Phase 1 Safety, Pharmacokinetics, and Fluorescence Imaging Study of Tozuleristide (BLZ-100) in Adults With Newly Diagnosed or Recurrent Gliomas.

BACKGROUND: Fluorescence-guided surgery (FGS) can improve extent of resection in gliomas. Tozuleristide (BLZ-100), a near-infrared imaging agent composed of the peptide chlorotoxin and a near-infrared fluorophore indocyanine green, is a candidate molecule for FGS of glioma and other tumor types. OBJECTIVE: To perform a phase 1 dose-escalation study to characterize the safety, pharmacokinetics, and fluorescence imaging of tozuleristide in adults with suspected glioma. METHODS: Patients received a single intravenous dose of tozuleristide 3 to 29 h before surgery. Fluorescence images of tumor and cavity in Situ before and after resection and of excised tissue ex Vivo were acquired, along with safety and pharmacokinetic measures. RESULTS: A total of 17 subjects received doses between 3 and 30 mg. No dose-limiting toxicity was observed, and no reported adverse events were considered related to tozuleristide. At doses of 9 mg and above, the terminal serum half-life for tozuleristide was approximately 30 min. Fluorescence signal was detected in both high- and low-grade glial tumors, with high-grade tumors generally showing greater fluorescence intensity compared to lower grade tumors. In high-grade tumors, signal intensity increased with increased dose levels of tozuleristide, regardless of the time of dosing relative to surgery. CONCLUSION: These results support the safety of tozuleristide at doses up to 30 mg and suggest that tozuleristide imaging may be useful for FGS of gliomas.

Polymalic acid chlorotoxin nanoconjugate for near-infrared fluorescence guided resection of glioblastoma multiforme.

Maximal surgical resection of glioma remains the single most effective treatment. Tools to guide the resection while avoiding removal of normal brain tissues can aid surgeons in achieving optimal results. One strategy to achieve this goal is to rely upon interoperative fluorescence staining of tumor cells in vivo, that can be visualized by the surgeon during resection. Towards this goal we have designed a biodegradable fluorescent mini nano imaging agent (NIA) with high specificity for U87MG glioma cells and previously unmet high light emission. The NIA is the conjugate of polymalic acid (PMLA) with chlorotoxin for tumor targeting, indocyanine green (ICG) for NIR fluorescence and the tri-leucin peptide as fluorescence enhancer. PMLA as a multivalent platform carries several molecules of ICG and the other ligands. The NIA recognizes multiple sites on glioma cell surface, demonstrated by the effects of single and combined competitors. Systemic IV injection into xenogeneic mouse model carrying human U87MG glioblastoma indicated vivid tumor cell binding and internalization of NIA resulting in intensive and long-lasting tumor fluorescence. The NIA is shown to greatly improve tumor removal supporting its utility in clinical applications.

Real-time Visualization of Breast Carcinoma in Pathology Specimens From Patients Receiving Fluorescent Tumor-Marking Agent Tozuleristide.

CONTEXT.­: Resection of breast carcinoma with adequate margins reduces the risk of local recurrence and reoperation. Tozuleristide (BLZ-100) is an investigational peptide-fluorophore agent that may aid in intraoperative tumor detection and margin assessment. In this study, fluorescence imaging was conducted ex vivo on gross breast pathology specimens. OBJECTIVES.­: To determine the potential of tozuleristide to detect breast carcinoma in fresh pathology specimens and the feasibility of fluorescence-guided intraoperative pathology assessment of surgical margins. DESIGN.­: Twenty-three patients received an intravenous bolus dose of 6 or 12 mg of tozuleristide at least 1 hour before surgery. Fifteen lumpectomy and 12 mastectomy specimens were evaluated for fluorescence by the site's clinical pathology staff using the SIRIS, an investigational near-infrared imaging device. The breast tissue was then processed per usual procedures. Fluorescent patterns were correlated with the corresponding hematoxylin-eosin-stained sections. Clinical pathology reports were used to correlate fluorescent signal to grade, histotype, prognostic marker status, and margin measurements. RESULTS.­: Tozuleristide fluorescence was readily observed in invasive and in situ breast carcinoma specimens. Most invasive carcinomas were bright and focal, whereas in situ lesions demonstrated a less intense, more diffuse pattern. Tozuleristide was detected in ductal and lobular carcinomas with a similar fluorescent pattern. Fluorescence was detected in high- and low-grade lesions, and molecular marker/hormone receptor status did not affect signal. Fluorescence could be used to identify the relationship of carcinoma to margins intraoperatively. CONCLUSIONS.­: Tumor targeting with tozuleristide allowed visual real-time distinction between pathologically confirmed breast carcinoma and normal tissue.

Real time optical Biopsy: Time-resolved Fluorescence Spectroscopy instrumentation and validation.

The Time-resolved fluorescence spectroscopy (TR-FS) has the potential to differentiate tumor and normal tissue in real time during surgical excision. In this manuscript, we describe the design of a novel TR-FS device, along with preliminary data on detection accuracy for fluorophores in a mixture. The instrument is capable of near real-time fluorescence lifetime acquisition in multiple spectral bands and analysis. It is also able to recover fluorescence lifetime with sub-20ps accuracy as validated with individual organic fluorescence dyes and dye mixtures yielding lifetime values for standard fluorescence dyes that closely match with published data. We also show that TR-FS is able to quantify the relative concentration of fluorescence dyes in a mixture by the unmixing of lifetime decays. We show that the TR-FS prototype is able to identify in near-real time the concentrations of dyes in a complex mixture based on previously trained data. As a result, we demonstrate that in complex mixtures of fluorophores, the relative concentration information is encoded in the fluorescence lifetime across multiple spectral bands. We show for the first time the temporal and spectral measurements of a mixture of fluorochromes and the ability to differentiate relative concentrations of each fluorochrome mixture in real time.

A testbed for wide-field, high-resolution, gigapixel-class cameras.

The high resolution and wide field of view (FOV) of the AWARE (Advanced Wide FOV Architectures for Image Reconstruction and Exploitation) gigapixel class cameras present new challenges in calibration, mechanical testing, and optical performance evaluation. The AWARE system integrates an array of micro-cameras in a multiscale design to achieve gigapixel sampling at video rates. Alignment and optical testing of the micro-cameras is vital in compositing engines, which require pixel-level accurate mappings over the entire array of cameras. A testbed has been developed to automatically calibrate and measure the optical performance of the entire camera array. This testbed utilizes translation and rotation stages to project a ray into any micro-camera of the AWARE system. A spatial light modulator is projected through a telescope to form an arbitrary object space pattern at infinity. This collimated source is then reflected by an elevation stage mirror for pointing through the aperture of the objective into the micro-optics and eventually the detector of the micro-camera. Different targets can be projected with the spatial light modulator for measuring the modulation transfer function (MTF) of the system, fiducials in the overlap regions for registration and compositing, distortion mapping, illumination profiles, thermal stability, and focus calibration. The mathematics of the testbed mechanics are derived for finding the positions of the stages to achieve a particular incident angle into the camera, along with calibration steps for alignment of the camera and testbed coordinate axes. Measurement results for the AWARE-2 gigapixel camera are presented for MTF, focus calibration, illumination profile, fiducial mapping across the micro-camera for registration and distortion correction, thermal stability, and alignment of the camera on the testbed.

Coded aperture compressive temporal imaging.

We use mechanical translation of a coded aperture for code division multiple access compression of video. We discuss the compressed video's temporal resolution and present experimental results for reconstructions of > 10 frames of temporal data per coded snapshot.

Autofocus for a multiscale gigapixel camera.

In recent studies, the advanced wide field of view architectures for image reconstruction and exploitation (AWARE) multiscale camera, which is composed of a monocentric objective lens and an array of microcameras, was developed for the realization of snapshot wide field of view and high resolution imaging. This paper describes accelerated autofocus (AF) methods for the AWARE system based on a hierarchical spatial algorithm and an iterative temporal algorithm. In the algorithms, sensor positions of each microcamera are hierarchically scanned with contrast detection to effectively search for a focusing distance. The positions are then updated iteratively for dynamic scenes using temporal information. The algorithms are theoretically analyzed and experimentally demonstrated. The developed AF methods can be used for the realization of the temporal gigapixel imaging by the AWARE system.

Joint segmentation and reconstruction of hyperspectral data with compressed measurements.

This work describes numerical methods for the joint reconstruction and segmentation of spectral images taken by compressive sensing coded aperture snapshot spectral imagers (CASSI). In a snapshot, a CASSI captures a two-dimensional (2D) array of measurements that is an encoded representation of both spectral information and 2D spatial information of a scene, resulting in significant savings in acquisition time and data storage. The reconstruction process decodes the 2D measurements to render a three-dimensional spatio-spectral estimate of the scene and is therefore an indispensable component of the spectral imager. In this study, we seek a particular form of the compressed sensing solution that assumes spectrally homogeneous segments in the two spatial dimensions, and greatly reduces the number of unknowns, often turning the underdetermined reconstruction problem into one that is overdetermined. Numerical tests are reported on both simulated and real data representing compressed measurements.

Multiframe image estimation for coded aperture snapshot spectral imagers.

A coded aperture snapshot spectral imager (CASSI) estimates the three-dimensional spatiospectral data cube from a snapshot two-dimensional coded projection, assuming that the scene is spatially and spectrally sparse. For less spectrally sparse scenes, we show that the use of multiple nondegenerate snapshots can make data cube recovery less ill-posed, yielding improved spatial and spectral reconstruction fidelity. Additionally, data acquisition can be easily scaled to meet the time/resolution requirements of the scene with little modification or extension of the original CASSI hardware. A multiframe reconstruction of a 640 × 480 × 53 voxel datacube with 450-650 nm white-light illumination of a scene reveals substantial improvement in the reconstruction fidelity, with limited increase in acquisition and reconstruction time.

Technical note: rapid prototyping of 3D grid arrays for image guided therapy quality assurance.

Three dimensional grid phantoms offer a number of advantages for measuring imaging related spatial inaccuracies for image guided surgery and radiotherapy. The authors examined the use of rapid prototyping technology for directly fabricating 3D grid phantoms from CAD drawings. We tested three different fabrication process materials, photopolymer jet with acrylic resin (PJ/AR), selective laser sintering with polyamide (SLS/P), and fused deposition modeling with acrylonitrile butadiene styrene (FDM/ABS). The test objects consisted of rectangular arrays of control points formed by the intersections of posts and struts (2 mm rectangular cross section) and spaced 8 mm apart in the x, y, and z directions. The PJ/AR phantom expanded after immersion in water which resulted in permanent warping of the structure. The surface of the FDM/ABS grid exhibited a regular pattern of depressions and ridges from the extrusion process. SLS/P showed the best combination of build accuracy, surface finish, and stability. Based on these findings, a grid phantom for assessing machine-dependent and frame-induced MR spatial distortions was fabricated to be used for quality assurance in stereotactic neurosurgical and radiotherapy procedures. The spatial uniformity of the SLS/P grid control point array was determined by CT imaging (0.6 x 0.6 x 0.625 mm3 resolution) and found suitable for the application, with over 97.5% of the control points located within 0.3 mm of the position specified in CAD drawing and none of the points off by more than 0.4 mm. Rapid prototyping is a flexible and cost effective alternative for development of customized grid phantoms for medical physics quality assurance.