Real-Time, in Vivo Correlation of Molecular Structure with Drug Distribution in the Brain Striatum Following Convection Enhanced Delivery.

The blood-brain barrier (BBB) represents a major obstacle in delivering therapeutics to brain lesions. Convection-enhanced delivery (CED), a method that bypasses the BBB through direct, cannula-mediated drug delivery, is one solution to maintaining increased, effective drug concentration at these lesions. CED was recently proven safe in a phase I clinical trial against diffuse intrinsic pontine glioma (DIPG), a childhood cancer. Unfortunately, the exact relationship between drug size, charge, and pharmacokinetic behavior in the brain parenchyma are difficult to observe in vivo. PET imaging of CED-delivered agents allows us to determine these relationships. In this study, we label different modifications of the PDGFRA inhibitor dasatinib with fluorine-18 or via a nanofiber-zirconium-89 system so that the effect of drug structure on post-CED behavior can accurately be tracked in vivo, via PET. Relatively unchanged bioactivity is confirmed in patient- and animal-model-derived cell lines of DIPG. In naïve mice, significant individual variability in CED drug clearance is observed, highlighting a need to accurately understand drug behavior during clinical translation. Generally, the half-life for a drug to clear from a CED site is short for low molecular weight dasatinib analogs that bare different charge; 1-3 (1, 32.2 min (95% CI: 27.7-37.8), 2, 44.8 min (27.3-80.8), and 3, 71.7 min (48.6-127.6) minutes) and is much longer for a dasatinib-nanofiber conjugate, 5, (42.8-57.0 days). Positron emission tomography allows us to accurately measure the effect of drug size and charge in monitoring real-time drug behavior in the brain parenchyma of live specimens.

Combined targeting of PI3K and MEK effector pathways via CED for DIPG therapy.

BACKGROUND: Midline gliomas like diffuse intrinsic pontine glioma (DIPG) carry poor prognosis and lack effective treatment options. Studies have implicated amplifications in the phosphatidylinositol 3-kinase (PI3K) signaling pathway in tumorigenesis; compensatory activation of parallel pathways (eg, mitogen-activated protein kinase [MEK]) may underlie the resistance to PI3K inhibition observed in the clinic. METHODS: Three patient-derived cell lines (SU-DIPG-IV, SU-DIPG-XIII, and SF8628) and a mouse-derived brainstem glioma cell line were treated with PI3K (ZSTK474) and MEK (trametinib) inhibitors, alone or in combination. Synergy was analyzed using Chou-Talalay combination index (CI). These agents were also used alone or in combination in a subcutaneous SU-DIPG-XIII tumor model and in an intracranial genetic mouse model of DIPG, given via convection-enhanced delivery (CED). RESULTS: We found that these agents abrogate cell proliferation in a dose-dependent manner. Combination treatments were found to be synergistic (CI < 1) across cell lines tested. They also showed significant tumor suppression when given systemically against a subcutaneous DIPG model (alone or in combination) or when given via direct intracranial injection (CED) in a intracranial DIPG mouse model (combination only, median survival 47 vs 35 days post-induction, P = .038). No significant short- or long-term neurotoxicity of ZSTK474 and trametinib delivered via CED was observed. CONCLUSIONS: Our data indicate that ZSTK474 and trametinib combinatorial treatment inhibits malignant growth of DIPG cells in vitro and in vivo, prolonging survival. These results suggest a promising new combinatorial approach using CED for DIPG therapy, which warrants further investigation.

Volume of distribution and clearance of peptide-based nanofiber after convection-enhanced delivery.

OBJECTIVE Drug clearance may be a limiting factor in the clinical application of convection-enhanced delivery (CED). Peptide-based nanofibers (NFPs) have a high aspect ratio, and NFPs loaded with drugs could potentially maintain effective drug concentrations for an extended period sufficient for cancer therapy. The objective of this study was to assess the volume of distribution (Vd) and clearance of variable lengths of NFPs when administered using CED. METHODS NFPs composed of multiple methoxypolyethylene glycol (mPEG)-conjugated constructs (mPEG2000-KLDLKLDLKLDL-K( FITC)-CONH2, for which FITC is fluorescein isothiocyanate) were assembled in an aqueous buffer. The NFPs were approximately 5 nm in width and were formulated into different lengths: 100 nm (NFP-100), 400 nm (NFP-400), and 1000 nm (NFP-1000). The NFP surface was covalently conjugated with multiple Cy5.5 fluorophores as the optical reporters to track the post-CED distribution. Forty-two 6- to 8-week-old Ntv-a;p53fl/fl mice underwent CED to the striatum. Animals were killed immediately, 24 hours or 72 hours after CED. The brains were extracted and sectioned for assessing NFP Vd to volume of infusion (Vi) ratio, and clearance using fluorescence microscopy. RESULTS CED of NFPs was well tolerated by all the animals. The average Vd/Vi ratios for NFP-100, NFP-400, NFP-1000, and unconjugated positive control (free Cy5.5) were 1.87, 2.47, 1.07, and 3.0, respectively, which were statistically different (p = 0.003). The percentages remaining of the original infusion volume at 24 hours for NFP-100, -400, and -1000 were 40%, 90%, and 74%, respectively. The percentages remaining at 72 hours for NFP-100, -400, and -1000 were 15%, 30%, and 46%, respectively. Unconjugated Cy5.5 was not detected at 24 or 72 hours after CED. CONCLUSIONS CED of NFPs is feasible with Vd/Vi ratios and clearance rates comparable to other nanocarriers. Of the 3 NFPs, NFP-400 appears to provide the best distribution and slowest clearance after 24 hours. NFP provides a dynamic theranostic platform, with the potential to deliver clinically efficacious drug payload to brain tumor after CED.

A Murine Model for Quantitative, Real-Time Evaluation of Convection-Enhanced Delivery (RT-CED) Using an 18[F]-Positron Emitting, Fluorescent Derivative of Dasatinib.

The blood brain barrier can limit the efficacy of systemically delivered drugs in treating neurological malignancies; therefore, alternate routes of drug administration must be considered. The Abl-kinase inhibitor, dasatinib, is modified to give compound 1 ([18F]-1) so that 18F-positron emission tomography (PET) and fluorescent imaging can both be used to observe drug delivery to murine orthotopic glioma. In vitro Western blotting, binding studies (IC50 = 22 ± 5 nmol/L), and cell viability assays (IC50 = 46 ± 30 nmol/L) confirm nanomolar, in vitro effectiveness of [18F]-1, a dasatinib derivative that is visible by 18F-PET and fluorescence. [18F]-1 is used to image dynamic direct drug delivery via two different drug delivery techniques to orthotopic murine brainstem glioma (mBSG) bearing mice. Convection enhanced delivery (CED) delivers higher concentrations of drug to glioma-containing volumes versus systemic, tail-vein delivery. Accurate delivery and clearance data pertaining to dasatinib are observed, providing personalized information that is important in dosimetry and redosing. Cases of missed drug delivery are immediately recognized by PET/CT, allowing for prompt intervention in the case of missed delivery. Mol Cancer Ther; 16(12); 2902-12. ©2017 AACR.