靶向间皮瘤中的自噬和细胞周期

Introduction. Mesothelioma is considered incurable for the majority of patients, and current systemic therapies are inadequate for most patients. Targeting 1) autophagy and 2) autophagy in combination with the inhibition of select cyclin-dependent kinases (CDKs) have potential for mesothelioma, as the cell cycle is altered in the majority of mesotheliomas. Methods. We conducted cell proliferation assays in mesothelioma cell lines to determine the IC 50 for the autophagy inhibitors ULK-101 and hydroxychloroquine (HCQ). ULK-101, HCQ, or control were incubated with mesothelioma cells for 72 hours, and cell viability was determined using a Cell Counting Kit-8 spectrophotometric assay. We then determined mRNA expression via RNA-sequencing (RNA-Seq) after CDK4/6 inhibition with abemaciclib. Subsequently, we conducted cell proliferation assays to determine if the combination of ULK-101 plus abemaciclib or HCQ plus abemaciclib could be additive or synergistic. Lastly, we obtained micrographs of H2452 cells treated with abemaciclib and subsequent organelle labeling stain (LysoTracker Red and mitoview green). Results. ULK-101 exhibited potency against mesothelioma cell lines (H2373 and H2452) with IC 50 values ranging from 11.9 μM to 13.3 μM, while the IC 50 values for HCQ against these cell lines were 32.5 μM and 45 μM. When H2373 and H2452 cells were exposed to varying combinations of abemaciclib plus ULK-101, or H2452 cells were exposed to varying combinations of HCQ plus abemaciclib, synergy was exhibited at several combinations of concentrations, with several Combination Indices < 1 as calculated via the Chou-Talalay method. This suggests a synergistic effect of these combination of drugs on decreasing cell proliferation. After exposure to abemaciclib, ULK1 (ATG1) and ATG5 (role in autophagy activation) transcript expression increased as identified via RNA-Seq. Micrographs of organelle tracking after exposure to abemaciclib demonstrated increased lysosome size and number, suggesting autophagy activation. Conclusions. ULK-101 is active against mesothelioma cells, while ULK-101 plus abemaciclib act synergistically against mesothelioma. Also, abemaciclib was associated with increased lysosome size and number, suggesting autophagy activation. Collectively, the results suggest that autophagy and the cell cycle pathways may be targeted in mesothelioma and could be co-targeted in a synergistic fashion.