Diffuse reflectance spectroscopy for colorectal cancer surgical guidance

Introduction

In 2020, colorectal cancer (CRC) accounted for 11.3% of diagnosed cancer cases and caused 10.2% (0.88 million) of cancer-related deaths. The significant morbidity and mortality associated with CRC stem from factors such as cancer recurrence, postoperative complications, and pre-existing health conditions in patients. Enhanced survival rates are observed when surgeries achieve complete resection aided by real-time surgical guidance tools for precise intraoperative cancer margin delineation. Laparoscopic surgery (LS) holds promise in reducing long-term surgical complications compared to open surgery (OS), which is prone to short-term complications.

Currently, surgical decisions rely on tissue structure analysis, yet greater accuracy could be achieved by molecular-sensitive tissue identification methods. Development focus lies in automated methods for intraoperative guidance, providing surgeons with real-time probability scores for tissue types. Surgeons can then perform biopsies from suspected cancer-containing locations post-resection, ensuring complete removal of cancer cells from every patient. Among the most promising real-time and molecular-sensitive tools are those utilizing optical spectroscopy methods, particularly diffuse reflectance spectroscopy (DRS).

Methods of implementation

DRS leverages diffuse reflected light within tissue to extract structural and biochemical information based on light scattering and absorption properties, respectively. These properties include scattering amplitude α′, Mie scattering power bMie, and the contribution of Rayleigh scattering fRay to total Mie scattering, associated with tissue microstructures such as organelles and membranes. Absorption properties encompass volume fractions of tissue chromophores, including β-carotene, bile, bilirubin, and others.

Advantages of DRS in colorectal cancer surgery are non-invasive does not require tissue excision, reducing patient discomfort and risk of complications, provides immediate feedback to surgeons during the procedure, aiding in decision-making, quantitative measurements reduce subjectivity in tumor identification and margin assessment.

Our DRS system comprised a tungsten-halogen broadband light source with emission between 350 and 2400 nm (HL-2000-HP, Ocean Optics), connected with two fiber-optic probes to send light from the source to the tissue and collect the tissue reflected light to be detected by two spectrometers. One spectrometer detected the reflected light intensity at the wavelength ranging between 350 and 1140 nm (QE-Pro, Ocean Optics) and another detected intensity in the wavelength range between 1090 and 1920 nm (NIR-Quest, Ocean Optics). Overlapping wavelengths between 1090 and 1140 nm were utilized to combine the two collected DRS spectra into a single broadband spectrum (350-1920 nm) for each measured tissue site.

Figure 1: Schematic of DRS system.

• HL-2000-HP light source:

The HL-2000-HP is a high-performance tungsten-halogen light source designed to provide a broad emission spectrum ranging from 350 to 2400 nm. This versatile light source is part of Ocean Insight's HL-2000 product family, known for its reliability and flexibility in laboratory settings.

Key features of the HL-2000-HP include:

•  High-powered Tungsten Halogen bulb with a nominal power of 20 W.

•  A typical output power of 8.8 mW and a color temperature of 3000 K.

•  An optical output drift of less than 0.1% per hour, ensuring stable performance over time.

•  A bulb life expectancy of approximately 1000 hours, providing long-term usability.

•  Warm-up time of about 10 minutes, after which it reaches optimal stability.

The HL-2000-HP is equipped with an integrated fan to maintain a cool and stable operation and includes a built-in holder for filters to condition the light as needed. Its design is optimized for fiber optic applications, with adjustable focus and alignment of the SMA 905 connector to maximize light throughput.

Figure 2: HL-2000-HP light source.

• QEPro spectrometer

The QE Pro is a high-sensitivity spectrometer that stands out for its low stray light performance and is well-suited for a variety of low light level applications. With a wavelength range of 350 to 1140 nm, it is particularly effective for tasks such as fluorescence, DNA sequencing, and Raman analysis.

Key features of the QE Pro include:

•  A back-thinned CCD detector with high quantum efficiency.

•  Onboard spectral buffering to ensure data integrity at high collection rates.

•  Thermoelectric cooling for thermal stability, which is crucial for precise measurements.

•  A dynamic range of approximately 85000:1, allowing for detection of both strong and weak signals.

•  A signal-to-noise ratio greater than 1000:1 on single acquisition, ensuring clear and accurate data.

The QE Pro's design allows for flexibility and customization, with options for interchangeable slits and an optional internal shutter. This makes it a versatile tool for researchers and professionals who require a reliable and high-performance spectrometer for their specialized applications.

Figure 3: QEPro spectrometer.

• NIR-Quest spectrometer

The NIRQuest spectrometer series is a family of near-infrared spectrometers designed for high-sensitivity applications across a variety of industries. The NIRQuest spectrometers are known for their robust performance, particularly in the wavelength range of 1090 to 1920 nm, which makes them suitable for applications such as moisture content analysis in agriculture, plastics identification in recycling, and chemical concentration measurements.

Key features of the NIRQuest spectrometers include:

•  High sensitivity due to an enhanced optical bench and aperture design, allowing for lower limits of detection.

•  Thermal stability with thermoelectric cooling to -20 °C, ensuring low dark current and reliable performance over time.

•  Flexible applications, thanks to their compatibility with a comprehensive line of fiber optic light sources, optical fibers, and sampling accessories.

•  Preconfigured models that cover different wavelength ranges, including the specific 1090-1920 nm range, to cater to various NIR applications.

The NIRQuest series is part of Ocean Insight's commitment to providing advanced spectroscopy solutions, and it represents the next generation of NIR spectrometers with improved performance and versatility.

Figure 4: NIRQuest spectrometer.

Conclusions

By using broadband diffuse reflectance spectroscopy (DRS) method we assessed key structural and biochemical parameters for surgical colorectal cancer (CRC) delineation. We categorized CW, fat, and tumor tissues based on threshold values of f_lipid, R_vessel, α', and b_Mie, which were identified as important parameters in both probe 1 and probe 2 decision trees. Probe 1 targeted superficial tissues (0.5–1 mm deep), while probe 2 focused on deeper tissue layers (0.5–2 mm deep).

Our results demonstrated high classification performance, with probe 1 achieving 95.9 ± 0.7% sensitivity, 98.9 ± 0.3% specificity, 90.2 ± 0.4% accuracy, and 95.5 ± 0.3% AUC, and probe 2 achieving 96.9 ± 0.8% sensitivity, 98.9 ± 0.2% specificity, 94.0 ± 0.4% accuracy, and 96.7 ± 0.4% AUC. Future studies should validate these metrics in vivo using DRS and/or hyperspectral imaging techniques during laparoscopy, open surgery, and robotic surgery.