Chemical Library Screening and Structure-Function Relationship Studies Identify Bisacodyl as a Potent and Selective Cytotoxic Agent Towards Quiescent Human Glioblastoma Tumor Stem-Like Cells

Abstract

Cancer stem-like cells reside in hypoxic and slightly acidic tumor niches. Such microenvironments favor more aggressive undifferentiated phenotypes and a slow growing "quiescent state" which preserves them from chemotherapeutic agents that essentially target proliferating cells. Our objective was to identify compounds active on glioblastoma stem-like cells, including under conditions that mimick those found in vivo within this most severe and incurable form of brain malignancy. We screened the Prestwick Library to identify cytotoxic compounds towards glioblastoma stem-like cells, either in a proliferating state or in more slow-growing "quiescent" phenotype resulting from non-renewal of the culture medium in vitro. Compound effects were assessed by ATP-level determination using a cell-based assay. Twenty active molecules belonging to different pharmacological classes have thus been identified. Among those, the stimulant laxative drug bisacodyl was the sole to inhibit in a potent and specific manner the survival of quiescent glioblastoma stem-like cells. Subsequent structure-function relationship studies led to identification of 4,4'-dihydroxydiphenyl-2-pyridyl-methane (DDPM), the deacetylated form of bisacodyl, as the pharmacophore. To our knowledge, bisacodyl is currently the only known compound targeting glioblastoma cancer stem-like cells in their quiescent, more resistant state. Due to its known non-toxicity in humans, bisacodyl appears as a new potential anti-tumor agent that may, in association with classical chemotherapeutic compounds, participate in tumor eradication.

Research progress on therapeutic targeting of quiescent cancer cells

“…injury or tumor acidification) and cell-intrinsic regulatory mechanisms [1,2]. Scientific literature to date supports the existence of quiescent cancer cells in many tumor types, including breast, liver and pancreatic cancer, acute myeloid leukaemia, melanoma, and glioblastoma [3,4]. In earlier studies, cells were induced to quiescence via serum starvation (whereby cells at low density were grown in serum-free medium) or contact inhibition (cells were grown in 10% serum until complete confluence) [5][6][7].…”

“…In earlier studies, cells were induced to quiescence via serum starvation (whereby cells at low density were grown in serum-free medium) or contact inhibition (cells were grown in 10% serum until complete confluence) [5][6][7]. Quiescent cells were confirmed based on the lack of expression of Ki67 (a cell proliferation marker), negative EDU incorporation [7] or low rate of BrdU incorporation [3], and low Erk 1/2: p38 MAPK ratio [6][7][8]. Moreover, mVenus-p27K À , a fusion protein composed of the fluorescent protein mVenus and a defective mutant of p27 combined with Fucci probe could effectively recognize and distinguish cells at G0 from those at G1 during the G0-G1 transition state [9,10].…”

“…Quiescent cells initiate a series of programs that protect against cellular stress and toxicity. Although the protected state is favorable for long-term maintenance of the quiescent state, activation of these pathways in tumor cells may play a crucial role in higher resistance to conventional cancer therapies that largely target proliferating cells [3,5,11,12]. Quiescent tumor cells are considered significantly less radio/ chemosensitive with a greater repair capacity than cycling cells [11,13,14].…”

“…Quiescent tumor cells are considered significantly less radio/ chemosensitive with a greater repair capacity than cycling cells [11,13,14]. Resistance of G0 cells from breast, pancreatic, ovarian tumors and squamous cell carcinomas to traditional therapeutic patterns has been reported [3,15]. In addition, quiescent mesenchymal stem cells are more refractory to heat stress (HS) than growing cells.…”

“…The laxative bisacodyl/DDPM is the first small molecule identified displaying cytotoxicity to slow-growing cancer cells [3]. Accumulating evidence indicates that DDPM, the active metabolite of bisacodyl, induces necrosis of quiescent human glioblastoma stem-like cells (GSCs) in vivo and in vitro without exerting deleterious effects on non-cancer neural cells [5,66].…”