Evaluation and Validation of Plasma Proteins Using Two Different Protein Detection Methods for Early Detection of Colorectal Cancer

Abstract

Objective: Plasma protein biomarkers could be an efficient alternative for population-based screening for early detection of colorectal cancer (CRC). The objective of this study was to evaluate and validate plasma proteins individually and as a signature for early detection of CRC. Methods: In a three-stage design, proteins were measured firstly by liquid chromatography/multiple reaction monitoring-mass spectrometry (LC/MRM-MS) and later by proximity extension assay (PEA) in a discovery set consisting of 96 newly diagnosed CRC cases and 94 controls free of neoplasms at screening colonoscopy. Two algorithms (one for each measurement method) were derived by Lasso regression and .632+ bootstrap based on 11 proteins that were included in both the LC/MRM-MS and PEA measurements. Additionally, another algorithm was constructed from the same eleven biomarkers plus amphireglin, the most promising protein marker in the PEA measurements that had not been available from the LC/MRM-MS measurements. Lastly the three prediction signatures were validated with PEA in independent samples of participants of screening colonoscopy (CRC (n = 56), advanced adenoma (n = 101), and participants free of neoplasm (n = 102)). Results: The same four proteins were included in all three prediction signatures; mannan binding lectin serine protease 1, osteopontin, serum paraoxonase lactonase 3 and transferrin receptor protein 1, and the third prediction signature additionally included amphiregulin. In the independent validation set from a true screening setting, the five-marker blood-based signature including AREG presented areas under the curves of 0.82 (95% CI, 0.74-0.89), 0.86 (95% CI, 0.77-0.92) and 0.76 (95% CI, 0.64-0.86) for all, early and late stages CRC, respectively. Conclusion: Two different measurement methods consistently identified four protein markers and an algorithm additionally including amphiregulin, a marker measured by PEA only, showed promising performance for detecting early stage CRC in an independent validation in a true screening setting. These proteins may be potential candidates for blood-based tests for early detection of CRC.

Clinical Assay for the Early Detection of Colorectal Cancer Using Mass Spectrometric Wheat Germ Agglutinin Multiple Reaction Monitoring

“…Another study carried out by Xie et al used CELA1, CEL2A, CTRL, and TRY2 with a logistic regression and achieved an AUC of 0.90 (with a sensitivity of 86.7% and a specificity of 83.3%) in CRC diagnoses [ 52 ]. Bhardwaj et al utilized AREG, MASP1, OPN, PON3, and TR with the least absolute shrinkage and selection operator (LASSO) and achieved an AUC of 0.86 (with a sensitivity of 83% and a specificity of 80%) in diagnosing early-stage CRC [ 53 ]. In another study from Bhardwaj et al, they combined HP, LRG1, and PON3 with LASSO and achieved an AUC of 0.83 (with a sensitivity of 67% and a specificity of 80%) in diagnosing early-stage CRC; they also utilized eight biomarkers with LASSO and achieved an AUC of 0.96 (with a sensitivity of 93% and a specificity of 80%) in diagnosing late-stage CRC [ 54 ].…”

Comparison of Proteomic Technologies for Blood-Based Detection of Colorectal Cancer

“…In a previous study from our group, 11 protein biomarkers assayed using PEA and LC/MRM-MS were compared in plasma samples from clinically recruited CRC cases [ 16 ]; in that study, a good correlation was found for 8 out of 11 biomarkers. In the current study, good correlations could be observed for four of the nine biomarkers measured using PEA and LC/MRM-MS in samples from participants of screening colonoscopy.…”

“…Even though many blood-based protein signatures have been identified and numerous proteomic technologies are available, direct comparisons of the diagnostic performance of the technologies for detection of cancer in screening settings are scarce. Previous studies from our group utilizing proximity extension assays (PEA) and liquid chromatography/multiple reaction monitoring–mass spectrometry (LC/MRM-MS) have identified several individual markers and multimarker protein signatures for the detection of CRC [ 13 , 14 , 15 , 16 , 17 , 18 ]. The objective of the current study was to evaluate and compare the diagnostic potential of overlapping protein biomarkers measured with PEA, LC/MRM-MS, quantibody microarrays (QMAs) and immunome full-length functional protein arrays (IpAs).…”

Fecal DNA methylation markers for detecting stages of colorectal cancer and its precursors: a systematic review

“…Compared to molecular markers such as methylation of SEPT9 [19,65,66] in tumor-derived cell-free DNA, microRNA signatures in various biofluids (plasma, serum, or stool) [67][68][69], genetic [70,71], or proteomic markers [72][73][74] performance of methylation markers in stool DNA for detecting various stages of CRC seems poorer. In the reviewed studies, methylation of several genes was associated with increased risk of NAA/AA/Ad, early and late stages of CRC, supporting a role of DNA methylation at all stages of CRC, and suggesting potential use of these biomarkers for risk stratification in CRC screening.…”