Proteomic changes in cerebrospinal fluid from primary central nervous system lymphoma patients are associated with protein ectodomain shedding

Abstract

Primary central nervous system lymphomas (PCNSLs) are mature B-cell lymphomas confined to the central nervous system (CNS). Blood-brain barrier (BBB) dysfunction drastically alters the cerebrospinal fluid (CSF) proteome in PCNSL patients. To reveal the interaction of PCNSL tumors with CNS structures and the vasculature, we conducted a whole-proteome analysis of CSF from PCNSL patients (n = 17 at initial diagnosis) and tumor-free controls (n = 10) using label-free quantitative mass spectrometry. We identified 601 proteins in the CSF proteome using a one-step approach without further prefractionation, and quantified 438 proteins in detail using the Hi-N method. An immunoassay revealed that 70% of the patients in our unselected PCNSL patient cohort had BBB dysfunction. Correlation analysis indicated that 127 (30%) of the quantified proteins were likely increased in PCSNL patients due to BBB dysfunction. After the exclusion of these proteins, 66 were found to differ in abundance (fold-change > 2.0, p < 0.05) between PCNSL and control CSF proteomes, and most of those were associated with the CNS. These data also provide the first evidence that proteomic changes in CSF from PCNSL patients are mainly associated with protein ectodomain shedding, and that shedding of human leukocyte antigen class 2 proteins is a mechanism of tumor-cell immune evasion.

A Multiplex Assay for the Stratification of Patients with Primary Central Nervous System Lymphoma Using Targeted Mass Spectrometry

“…Removal of 375 plasma proteins which likely appear as a result of plasma leakage due to BBB disruption allowed us to avoid false-positive candidate biomarkers [17]. Detailed data analysis and immunohistochemistry confirmed the results from a previous study which showed that more than half of the proteins with significant abundance changes in the CSF of PCNSL patients originated mainly from the surrounding CNS tissue instead of the lymphoma tissue [13]. Here, we speculate that the lymphoma tissue interacts and communicates with its environment (CNS tissue) that causes the release of proteinaceous factors by different secretory pathways (ectodomain shedding).…”

“…This holds also true for the comparison of PCNSL with SCNSL (51%), PCNSL with gliomas (56%), PCNSL with "other tumors" (55%) and PCNSL with MS (52%) ( Table 2). We also identified a high number of secreted and membrane proteins among the differentially abundant proteins which is in concordance with our former observations [13]. As the second criterion for the selection of candidate biomarkers, we considered the diagnostic accuracy determined by the ROC analysis of the individual peptide intensities determined by label-free mass spectrometry (Table 4, Figure S1A-E).…”

“…Frequently, PCNSL and other brain tumors cause blood-brain barrier (BBB) dysfunction, which leads to plasma protein leakage into the CSF [13,16,17]. To reveal the extent of BBB dysfunction in individual patients, we determined the CSF/serum quotients of albumin, IgG, IgA and IgM concentration ( Figure 1 and Table 2).…”

“…The analysis of putative biomarkers in serum (IL-6, IL-8, IL-10, CXCL-12 and CXCL 13) and CSF (IL-10, CXCL-12 and CXCL 13) revealed aberrant protein levels in association with PCNSL [8][9][10][11][12]. Furthermore, the occurrence of ectodomains from transmembrane proteins such as CD27 and sIL-2R were discovered in the CSF of PCNSL patients, which may represent candidate biomarkers [9,11,[13][14][15].…”

Degradomics in Biomarker Discovery

“…In clinical samples, several types of shed ectodomains have been identified in the biological fluids of cancer patients. A study performed on the cerebrospinal fluid proteome of patients with primary central nervous system lymphoma showed increased presence of 36 shed membrane proteins, which could be further examined and validated as possible biomarkers . In general, many of the disease‐related shed proteins are various types of adhesion molecules such as CAM proteins, cadherins, and CD44.…”