Objective: Glioblastoma (GBM) is one of the deadliest brain tumors both in adults and children. While next-generation sequencing rapidly identifies whole exome/genome gene mutations, the efficacy of targeting mutated genes/pathways has not been systematically analyzed. Additionally, it remains unknown if neurospheres (enriched with cancer stem cells) and non-stem monolayer tumor cells will respond equally to the same target therapies.
Methods: Matching pairs of neurosphere and monolayer cell cultures from 4 molecularly characterized PDOX mouse models established from distinct clinical stages of pediatric glioma (treatment-nave, early recurrent, terminal, and radiation-induced) were treated with 7,913 drugs (mostly in clinical use or trials) at 4 doses (0-10 uM) for 3 and 7 days. Dose-response curves and active area under the curve (AAUC) were constructed to identify active drugs and correlate with model-specific gene mutations.
Results: Relatively long-term treatment (7 day screening) was achieved for high-throughput drug screening and revealed time- and dose-dependent effects in most of the 4,629 compounds that were active in at least one culture at one time point. When gene mutations were druggable, not all inhibitors of the same family were active. In IC-4687GBM (a treatment-nave GBM), high NF1 mutation (<76-99% allele) frequency was found in patient tumor, xenografts, and cultured cells, but only 3/17 MEK inhibitors were active in neurospheres and 2/17 in monolayer cells (GDC0980 and PI-103 were active in both) on day 7. In IC-R0315GBM (from an autopsied terminal GBM) that carried PI3KCA mutation (allele frequency 22-27%), 5/33 PI3K inhibitors were active in neurospheres and 8/33 in monolayer cells (only Tremetinib was active in both) after 7 days. In IC-3752GBM (recurrent GBM) and ICb-1127AA (radiation-induced anaplastic astrocytoma), no druggable mutations were detected. The number of active drugs on day 7 was 366 in IC-4768GBM, 406 in IC-3752GBM, 284 in IC-R0315GBM, and 305 in ICb-1277AA. When the 4 matching pairs of neurospheres and monolayers were compared, the agents active in both cultures ranged from 36% to 60%, active only in neurosphere from 10.3% to 25%, and active only in monolayer cells from 14.8% to 53%. Subsequent in vivo validation using MLN8327 in IC-4687GBM and IC-R0315GBM showed that effective targeting of both neurosphere and monolayer was required for significantly-improved animal survival times.
Conclusion: We showed that long-term treatment is feasible for high-throughput drug screening. Targeting druggable mutations can be achieved but only by a fraction of specific agents. Neurospheres and monolayer cells do not always respond equally toward the same drugs, and effective targeting of both subpopulations is needed to generate prolonged animal survival times.
Citation Format: Lin Qi, Yuchen Du, Mari Kogiso, Goeun Bae, Frank Braun, Holly Lindsay, Huiyuan Zhang, Sibo Zhao, Sarah Injac, Patricia Baxter, Jack Su, Michael Mancini, Oliver Hampton, William Parsons, Murali Chintagumpala, Clifford Stephan, Peter Davies, Xiao-Nan Li, Xiao-Nan Li. Identification of personalized active agents in pediatric gliomas through high-throughput drug screening in matching pairs of patient derived orthotopic xenograft (PDOX) neurosphere and monolayer cells [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 4214. doi:10.1158/1538-7445.AM2017-4214