Cryopreservation of neurospheres derived from human glioblastoma multiforme.
Cancer stem cells have been shown to initiate and sustain tumor growth. In many instances, clinical material is limited, compounded by a lack of methods to preserve such cells at convenient time points. Although brain tumor-initiating cells grown in a spheroid manner have been shown to maintain their integrity through serial transplantation in immune-compromised animals, practically, it is not always possible to have access to animals of suitable ages to continuously maintain these cells. We therefore explored vitrification as a cryopreservation technique for brain tumor-initiating cells. Tumor neurospheres were derived from five patients with glioblastoma multiforme (GBM). Cryopreservation in 90% serum and 10% dimethyl sulfoxide yielded greatest viability and could be explored in future studies. Vitrification yielded cells that maintained self-renewal and multipotentiality properties. Karyotypic analyses confirmed the presence of GBM hallmarks. Upon implantation into NOD/SCID mice, our vitrified cells reformed glioma masses that could be serially transplanted. Transcriptome analysis showed that the vitrified and nonvitrified samples in either the stem-like or differentiated states clustered together, providing evidence that vitrification does not change the genotype of frozen cells. Upon induction of differentiation, the transcriptomes of vitrified cells associated with the original primary tumors, indicating that tumor stem-like cells are a genetically distinct population from the differentiated mass, underscoring the importance of working with the relevant tumor-initiating population. Our results demonstrate that vitrification of brain tumor-initiating cells preserves the biological phenotype and genetic profiles of the cells. This should facilitate the establishment of a repository of tumor-initiating cells for subsequent experimental designs.
Chong
Yuk-Kien
YK
Singapore Institute for Clinical Sciences, Genome Institute of Singapore, Agency for Science, Technology and Research, Singapore.
Toh
Tan-Boon
TB
Zaiden
Norazean
N
Poonepalli
Anuradha
A
Leong
Siew Hong
SH
Ong
Catherine Ee Ling
CE
Yu
Yiting
Y
Tan
Patrick B
PB
See
Siew-Ju
SJ
Ng
Wai-Hoe
WH
Ng
Ivan
I
Hande
Manoor P
MP
Kon
Oi Lian
OL
Ang
Beng-Ti
BT
Tang
Carol
C
D000071916:AC133 Antigen; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D002449:Cell Aggregation; D002454:Cell Differentiation; D049109:Cell Proliferation; D048430:Cell Shape; D002470:Cell Survival; D015925:Cryopreservation; D015972:Gene Expression Regulation, Neoplastic; D005909:Glioblastoma / Q000235:genetics / Q000473:pathology*; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D007621:Karyotyping; D051379:Mice; D016513:Mice, SCID; D039902:Multipotent Stem Cells / Q000473:pathology; D014411:Neoplastic Stem Cells / Q000473:pathology; D009474:Neurons / Q000473:pathology*; D010455:Peptides / Q000378:metabolism; D018874:Spheroids, Cellular / Q000473:pathology*; D014407:Tumor Cells, Cultured; D023041:Xenograft Model Antitumor Assays
D000071916:AC133 Antigen; D015703:Antigens, CD; D006023:Glycoproteins; D010455:Peptides