, 13:136 | Cite as

Identification of putative biomarkers for leptomeningeal invasion in B-cell non-Hodgkin lymphoma by NMR metabolomics

  • Gonçalo Graça
  • Joana Desterro
  • Joana Sousa
  • Carlos Fonseca
  • Margarida Silveira
  • Jacinta Serpa
  • Tânia Carvalho
  • Maria G. da Silva
  • Luís G. Gonçalves
Original Article



B-cell non-Hodgkin lymphoma (B-NHL) is the most common hematological malignancy and different genetic alterations are frequently detected in transformed B lymphocytes. Within this heterogeneous disease, certain aggressive subgroups have an increased risk of central nervous system (CNS) involvement at diagnosis and/or relapse, resulting in parenchymal or leptomeningeal infiltration (LI) in 5–15% of cases. The current sensitivity limitations of cerebrospinal fluid (CSF) cytology and contrast-enhanced MRI for CNS involvement, mainly at early stages, motivates the search for alternative diagnostic methods.


Here we aim at using untargeted 1H-NMR metabolomics to identify putative biomarkers for LI in B-NHL patients.


CSF and peripheral blood samples were obtained from B-NHL patients with a positive (n = 7, LI group) or negative LI diagnostic (n = 13, control group). For seven patients, CSF samples were collected during the course of intrathecal chemotherapy, making it possible to assess the patient´s response to treatment. 1H-NMR spectra were acquired and statistical multivariate and univariate analysis were performed to identify significant alterations.


Significant metabolite differences were found between LI and control groups in CSF, but not in serum. A predictive PLS-DA cross-validated model identified significant pool changes in glycine, alanine, pyruvate, acetylcarnitine, carnitine, and phenylalanine. Additionally, increments in protein signals were detected in the LI group. Significantly, the PLS-DA model predicted correctly all samples obtained from the group of patients in remission during LI treatment.


The results show that the CSF NMR-metabolomics approach is a promising complementary method in clinical diagnosis and treatment follow-up of LI in B-NHL patients.


NMR metabolomics B cell non-Hodgkin lymphoma Leptomeningeal infiltration Cerebrospinal fluid Serum 



The authors want to acknowledge Prof. Helena Santos for her support, involvement and contribution to the project. The NMR data was acquired at CERMAX (Centro de Ressonância Magnética António Xavier) and at CICS-UBI which are members of the Portuguese NMR network.


This work was supported by project PTDC/BIM-ONC/1242/2012 from Fundação para a Ciência e a Tecnologia (FCT), Portugal; project LISBOA-01-0145-FEDER-007660 (Microbiologia Molecular, Estrutural e Celular) and iNOVA4Health—UID/Multi/04462/2013 funded by FEDER through COMPETE2020—POCI and by national funds through FCT. GG and LGG were recipients of post-doc Grants, SFRH/BPD/93752/2013 and SFRH/BPD/111100/2015, awarded by FCT.

Author contributions

GG, LGG, and MGS wrote the manuscript. GG, assisted by JSo, prepared samples and acquired the NMR spectra. GG with assistance of LGG performed the analysis of spectra, produced the statistical models and interpreted results. JD, CF, MGS and MS collected CSF and blood samples, supervised the biochemical analyses, and gathered the clinical data. GG, LGG, TC, JSe and MGS designed the study. All authors contributed to the revision of the manuscript.

Compliance with ethical standards

Conflict of interest

All authors declare that they have no conflicts of interest.

Ethical approval

This study was reviewed and approved by the ethical committee of the Portuguese Oncology Institute Francisco Gentil, Lisbon (Approval Number: GIC/733 + UIC/660) and performed in accordance with the 1964 Helsinki declaration and its later amendments.

Informed consent

Serum and CSF samples were collected for routine clinical procedures and analyzed retrospectively; therefore in this study a formal consent is not required.

Supplementary material

11306_2017_1269_MOESM1_ESM.docx (168 kb)
Supplementary material 1 (DOCX 167 KB)


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Copyright information

© Springer Science+Business Media, LLC 2017

Authors and Affiliations

  • Gonçalo Graça
    • 1
  • Joana Desterro
    • 2
  • Joana Sousa
    • 1
  • Carlos Fonseca
    • 2
  • Margarida Silveira
    • 2
  • Jacinta Serpa
    • 2
    • 3
  • Tânia Carvalho
    • 4
  • Maria G. da Silva
    • 2
  • Luís G. Gonçalves
    • 1
  1. 1.ITQB NOVA, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de LisboaOeirasPortugal
  2. 2.IPOLFG, Instituto Português de Oncologia de Lisboa Francisco GentilLisbonPortugal
  3. 3.CEDOC, Centro de Investigação em Doenças Crónicas, NOVA Medical School, Faculdade de Ciências Médicas, Universidade Nova de LisboaLisbonPortugal
  4. 4.IMM, Instituto de Medicina Molecular, Universidade de LisboaLisbonPortugal

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