Tumor and Cerebrospinal Fluid microRNAs in Primary Central Nervous System Lymphomas

Abstract

Primary central nervous system lymphoma (PCNSL) is a rare, highly aggressive, extranodal form of non-Hodgkin lymphoma, predominantly diagnosed as primary diffuse large B-cell lymphoma of the central nervous system (CNS DLBCL). Fast and precise diagnosis of PCNSL is critical yet challenging. microRNAs, important regulators in physiology and pathology are potential biomarkers. In 131 patients with CNS DLBCL and with non-malignant brain lesions (n-ML), miR-21, miR-19b and miR-92a, miR-155, miR-196b, miR-let-7b, miR-125b, and miR-9 were examined by RT-qPCR in brain biopsy samples (formalin-fixed paraffin-embedded tissues, FFPET; CNS DLBCL, n = 52; n-ML, n = 42) and cerebrospinal fluid samples (CSF; CNS DLBCL, n = 30; n-ML, n = 23) taken for routine diagnosis. FFPET samples were split into study and validation sets. Significantly higher CSF levels of miR-21, miR-19b, and miR-92a were identified in PCNSL but not in n-ML, and differentiated PCNSL from n-ML with 63.33% sensitivity and 80.77% specificity. In FFPETs, miR-155 and miR-196b were significantly overexpressed and miR-let-7b, miR-125b, and miR-9 were downregulated in PCNSL as compared to n-ML. Combined miR-155 and miR-let-7b expression levels in FFPETs discriminated PCNSL and n-ML with a 97% accuracy. In conclusion, tissue miR-155, miR-196b, miR-9, miR-125b, and miR-let-7b expression profiles differentiate PCNSL from n-ML. PCNSL CSFs and the relevant biopsy samples are characterized by specific, different microRNA profiles. A logistic regression model is proposed to discriminate between PCNSL and non-malignant brain lesions. None of the examined microRNAs influenced overall survival of PCNSL patients. Further ongoing developments involve next generation sequencing-based profiling of biopsy and CSF samples.

A Set of 17 microRNAs Common for Brain and Cerebrospinal Fluid Differentiates Primary Central Nervous System Lymphoma from Non-Malignant Brain Tumors

“…The utility of CSF miR-21, miR-19b, and miR-92a as PCNSL markers has been suggested [ 12 ]. We have recently confirmed [ 13 ] that the miR-21, miR-19b, and miR-92a levels were significantly higher in CSFs of patients with CNS DLBCL than of patients with non-malignant brain tumors; however, the diagnostic accuracy of those miRs was found to be lower than previously suggested. We also demonstrated that CNS DLBCL CSFs and the relevant brain biopsy samples are characterized by specific, different miR profiles [ 13 ].…”

“…For HP examination, hematoxylin and eosin staining was performed. For IHC, tissue sections were incubated with the diluted antibodies for 1 h in an autostainer (Dako, Carpinteria, CA, USA) following the antigen-retrieval technique, if necessary, using the EnVision™ Detection Systems FLEX kit (Dako Corp., Carpinteria, CA, USA, code K 8000) and monoclonal antibodies (MoAbs) specific for CD20, CD10, BCL6, MUM1, BCL2, CD5, and Ki-67, as previously described [ 13 ]. A reaction for CD20, BCL6, MUM1, and CD5 was considered positive if at least 20% of the CNS DLBCL cells showed staining, while for CD10 if any cell showed staining.…”

“…Immunophenotyping of CSF samples was performed by FCM. Concentrated cells isolated from CSFs by centrifugation were incubated with a panel of MoAbs (for the staining procedure, see [ 14 ], and for a list of MoAbs, see [ 13 ]). The expression of B-cell antigens CD(45/19/20/10/HLADR), T-cell antigens CD(45/3/4/8/5/43), and macrophage antigens (CD14/CD4/43/HLADR) was quantified on FACSCalibur and FACSCanto II cytometers (Becton Dickinson, BD, San Jose, CA, USA) and samples were categorized into three groups, according to the percentages of positive cells: (−), no expression (<20% of positive neoplastic cells); (+/−), expression in ≥20%<100% of cells; and (+), expression in 100% of cells.…”

“…Total RNA was isolated from CSF and tumor samples according to the protocols described earlier by Zajdel et al [ 13 ]. Briefly, total RNA was isolated from CSF samples with the Gene Matrix Universal RNA/miRNA Purification Kit (EURx, Gdansk, Poland), according to the manufacturer’s instructions.…”

Liquid Biopsy and Other Non-Invasive Diagnostic Measures in PCNSL

“…Notably, these findings could be reproduced (sensitivity 97.4%) in an enlarged cohort ( n = 39) of PCNSL patients [ 50 ]. Furthermore, these data were confirmed by Zajdel et al with a lower diagnostic accuracy, which can be explained by the selection of a different reference group [ 27 ].…”