Donor-derived brain tumor following neural stem cell transplantation in an ataxia telangiectasia patient

Abstract

Background: Neural stem cells are currently being investigated as potential therapies for neurodegenerative diseases, stroke, and trauma. However, concerns have been raised over the safety of this experimental therapeutic approach, including, for example, whether there is the potential for tumors to develop from transplanted stem cells.

Methods and findings: A boy with ataxia telangiectasia (AT) was treated with intracerebellar and intrathecal injection of human fetal neural stem cells. Four years after the first treatment he was diagnosed with a multifocal brain tumor. The biopsied tumor was diagnosed as a glioneuronal neoplasm. We compared the tumor cells and the patient's peripheral blood cells by fluorescent in situ hybridization using X and Y chromosome probes, by PCR for the amelogenin gene X- and Y-specific alleles, by MassArray for the ATM patient specific mutation and for several SNPs, by PCR for polymorphic microsatellites, and by human leukocyte antigen (HLA) typing. Molecular and cytogenetic studies showed that the tumor was of nonhost origin suggesting it was derived from the transplanted neural stem cells. Microsatellite and HLA analysis demonstrated that the tumor is derived from at least two donors.

Conclusions: This is the first report of a human brain tumor complicating neural stem cell therapy. The findings here suggest that neuronal stem/progenitor cells may be involved in gliomagenesis and provide the first example of a donor-derived brain tumor. Further work is urgently needed to assess the safety of these therapies.

Figure 1. Pre-Operative MRI

(A) Brain MRI (T2) demonstrating a lesion (arrow) based on the tentorium, next to the brain stem (BS). (B) Spinal-lumbar MRI (T2) showing intradural lesion (arrow), at the level of the L4 vertebra.

Figure 2. Intra-Operative View

(A) The main lesion situated within the cauda equina nerve roots (I, the main tumor bulk; II, nerve root entering the tumor; III, normal nerve root). (B) One lumbar level above the main lesion, several “satellite” lesions are seen adherent to the caudal nerve roots (I, II, III, satellite tumors rostral to the main tumor bulk).

Figure 3. Hematoxylin–Eosine Staining of the Tumor

(A) The bulk of the specimen consisted of ill-defined lobules of glioneuronal cells. (B) These cells are best seen on Masson trichome stain. (C) At medium power glioneuronal cell circumscription by astroglial cells is apparent. (D) The centers of the lobules consist of two cell types, including pale neurons with large round nuclei, open chromatin, and a distinct nucleolus as well as smaller, astrocytic cells with more hyperchromatic nuclei and pale cytoplasm.

Figure 4. Immunohistochemistry of the Tumor

(A) Immunohistochemical staining of the cells comprising the nodules show abundance of neurons as evidenced on synaptophysin, (B) neu N, and (C) neurofilament protein preparations. (D) The stromal astrocytes show cytoplasmic reactivity for glial fibrillary acidic protein.

Figure 5. Histology and Immunohistochemistry of the Tumor

(A) The minor element of the tumor consists of markedly elongate ependymal cells (tanycytes) with refractile, fibril-rich processes terminating upon the stroma. (B) Note strong staining for S-100 protein, and (C) glial fibrillary acidic protein. (D) Scattered dot-like paranuclear microlumens, a feature of ependymal cells, show epithelial membrane antigen reactivity.

Figure 6. Analysis of X and Y Chromosomes of the Tumor Cells Using I-FISH and PCR Analysis of the Amelogenin X- and Y-Specific Alleles Showing the Presence of Female Cells in the Tumor

(A) Combined analysis of morphology and I-FISH with X chromosome (SpectrumGreen) and Y chromosome (SpectrumOrange) probes on tumor touch preps. Upper panels, May Grunwald Giemsa-stained cells; lower panels, I-FISH analysis of the cells appearing in the upper panel. Two green signals indicate female cells (XX). One green and one orange signal indicate male cells (XY). (B) PCR analysis of the X chromosome and Y chromosome amelogenin homologous alleles in the patient's PB and tumor (T).

Figure 7. Analysis of the ATM C103T Mutation and the VKORC1 G9041A (rs7294) Polymorphism Showing Foreign Origin of the Tumor Cells

(A) MALDI-TOF mass spectrometry of nucleotide 103 of the ATM gene using the SEQUENOM MassArray. C, wild type allele; T, the North African Jewish ATM mutated allele. F, father's PB; M, mother's PB; P-PB, patient's PB; T, tumor; T-LCM, isolated tumor cells obtained by LCM. (B) MALDI-TOF mass spectrometry of VKORC1 9041G/A (rs7294) polymorphism using the SEQUENOM MassArray. G/A, SNP at position 9041 (rs7294) of the VKORC1 gene. F, father's PB; M, mother's PB; P-PB, patient's PB; T, undissected tumor; T-LCM, isolated tumor cells obtained by LCM.

Figure 8. Analysis of DNA Tetranucleotide Repeats by Fluorescent PCR and Indicating the Origin of Tumor Cells from at Least Two Donors

(A) Polymorphic pattern of the DNA tetranucleotide repeats of the D18S51, D21S11, D8S117, vWA, D2S1338, and FGA loci as detected by fluorescent PCR. PB, patient PB; T, tumor. (B) HLA-A*, B*, and DRB1* typing obtained by reverse-sequence specific oligonucleotide probes (SSOP) using the Luminex platform. The patient's germline HLA (determined in PB and BS) are highlighted in red. Nonself alleles are highlighted in black.