The Application Of Vitrification-Based Cryopreservation Combining Slice Culturing Technology In Drug Testing Research

Purpose: The aim of this study was to propose a novel method combining vitrificationbased cryopreservation of tumor biopsy and precision-cut slice cultivation,to evaluate the efficacy of this experimental model in assessment of anticancer drug response.Methods: Biopsy tissues of rectal cancer liver metastasis were cryopreserved.Warmed tissues were used to establish patient-derived xenograft models and went down to the future generation.Xenografts were cut into 300?m thick slices.Tissue slices(2mm in diameter)were maintained on transwell inserts and treated with oxaliplatin and regorafenib at the concentration of 20 ?M,respectively.Mice were injected with oxaliplatin in 200 ?ml PBS at a dose of 5mg/kg and regorafenib in 200 ?ml PBS at a dose of 30mg/kg,respectively.CCK-8 assay,Calcein-AM cell viability assay/Hoechst 33342 staining and hematoxylin and eosin/immunohistochemistry staining were performed to evaluate the drug response.Results: Biological characteristics of rectal cancer liver metastasis biopsy could be retained by vitrification-based cryopreservation.Precision-cut slice provided complete three-dimensional architecture and full heterogeneity of the original tumor.The patientderived xenograft models were successfully established by both fresh and warmed biopsy tissues.The positive drug responses in xenograft model were consistent with slice culture in vitro.Conclusions: Tumor biopsy could be effectively cryopreserved,of which biological characteristics were well retained.Besides,precision-cut slices derived from the warmed tissues provided an efficient tool to assess anticancer drug response in vitro.