Kristen Jones

Glioblastoma Eradication Following Immune Checkpoint Blockade in an Orthotopic, Immunocompetent Model

Glioblastoma has been especially challenging to treat. In a systematic analysis of combinations of checkpoint therapies in a murine model, some single and dual immunotherapies increased intratumoral effectors, reduced suppressors, and eliminated the tumors. Inhibition of immune checkpoints, including cytotoxic T-lymphocyte antigen-4 (CTLA-4), programmed death-1 (PD-1), and its ligand PD-L1, has demonstrated exciting and durable remissions across a spectrum of malignancies. Combinatorial regimens blocking complementary immune checkpoints further enhance the therapeutic benefit. The activity of these agents for patients with glioblastoma, a generally lethal primary brain tumor associated with significant systemic and microenvironmental immunosuppression, is not known. We therefore systematically evaluated the antitumor efficacy of murine antibodies targeting a broad panel of immune checkpoint molecules, including CTLA-4, PD-1, PD-L1, and PD-L2 when administered as single-agent therapy and in combinatorial regimens against an orthotopic, immunocompetent murine glioblastoma model. In these experiments, we observed long-term tumor-free survival following single-agent anti–PD-1, anti–PD-L1, or anti–CTLA-4 therapy in 50%, 20%, and 15% of treated animals, respectively. Combination therapy of anti–CTLA-4 plus anti–PD-1 cured 75% of the animals, even against advanced, later-stage tumors. In long-term survivors, tumor growth was not seen upon intracranial tumor rechallenge, suggesting that tumor-specific immune memory responses were generated. Inhibitory immune checkpoint blockade quantitatively increased activated CD8+ and natural killer cells and decreased suppressive immune cells in the tumor microenvironment and draining cervical lymph nodes. Our results support prioritizing the clinical evaluation of PD-1, PD-L1, and CTLA-4 single-agent targeted therapy as well as combination therapy of CTLA-4 plus PD-1 blockade for patients with glioblastoma. Cancer Immunol Res; 4(2); 124–35. ©2015 AACR.

D-2-hydroxyglutarate produced by mutant IDH2 causes cardiomyopathy and neurodegeneration in mice

Mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) have been discovered in several cancer types and cause the neurometabolic syndrome D2-hydroxyglutaric aciduria (D2HGA). The mutant enzymes exhibit neomorphic activity resulting in production of D2-hydroxyglutaric acid (D-2HG). To study the pathophysiological consequences of the accumulation of D-2HG, we generated transgenic mice with conditionally activated IDH2(R140Q) and IDH2(R172K) alleles. Global induction of mutant IDH2 expression in adults resulted in dilated cardiomyopathy, white matter abnormalities throughout the central nervous system (CNS), and muscular dystrophy. Embryonic activation of mutant IDH2 resulted in more pronounced phenotypes, including runting, hydrocephalus, and shortened life span, recapitulating the abnormalities observed in D2HGA patients. The diseased hearts exhibited mitochondrial damage and glycogen accumulation with a concordant up-regulation of genes involved in glycogen biosynthesis. Notably, mild cardiac hypertrophy was also observed in nude mice implanted with IDH2(R140Q)-expressing xenografts, suggesting that 2HG may potentially act in a paracrine fashion. Finally, we show that silencing of IDH2(R140Q) in mice with an inducible transgene restores heart function by lowering 2HG levels. Together, these findings indicate that inhibitors of mutant IDH2 may be beneficial in the treatment of D2HGA and suggest that 2HG produced by IDH mutant tumors has the potential to provoke a paraneoplastic condition.

Preclinical antitumor efficacy of selective exportin 1 inhibitors in glioblastoma

BACKGROUND Glioblastoma (GBM) is poorly responsive to current chemotherapy. The nuclear transporter exportin 1 (XPO1, CRM1) is often highly expressed in GBM, which may portend a poor prognosis. Here, we determine the efficacy of novel selective inhibitors of nuclear export (SINE) specific to XPO1 in preclinical models of GBM. METHODS Seven patient-derived GBM lines were treated with 3 SINE compounds (KPT-251, KPT-276, and Selinexor) in neurosphere culture conditions. KPT-276 and Selinexor were also evaluated in a murine orthotopic patient-derived xenograft (PDX) model of GBM. Cell cycle effects were assayed by flow cytometry in vitro and immunohistochemistry in vivo. Apoptosis was determined by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and caspase 3/7 activity assays. RESULTS Treatment of GBM neurosphere cultures with KPT-276, Selinexor, and KPT-251 revealed dose-responsive growth inhibition in all 7 GBM lines [range of half-maximal inhibitory concentration (IC50), 6-354 nM]. In an orthotopic PDX model, treatment with KPT-276 and Selinexor demonstrated pharmacodynamic efficacy, significantly suppressed tumor growth, and prolonged animal survival. Cellular proliferation was not altered with SINE treatment. Instead, induction of apoptosis was apparent both in vitro and in vivo with SINE treatment, without overt evidence of neurotoxicity. CONCLUSIONS SINE compounds show preclinical efficacy utilizing in vitro and in vivo models of GBM, with induction of apoptosis as the mechanism of action. Selinexor is now in early clinical trials in solid and hematological malignancies. Based on these preclinical data and excellent brain penetration, we have initiated clinical trials of Selinexor in patients with relapsed GBM.

Abstract 572: Inhibition of IDO1 with epacadostat enhances anti-tumor efficacy of PD-1 blockade in a syngeneic glioblastoma (GBM) model

Purpose: To determine if epacadostat, an oral indoleamine 2,3-dioxygenase (IDO1) inhibitor has therapeutic benefit against GBM when administered as single agent and with PD-1 blocking antibody. Methods: An initial survival experiment was performed to assess efficacy and was followed by an identical repeat experiment for validation. 1X10 5 luciferized GL261 cells, a murine GBM tumor line derived from intracerebral methylcholanthrene implantation, were stereotactically implanted intracranially in albino syngeneic C57BL/6 mice. Mice with increasing bioluminescence on days 3 and 6 were randomized (n=8/group) to receive treatment beginning on day 6: anti-PD-1 (332.8H3, mouse IgG1; 500 μg intraperitoneal (IP) on day 6, 250 μg q 3 days X 7); epacadostat (Incyte Corporation, orally dosed at 300 mg/kg/day for 5 days on/2days off for 3 weeks); anti-PD-1 + epacadostat; and control therapy (isotype IgG antibody IP and 0.5% methocel in water). Tumor response assessments were performed by quantifying bioluminescence and survival. A re-challenge experiment was performed in long-term survivors to assess for tumor immune responses capable of preventing relapse. All long-term surviving mice (defined as ≥ 100 days) from the efficacy experiment were injected with 1X10 5 GL261 cells in the contralateral hemisphere and followed for survival. Results: In both preclinical efficacy experiments, median survival in the epacadostat monotherapy group did not differ from controls (approximately 30 days). Four of 8 mice (50%) treated with anti-PD-1 were long-term survivors in both efficacy experiments. In the epacadostat plus anti-PD-1 combination group, 81% of the mice were long-term survivors (7 of 8 in experiment 1 and 6 of 8 in experiment 2). Of note, none of the long-term surviving mice developed evidence of tumor; thus the median survival among the anti-PD-1 and epacadostat plus anti-PD-1 combination groups were both > 100 days. In the re-challenge study, all of the mice who underwent GL261 re-inoculation survived > 100 more days with no evidence of tumor recurrence. Conclusions: IDO1 inhibition with epacadostat increased the eradication rate of anti-PD-1 therapy in an orthotopic syngeneic GBM model and long term survivors rejected tumor following orthotopic re-challenge. Further combinatorial studies incorporating IDO inhibitor therapy for GBM, including mechanistic studies, are warranted. Citation Format: David A. Reardon, Prafulla C. Gokhale, Sarah R. Klein, Kristen L. Jones, Paul T. Kirschmeier, Maria Speranza, Holly Koblish, Peggy Scherle, Lance Leopold, Robert Newton, Gordon J. Freeman. Inhibition of IDO1 with epacadostat enhances anti-tumor efficacy of PD-1 blockade in a syngeneic glioblastoma (GBM) model [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 572. doi:10.1158/1538-7445.AM2017-572

Development and validation of a new MRI simulation technique that can reliably estimate optimal in vivo scanning parameters in a glioblastoma murine model

Background Magnetic Resonance Imaging (MRI) relies on optimal scanning parameters to achieve maximal signal-to-noise ratio (SNR) and high contrast-to-noise ratio (CNR) between tissues resulting in high quality images. The optimization of such parameters is often laborious, time consuming, and user-dependent, making harmonization of imaging parameters a difficult task. In this report, we aim to develop and validate a computer simulation technique that can reliably provide “optimal in vivo scanning parameters” ready to be used for in vivo evaluation of disease models. Methods A glioblastoma murine model was investigated using several MRI imaging methods. Such MRI methods underwent a simulated and an in vivo scanning parameter optimization in pre- and post-contrast conditions that involved the investigation of tumor, brain parenchyma and cerebrospinal fluid (CSF) CNR values in addition to the time relaxation values of the related tissues. The CNR tissues information were analyzed and the derived scanning parameters compared in order to validate the simulated methodology as a reliable technique for “optimal in vivo scanning parameters” estimation. Results The CNRs and the related scanning parameters were better correlated when spin-echo-based sequences were used rather than the gradient-echo-based sequences due to augmented inhomogeneity artifacts affecting the latter methods. “Optimal in vivo scanning parameters” were generated successfully by the simulations after initial scanning parameter adjustments that conformed to some of the parameters derived from the in vivo experiment. Conclusion Scanning parameter optimization using the computer simulation was shown to be a valid surrogate to the in vivo approach in a glioblastoma murine model yielding in a better delineation and differentiation of the tumor from the contralateral hemisphere. In addition to drastically reducing the time invested in choosing optimal scanning parameters when compared to an in vivo approach, this simulation program could also be used to harmonize MRI acquisition parameters across scanners from different vendors.

Abstract 4560: Anti-tumor activity of a fully human anti-CD27 monoclonal antibody in a transgenic mouse model

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL The costimulatory molecule CD27 is a member of the TNF receptor superfamily, and is constitutively expressed on the majority of mature T cells, memory B cells, and a portion of NK cells. The interaction of CD27 with its ligand CD70 plays an important role in the activation, proliferation, survival, and maturation of effector capacity and memory in T cells; in clonal B cell expansion and germinal center formation; and in NK cell cytolytic activity. Previous published work demonstrated that agonistic anti-mouse CD27 mAbs, given without a DC maturation signal, have potent anti-tumor activity through boosting of T cell immunity. To explore the therapeutic potential of this target, a panel of fully human antibodies recognizing human CD27 was generated using human Ig transgenic mice immunized with recombinant CD27. These anti-CD27 mAbs showed specific and high affinity binding to recombinant CD27 and CD27-expressing lymphoma cells. Human CD27-transgenic (hCD27-Tg) mice were generated to evaluate the anti-CD27 mAbs in vivo. The CD27-Tg mouse line was back-crossed onto a Balb/c background in order to assess anti-tumor activity using the BCL1 B-lymphoma line. Approximately 107 BCL1 cells were delivered i.v. to backcrossed animals on day 0. Doses of 150 to 600 ug of our lead anti-CD27 mAb (clone 1F5) were delivered to mice i.p. on days 3, 5, 7, 9 and 11. In multiple experiments, tumor growth in mice treated with the anti-CD27 mAb 1F5 was substantially delayed compared to that of control-treated mice. Further, many anti-CD27 treated animals did not show evidence of tumor growth several weeks past the point where all control animals had been euthanized. These data confirm that anti-human CD27 mAbs can mediate similar anti-tumor effects as the anti-mouse CD27 agonist antibodies. The 1F5 antibody was also demonstrated to cross-react with CD27 from cynomolgus monkeys, and in a small pilot study was well tolerated and without evidence of significant lymphocyte activation. Taken together the data support the therapeutic potential of this anti-CD27 mAb to enhance immune responses in cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4560. doi:10.1158/1538-7445.AM2011-4560