Abstract 5289: Pre-clinical evaluation of CDK4/6 inhibitor resistance and immunologic modification in liposarcoma
Authors: Musi Elgilda , Patwardhan Parag P. , Ingham Matthew , Schwartz Gary K. 
Published: September 2020

Abstract
Soft tissue sarcomas are heterogeneous mesenchymal neoplasms and account for 1% of all cancers in adults. One of the most common diagnosed soft tissue sarcomas is well-differentiated (WD)/dedifferentiated (DD) liposarcoma which is characterized by amplification of CDK4. We have previously reported that the CDK4 inhibitor palbociclib induces cell cycle arrest in the CDK4 amplified, LS141 and DDLS liposarcoma cell lines in vitro (Barretina J. et al Nature Genet. 2010) and in clinical trial results palbociclib results in prolonged progression free survival in patients with WD/DD-liposarcoma (Dickson M Et al. J Clin Oncol. 2013). We have now studied the mechanisms of CDK4/6 resistance in liposarcoma by exposing LS141 and DDLS cell lines to increasing concentrations of the CDK4/6 inhibitor, ribociclib, for an extended time period of 1-2 months. Cell survival, as measured by cell proliferation assays, indicated IC50s of 10µM for CDK4/6 Resistant LS141 and 15 µM for CDK4/6 Resistant DDLS, which is more than ten times higher than the sensitive parental cells. We also compared the established CDK4/6 resistant cell lines to palbociclib and found that resistance was similar to the ribociclib effect. Cell cycle changes to CDK4/6 resistant cell lines exhibited no effect of G1 arrest when exposed to higher concentrations of the drug. We also examined the effect of ribociclib and palbociclib on p-RB after treatment in sensitive parental and CDK4/6 resistant liposarcoma cell lines and showed that inhibition of p-Rb occurred at lower doses of 250-500 nM in the sensitive cell lines compared to higher doses of 2-8 µM in the resistant cell lines. Overexpression of cyclin E1 and p-CDK2 were prominent in the resistant cell lines when examined by western blot. siRNA suppression of cyclin E showed a reversal of resistance as determined by proliferation assays. We also observed induction of immunological markers including PD-L1, HLA-A2 and B2M (MHC-1 class) with CDK4/6 inhibitor treatment in the sensitive cell lines. Flow cytometry showed HLA-A2 and B2M increased in the resistant LS141 while B2M only increased in the resistant DDLS compared to their parental cell lines. We also observed PD-L1 increased in the resistant LS141 while it decreased in the resistant DDLS when exposed to the drug over long periods of time. Cytokine production in the CDK4/6 resistant liposarcoma was also seen with IL-8 increasing in resistant LS141 and IL-6 increasing in resistant DDLS cell line. Taken together these results indicate that cyclin E1/CDK2 activation is a mechanism of resistance to CDK4/6 inhibitors in WD/DD-liposarcoma and that CDK4/6 inhibition induces immunologic effects including induction of PD-L1 that are modified over time in CDK4/6 resistant liposarcoma cells. These results would support CDK2/4/6 targeting in this disease and the development of combination trials with PD-L1 inhibitors.
Citation Format: Elgilda Musi, Parag P. Patwardhan, Matthew Ingham, Gary K. Schwartz. Pre-clinical evaluation of CDK4/6 inhibitor resistance and immunologic modification in liposarcoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5289.