Real‐time detection of breast cancer at the cellular level

Abstract

Novel optoelectronic instrumentation has been developed for the multispectral imaging of autofluorescence emitted by metabolic fluorophores. The images resolve individual cells while spectra are collected for each pixel in the images. These datacubes are generated at a rate of 10 per second-fast enough for surgical guidance. The data is processed in real time to provide a single color-coded image to the surgeon. To date, the system has been applied to fresh, ex vivo, human surgical specimens and has distinguished breast cancer from benign tissue. The approach is applicable to in vivo measurements of surgical margins and needle-based optical biopsies. Ongoing work demonstrates that the system has great potential for translation to a hand-held probe with high sensitivity and specificity. K E Y W O R D S autofluorescence, breast cancer, multispectral imaging, surgical margin more prevalent than typical aerobic cellular respiration through the electron-transport chain. This in turn leads to an increase in the reduced form of nicotinamide adenine dinucleotide (NADH)/flavin adenine dinucleotide (FAD) ratio (or the redox ratio). Simultaneously, hemoglobin oxygenation near the tumor decreases as the growing J Cell Physiol. 2019;234:5413-5419. wileyonlinelibrary.com/journal/jcp © 2018 Wiley Periodicals, Inc. | 5413 CARVER ET AL. | 5417Once fully translated, our optics will be redesigned into an ergonometric hand-held probe tethered to a cart supporting a colorcoded display for use by surgeons in the operating room. This new capability will shift clinical practice by enabling surgeons to assess tumor margins in real time to inform the extent of surgical resection.Color-coded images will cover a 5 mm field of view in 0.1 s. Translation of this capability to a hand-held probe is crucial for implementation by surgeons (You et al., 2018). Validation studies will be extended beyond correlations with H&E staining. We will also harvest cancer and normal cells identified by our optical system via LCM for subsequent analysis by genetic (DNA-encoded regulatory information) and epigenetic (non-DNA-encoded regulatory information) approaches. These efforts are expected to show that our optical contrast is related to cellular metabolism rather than blood and oxygenation issues. It is anticipated that the multispectral imaging approach we have developed for breast cancer will support optical imaging of other solid tumors.Our results were obtained with spectral vectors that were obtained with a software tool that averages the spectra of all pixels in the given image. As a result, a cancer vector can be "polluted" with noncancerous pixels. Even under that circumstance, the clear differences between the two images in Figure 3 are still observed.Ongoing work will "purify" the vectors by averaging over restricted regions-of-interest rather than the full images. The vectors can also be improved by harvesting more wavelengths in the NADH band, and using fewer, wider bins in the FAD band. These enhancements should allow for the straightforward s...

Intraoperative microscopic autofluorescence detection and characterization in brain tumors using stimulated Raman histology and two-photon fluorescence

“… Emission spectra of endogenous fluorophores [collagen, melanin, elastin, NADH, FAD, lipofuscin (-like pigments), porphyrins] ( 26 – 32 ) that may interfere with the overlapping emission of the exogenous fluorophores fluorescein and PPIX ( 33 ) commonly applied in neurooncological surgery. The corresponding range focused filter setup designed in the dual modal SRS and two-photon fluorescence microscope applied in this study also overlaps with the emission detection filters commonly applied in FGS, (here, YELLOW560© and BLUE400©, Carl Zeiss Meditec, Oberkochem, Germany) ( 34 , 35 ) the values are normalized to the maximum intensity in the range between 450 and 750 nm and emission was observed with one-photon excitation between 330 and 405 nm for the endogenous fluorophores as well as PPIX and 480 nm for fluorescein.…”

Emerging clinical applications in oncology for non‐invasive multi‐ and hyperspectral imaging of cell and tissue autofluorescence

“…The real-time detection of breast cancer was investigated by Carvar et al [56] who created single colour-coded images for the assessment of surgical margins and needle-based biopsies using data cubes with excitations at 375, 405 and 488 nm with 10 spectral bins bounded by two of the emission wavelengths. By assessing cellular concentrations of NADH and FAD they found definable differences between cancer and a benign condition fibroadenoma for which differential diagnosis is needed.…”

“…Cancers that have been investigated using hyper/ multispectral imaging include colon [35][36][37][38][39], oral [40][41][42][43][44][45][46][47][48], head and neck [49][50][51][52][53], skin [54,55], breast [56,57], cervical [58], gastric [59], ocular [17,20], bladder [60], lung [61], brain [62,63], and ovarian [64] (Table 1 It is readily apparent that work has focused on accessible tumours that can be reached endoscopically (e.g., colon and oral) or external tumours (e.g., ocular and skin). This is unsurprising as this optical imaging technology is best suited for application in areas of the body which do not require surgery to access.…”

“…Cancers that have been investigated using hyper/multispectral imaging include colon [35–39], oral [40–48], head and neck [49–53], skin [54, 55], breast [56, 57], cervical [58], gastric [59], ocular [17, 20], bladder [60], lung [61], brain [62, 63], and ovarian [64] (Table 1). Research interest in this area is intensifying, with seven publications published 4 years covered by this scoping review, eight in the second, and 16 in the most recent.…”

Biomedical Applications of Translational Optical Imaging: From Molecules to Humans

“…Mapping spectral bands into the time domain can be applicable to microscopy, endoscopy, and cytometry. One can imagine other applications involving fluorescence, two-photon spectroscopy, and Raman spectroscopy with a future impact on cytomics, histomics, and clinical settings [208,209,212].…”