Characterizing variability of in vivo Raman spectroscopic properties of different anatomical sites of normal colorectal tissue towards cancer diagnosis at colonoscopy

This study aims to characterize the in vivo Raman spectroscopic properties of normal colorectal tissues and to assess distinctive biomolecular variations of different anatomical locations in the colorectum for cancer diagnosis. We have developed a novel 785 nm excitation fiber-optic Raman endoscope that can simultaneously acquire in vivo fingerprint (FP) spectra (800-1800 cm^-1) and high-wavenumber (HW) Raman spectra (2800-3600 cm^-1) from the subsurface of colorectal tissue. We applied the FP/HW Raman endoscope for in vivo tissue Raman measurements of various normal colorectal anatomical locations (i.e., ascending colon (n = 182), transverse colon (n = 249), descending colon (n = 124), sigmoid (n = 212), and rectum (n = 362)) in 50 subjects. Partial least-squares (PLS)-discriminant analysis (DA) was employed to evaluate the interanatomical variability. The normal colorectal tissue showed a subtle interanatomical variability in molecular constituents (i.e., proteins, lipids, and water content) and could be divided into three major clusterings: (1) ascending colon and transverse colon, (2) descending colon, and (3) sigmoid and rectum. The PLS-DA multiclass algorithms were able to identify different tissue sites with varying sensitivities (SE) and specificities (SP) (ascending colon: SE: 1.10%, SP: 91.02; transverse colon: SE: 14.06%, SP: 78.78; descending colon: SE: 40.32%, SP: 81.99; sigmoid: SE: 19.34%, SP: 87.90; rectum: SE: 71.55%, SP: 77.84). The interanatomical molecular variability was orders of magnitude less than neoplastic tissue transformation. Further PLS-DA modeling on in vivo FP/HW tissue Raman spectra yielded a diagnostic accuracy of 88.8% (sensitivity: 93.9% (93/99); specificity 88.3% (997/1129) for colorectal cancer detection. This work discloses that interanatomical Raman spectral variability of normal colorectal tissue is subtle compared to cancer tissue, and the simultaneous FP/HW Raman endoscopic technique has promising potential for real-time, in vivo diagnosis of colorectal cancer at the molecular level.