Abstract LB-117: Dasatinib for the treatment of patients with non-small cell lung cancer harboring YES1 amplification
Authors: Garmendia Irati , Agorreta Jackeline , Pajares María J , Ajona Daniel , Alameda Daniel , Behrens Carmen , Wistuba Ignacio I. , Pio Ruben , Montuenga Luis 
Published: July 2017

Introduction

The characterization of still unknown genetic alterations is essential to develop novel and more effective treatments in lung cancer. The analysis of tumor molecular profiles from patients with NSCLC allowed us to identify the amplification of YES1 as a potential relevant molecular alteration in NSCLC. The aim of this project is the validation of this alteration as a therapeutic target in lung cancer.

Experimental procedures

Seven lung cancer cell lines of different histological subtypes with different YES1 gene copy number and mRNA expression levels were selected for the following assays. siRNA technology was used to specifically knockdown expression of YES1. Cell proliferation and survival were assessed by MTT and clonogenic assays. Apoptosis was analyzed by cell cytometry.

YES1 was overexpressed in A549 cell line (low-expressor) (A549-YES1). A549 cells transfected with empty vector were used as control (A549-Ø). Overexpression was confirmed by qPCR and Western blot. Cell proliferation was analyzed by MTT assays. Apoptosis was studied by cytometry after treatment with staurosporine.

14 NSCLC cell lines with high or low YES1 expression were treated with dasatinib (0, 0.01, 0.1, 1, 10 and 50 μM). Cell proliferation was assessed by MTT assays.

A549-YES1 and A549- Ø cells were subcutaneously injected in the flanks of 7-8-week-old male BALB/c-Rag2−/−-IL2γc−/− immunodeficient mice. Tumor volume was calculated.

YES1 expression was assessed using immunohistochemical techniques in 96 lung cancer patient samples and its corresponding paired non-tumoral tissue. For survival analysis, Kaplan-Meier survival curves and the log-rank test were used to examine differences in recurrence-free survival and overall survival. The results were validated in an independent series of 500 NSCLC patients.

Results

YES1 knockdown by siRNAs decreased proliferation and survival only in cell lines harboring YES1 amplification. YES1 downregulation led to apoptosis only in those cells. Constitutive overexpression of YES1 in A549 stimulated its proliferation and decreased apoptosis.

Dasatinib treatment dramatically inhibited proliferation in high YES1-expressing cell lines, whereas low YES1 cells were more resistant to dasatinib treatment (GI50s were four orders of magnitude higher in resistant cells).

Mice injected with A549-YES1 cells presented larger tumor volumes than mice injected with A549-Ø cells.

We evaluated the prognostic role of YES1 protein expression in two independent series of NSCLC patients. In both series, the multivariate analysis revealed that high YES1 expression is an independent poor prognostic factor for overall survival.

Conclusions

Our results indicate that YES1 is a novel therapeutic target in NSCLC. Moreover, amplification and high expression of YES1 may define the subset of patients that can potentially derive benefit from dasatinib.

Citation Format: Irati Garmendia, Jackeline Agorreta, María J Pajares, Daniel Ajona, Daniel Alameda, Carmen Behrens, Ignacio I. Wistuba, Ruben Pio, Luis Montuenga. Dasatinib for the treatment of patients with non-small cell lung cancer harboring YES1 amplification [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-117. doi:10.1158/1538-7445.AM2017-LB-117