAR and CD44 promoters in olaparib treated cells whereas enrichment was lowest in combination phenelzine/olaparib group. PARP1 depletion upregulated PD-L1 mRNA expression, however concurrent LSD1 depletion with siRNA or phenelzine downregulated the PD-L1 mRNA. PD-L1 protein was downregulated in phenelzine (but not with GSK2879552) treated (ex-vivo) circulating breast cancer tumour cells. Subsequent genome wide analysis has identified novel epigenetic tags and pathways. Phenelzine/olaparib combination reduced tumour growth in mice xenograft model and reduced the number highly aggressive CSV/LSD1/PARP1 positive cells in ipsilateral lymph nodes.

Conclusion: Combination olaparib/phenelzine synergistically inhibited CSCs in MDA-MB-231. LSD1 negatively regulated PD-L1 expression and reversed the effect of PARP1 on PDL1 status in this breast cancer model. These data support a novel combinatory approach to improve PARPi efficiency.