Yancey Gillespie

A phase I trial of intratumoral administration of reovirus in patients with histologically confirmed recurrent malignant gliomas.

Reovirus is an oncolytic virus with activity in in vivo models of malignant gliomas (MGs). The primary aims were to determine the dose-limiting toxicity (DLT) and maximum tolerated dose (MTD) of intratumoral administration of reovirus in patients with recurrent MGs. Response, survival, and time to progression (TTP) were secondary aims. Patients were adults, had Karnofsky Performance score > or = 60, received prior radiotherapy with or without chemotherapy, and had up to the third recurrence of MG. Reovirus was administered intratumorally stereotactically at 1 x 10(7), 1 x 10(8), or 1 x 10(9) tissue culture infectious dose 50 (TCID50) in a volume of 0.9 ml. Twelve patients were treated at three dose levels (3, 6, and 3 patients, respectively). Seven were men, median Karnofsky Performance score was 80, and median age was 53.5 years. There were no grade III or IV adverse events (AEs) definitely or probably related to treatment. Ten patients had tumor progression, one had stabilization, and one was not evaluable for response. Median survival was 21 weeks (range, 6-234), and one is alive 54 months after treatment. Median TTP was 4.3 weeks (range, 2.6-39). An MTD was not reached. The intratumoral administration of the genetically unmodified reovirus was well tolerated using these doses and schedule, in patients with recurrent MG.

Reactive species balance via GTP cyclohydrolase I regulates glioblastoma growth and tumor initiating cell maintenance

Background Depending on the level, differentiation state, and tumor stage, reactive nitrogen and oxygen species inhibit or increase cancer growth and tumor initiating cell maintenance. The rate-limiting enzyme in a pathway that can regulate reactive species production but has not been thoroughly investigated in glioblastoma (GBM; grade IV astrocytoma) is guanosine triphosphate (GTP) cyclohydrolase 1 (GCH1). We sought to define the role of GCH1 in the regulation of GBM growth and brain tumor initiating cell (BTIC) maintenance. Methods We examined GCH1 mRNA and protein expression in patient-derived xenografts, clinical samples, and glioma gene expression datasets. GCH1 levels were modulated using lentiviral expression systems, and effects on cell growth, self-renewal, reactive species production, and survival in orthotopic patient-derived xenograft models were determined. Results GCH1 was expressed in GBMs with elevated but not exclusive RNA and protein levels in BTICs in comparison to non-BTICs. Overexpression of GCH1 in GBM cells increased cell growth in vitro and decreased survival in an intracranial GBM mouse model. In converse experiments, GCH1 knockdown with short hairpin RNA led to GBM cell growth inhibition and reduced self-renewal in association with decreased CD44 expression. GCH1 was critical for controlling reactive species balance, including suppressing reactive oxygen species production, which mediated GCH1 cell growth effects. In silico analyses demonstrated that higher GCH1 levels in glioma patients correlate with higher glioma grade, recurrence, and worse survival. Conclusions GCH1 expression in established GBMs is pro-tumorigenic, causing increased growth due, in part, to promotion of BTIC maintenance and suppression of reactive oxygen species.

Abstract 1925: Modeling physiologic microenvironments in three-dimensional microtumors facilitates brain tumor initiating cell maintenance

Development of effective novel anti-tumor treatments will require improved in vitro models that incorporate physiologic microenvironments and maintain intratumoral heterogeneity including tumor initiating cells. Brain tumor initiating cells (BTIC) are a target for cancer therapy because they are highly tumorigenic and contribute to tumor angiogenesis, invasion, and therapeutic resistance. Current leading studies rely on BTIC isolation from patient-derived xenografts followed by propagation as neurospheres. As this process is expensive and time-consuming, we determined whether three-dimensional microtumors were an alternative in vitro method for modeling tumor growth via BITC maintenance and/or enrichment. Brain tumor cells were grown as neurospheres or as microtumors produced using a human-derived biomatrix HuBiogelTM and maintained with physiologically relevant microenvironments. Percentages of BITCs were determined based on cell surface marker expression (CD133), label retention (carboxyfluorescein succinimidyl ester; CFSE), and tumorsphere formation capacity. Our data demonstrate that expansion of brain tumor cells as hypoxic and nutrient restricted microtumors significantly increased the percentage of both CD133+ and CFSE+ cells. We further demonstrate that BTIC-marker positive cells isolated from microtumors maintain neurosphere formation capacity in the in vitro limiting dilution assay and tumorigenic potential in vivo. These data demonstrate that microtumors can be a useful three-dimensional biological model for the study of BTIC maintenance and targeting. Citation Format: Ashley Gilbert, Kiera Walker, Anh Tran, Yancey Gillespie, Raj Singh, Anita B. Hjelmeland. Modeling physiologic microenvironments in three-dimensional microtumors facilitates brain tumor initiating cell maintenance [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 1925. doi:10.1158/1538-7445.AM2017-1925

Abstract 1679: Combined efficacy of Cediranib and Quinacrine in glioma is enhanced by hypoxia and is associated with autophagic vacuole accumulation.

Despite robust vascularity of malignant gliomas, anti-angiogenic therapy largely fails to induce durable responses. We have previously reported that efficacy with Cediranib (Ced), a VEGF/PDGF receptor tyrosine kinase inhibitor, is synergistically enhanced via combination with late-stage autophagy inhibitor quinacrine (Quin), in intracranial 4C8 mouse glioma, resulting in decreased tumor vascularization and growth, increased tumor necrosis and improved mouse survival (#1905, AACR 2012). Our present study was aimed at investigating the role of autophagy (a cellular catabolic pathway that promotes tumor cell survival under hypoxic/nutrient stress), in this synergistic efficacy. MTS assays revealed dose-dependent reductions in cell viability for Ced and Quin under normal (Nrm) and hypoxic (0.5%O2, Hyp) conditions: IC50s(μM) under Nrm and Hyp were 2.7±0.1 and 2.4±0.2 for Ced, and 3.2±0.2 and 2.4±0.04 for Quin, respectively. Greater than additive combined efficacy for Ced+Quin occurred only under Hyp (cell viability reductions for 1μM C + 2.5μM Q: 78±7%(Hyp) vs. 31±3%(Nrm), p Citation Format: Merryl Lobo, Peter Kurre, Matthias Schabel, Yancey Gillespie, Randall Woltjer, Martin Pike. Combined efficacy of Cediranib and Quinacrine in glioma is enhanced by hypoxia and is associated with autophagic vacuole accumulation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1679. doi:10.1158/1538-7445.AM2013-1679