FASEB J. 2022 May;36 Suppl 1
Lea Stitzlein,
Achintyan Gangadharan,
Leslie Grasse,
Alexsandra Espejo,
Melissa Singh,
Yue Lu,
Ravesanker Ezhilarasan,
Joy Gumin,
Erik Sulman,
Frederick Lang,
Joya Chandra.
Epigenetically directed cancer therapeutics, such as targeting the histone demethylase LSD1, are an emerging approach to the treatment of glioblastoma (GBM), with dozens of candidates in preclinical development. To date, knockdown (KD) of LSD1 exerts effects on proliferation, but an evaluation and comparison of pharmacological LSD1 inhibitors in GBM is lacking. Therefore, the present study evaluated the effect of LSD1 KD compared with pharmacological inhibition and studied four LSD1 inhibitors in patient derived GSCs in vitroand in vivo for therapeutic efficacy and genes associated with resistance. Changes in gene expression were observed upon KD of LSD1 and compared with pharmacological inhibition of LSD1 in GBM cells. Four LSD1 inhibitors were assessed in a panel of nine GSC lines for their effects on cell viability and colony formation. The radioresistant GSC line, GSC 20, was used to establish an orthotopic xenograft model to evaluate the in vivo activity of LSD1 inhibition. In parallel, we identified five genes associated with resistance to LSD1 inhibition using RNA-seq comparing non-transformed and isogenic resistant GBM lines. Lastly, we probed for the presence of this gene set in our in vitro and in vivo GSC experiments with LSD1 inhibitors and in GBM patient exome data. In GBM cell lines, we identified an upregulation of genes associated with cell death, regulation of cell proliferation, and regulation of intracellular signaling upon both LSD1 KD and LSD1 inhibition using tranylcypromine treatment. Additional, more selective, pharmacological inhibitors of LSD1 (GSK-LSD1, RN-1 and SP-2577) in vitro had differential responses in the GSCs that were more varied, and responses were dependent on the specific LSD1 inhibitor and GSC line. Importantly, the response to LSD1 inhibition in vitro was not dependent on the GSC radiosensitivity or molecular subtype indicating unique determinants of sensitivity. When assessing in vivoefficacy, the GSC tumor-bearing mice treated with GSK-LSD1 had a strong reduction in tumor burden that occurred seven weeks into treatment. However, tumor regrowth occurred and overall survival was not significantly better in LSD1 inhibitor treated mice. To understand mechanisms associated with tumor regrowth, we mined our existing data on selectivity and sensitivity to LSD1 blockade and identified five genes potentially associated with resistance to LSD1 inhibition, including HKDC1, RAB3IL1, RAB39B, FTH1, and FAM213A. These genes were found to be upregulated in treatment resistant GSCs and in the brain tissue of GSK-LSD1 treated mice after they succumbed to tumor burden. Finally, the resistant-associated genes were present in exome sequencing data from GBM patients where they showed an inverse relationship with LSD1 expression. Future studies will focus on designing combination therapies with LSD1 inhibitors that will enhance their effects and overcome resistance associated with treatment by targeting pathways associated with the genes we have uncovered.