Journal: Science Advances

Article Title: FSP1 and histone deacetylases suppress cancer persister cell ferroptosis

doi: 10.1126/sciadv.aea8771

Figure Lengend Snippet: ( A ) UMAP of PC9 parental and persister cells treated with or without 1 μM RSL3 for 24 hours. ( B ) Enriched Hallmarks gene sets in persister cells treated with or without RSL3. Positive normalized enrichment score (NES) values indicate enrichment in RSL3-treated persister cells. TNFα, tumor necrosis factor–α; NF-κB, nuclear factor κB; IL-6, interleukin-6; JAK, Janus kinase; STAT3, signal transducer and activator of transcription 3; UV, ultraviolet. ( C ) UMAP of PC9 persister cells treated with and without RSL3. ( D ) Pseudotime analysis of PC9 persister cells treated with and without RSL3. Solid black line indicates the estimated trajectory across cell states. ( E ) UMAP of PC9 persister cells treated with and without RSL3 colored by cluster. ( F ) Ferroptosis driver gene set signature score across clusters in (E). ( G ) Ferroptosis suppressor gene set signature score across clusters in (E). [(F) and (G)] P values calculated with Mann-Whitney test. ( H ) PC9 parental, persister, and persister cells treated with 500 nM RSL3 for 24 hours were analyzed for oxygen consumption rate (OCR). A total of 1.25 μM oligomycin (Oligo), 1 μM carbonyl cyanide p -trifluoromethoxyphenylhydrazone (FCCP), and 1 μM rotenone plus 1 μM antimycin A (R + A). n = 3 biological replicates; mean ± SEM is shown; P values calculated between parental and persister cell conditions (stars) or between persister cells and RSL3-treated persister cells (crosses) using two-tailed Student’s t test. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, †† P < 0.01, and ††† P < 0.001. ( I ) PC9 parental and persister cells derived from 2.5 μM erlotinib analyzed for mitochondrial ROS. Par, parental; pers, persister. ( J and K ) PC9 persister cells derived from 2.5 μM erlotinib (J) and A375 persister cells derived from 250 nM dabrafenib and 25 nM trametinib (K) in combination with mitoTEMPO were treated with 500 nM RSL3 for 24 hours. Viability was normalized to the respective mitoTEMPO-treated persister cells without RSL3 treatment. [(I) to (K)] n = 3 biological replicates; mean ± SD is shown; P values calculated with two-tailed Student’s t test.

Article Snippet: Vorinostat (HY-10221), dabrafenib (HY-14660), trametinib (HY-10999), metformin (HY-B0627), mitoTEMPO (HY-112879), IACS-010759 (HY-112037), FCCP (HY-100410), and BAY2402234 (HY-112645) were purchased from MedChemExpress.

Techniques: MANN-WHITNEY, Two Tailed Test, Derivative Assay

Journal: Science Advances

Article Title: FSP1 and histone deacetylases suppress cancer persister cell ferroptosis

doi: 10.1126/sciadv.aea8771

Figure Lengend Snippet: ( A to F ) Synergy heatmaps between GPX4 inhibitor RSL3 and HDAC inhibitors panobinostat and vorinostat following 24-hour cotreatment. Bliss synergy score calculated with SynergyFinder 3.0. Red color and positive scores indicate synergy, and green color and negative scores indicate buffering. [(A) and (B)] PC9 parental cells and persister cells derived from 50 nM erlotinib. [(C) and (D)] A375 parental cells and persister cells derived from 10 nM dabrafenib with 1 nM trametinib. [(E) and (F)] BT474 parental cells and persister cells derived from 2 μM lapatinib. ( G to J ) Prederived PC9 or A375 persister cells were treated for 48 hours with a nontoxic concentration of HDAC inhibitor (see fig. S7), rinsed, and then treated with RSL3 for 24 hours while maintained under targeted therapy treatment. Data normalized to untreated persister cells. Concentration and HDAC inhibitor used were as follows: (G) 7.5 nM panobinostat, (H) 5 nM panobinostat, (I) 100 nM vorinostat, and (J) 1 μM vorinostat. RSL3 concentrations used were as follows: (G) 150 nM, (H) 100 nM, (I) 150 nM, and (J) 80 nM. n = 3 biological replicates; mean ± SD is shown; P values calculated with two-tailed Student’s t test.

Article Snippet: Vorinostat (HY-10221), dabrafenib (HY-14660), trametinib (HY-10999), metformin (HY-B0627), mitoTEMPO (HY-112879), IACS-010759 (HY-112037), FCCP (HY-100410), and BAY2402234 (HY-112645) were purchased from MedChemExpress.

Techniques: Derivative Assay, Concentration Assay, Two Tailed Test

Journal: Science Advances

Article Title: FSP1 and histone deacetylases suppress cancer persister cell ferroptosis

doi: 10.1126/sciadv.aea8771

Figure Lengend Snippet: ( A ) UMAP of PC9 parental and persister cells treated with or without HDAC inhibitor panobinostat for 48 hours. ( B ) Enriched Hallmarks gene sets between persister cells treated with and without panobinostat. Positive NES values indicate gene sets enriched in persister cells treated with panobinostat. ( C and D ) Treatment with panobinostat does not decrease GSH levels in PC9 (7.5 nM panobinostat) or A375 (5 nM panobinostat) persister cells. Buthionine sulfoximine (BSO; 1 mM) was used as a positive control for GSH depletion. ( E ) Ferroptosis sensitization of PC9 persister cells from treatment with panobinostat is not inhibited by GSH ethyl ester (GSHee, 1 mM). ( F ) PC9 persister cell treatment with panobinostat decreases rather than increases intracellular iron. ( G and H ) Panobinostat treatment of PC9 persister cells increases total cellular ROS (G) and mitochondrial ROS (H). ( I and J ) PC9 persister cells derived from 2.5 μM erlotinib (I) and A375 persister cells derived from 250 nM dabrafenib and 25 nM trametinib (J) were cotreated with mitoTEMPO and were treated with 7.5 and 5 nM panobinostat, respectively, for 48 hours with and without 500 nM RSL3 for 24 hours. Viability was normalized to the viability of targeted therapy with mitoTEMPO and panobinostat without RSL3. [(C) to (J)] n = 3 biological replicates; mean ± SD is shown; P values calculated with two-tailed Student’s t test.

Article Snippet: Vorinostat (HY-10221), dabrafenib (HY-14660), trametinib (HY-10999), metformin (HY-B0627), mitoTEMPO (HY-112879), IACS-010759 (HY-112037), FCCP (HY-100410), and BAY2402234 (HY-112645) were purchased from MedChemExpress.

Techniques: Positive Control, Derivative Assay, Two Tailed Test