Biological characteristics comparison of glioblastoma tumorsphere obtained from the fresh and cryopreservation sample

Abstract

Background: Glioblastoma (GBM) is the most common and aggressive form of a primary tumor with poor prognosis. Tumorsphere (TS) isolated from human GBM have been shown to possess the potential for studying cancer biology, preclinical in vitro screening of anticancer drugs and use of patient-derived xenograft (PDX) models. Several studies have shown the high clinical significance of TS, but the storing of patient samples at an appropriate time point after surgical treatment is very limited. Recently some studies have demonstrated the benefits of cryopreservation, but to date, there have been no studies comparing the molecular comparison of Fresh and Cryo TS in detail. In this study, we tried to compare and analyze whether TS isolated by cryopreservation really maintains the same pattern as fresh sample devised from various aspects. Methods: Surgically obtained primary GBM tissues were divided into a fresh and cryopreserved group. Each tissue was chopped and further processed to cells isolation with or without cryopreservation media. Acquired TSs with two different culture method were analyzed for transcriptome analysis, stem cell characterization, neurosphere formation, invasive properties, Temozolomide (TMZ) response and mouse orthotopic models. Results: A total of 40 diagnosed primary GBM patient samples were used in this study. TS isolation success rate were 65% in fresh and 60% in cryopreserved group, respectively. There were no significant differences in neurosphere formation, stemness, differentiation between fresh and cryopreserved groups. TSs in two different environments featured the similar invasiveness, TMZ responsiveness and animal results highly showed similar capacity of tumor generation. In the RNA sequencing, the cancer subtypes between different TSs may be different, but the similarity between fresh and cryopreserved TSs did not change. Conclusions: These results showed that cryopreservation cells can be used for experiment as primary cultured cells, suggesting that cryopreservation is the alternative method when TS cannot be isolated immediately after tumor resection.