Cerebrospinal fluid liquid biopsy for detecting somatic mosaicism in brain

doi:10.1093/braincomms/fcaa235

. 2021 Jan 21;3(1):fcaa235.

doi: 10.1093/braincomms/fcaa235. eCollection 2021.

Cerebrospinal fluid liquid biopsy for detecting somatic mosaicism in brain

Zimeng Ye¹, Zac Chatterton², Jahnvi Pflueger^{3

4}, John A Damiano¹, Lara McQuillan¹, Anthony Simon Harvey^{5

6

7}, Stephen Malone⁸, Hongdo Do^{9

10

11}, Wirginia Maixner¹², Amy Schneider¹, Bernadette Nolan⁸, Martin Wood¹³, Wei Shern Lee^{5

6}, Greta Gillies^{5

6}, Kate Pope⁵, Michael Wilson^{5

6}, Paul J Lockhart^{5

6}, Alexander Dobrovic^{10

11

14}, Ingrid E Scheffer^{1

5

6

7}, Melanie Bahlo^{15

16}, Richard J Leventer^{5

6

7}, Ryan Lister^{3

4}, Samuel F Berkovic¹, Michael S Hildebrand^{1

5}

Affiliations

¹ Department of Medicine (Austin Health), University of Melbourne, Melbourne, Victoria 3084, Australia.
² Brain and Mind Centre, Sydney Medical School, The University of Sydney, Sydney, New South Wales 2050, Australia.
³ Australian Research Council Centre of Excellence in Plant Energy Biology, School of Molecular Sciences, The University of Western Australia, Perth, Western Australia 6009, Australia.
⁴ Harry Perkins Institute of Medical Research, Perth, Western Australia 6150, Australia.
⁵ Murdoch Children's Research Institute, Melbourne, Victoria 3052, Australia.
⁶ Department of Paediatrics, University of Melbourne, Royal Children's Hospital, Melbourne, Victoria 3052, Australia.
⁷ Department of Neurology, Royal Children's Hospital, Melbourne, Victoria 3052, Australia.
⁸ Department of Neurosciences, Queensland Children's Hospital, Brisbane, Queensland 4101, Australia.
⁹ Department of Anatomical Pathology, St. Vincent's Hospital, Melbourne, Victoria 3065, Australia.
¹⁰ School of Cancer Medicine, La Trobe University, Melbourne, Victoria 3086, Australia.
¹¹ Department of Clinical Pathology, University of Melbourne, Melbourne, Victoria 3010, Australia.
¹² Department of Neurosurgery, Royal Children's Hospital, Melbourne, Victoria 3052, Australia.
¹³ Neurosurgical Department, Queensland Children's Hospital, Brisbane, Queensland 4101, Australia.
¹⁴ Translational Genomics and Epigenomics Laboratory, Olivia Newton-John Cancer Research Institute, Melbourne, Victoria 3084, Australia.
¹⁵ Population Health and Immunity Division, The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria 3052, Australia.
¹⁶ Department of Medical Biology, University of Melbourne, Melbourne, Victoria 3010, Australia.

PMID: 33738444
PMCID: PMC7954394
DOI: 10.1093/braincomms/fcaa235

Cerebrospinal fluid liquid biopsy for detecting somatic mosaicism in brain

Zimeng Ye et al. Brain Commun. 2021.

. 2021 Jan 21;3(1):fcaa235.

doi: 10.1093/braincomms/fcaa235. eCollection 2021.

Authors

Affiliations

¹ Department of Medicine (Austin Health), University of Melbourne, Melbourne, Victoria 3084, Australia.
² Brain and Mind Centre, Sydney Medical School, The University of Sydney, Sydney, New South Wales 2050, Australia.
³ Australian Research Council Centre of Excellence in Plant Energy Biology, School of Molecular Sciences, The University of Western Australia, Perth, Western Australia 6009, Australia.
⁴ Harry Perkins Institute of Medical Research, Perth, Western Australia 6150, Australia.
⁵ Murdoch Children's Research Institute, Melbourne, Victoria 3052, Australia.
⁶ Department of Paediatrics, University of Melbourne, Royal Children's Hospital, Melbourne, Victoria 3052, Australia.
⁷ Department of Neurology, Royal Children's Hospital, Melbourne, Victoria 3052, Australia.
⁸ Department of Neurosciences, Queensland Children's Hospital, Brisbane, Queensland 4101, Australia.
⁹ Department of Anatomical Pathology, St. Vincent's Hospital, Melbourne, Victoria 3065, Australia.
¹⁰ School of Cancer Medicine, La Trobe University, Melbourne, Victoria 3086, Australia.
¹¹ Department of Clinical Pathology, University of Melbourne, Melbourne, Victoria 3010, Australia.
¹² Department of Neurosurgery, Royal Children's Hospital, Melbourne, Victoria 3052, Australia.
¹³ Neurosurgical Department, Queensland Children's Hospital, Brisbane, Queensland 4101, Australia.
¹⁴ Translational Genomics and Epigenomics Laboratory, Olivia Newton-John Cancer Research Institute, Melbourne, Victoria 3084, Australia.
¹⁵ Population Health and Immunity Division, The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria 3052, Australia.
¹⁶ Department of Medical Biology, University of Melbourne, Melbourne, Victoria 3010, Australia.

PMID: 33738444
PMCID: PMC7954394
DOI: 10.1093/braincomms/fcaa235

Erratum in

Corrigendum to: Cerebrospinal fluid liquid biopsy for detecting somatic mosaicism in brain.
[No authors listed] [No authors listed] Brain Commun. 2021 Jun 17;3(2):fcab103. doi: 10.1093/braincomms/fcab103. eCollection 2021. Brain Commun. 2021. PMID: 34151265 Free PMC article.

Abstract

Brain somatic mutations are an increasingly recognized cause of epilepsy, brain malformations and autism spectrum disorders and may be a hidden cause of other neurodevelopmental and neurodegenerative disorders. At present, brain mosaicism can be detected only in the rare situations of autopsy or brain biopsy. Liquid biopsy using cell-free DNA derived from cerebrospinal fluid has detected somatic mutations in malignant brain tumours. Here, we asked if cerebrospinal fluid liquid biopsy can be used to detect somatic mosaicism in non-malignant brain diseases. First, we reliably quantified cerebrospinal fluid cell-free DNA in 28 patients with focal epilepsy and 28 controls using droplet digital PCR. Then, in three patients we identified somatic mutations in cerebrospinal fluid: in one patient with subcortical band heterotopia the LIS1 p. Lys64* variant at 9.4% frequency; in a second patient with focal cortical dysplasia the TSC1 p. Phe581His*6 variant at 7.8% frequency; and in a third patient with ganglioglioma the BRAF p. Val600Glu variant at 3.2% frequency. To determine if cerebrospinal fluid cell-free DNA was brain-derived, whole-genome bisulphite sequencing was performed and brain-specific DNA methylation patterns were found to be significantly enriched (P = 0.03). Our proof of principle study shows that cerebrospinal fluid liquid biopsy is valuable in investigating mosaic neurological disorders where brain tissue is unavailable.

Keywords: cell-free DNA; cerebrospinal fluid; focal epilepsy; liquid biopsy; somatic mutations.

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Figures

**Figure 1**
**ddPCR 2D-plot for molecular diagnosis in CSF from three patients with lesional focal epilepsy.** (A). 2D-Plot showing droplets positive (blue, upper left quadrant) for *LIS1* Lys64* in patient 1. CSF cell-free DNA was tested twice with similar VAF, then data were combined. Mutant concentration 3.61 copies/μl; wild-type concentration 35 copies/μl; VAF: 9.4%. (B) 2D-plot showing droplets positive for *TSC1* p. Phe581His*6 in patient 2. CSF cell-free DNA: mutant concentration 0.47 copies/μl; wild-type concentration 5.58 copies/μl; VAF: 7.8%. (C) 2D-plot showing droplets positive for *BRAF* Val600Glu in patient 3. CSF cell-free DNA: mutant concentration 0.42 copies/μl; wild-type concentration 12.7 copies/μl; VAF: 3.2%. Green droplets are wild-type copies, orange droplets are double-positive copies and grey droplets are empty. VAF: variant allele frequency.

**Figure 2**
**Enrichment of CNS-derived cell-free DNA in CSF samples.** (A) Unique pseudo-alignment of cell-free DNA from CSF and plasma to reference sequences/k-mers from 11 tissues. (B) Unique sequence pseudo-alignments of cell-free DNA from CSF and plasma to brain and blood cells. CSF-derived cell-free DNA has significantly higher number of unique reads pseudo-aligned to brain compared to plasma-derived cell-free DNA (P = 0.03), whereas plasma-derived cell-free DNA was significantly enriched for unique reads pseudo-aligned to blood cells (P = 0.04).

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References

1. Allen AS, Berkovic SF, Cossette P, Delanty N, Dlugos D, Eichler EE, et al. De novo mutations in epileptic encephalopathies. Nature 2013; 501: 217–21. - PMC - PubMed
1. Berghoff AS, Preusser M.. BRAF alterations in brain tumours: molecular pathology and therapeutic opportunities. Curr Opin Neurol 2014; 27: 689–96. - PubMed
1. Bianchi DW, Chiu RWK.. Sequencing of circulating cell-free DNA during pregnancy. N Engl J Med 2018; 379: 464–73. - PMC - PubMed
1. Bray NL, Pimentel H, Melsted P, Pachter L.. Near-optimal probabilistic RNA-seq quantification. Nat Biotechnol 2016; 34: 525–7. - PubMed
1. Chandrananda D, Thorne NP, Bahlo M.. High-resolution characterization of sequence signatures due to non-random cleavage of cell-free DNA. BMC Med Genomics 2015; 8: 29. - PMC - PubMed

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[1] Allen AS, Berkovic SF, Cossette P, Delanty N, Dlugos D, Eichler EE, et al. De novo mutations in epileptic encephalopathies. Nature 2013; 501: 217–21. - PMC - PubMed

[2] Allen AS, Berkovic SF, Cossette P, Delanty N, Dlugos D, Eichler EE, et al. De novo mutations in epileptic encephalopathies. Nature 2013; 501: 217–21. - PMC - PubMed

[3] Berghoff AS, Preusser M.. BRAF alterations in brain tumours: molecular pathology and therapeutic opportunities. Curr Opin Neurol 2014; 27: 689–96. - PubMed

[4] Berghoff AS, Preusser M.. BRAF alterations in brain tumours: molecular pathology and therapeutic opportunities. Curr Opin Neurol 2014; 27: 689–96. - PubMed

[5] Bianchi DW, Chiu RWK.. Sequencing of circulating cell-free DNA during pregnancy. N Engl J Med 2018; 379: 464–73. - PMC - PubMed

[6] Bianchi DW, Chiu RWK.. Sequencing of circulating cell-free DNA during pregnancy. N Engl J Med 2018; 379: 464–73. - PMC - PubMed

[7] Bray NL, Pimentel H, Melsted P, Pachter L.. Near-optimal probabilistic RNA-seq quantification. Nat Biotechnol 2016; 34: 525–7. - PubMed

[8] Bray NL, Pimentel H, Melsted P, Pachter L.. Near-optimal probabilistic RNA-seq quantification. Nat Biotechnol 2016; 34: 525–7. - PubMed

[9] Chandrananda D, Thorne NP, Bahlo M.. High-resolution characterization of sequence signatures due to non-random cleavage of cell-free DNA. BMC Med Genomics 2015; 8: 29. - PMC - PubMed

[10] Chandrananda D, Thorne NP, Bahlo M.. High-resolution characterization of sequence signatures due to non-random cleavage of cell-free DNA. BMC Med Genomics 2015; 8: 29. - PMC - PubMed

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Cerebrospinal fluid liquid biopsy for detecting somatic mosaicism in brain

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Cerebrospinal fluid liquid biopsy for detecting somatic mosaicism in brain

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