Quantitative proteomic analysis of oral brush biopsies identifies secretory leukocyte protease inhibitor as a promising, mechanism-based oral cancer biomarker.

Ya Yang,Yaqin Zhu,Nelson L. Rhodus,Timothy J. Griffin,Beverly R. K. Wuertz,Xiaobing Chen,Frank G. Ondrey,John M. Koomen

doi:10.1371/journal.pone.0095389

Abstract

A decrease in the almost fifty percent mortality rate from oral cancer is needed urgently. Improvements in early diagnosis and more effective preventive treatments could affect such a decrease. Towards this end, we undertook for the first time an in-depth mass spectrometry-based quantitative shotgun proteomics study of non-invasively collected oral brush biopsies. Proteins isolated from brush biopsies from healthy normal tissue, oral premalignant lesion tissue (OPMLs), oral squamous cell carcinoma (OSCC) and matched control tissue were compared. In replicated proteomic datasets, the secretory leukocyte protease inhibitor (SLPI) protein stood out based on its decrease in abundance in both OPML and OSCC lesion tissues compared to healthy normal tissue. Western blotting in additional brushed biopsy samples confirmed a trend of gradual decreasing SLPI abundance between healthy normal and OPML tissue, with a larger decrease in OSCC lesion tissue. A similar SLPI decrease was observed in-vitro comparing model OPML and OSCC cell lines. In addition, exfoliated oral cells in patients’ whole saliva showed a loss of SLPI correlated with oral cancer progression. These results, combined with proteomics data indicating a decrease in SLPI in matched healthy control tissue from OSCC patients compared to tissue from healthy normal tissue, suggested a systemic decrease of SLPI in oral cells correlated with oral cancer development. Finally, in-vitro experiments showed that treatment with SLPI significantly decreased NF-kB activity in an OPML cell line. The findings indicate anti-inflammatory activity in OPML, supporting a mechanistic role of SLPI in OSCC progression and suggesting its potential for preventative treatment of at-risk oral lesions. Collectively, our results show for the first time the potential for SLPI as a mechanism-based, non-invasive biomarker of oral cancer progression with potential in preventive treatment.

Highlights

The survival rate for people diagnosed with oral cancer, predominantly in the form of oral squamous cell carcinoma (OSCC), is only slightly better than 50%[1]
The first compared separate protein mixtures pooled from two oral premalignant lesion tissue (OPMLs) tissues, two matched OPML control tissues, two OSCC tissues and two healthy normal tissues
The second compared separate protein mixtures pooled from four OPML tissues, four OSCC tissues, four matched OSCC control tissues and four healthy normal tissues

Summary

Introduction

The survival rate for people diagnosed with oral cancer, predominantly in the form of oral squamous cell carcinoma (OSCC), is only slightly better than 50%[1]. Preventive treatments are more effective, increasing the survival rate to 80% or better[4]. There is a pressing need for better ways to diagnose and treat at-risk OPML and/or early-stage OSCC oral lesions[2]. Invasive incisional biopsy followed by histopathology is the current gold standard for oral cancer diagnosis[5]. Scalpel biopsy is prone to undersampling of lesions[8,9], thereby leading to errors in diagnosis

Methods

Results

Conclusion