Differential extracellular matrix proteomic signatures in colorectal tumors from Appalachian and non‑Appalachian patients

Alexander T. Sougiannis; Harrison B. Taylor; Stephen C. Zambrzycki; Lindsey Conroy; Rachel Strubler; Christin Edge; Richard R. Drake; Kristin Wallace; Derek Allison; Eun Lee; Ramon C. Sun; Peggi M. Angel

doi:10.3892/ol.2025.15159

Differential extracellular matrix proteomic signatures in colorectal tumors from Appalachian and non‑Appalachian patients

Alexander T. Sougiannis, Harrison B. Taylor, Stephen C. Zambrzycki, Lindsey Conroy, Rachel Strubler, Christin Edge, Richard R. Drake, Kristin Wallace, Derek Allison, Eun Lee, Ramon C. Sun, Peggi M. Angel

Bioanalytical Mass Spectrometry

Research output: Contribution to journal › Article › peer-review

Abstract

Emerging evidence reports that regulation of the extracellular matrix influences the progression of colorectal cancer (CRC). The present study investigated regulation of the extracellular matrix proteome in colorectal malignancy within a high‑risk Appalachian population compared with non‑Appalachian populations. A targeted mass spectrometry imaging proteomic method directed at collagen regulation was used. Tissue microarrays (TMAs) comprising of matched CRC with adjacent normal to tumor (NAT) from 45 patients were constructed into 86 samples to evaluate the extracellular matrix proteome (ECM). A total of five specific peaks were discov‑ ered to differ between NAT and tumor with high sensitivity and specificity by receiver operating characteristic (AUROC) ≥0.7, Wilson/Brown P<0.0002. Evaluation of patient TMA cores showed increased levels of combined ECM peptides in advanced stage Appalachian CRC (III + IV) compared with early staged CRC (I + II) (AUROC 0.8595; 95% confi‑ dence interval, 0.8190‑0.8999; Wilson/Brown P<1.0x10^‑15), contrasting with the non‑Appalachian tumors, which showed a decreased ability to discriminate between early and late stage (AUROC 0.6618; 95% confidence interval, 0.6126‑0.7110; Wilson/Brown P<1.0x10^‑9). Comparison of advanced stage CRCs between Appalachian and non‑Appalachian popula‑ tions showed high sensitivity and specificity in distinguishing the populations (AUROC 0.7612; 95% confidence interval, 0.7109‑0.8114; Wilson/Brown P<3.0x10^‑15). History of smoking, sex and tumor origin location did not show signifi‑ cant ability to distinguish by AUROC. A combination of high mass resolution, high mass accuracy spatial proteomics and sequencing proteomics by liquid chromatography coupled to tandem mass spectrometry revealed that fibrillar collagens were spatially regulated within the CRC tumor microenviron‑ ment. Fibrillar collagen post‑translational modifications of hydroxylated proline revealed distinct spatial separation based on the presence of a number of hydroxylated proline sites. The present study highlighted that the targeted mass spectrometry imaging of the ECM proteome may provide new insight and novel predictive tools for understanding CRC, particularly among Appalachian patients.

Original language	English
Article number	413
Journal	Oncology Letters
Volume	30
Issue number	3
DOIs	https://doi.org/10.3892/ol.2025.15159
State	Published - Sep 2025

Funding

The authors would like to thank Mr. William H. Angel (Consultant, Charleston, SC, USA) for their assistance in creating county maps. This study was supported by the National Institutes of Health/National Cancer Institute (grant nos. R01CA253460, R21CA240148 and R01CA226086); the Biorepository and Tissue Analysis Shared Resource, Hollings Cancer Center, Medical University of South Carolina; the MUSC Digestive Disease Research Core (grant no. P30DK123704); the South Carolina Centers of Economic Excellence SmartState program; the Biospecimen Procurement and Translational Pathology Shared Resource Facility of the University of Kentucky Markey Cancer Center (grant no. P30CA177558). In addition, mass spectrometry data were collected in the University of Cincinnati Proteomics Laboratory under the direction of KDG on a Thermo Orbitrap Eclipse instrument purchased in part through an NIH instrumentation (grant no. 1S10OD026717). The MUSC Mass Spectrometry Facility and Redox Proteomics Core is supported by the Medical University of South Carolina and grant no. P20GM103542 (from NIH/NIGMS) with shared instrumentation S10OD010731, S10OD025126 and S10 0D030212 (from NIH/OD). This study was supported by the National Institutes of Health/National Cancer Institute (grant nos. R01CA253460, R21CA240148 and R01CA226086); the Biorepository and Tissue Analysis Shared Resource, Hollings Cancer Center, Medical University of South Carolina; the MUSC Digestive Disease Research Core (grant no. P30DK123704); the South Carolina Centers of Economic Excellence SmartState program; the Biospecimen Procurement and Translational Pathology Shared Resource Facility of the University of Kentucky Markey Cancer Center (grant no. P30CA177558). In addition, mass spectrometry data were collected in the University of Cincinnati Proteomics Laboratory under the direction of KDG on a Thermo Orbitrap Eclipse instrument purchased in part through an NIH instrumentation (grant no. 1S10OD026717). The MUSC Mass Spectrometry Facility and Redox Proteomics Core is supported by the Medical University of South Carolina and grant no. P20GM103542 (from NIH/NIGMS) with shared instrumentation S10OD010731, S10OD025126 and S10 0D030212 (from NIH/OD).

Keywords

cancer
collagen
colorectal cancer
extracellular matrix
imaging
mass spectrometry imaging
spatial proteomics

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10.3892/ol.2025.15159

Cite this

Sougiannis, A. T., Taylor, H. B., Zambrzycki, S. C., Conroy, L., Strubler, R., Edge, C., Drake, R. R., Wallace, K., Allison, D., Lee, E., Sun, R. C., & Angel, P. M. (2025). Differential extracellular matrix proteomic signatures in colorectal tumors from Appalachian and non‑Appalachian patients. Oncology Letters, 30(3), Article 413. https://doi.org/10.3892/ol.2025.15159

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AU - Sougiannis, Alexander T.

AU - Taylor, Harrison B.

AU - Zambrzycki, Stephen C.

AU - Conroy, Lindsey

AU - Strubler, Rachel

AU - Edge, Christin

AU - Drake, Richard R.

AU - Wallace, Kristin

AU - Allison, Derek

AU - Lee, Eun

AU - Sun, Ramon C.

AU - Angel, Peggi M.

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N2 - Emerging evidence reports that regulation of the extracellular matrix influences the progression of colorectal cancer (CRC). The present study investigated regulation of the extracellular matrix proteome in colorectal malignancy within a high‑risk Appalachian population compared with non‑Appalachian populations. A targeted mass spectrometry imaging proteomic method directed at collagen regulation was used. Tissue microarrays (TMAs) comprising of matched CRC with adjacent normal to tumor (NAT) from 45 patients were constructed into 86 samples to evaluate the extracellular matrix proteome (ECM). A total of five specific peaks were discov‑ ered to differ between NAT and tumor with high sensitivity and specificity by receiver operating characteristic (AUROC) ≥0.7, Wilson/Brown P<0.0002. Evaluation of patient TMA cores showed increased levels of combined ECM peptides in advanced stage Appalachian CRC (III + IV) compared with early staged CRC (I + II) (AUROC 0.8595; 95% confi‑ dence interval, 0.8190‑0.8999; Wilson/Brown P<1.0x10‑15), contrasting with the non‑Appalachian tumors, which showed a decreased ability to discriminate between early and late stage (AUROC 0.6618; 95% confidence interval, 0.6126‑0.7110; Wilson/Brown P<1.0x10‑9). Comparison of advanced stage CRCs between Appalachian and non‑Appalachian popula‑ tions showed high sensitivity and specificity in distinguishing the populations (AUROC 0.7612; 95% confidence interval, 0.7109‑0.8114; Wilson/Brown P<3.0x10‑15). History of smoking, sex and tumor origin location did not show signifi‑ cant ability to distinguish by AUROC. A combination of high mass resolution, high mass accuracy spatial proteomics and sequencing proteomics by liquid chromatography coupled to tandem mass spectrometry revealed that fibrillar collagens were spatially regulated within the CRC tumor microenviron‑ ment. Fibrillar collagen post‑translational modifications of hydroxylated proline revealed distinct spatial separation based on the presence of a number of hydroxylated proline sites. The present study highlighted that the targeted mass spectrometry imaging of the ECM proteome may provide new insight and novel predictive tools for understanding CRC, particularly among Appalachian patients.

AB - Emerging evidence reports that regulation of the extracellular matrix influences the progression of colorectal cancer (CRC). The present study investigated regulation of the extracellular matrix proteome in colorectal malignancy within a high‑risk Appalachian population compared with non‑Appalachian populations. A targeted mass spectrometry imaging proteomic method directed at collagen regulation was used. Tissue microarrays (TMAs) comprising of matched CRC with adjacent normal to tumor (NAT) from 45 patients were constructed into 86 samples to evaluate the extracellular matrix proteome (ECM). A total of five specific peaks were discov‑ ered to differ between NAT and tumor with high sensitivity and specificity by receiver operating characteristic (AUROC) ≥0.7, Wilson/Brown P<0.0002. Evaluation of patient TMA cores showed increased levels of combined ECM peptides in advanced stage Appalachian CRC (III + IV) compared with early staged CRC (I + II) (AUROC 0.8595; 95% confi‑ dence interval, 0.8190‑0.8999; Wilson/Brown P<1.0x10‑15), contrasting with the non‑Appalachian tumors, which showed a decreased ability to discriminate between early and late stage (AUROC 0.6618; 95% confidence interval, 0.6126‑0.7110; Wilson/Brown P<1.0x10‑9). Comparison of advanced stage CRCs between Appalachian and non‑Appalachian popula‑ tions showed high sensitivity and specificity in distinguishing the populations (AUROC 0.7612; 95% confidence interval, 0.7109‑0.8114; Wilson/Brown P<3.0x10‑15). History of smoking, sex and tumor origin location did not show signifi‑ cant ability to distinguish by AUROC. A combination of high mass resolution, high mass accuracy spatial proteomics and sequencing proteomics by liquid chromatography coupled to tandem mass spectrometry revealed that fibrillar collagens were spatially regulated within the CRC tumor microenviron‑ ment. Fibrillar collagen post‑translational modifications of hydroxylated proline revealed distinct spatial separation based on the presence of a number of hydroxylated proline sites. The present study highlighted that the targeted mass spectrometry imaging of the ECM proteome may provide new insight and novel predictive tools for understanding CRC, particularly among Appalachian patients.

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KW - mass spectrometry imaging

KW - spatial proteomics

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ER -

Differential extracellular matrix proteomic signatures in colorectal tumors from Appalachian and non‑Appalachian patients

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