Proteomic characterization of LIN28A-driven resistance to imatinib in Chronic Myeloid Leukemia
Abstract
Chronic Myeloid Leukemia (CML) accounts for 15% of new leukemia cases in adults. In the United States alone, CML cases have increased steadily from 30,000 in 2000 to 150,000-180,000 in 2020. CML is characterized by a fusion protein called BCR-ABL, resulting from chromosomes 9 and 22 translocating. Imatinib, a BCR-ABL kinase inhibitor, was approved in 2001 and has improved CML patients' survival and quality of life. However, over 25% of patients will have to switch to other tyrosine kinase inhibitors (TKIs) after imatinib due to resistance or intolerance.
To better understand the resistance mechanism in CML, we used proteomics and phosphoproteomics to study parental and imatinib-resistant cell lines with and without imatinib treatment. We observed an enrichment of pathways involved in oxidative phosphorylation, fatty acid metabolism, and translation in the resistance clones. Kinase-substrate prediction analysis showed increased activity for the kinase AKT1. Furthermore, we found that LIN28A, an mRNA-binding protein that regulates the AKT signalling, was upregulated in imatinib-resistant cells and had increased phosphorylation at S200. Previous studies have shown that LIN28A/B increases in later and more resistant stages of CML. Additionally, LIN28A is involved in the resistance of breast, ovarian and pancreatic cancers to multiple therapies. To confirm the activity of LIN28A in our resistant cells, we measured the expression of let-7 family miRNAs and found significantly lower levels for the majority of the family in the resistant cells. LIN28A knockdowns resensitized the resistant clone to imatinib, and overexpression of LIN28A promoted resistance to imatinib. Using phosphoproteomics, we identified AKT, RPS6K, and PKC drivers of LIN28A resistance. With the use of midostaurin, a PKC inhibitor and AKTi-1/2, an AKT inhibitor, we were able to overcome the resistance.
The recent development of LIN28 inhibitors has been used to overcome resistance in other cancers. Therefore, we screened three commercially available LIN28 inhibitors, identifying LIN28i-1632 to be synergistic with imatinib. The synergistic combination led to changes in G2M checkpoint apoptosis and AKT signalling. These results show that targeting LIN28A both pharmacologically and genetically overcomes resistance to imatinib in CML.