Fluorescence-guided optical coherence tomography imaging for colon cancer screening: a preliminary mouse study

Abstract

A new concept for cancer screening has been preliminarily investigated. A cancer targeting agent loaded with a near-infrared (NIR) dye was topically applied on the tissue to highlight cancer-suspect locations and guide optical coherence tomography (OCT) imaging, which was used to further investigate tissue morphology at the micron scale. A pilot study on ApcMin mice has been performed to preliminarily test this new cancer screening approach. As a cancer-targeting agent, poly(epsilon-caprolactone) microparticles (PCLMPs), labeled with a NIR dye and functionalized with an RGD (argenine-glycine-aspartic acid) peptide, were used. This agent recognizes the α(ν)β(3) integrin receptor (ABIR), which is over-expressed by epithelial cancer cells. The contrast agent was administered topically in vivo in mouse colon. After incubation, the animals were sacrificed and fluorescence-guided high resolution optical coherence tomography (OCT) imaging was used to visualize colon morphology. The preliminary results show preferential staining of the abnormal tissue, as indicated by both microscopy and laser-induced fluorescence imaging, and OCT's capability to differentiate between normal mucosal areas, early dysplasia, and adenocarcinoma. Although very preliminary, the results of this study suggest that fluorescence-guided OCT imaging might be a suitable approach for cancer screening. If successful, this approach could be used by clinicians to more reliably diagnose early stage cancers in vivo.

Keywords: (170.0170) Medical optics and biotechnology; (170.4500) Optical coherence tomography; (170.4580) Optical diagnostics for medicine; (170.6935) Tissue characterization.

Fig. 1

Scanning electron microscope images of the PCLMPs. Gold coverage with 10-20 nm nanobeads can be observed in the left side higher resolution image.

Fig. 2

Schematic (A) and photographs (B,C) of the SS OCT system and imaging probe.

Fig. 3

Bright field and fluorescence images of HT-29 cells incubated with PCLMPs. A, A’ RGD-functionalized PCLMPs; B, B’ Blank PCLMPs. Magnification 20X.

Fig. 4

Binding statistics and incubation time for the HT-29 and SW-480 cell lines

Fig. 5

(A) Fluorescence image of the flattened colon showing very low attachment of the PCLMPs, as indicated by random bright spots. The vertical line indicates the position of the OCT scan, while the dashed line indicates the direction of this scan. (B) Cross-sectional OCT image showing the morphology of the colon layers; The OCT image displays the normal mucosa (M), the thin submucosal (SM) layer, and the muscularis propria (MP). The boundaries between these layers are well demarcated (see M/SM and SM/MP). (C) Colon histology within the ROI indicated in B.

Fig. 6

(Media 1). Fly-through video of both fluorescence and OCT measurements. (A) Fluorescence imaging showing the OCT raster scan, as indicated by the aiming beam. (B) Combined enface and cross-sectional OCT imaging. The en face view was taken at a depth indicated by the line from the cross-sectional image. The raw fluorescence video was acquired at 5 fps, while the raw OCT video was acquired at 20 fps. OCT scale bar = 250 μm.

Fig. 7

(A) Fluorescence, (B) OCT, and (C) histology of adenoma (AD) in mouse upper colon. The adenoma almost triples the thickness of the mucosal layer in the inflamed area (mucosal thickening-MT). The adjacent normal mucosa (M), submucosa (SM), and muscularis propria (MP) are labeled. Accumulation of PCL microparticles (PCLMPs) in the adenoma area is visible in the OCT image and confirmed by histology. Individual PCLMPs are discernable in the magnified (x40) histology area. OCT scale bar: 250 μm; Histology magnification: 5x and 40x, respectively

Fig. 8

(A) Fluorescence image of a colon tissue specimen, (B) En face OCT image of the ROI indicated in A by a dotted rectangle. Cross-sectional images (B′ & B″) along the dotted lines from B show evident changes in the morphology of the mucosal layer, which are characteristic to adenomas.

Fig. 9

OCT image and histopathology of adenomatous polyp. (A). OCT image of adenoma displays a markedly expanded mucosa (development of a large polyp) and not very well defined tissue boundaries underlying the lesion. B. Corresponding histopathology; B’. Magnified histology on the polyp area confirms the presence of adenoma. M- mucosa; SM-submucosa; MP-muscularis propria; AD: adenoma; OCT scale bar: 500 μm; Histology magnification: 5x and 20x, respectively.

Fig. 10

Fluorescence (A), OCT image (B). and histopathology (C) of adenocarcinoma in mouse colorectal region. As observed, adenocarcinoma (ADC) was invading and replacing the all three layers: mucosa (M), Submicosa (SM) and muscularis propria (MP). OCT scale bar: 250 μm; Histology magnification: x10 and x40, respectively.

Fig. 11

(Media 2). Fly-through video of both fluorescence and OCT measurements. (A) Fluorescence imaging showing the OCT raster scan, as indicated by the aiming beam. (B) Combined enface and cross-sectional OCT imaging. The en face view was taken at a depth indicated by the line from the cross-sectional image. The raw fluorescence video was acquired at 10 fps, while the raw OCT video was acquired at 20 fps. OCT scale bar: 250 μm.

Fig. 12

(A, B, C) Representative fluorescence images for normal colon, colon adenoma, and colon adenocarcinoma. (A′, B′, C′) Processed images with removed background noise. (D) Integrated fluorescence signal for 11 specimens that were assigned to the three pathologies.

Fig. 13

OCT image (A) and histology (B) of normal colon. The OCT image displays the normal mucosa (M), the thin submucosal (SM) layer, the muscularis propria (MP), and the subserosa (SS) boundary with some underlying fat (F) or connective tissue layers. OCT scale bar: 250 μm; Histology magnification: 5x.

Fig. 14

OCT (A) and histopathology (B) of adenoma in mouse upper colon. The adenoma almost triples the thickness of the mucosal layer in the inflamed area (mucosal thickening-MT). The adjacent normal mucosa (M), submucosa (SM), and muscularis propria (MP) are labeled. OCT scale bar: 250 μm; Histology magnification: 5x and 20x, respectively.