Abstract 2292: Tyrosine kinome profiling of renal cell carcinoma (RCC)
Authors: Puco Katarina , Tahiri Andliena , Naji Faris , Heinrich Daniel , Kristenssen Vessela N. , Alfsen Glenny C. , Farkas Lorant , Nilsen Frode S. , Müller Stig , Oldenburg Jan , Geisler Jürgen 
Published: July 2021

Abstract
Background: Tyrosine kinase inhibitors (TKIs) constitute the backbone of treatment of metastatic (m) RCC. TKIs show great inter-individual variability of both efficacy and toxicity. The aim of this study was to compare the tyrosine kinase activity in normal and malignant kidney tissue and explore TKI effect on tumor tissue ex vivo.
Materials and methods: Human nephrectomy samples (n = 295) consecutively collected at Akershus University Hospital from 2013 to 2019 were available. For this study, 22 samples of patients with mRCC (twenty-one clear cell and one chromophobe histology) and 3 with non-metastatic disease (two clear cell and one papillary histology) were selected. Tumor and normal tissue from each patient were prepared and assessed by an experienced pathologist, and 5µg of protein lysates was extracted. Kinase activity profiling of 144 tyrosine kinase peptides was performed using PamChip Arrays for PAMSTATION12. Protein lysates from tumors were in addition treated with four different TKIs ex vivo, based on IC50; sunitinib (2.5 µM), cabozantinib (2.5 µM), pazopanib (10 µM) and tivozanib (2 µM). All data was processed and analyzed in Bionavigator V.6 (PamGene).
Results: The overall tyrosine kinase activity was increased in tumor tissue compared to normal tissue in all samples. The activity of 36 tyrosine kinases differed significantly (FDR &lt; 0.05), with 20 kinases showing higher and 16 showing lower phosphorylation in tumor compared to normal tissue. Furthermore, adding TKIs to tumor samples ex vivo revealed different potencies of the TKIs. Tivozanib and cabozantinib showed higher inhibition potency compared to sunitinib and pazopanib. Two well-defined peptide clusters were especially affected by tivozanib and cabozantinib exposure. Compared to untreated tumor tissue, the activity of 29 kinases including common TKI targets MET, EGFR, PDGFR and FGFR was significantly reduced in cluster 1. However, we also identified reduced activity of several other kinases i.e. RB, CDK2, CLB, CAM, ZAP70 and PDK1 involved in RAS, RAP1, PI3K-AKT and prostate cancer signaling pathways. In cluster 2, 21 kinases showed increased activity upon TKI exposure, amongst them JNK, ERK, erbB1/erbB2, Shp2 and PLCγ involved in MAPK, ERBB and proteoglycans signaling pathways, and possibly accounting for subsequent TKI resistance.
Conclusion: Kinase activity differs significantly between malignant and normal kidney tissue. Furthermore, we have demonstrated different inhibitory effects of TKIs ex-vivo and identified two peptide clusters significantly affected by TKI. Further analysis of affected kinases and signaling pathways as well as correlations with clinical data for the same group of patients will be performed. To our knowledge, similar systematic comprehensive analysis of tyrosine kinase activity and correlation to clinical data has not been reported previously.
Citation Format: Katarina Puco, Andliena Tahiri, Faris Naji, Daniel Heinrich, Vessela N. Kristenssen, Glenny C. Alfsen, Lorant Farkas, Frode S. Nilsen, Stig Müller, Jan Oldenburg, Jürgen Geisler. Tyrosine kinome profiling of renal cell carcinoma (RCC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2292.