Abstract
The phosphoinositide 3-kinase (PI3K) pathway is believed to be of key importance in pediatric glioblastoma. Novel inhibitors of the PI3K pathway are being developed and are entering clinical trials. Our aim is to identify potential non-invasive biomarkers of PI3K signaling pathway inhibition in pediatric glioblastoma using in vitro nuclear magnetic resonance (NMR) spectroscopy, to aid identification of target inhibition and therapeutic response in early phase clinical trials of PI3K inhibitors in childhood cancer. Treatment of SF188 and KNS42 human pediatric glioblastoma cell lines with the dual pan-Class I PI3K/mTOR inhibitor PI-103, inhibited the PI3K signaling pathway and resulted in a decrease in phosphocholine (PC), total choline (tCho) and lactate levels (p<0.02) as detected by phosphorus (31P)- and proton (1H)-NMR. Similar changes were also detected using the pan–Class I PI3K inhibitor GDC-0941 which lacks significant mTOR activity and is entering Phase II clinical trials. In contrast, the DNA damaging agent temozolomide (TMZ), which is used as current frontline therapy in the treatment of glioblastoma postoperatively (in combination with radiotherapy), increased PC, glycerophosphocholine (GPC) and tCho levels (p<0.04). PI-103-induced NMR changes were associated with alterations in protein expression levels of regulatory enzymes involved in glucose and choline metabolism including GLUT1, HK2, LDHA and CHKA. Our results show that by using NMR we can detect distinct biomarkers following PI3K pathway inhibition compared to treatment with the DNA-damaging anti-cancer agent TMZ. This is the first study reporting that lactate and choline metabolites are potential non-invasive biomarkers for monitoring response to PI3K pathway inhibitors in pediatric glioblastoma.
Highlights
40% of all pediatric brain tumors are astrocytomas, and of these some 15–20% are malignant gliomas, i.e. high-grade (WHO grade III and IV) tumors [1,2]
We report distinct metabolic changes including a decrease in the levels of lactate, phosphocholine (PC) and total choline following phosphoinositide 3-kinase (PI3K) pathway inhibition with PI-103 or GDC-0941, whereas there was an increase in PC, glycerophosphocholine (GPC) and tCho following treatment with TMZ
Treatment of the pediatric glioblastoma cell line SF188 with the dual pan-Class I PI3K/mTOR inhibitor PI-103 [14,15,16] for 8, 16 and 24 hours at pharmacologically active concentrations corresponding to 56GI50 (GI50 = 0.2 mM) resulted in an increase in G1 cell population and a decrease in S phase relative to control cells (Figure 1A, Table 1)
Summary
40% of all pediatric brain tumors are astrocytomas (gliomas), and of these some 15–20% are malignant gliomas, i.e. high-grade (WHO grade III and IV) tumors [1,2]. As well as numerous qualitative and quantitative differences in DNA copy number abnormalities between pediatric and adult HGG [3], childhood tumors are defined in part by the presence of specific somatic mutations in the gene encoding the histone H3.3 variant, H3F3A [7]. The K27 mutations are typically seen in younger pediatric patients with tumors arising in central locations e.g. brainstem or thalamus, whereas the G34 mutations arise in older pediatric/adolescent patients with tumors arising in the supratentorial (typically cerebral hemispheric) locations [3,7,8] These histone H3 mutations are not seen in adult HGG beyond approximately 30 years of age and are mutually exclusive with the high frequency of IDH1 mutations seen in adult populations between 35–45 years of age
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