Oncogenic Micrornas In Cerebrospinal Fluid and Sera Reflect Therapy Efficacy and Their Reappearance Precedes Clinical Relapse In Primary and Secondary CNS Lymphoma

MicroRNAs (miRNAs) are single-stranded non-coding RNAs (21-25nt) that negatively regulate gene expression at post-transcriptional level by blocking translation and mRNA destabilization. Diffuse large B cell lymphomas (DLBCL) overexpress oncogenic microRNAs (including miR-17-92 and miR-155) that regulate tumor growth and spreading. Unlike longer species of RNA, microRNAs are stable and detectable in body fluids including serum/plasma and cerebrospinal fluid (CSF), however their kinetics in body fluids upon course the therapy of DLBCL has not been fully explored. Notably DLBCLs with primary (PCNSL) or secondary CNS involvement display poor prognosis, short overall survival and their current diagnosis and disease monitoring is based on conventiontional methods (magnetic resonance imaging, histopathology, CSF cytology and flow cytometry) with unsatisfactory sensitivity.

We investigated abundance of microRNAs in the CSF and sera of DLBCL patients with (N=18) or without (N=30) CNS involvement and compared their levels at diagnosis and during treatment-response phase.

Extensive time-points to collect CSF and sera (5-20) were used during and after administration of therapy. Twenty candidate miRs from DLBCL patients were quantitated by TaqMan RT-PCR and their levels adjusted to those of control miRs and normal donors.

Analysis of CSF at diagnosis revealed that most prominently deregulated miRs (members of oncogenic miR-17-92 cluster and miR-21) were specificaly elevated in both PCNSL (N=11) and systemic lymphomas with CNS involvement (SCNSL, N=7) to similar extent while in systemic lymphomas without CNS involvement (N=30) their levels were significantly lower. Analysis of sera at diagnosis revealed that all SCNSL patients and ∼50% of systemic lymphomas without CNS involvement expressed high levels of the oncomiRs. Surprisingly, also a subset of PCNSL patients had elevated these miRs in the sera and in turn, a subset of systemic lymphomas without CNS involvement showed moderately elevated oncomiRs in the CSF. These data indicate that 1) lymphoma patients with CNS involvement (PCNSL or SCNSL) are characterized by increase of oncogenic microRNAs in CSF, 2) blood brain barrier is likely permissive for miRs in both directions.

Next, we investigated the dynamics of abundance of the oncomiRs in CSF upon course of the treatment in 8 patients with PCNSL or SCNSL. In therapy-responders (N=4) the oncomiRs gradually decreased in CSF. However, their levels were not completely normalized as compared to normal individuals. In contrast, refractory lymphomas (N=2) displayed gradual increase of these miRs in CSF regardless the treatment. Interestingly, clinical relapse within CNS as confirmed by MRI and cytology (N=2) was preceded (>3 months) by significant and gradual elevation of miRs in CSF. In systemic lymphoma (N=2) increased levels of studied oncomiRs in CSF preceded the CNS progression. In all cases high levels of miRs in CSF preceded and positively correlated with data from flow cytometry and cytology.

The measurement of oncomiRs in CSF and sera represents sensitive tool for detection of CNS lymphoma, for monitoring and estimation of therapy efficacy and prediction of the CNS relapse. Furthermore, our data indicate that elevated expression of oncomiRs in CSF and sera in systemic lymphomas may become helpful to predict CNS lymphoma involvement. Grants: GACR305 13-12449P, UNCE204021, PRVOUK: P24/LF1/3, P27/LF1/1, P 27/2012 LF3

Trneny: Roche: Honoraria, Research Funding.