Project Title: DEVELOPING AND CHARACTERIZING MODELS OF RESISTANCE TO CYCLIN-DEPENDENT-KINASE 4 (CDK4) INHIBITION IN THE CDK4 AMPLIFIED CANCER, DEDIFFERENTIATED LIPOSARCOMA
Project Mentor: James Chen – Internal Medicine
Dedifferentiated liposarcomas (DDLPS) are malignant adipocytic cancers characterized by an amplification of the cell cycle regulatory gene, CDK4. Palbociclib, a CDK4/6-inhibitor, is rapidly becoming a standard of care treatment; however, innate and acquired resistance to the drug limits efficacy in patients with DDLPS. Molecular mechanisms and biomarkers of palbociclib resistance have been posited in non-mesenchymal tumors, but are poorly understood in DDLPS. To this end, we developed palbociclib-resistant DDLPS cell lines derived from several patient DDLPS samples to better characterize these phenotypic and genomic differences.
Three human CDK4 amplified DDLPS cell lines (224A, 246, 863) and one liposarcoma control cell line without CDK4 amplification and RB loss (LiSa2) were brought into culture. Developing resistant cells were intermittently treated at palbociclib’s IC25 and surviving cells were allowed to recover before retreatment. Parental lines were grown as controls in tandem with the developing resistant lines to compare molecular changes in response to palbociclib. Cell viability was measured by an XTT assay and protein expression was measured using whole cell lysates for Western blotting.
Parental IC50 of the three CDK4 amplified DDLPS cell lines ranged from 8-12 µM and the cell line without CDK4 amplification had an IC50 of 12 µM. Resistant cell strains demonstrated minimal changes in the measured IC50. Initial response to palbociclib decreased expression of cell cycle proteins, CDK4 and pRb, and increased Cyclin E, CDK2, and CDK6. After 6 months of treatment, a statistically significant decrease in Cyclin E, CDK2, and pRb, was observed in the resistant cell lines and no changes were seen in CDK4, Cyclin D, and p16. (all p<0.01).
Here we present the development of novel, palbociclib resistant, DDLPS cell lines. While the initial response to combat palbociclib treatment increased Cyclin E and CDK2, these G1/S proteins are reduced in resistant lines. Resistant cell lines demonstrated decreased protein expression of Cyclin E, CDK2, and pRb. Clinically, DDLPS patients that exhibit innate dysregulation of RB1 or G1/S phase regulators may be more likely to demonstrate resistance to palbociclib. Next-generation sequencing of DNA and RNA expression in parent and resistant lines are underway.