Abstract 2501: Inhibition of the PI3K pathway potentiates temozolomide effects in pediatric glioblastoma and results in alterations in glucose and choline metabolism detected by MRS
Authors: Al-Saffar Nada , Marshall Lynley , Jackson Laura Elizabeth , Raynaud Florence , Clarke Paul , Jones Chris , Workman Paul , Pearson Andrew , Leach Martin 
Published: August 2012

Introduction: Children with high grade-glioma have a poor outcome with a 5-year survival rate of less than 20%. The use of adjuvant temozolomide (TMZ) has now been well accepted as a standard of care in glioblastoma. However, it does not seem to improve survival in children. We have recently shown that resistance to TMZ in pediatric glioma cell lines can be reversed by PI3K pathway inhibition. The aim of this study was to identify whether combination of PI3K pathway inhibition with TMZ would result in metabolic changes detectable by magnetic resonance spectroscopy (MRS), and potentially can be used as a noninvasive method of monitoring response in early stage clinical trials in children with glioma. Methods: The MGMT-independent TMZ-resistant pediatric glioblastoma cell line KNS42 was treated for 48h with dual PI3K/mTOR inhibitor PI-103 (2xGI50), TMZ (2xGI50) or with the combination of both (each at 2xGI50). Control and treated cells were extracted and the aqueous fractions were analyzed by 1H MRS. Results and Discussion: The number of treated KNS42 cells was significantly reduced relative to their controls, to 79±8% and 65±10% (P&lt;0.02) following treatment with PI-103 and TMZ respectively. Combination of PI-103 and TMZ further reduced the number of treated vs. control cells (down to 54±6%, P&lt;0.01). 1H MRS analysis showed that PI-103 treatment caused a decrease in lactate, phosphocholine (PC) and total choline (tCho) levels down to 48±17%, 52±11%, 64±14% (P&lt;0.02) respectively. In contrast, treament with TMZ resulted in an increase in glycerolphosphocholine (GPC) up to 176±43% (P=0.05). Combination of PI-103 with TMZ retained the combined metabolic effects of both agents including a decrease in the levels of lactate (49±12%, P&lt;0.01) and PC (71±11%, P&lt;0.02) while an increase in GPC (190±40%, P&lt;0.03) levels. tCho was normalized by the opposing changes in PC and GPC and hence was similar to controls. To explore potential mechanisms underlying the detected metabolic changes, immunoblotting was used to assess treatment effects on protein levels of enzymes involved in glycolysis and choline metabolism. The decrease in PC levels was associated with a decrease in the protein levels of Choline kinase (ChoK), the enzyme responsible for choline phosphorylation into PC. ChoK was not affected by TMZ single treatment leading to no change in PC levels. Furthermore, a decrease in hexokinase II protein was detected following PI-103 treatment and the combination, but not with TMZ alone, suggesting a mechanism for depletion of lactate. Conclusions: Our results show that using MRS we can detect distinct biomarkers following treatment with PI-103 compared to TMZ. Choline metabolites and lactate are potential noninvasive biomarkers for monitoring response to PI3K/mTOR inhibitors both as single agents and in combination with cytotoxic therapy in children with glioma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2501. doi:1538-7445.AM2012-2501