Maximilian Koch

Concurrent video-rate color and near-infrared fluorescence laparoscopy

Abstract. The visual identification and demarcation of tumors and tumor margins remains challenging due to the low optical contrast of cancer cells over surrounding tissues. Fluorescence molecular imaging was recently considered clinically for improving cancer detection during open surgery. We present herein a next step in the development of fluorescence molecular guidance by describing a novel video-rate imaging laparoscope capable of concurrently recording color and near-infrared fluorescence images and video. Video-rate operation is based on graphics processing unit-based image processing. We examine the optical characteristics of the system developed and provide performance metrics related to intra-operative endoscopic guidance, showcased on phantoms and endoscopic color and fluorescence molecular imaging of tumors in a mouse model of the human disease.

Advancing Surgical Vision with Fluorescence Imaging

Surgical success depends on the accuracy with which disease and vital tissue can be intraoperatively detected. However, the dominant visualization approach, i.e., human vision, does not see under the tissue surface and operates on low contrast between sites of disease, such as cancer, and the surrounding tissue. Intraoperative fluorescence imaging is emerging as a highly effective method to improve surgical vision and offers the potential to be intergrated seamlessly into the normal workflow of the operating room without causing disruption or undue delay. We review and compare two critical fluorescence imaging directions: one that uses nonspecific fluorescence dyes, addressing tissue perfusion and viability, and one that uses targeted agents, interrogating pathophysiological features of disease. These two approaches present detection sensitivity challenges that may differ by orders of magnitude and require different detection strategies. Nevertheless, fluorescence imaging provides the surgeon with previously unavailable real-time feedback that improves surgical precision and can become essential for interventional decision-making.

Enriching the Interventional Vision of Cancer with Fluorescence and Optoacoustic Imaging

Among several techniques considered for surgical and endoscopic imaging, novel optical methods are evolving as a promising approach for interventional guidance. Pilot clinical applications of fluorescence molecular imaging have demonstrated the benefits of using targeted fluorescent agents in cancer surgery. This premise can be extended broadly to interventional guidance through an increasing number of targeted agents and detection techniques. Beyond epi-illumination fluorescence imaging, optoacoustic (photoacoustic) methods are emerging to offer high-resolution cross-sectional optical imaging through several millimeters to centimeters of depth. We present an overview of key recent developments in optical interventional imaging and outline the potential for a paradigm shift in surgical and endoscopic visualization.

Molecular Fluorescence Endoscopy Targeting Vascular Endothelial Growth Factor A for Improved Colorectal Polyp Detection

Small and flat adenomas are known to carry a high miss-rate during standard white-light endoscopy. Increased detection rate may be achieved by molecular fluorescence endoscopy with targeted near-infrared (NIR) fluorescent tracers. The aim of this study was to validate vascular endothelial growth factor A (VEGF-A) and epidermal growth factor receptor (EGFR)–targeted fluorescent tracers during ex vivo colonoscopy with an NIR endoscopy platform. Methods: VEGF-A and EGFR expression was determined by immunohistochemistry on a large subset of human colorectal tissue samples—48 sessile serrated adenomas/polyps, 70 sporadic high-grade dysplastic adenomas, and 19 hyperplastic polyps—and tissue derived from patients with Lynch syndrome—78 low-grade dysplastic adenomas, 57 high-grade dysplastic adenomas, and 31 colon cancer samples. To perform an ex vivo colonoscopy procedure, 14 mice with small intraperitoneal EGFR-positive HCT116luc tumors received intravenous bevacizumab-800CW (anti-VEGF-A), cetuximab-800CW (anti-EGFR), control tracer IgG-800CW, or sodium chloride. Three days later, 8 resected HCT116luc tumors (2–5 mm) were stitched into 1 freshly resected human colon specimen and followed by an ex vivo molecular fluorescence colonoscopy procedure. Results: Immunohistochemistry showed high VEGF-A expression in 79%–96% and high EGFR expression in 51%–69% of the colorectal lesions. Both targets were significantly overexpressed in the colorectal lesions, compared with the adjacent normal colon crypts. During ex vivo molecular fluorescence endoscopy, all tumors could clearly be delineated for both bevacizumab-800CW and cetuximab-800CW tracers. Specific tumor uptake was confirmed with fluorescent microscopy showing, respectively, stromal and cell membrane fluorescence. Conclusion: VEGF-A is a promising target for molecular fluorescence endoscopy because it showed a high protein expression, especially in sessile serrated adenomas/polyps and Lynch syndrome. We demonstrated the feasibility to visualize small tumors in real time during colonoscopy using a NIR fluorescence endoscopy platform, providing the endoscopist a wide-field red-flag technique for adenoma detection. Clinical studies are currently being performed in order to provide in-human evaluation of our approach.

Towards clinically translatable NIR fluorescence molecular guidance for colonoscopy

White-light surveillance colonoscopy is the standard of care for the detection and removal of premalignant lesions to prevent colorectal cancer, and the main screening recommendation following treatment for recurrence detection. However, it lacks sufficient diagnostic yield, exhibits unacceptable adenoma miss-rates and is not capable of revealing functional and morphological information of the detected lesions. Fluorescence molecular guidance in the near-infrared (NIR) is expected to have outstanding relevance regarding early lesion detection and heterogeneity characterization within and among lesions in these interventional procedures. Thereby, superficial and sub-surface tissue biomarkers can be optimally visualized due to a minimization of tissue attenuation and autofluorescence by comparison with the visible, which simultaneously enhance tissue penetration and assure minimal background. At present, this potential is challenged by the difficulty associated with the clinical propagation of disease-specific contrast agents and the absence of a commercially available endoscope that is capable of acquiring wide-field, NIR fluorescence at video-rates. We propose two alternative flexible endoscopic fluorescence imaging methods, each based on a CE certified commercial, clinical grade endoscope, and the employment of an approved monoclonal antibody labeled with a clinically applicable NIR fluorophore. Pre-clinical validation of these two strategies that aim at bridging NIR fluorescence molecular guidance to clinical translation is demonstrated in this study.

Threshold Analysis and Biodistribution of Fluorescently Labeled Bevacizumab in Human Breast Cancer.

In vivo tumor labeling with fluorescent agents may assist endoscopic and surgical guidance for cancer therapy as well as create opportunities to directly observe cancer biology in patients. However, malignant and nonmalignant tissues are usually distinguished on fluorescence images by applying empirically determined fluorescence intensity thresholds. Here, we report the development of fSTREAM, a set of analytic methods designed to streamline the analysis of surgically excised breast tissues by collecting and statistically processing hybrid multiscale fluorescence, color, and histology readouts toward precision fluorescence imaging. fSTREAM addresses core questions of how to relate fluorescence intensity to tumor tissue and how to quantitatively assign a normalized threshold that sufficiently differentiates tumor tissue from healthy tissue. Using fSTREAM we assessed human breast tumors stained in vivo with fluorescent bevacizumab at microdose levels. Showing that detection of such levels is achievable, we validated fSTREAM for high-resolution mapping of the spatial pattern of labeled antibody and its relation to the underlying cancer pathophysiology and tumor border on a per patient basis. We demonstrated a 98% sensitivity and 79% specificity when using labeled bevacizumab to outline the tumor mass. Overall, our results illustrate a quantitative approach to relate fluorescence signals to malignant tissues and improve the theranostic application of fluorescence molecular imaging. Cancer Res; 77(3); 623-31. ©2016 AACR.

Wavelet-based data and solution compression for efficient image reconstruction in fluorescence diffuse optical tomography

Current fluorescence diffuse optical tomography (fDOT) systems can provide large data sets and, in addition, the unknown parameters to be estimated are so numerous that the sensitivity matrix is too large to store. Alternatively, iterative methods can be used, but they can be extremely slow at converging when dealing with large matrices. A few approaches suitable for the reconstruction of images from very large data sets have been developed. However, they either require explicit construction of the sensitivity matrix, suffer from slow computation times, or can only be applied to restricted geometries. We introduce a method for fast reconstruction in fDOT with large data and solution spaces, which preserves the resolution of the forward operator whilst compressing its representation. The method does not require construction of the full matrix, and thus allows storage and direct inversion of the explicitly constructed compressed system matrix. The method is tested using simulated and experimental data. Results show that the fDOT image reconstruction problem can be effectively compressed without significant loss of information and with the added advantage of reducing image noise.

Fluorescence-aided Tomographic Imaging of Synovitis in the Human Finger

PURPOSE To propose and evaluate indocyanine green (ICG)-enhanced tomographic optical imaging for detection and characterization of synovitis in affected finger joints of patients with rheumatoid arthritis and differentiation from healthy joints in comparison to 3-T magnetic resonance (MR) imaging. MATERIALS AND METHODS This prospective pilot study was approved by the institutional ethics committee. Six arthritic proximal interphalangeal (PIP) joints in six patients (five women and one man; mean age ± standard deviation, 62.6 years ± 13.3) with clinically determined rheumatoid arthritis and six healthy PIP joints from six volunteers (four women and two men; mean age, 41.5 years ± 20.2) were examined with an ICG-enhanced fluorescence molecular tomography (FMT) system and 3-T MR imaging as the standard of reference. The degree of inflammation was graded semiquantitatively on a four-point ordinate scale according to the Outcome Measures in Rheumatology Clinical Trials Rheumatoid Arthritis MR Imaging Score, or OMERACT RAMRIS. FMT reconstructions were coregistered with the MR images. Groups were compared by using a two-sided t test, and a weighted κ coefficient was used for comparing FMT and MR imaging semiquantitative scores, as well as assessing intrareader agreement. RESULTS FMT was used to detect synovitis in all arthritic joints. The reconstructed FMT signal correlated with MR imaging findings in intensity and spatial, transverse profile. Semiquantitative scoring of FMT correlated well with MR imaging findings (weighted κ coefficient = 0.90). The reconstructed quantitative FMT signal, denoting synovial hyperperfusion, was used to differentiate between synovitis and healthy joints (healthy joints, 1.25 ± 0.59; arthritic joints, 3.13 ± 1.03; P < .001). CONCLUSION FMT enhanced with ICG provided depth-resolved imaging of synovitis in PIP joints. FMT may help detect synovitis in patients with rheumatoid arthritis.

Comprehensive phantom for interventional fluorescence molecular imaging

Abstract. Fluorescence imaging has been considered for over a half-century as a modality that could assist surgical guidance and visualization. The administration of fluorescent molecules with sensitivity to disease biomarkers and their imaging using a fluorescence camera can outline pathophysiological parameters of tissue invisible to the human eye during operation. The advent of fluorescent agents that target specific cellular responses and molecular pathways of disease has facilitated the intraoperative identification of cancer with improved sensitivity and specificity over nonspecific fluorescent dyes that only outline the vascular system and enhanced permeability effects. With these new abilities come unique requirements for developing phantoms to calibrate imaging systems and algorithms. We briefly review herein progress with fluorescence phantoms employed to validate fluorescence imaging systems and results. We identify current limitations and discuss the level of phantom complexity that may be required for developing a universal strategy for fluorescence imaging calibration. Finally, we present a phantom design that could be used as a tool for interlaboratory system performance evaluation.

Near-infrared fluorescence cholangiopancreatoscopy: initial clinical feasibility results

BACKGROUND The recent clinical propagation of targeted fluorescence agents brings a promising alternative in endoscopy by complementing visual disease detection with molecular biomarkers. OBJECTIVE Development of near-infrared (NIR) fluorescence cholangiopancreatoscopy in real-time and validation of its clinical use. DESIGN Feasibility study. SETTING Tertiary referral center at a large university hospital. PATIENTS Patients with pancreatic and biliary diseases. INTERVENTIONS Routine cholangiopancreatoscopy with additional wide-field NIR fluorescence imaging. MAIN OUTCOME MEASUREMENTS We adapted a miniature cholangioscope for real-time concurrent wide-field color and NIR fluorescence imaging. Illumination is provided through a custom-designed fiber bundle, and the acquired images are relayed via a dichroic beam splitter to 2 charge-coupled devices for simultaneous measurement. We characterize the sensitivity and resolution and demonstrate the clinical feasibility by detecting indocyanine green localization in 2 patients. RESULTS A spatial optical resolution of approximately 50 μm was achieved, and fluorescent dye concentrations of 17.3 nM could be detected. Elevated fluorescence signals were detected during clinical measurements, and biopsy specimens confirmed the presence of malignancy in both patients. LIMITATIONS Feasibility study, limited number of patients. CONCLUSIONS The results demonstrate that real-time wide-field fluorescence detection in the NIR range is possible in humans by using adapted endoscopes. The feasibility of detecting indocyanine green in the pancreatobiliary ducts is verified, suggesting that cancer screening at a molecular level might play an increasingly important role in the future.