Multiple Treatment Cycles of Neural Stem Cell Delivered Oncolytic Adenovirus for the Treatment of Glioblastoma

Tumor tropic neural stem cells (NSCs) can improve the anti-tumor efficacy of oncovirotherapy agents by protecting them from rapid clearance by the immune system and delivering them to multiple distant tumor sites. We recently completed a first-in-human trial assessing the safety of a single intracerebral dose of NSC-delivered CRAd-Survivin-pk7 (NSC.CRAd-S-pk7) combined with radiation and chemotherapy in newly diagnosed high-grade glioma patients. The maximum feasible dose was determined to be 150 million NSC.CRAd-Sp-k7 (1.875 × 1011 viral particles). Higher doses were not assessed due to volume limitations for intracerebral administration and the inability to further concentrate the study agent. It is possible that therapeutic efficacy could be maximized by administering even higher doses. Here, we report IND-enabling studies in which an improvement in treatment efficacy is achieved in immunocompetent mice by administering multiple treatment cycles intracerebrally. The results imply that pre-existing immunity does not preclude therapeutic benefits attainable by administering multiple rounds of an oncolytic adenovirus directly into the brain.

Identifying the Involvement of Pro-Inflammatory Signal in Hippocampal Gene Expression Changes after Experimental Ischemia: Transcriptome-Wide Analysis

Acute cerebral ischemia induces distant inflammation in the hippocampus; however, molecular mechanisms of this phenomenon remain obscure. Here, hippocampal gene expression profiles were compared in two experimental paradigms in rats: middle cerebral artery occlusion (MCAO) and intracerebral administration of lipopolysaccharide (LPS). The main finding is that 10 genes (Clec5a, CD14, Fgr, Hck, Anxa1, Lgals3, Irf1, Lbp, Ptx3, Serping1) may represent key molecular links underlying acute activation of immune cells in the hippocampus in response to experimental ischemia. Functional annotation clustering revealed that these genes built the same clusters related to innate immunity/immunity/innate immune response in all MCAO differentially expressed genes and responded to the direct pro-inflammatory stimulus group. The gene ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway analyses also indicate that LPS-responding genes were the most abundant among the genes related to “positive regulation of tumor necrosis factor biosynthetic process”, “cell adhesion”, “TNF signaling pathway”, and “phagosome” as compared with non-responding ones. In contrast, positive and negative “regulation of cell proliferation” and “HIF-1 signaling pathway” mostly enriched with genes that did not respond to LPS. These results contribute to understanding genomic mechanisms of the impact of immune/inflammatory activation on expression of hippocampal genes after focal brain ischemia.

Neurofunctional and neuroimaging readouts for designing a preclinical stem-cell therapy trial in experimental stroke

With the aim of designing a preclinical study evaluating an intracerebral cell-based therapy for stroke, an observational study was performed in the rat suture model of ischemic stroke. Objectives were threefold: (i) to characterize neurofunctional and imaging readouts in the first weeks following transient ischemic stroke, according to lesion subtype (hypothalamic, striatal, corticostriatal); (ii) to confirm that intracerebral administration does not negatively impact these readouts; and (iii) to calculate sample sizes for a future therapeutic trial using these readouts as endpoints. Our results suggested that the most relevant endpoints were side bias (staircase test) and axial diffusivity (AD) (diffusion tensor imaging). Hypothalamic-only lesions did not affect those parameters, which were close to normal. Side bias in striatal lesions reached near-normal levels within 2 weeks, while rats with corticostriatal lesions remained impaired until week 14. AD values were decreased at 4 days and increased at 5 weeks post-surgery, with a subtype gradient: hypothalamic < striatal < corticostriatal. Intracerebral administration did not impact these readouts. After sample size calculation (18-147 rats per group according to the endpoint considered), we conclude that a therapeutic trial based on both readouts would be feasible only in the framework of a multicenter trial.

Intracerebral Administration of a Ligand-ASO Conjugate Selectively Reduces α-Synuclein Accumulation in Monoamine Neurons of Double Mutant Human A30P*A53T*α-Synuclein Transgenic Mice

α-Synuclein (α-Syn) protein is involved in the pathogenesis of Parkinson’s disease (PD). Point mutations and multiplications of the α-Syn, which encodes the SNCA gene, are correlated with early-onset PD, therefore the reduction in a-Syn synthesis could be a potential therapy for PD if delivered to the key affected neurons. Several experimental strategies for PD have been developed in recent years using oligonucleotide therapeutics. However, some of them have failed or even caused neuronal toxicity. One limiting step in the success of oligonucleotide-based therapeutics is their delivery to the brain compartment, and once there, to selected neuronal populations. Previously, we developed an indatraline-conjugated antisense oligonucleotide (IND-1233-ASO), that selectively reduces α-Syn synthesis in midbrain monoamine neurons of mice, and nonhuman primates. Here, we extended these observations using a transgenic male mouse strain carrying both A30P and A53T mutant human α-Syn (A30P*A53T*α-Syn). We found that A30P*A53T*α-Syn mice at 4–5 months of age showed 3.5-fold increases in human α-Syn expression in dopamine (DA) and norepinephrine (NE) neurons of the substantia nigra pars compacta (SNc) and locus coeruleus (LC), respectively, compared with mouse α-Syn levels. In parallel, transgenic mice exhibited altered nigrostriatal DA neurotransmission, motor alterations, and an anxiety-like phenotype. Intracerebroventricular IND-1233-ASO administration (100 µg/day, 28 days) prevented the α-Syn synthesis and accumulation in the SNc and LC, and recovered DA neurotransmission, although it did not reverse the behavioral phenotype. Therefore, the present therapeutic strategy based on a conjugated ASO could be used for the selective inhibition of α-Syn expression in PD-vulnerable monoamine neurons, showing the benefit of the optimization of ASO molecules as a disease modifying therapy for PD and related α-synucleinopathies.

Intranasal Oxytocin has sympathoexcitatory effects on vascular tone in healthy males

Objective: Oxytocin appears to be involved in the neuroendocrine regulation of sympathetic blood pressure (BP) homeostasis. In animals, intracerebral administration of oxytocin induces BP-relevant sympathetic activation. In humans, central nervous effects of oxytocin on BP regulation remain unclear. Intranasal administration supposedly delivers oligopeptides like oxytocin directly to the brain. We investigated the effects of intranasal oxytocin on sympathetic vascular baroreflex function in humans using microneurographic techniques. Methods: In a balanced, double-blind cross-over design oxytocin or placebo was administered intranasally to 12 lean healthy males (age 25±4 years). MSNA was assessed microneurographically before (pre), 30-45 (post-I) and 105-120 minutes (post-II) after oxytocin administration. Baroreflex was challenged via graded infusions of vasoactive drugs and correlation of BP with MSNA and heart rate (HR) defined baroreflex function. Experiments were conducted in the afternoon after a 5h fasting period. Results: After oxytocin, resting MSNA (burst rate and total activity) showed significant net-increases from pre to post-II compared to placebo (∆-increase: +4.3±1.2 (oxytocin) vs. +2.2±1.4 burst/min (placebo), ANOVA p<0.05; total activity 184±11.5% (oxytocin) vs. 121±14.3% (placebo), ANOVA; p=0.01). This was combined with a small but significant net-increase in resting diastolic BP, while systolic and mean arterial BP or HR as well as baroreflex sensitivity at vasoactive drug challenge were not altered. Conclusion: Intranasally administered oxytocin induced vasoconstrictory sympathoactivation in healthy male humans. The concomitant increase of diastolic BP was most likely attributable to increased vascular tone. This suggests oxytocin-mediated upward resetting of the vascular baroreflex setpoint at centers superordinate to the mere baroreflex-feedback-loop.

ATIM-38. GLITIPNI: A PHASE 1B CLINICAL TRIAL COMBINING SURGICAL RESECTION WITH DIRECT INTRACEREBRAL INJECTION OF IMMUNE CHECKPOINT INHIBITORS IN PATIENTS WITH RECURRENT GLIOBLASTOMA

INTRODUCTION Intravenous (iv) administration of PD-1 blocking mAb is largely ineffective for the treatment of recurrent glioblastoma (rGB). Combination of iv-ipilimumab (IPI) plus nivolumab (NIVO) is associated with a high incidence of irAE. Intracerebral (ic) administration of immune-checkpoint inhibiting mAb following the resection of rGB could be a more effective and safer alternative to iv-dosing. METHODS Patients underwent maximal safe resection of their rGB followed by ic-injection of 10mg IPI (cohort-1) or 5mg IPI plus 10mg NIVO (cohort-2) in the wall of the resection cavity. In both cohorts 10mg nivolumab was administered iv for a max of 6 doses, starting 1 day pre-operatively. RESULTS 21 pts were included (3 in C-1, 18 in C-2; 8F/13M; median age 56y [range 38–72]; 17 de novo GB, 4 secGB). All patients underwent maximal safe surgical resection followed by ic-injection of IPI and NIVO as planned. Median number of iv-administrations of NIVO was 5 (range 1–8). Treatment was generally well tolerated. Postoperatively, 2 patients experienced a G3 symptomatic increase in perilesional cerebral edema with neurological deterioration, reversible upon steroid treatment. One patient had worsening neurological symptoms related to an inflammatory intracerebral cyst at the resection site, requiring surgical decompression 4 months post-study treatment. Most frequent AEs were fatigue (2pts G3, 8pts G2), postoperative fever (11pts G1) and headache (3pts G2); 1pt developed G3 pneumonitis. No other immune-related AEs or treatment-related deaths occurred. After median follow-up of 60 weeks, median PFS is 14.4 weeks (95% CI 11.2–17.6); 11/21 patients are alive, and 1- and 2y-OS% are respectively 46% (95% CI 19- 73%), and15% (95% CI 0–42%). CONCLUSION This is the first study demonstrating the safety and activity of combined surgical resection of rGB with local intracerebral administration of immune checkpoint-inhibiting mAb. Survival compares favorably to historical controls justifying further investigation of this experimental therapy.

SCIDOT-30. GLITIPNI: A PHASE 1B CLINICAL TRIAL COMBINING SURGICAL RESECTION WITH DIRECT INTRACEREBRAL INJECTION OF IMMUNE CHECKPOINT INHIBITORS IN PATIENTS WITH RECURRENT GLIOBLASTOMA

INTRODUCTION Intravenous (iv) administration of PD-1 blocking mAb is largely ineffective for the treatment of recurrent glioblastoma (rGB). Combination of iv-ipilimumab (IPI) plus nivolumab (NIVO) is associated with a high incidence of irAE. Intracerebral (ic) administration of immune-checkpoint inhibiting mAb following the resection of rGB could be a more effective and safer alternative to iv-dosing. METHODS Patients underwent maximal safe resection of their rGB followed by ic-injection of 10mg IPI (cohort-1) or 5mg IPI plus 10mg NIVO (cohort-2) in the wall of the resection cavity. In both cohorts 10mg nivolumab was administered iv for a max of 6 doses, starting 1 day pre-operatively. RESULTS 21 pts were included (3 in C-1, 18 in C-2; 8F/13M; median age 56y [range 38–72]; 17 de novo GB, 4 secGB). All patients underwent maximal safe surgical resection followed by ic-injection of IPI and NIVO as planned. Median number of iv-administrations of NIVO was 5 (range 1–8). Treatment was generally well tolerated. Postoperatively, 2 patients experienced a G3 symptomatic increase in perilesional cerebral edema with neurological deterioration, reversible upon steroid treatment. One patient had worsening neurological symptoms related to an inflammatory intracerebral cyst at the resection site, requiring surgical decompression 4 months post-study treatment. Most frequent AEs were fatigue (2pts G3, 8pts G2), postoperative fever (11pts G1) and headache (3pts G2); 1pt developed G3 pneumonitis. No other immune-related AEs or treatment-related deaths occurred. After median follow-up of 60 weeks, median PFS is 14.4 weeks (95% CI 11.2–17.6); 11/21 patients are alive, and 1- and 2y-OS% are respectively 46% (95% CI 19- 73%), and15% (95% CI 0–42%). CONCLUSION This is the first study demonstrating the safety and activity of combined surgical resection of rGB with local intracerebral administration of immune checkpoint-inhibiting mAb. Survival compares favorably to historical controls justifying further investigation of this experimental therapy.