Hui Jun Lim1, Xiaochun Wang1, Philip Crowe1, David Goldstein1, Jia-Lin Yang2. 1. Sarcoma and Nano-oncology Group, Adult Cancer Program, Lowy Cancer Research Centre, Department of Medical Oncology and Surgery, Prince of Wales Clinical School, Faculty of Medicine, University of New South Wales, Randwick, NSW, Australia. 2. Sarcoma and Nano-oncology Group, Adult Cancer Program, Lowy Cancer Research Centre, Department of Medical Oncology and Surgery, Prince of Wales Clinical School, Faculty of Medicine, University of New South Wales, Randwick, NSW, Australia j.yang@unsw.edu.au.
Abstract
BACKGROUND/AIM: Sarcoma carries a poor prognosis prompting the need for targeted therapies aimed at deregulated signaling pathways. These include the PI3K/Akt/mTOR pathway commonly up-regulated in malignancies attributed to loss of PTEN expression. However, PTEN status and activation state of PI3K/Akt/mTOR pathway have not been comprehensively studied in sarcoma. The aims of this study were to characterise PTEN and Akt expression in a panel of sarcoma cell lines and then to examine mTOR inhibition using ridaforolimus. MATERIALS AND METHODS: PTEN genomic expression was analyzed using Sanger sequencing. PTEN, total Akt (tAkt) and phosphorylated Akt (pAkt) expression were quantified with western blot analysis. Antiproliferative effects of treatment regimens were designed using Chou & Talalay's isobologram and determined with crystal violet assay. RESULTS: Four cell lines had wild-type PTEN (exons 2 to 8), with normal protein expression. The GCT cell line had a missense mutation in exon 6 (C>T), associated with loss of PTEN protein expression. Increased pAkt expression was found in all cell lines following epidermal growth factor (EGF) stimulation, indicating that wild-type PTEN expression in four cell lines did not inhibit constitutive activation of PI3K/Akt/mTOR pathway. Nonetheless, all cell lines demonstrated sensitivity to ridaforolimus within a clinically relevant dose-range (half-maximal inhibitory concentration (IC50)=0.7-10 nM). CONCLUSION: PTEN mutation is rare in sarcoma cell lines and constitutive activation of PI3K/Akt/mTOR is independent of PTEN status. Copyright
BACKGROUND/AIM: Sarcoma carries a poor prognosis prompting the need for targeted therapies aimed at deregulated signaling pathways. These include the PI3K/Akt/mTOR pathway commonly up-regulated in malignancies attributed to loss of PTEN expression. However, PTEN status and activation state of PI3K/Akt/mTOR pathway have not been comprehensively studied in sarcoma. The aims of this study were to characterise PTEN and Akt expression in a panel of sarcoma cell lines and then to examine mTOR inhibition using ridaforolimus. MATERIALS AND METHODS:PTEN genomic expression was analyzed using Sanger sequencing. PTEN, total Akt (tAkt) and phosphorylated Akt (pAkt) expression were quantified with western blot analysis. Antiproliferative effects of treatment regimens were designed using Chou & Talalay's isobologram and determined with crystal violet assay. RESULTS: Four cell lines had wild-type PTEN (exons 2 to 8), with normal protein expression. The GCT cell line had a missense mutation in exon 6 (C>T), associated with loss of PTEN protein expression. Increased pAkt expression was found in all cell lines following epidermal growth factor (EGF) stimulation, indicating that wild-type PTEN expression in four cell lines did not inhibit constitutive activation of PI3K/Akt/mTOR pathway. Nonetheless, all cell lines demonstrated sensitivity to ridaforolimus within a clinically relevant dose-range (half-maximal inhibitory concentration (IC50)=0.7-10 nM). CONCLUSION:PTEN mutation is rare in sarcoma cell lines and constitutive activation of PI3K/Akt/mTOR is independent of PTEN status. Copyright
Authors: Paulina Jagodzińska-Mucha; Paweł Sobczuk; Michał Mikuła; Anna Raciborska; Anna Dawidowska; Maria Kulecka; Katarzyna Bilska; Anna Szumera-Ciećkiewicz; Anna Kluska; Magdalena Piątkowska; Anna Bałabas; Piotr Rutkowski; Iwona Ługowska Journal: Contemp Oncol (Pozn) Date: 2021-12-29