Deep sequencing and automated histochemistry of human tissue slice cultures improve their usability as preclinical model for cancer research

Susann Haehnel,Juergen Meixensberger,Sven-Holger Puppel,Ingo Bechmann,Kristin Reiche,Felicitas Rapp,Frank Gaunitz,Karsten Winter,Michael Rade,Dennis Loeffler,Andreas Horn,Friedemann Horn,Nicole Kaiser,Conny Blumert

doi:10.1038/s41598-019-56509-5

Abstract

Cancer research requires models closely resembling the tumor in the patient. Human tissue cultures can overcome interspecies limitations of animal models or the loss of tissue architecture in in vitro models. However, analysis of tissue slices is often limited to histology. Here, we demonstrate that slices are also suitable for whole transcriptome sequencing and present a method for automated histochemistry of whole slices. Tumor and peritumoral tissue from a patient with glioblastoma was processed to slice cultures, which were treated with standard therapy including temozolomide and X-irradiation. Then, RNA sequencing and automated histochemistry were performed. RNA sequencing was successfully accomplished with a sequencing depth of 243 to 368 x 106 reads per sample. Comparing tumor and peritumoral tissue, we identified 1888 genes significantly downregulated and 2382 genes upregulated in tumor. Treatment significantly downregulated 2017 genes, whereas 1399 genes were upregulated. Pathway analysis revealed changes in the expression profile of treated glioblastoma tissue pointing towards downregulated proliferation. This was confirmed by automated analysis of whole tissue slices stained for Ki67. In conclusion, we demonstrate that RNA sequencing of tissue slices is possible and that histochemical analysis of whole tissue slices can be automated which increases the usability of this preclinical model.

Highlights

Cancer research requires models closely resembling the tumor in the patient
Organotypic slice cultures derived from human tissues, including tumors, came into focus as an alternative model[2]
We report that organotypic slice cultures are suitable for automated histological analyses as well as whole transcriptome sequencing, thereby providing an adequate alternative with regard to individualized cancer research and therapy

Summary

Introduction

Cancer research requires models closely resembling the tumor in the patient. Human tissue cultures can overcome interspecies limitations of animal models or the loss of tissue architecture in in vitro models. We have already established slice cultures from human brains[3], Glioblastoma multiforme (GBM)[4,5], head and neck squamous cell carcinoma[6], human gastric and esophagogastric junction cancer[7], and colorectal carcinoma[8] Using these organotypic slice cultures, we tested, for example, effects of heavy ion therapy[5], polyethylenimine-based nanoparticles for siRNA delivery[9], and novel nanostructured scaffolds for cultivation[4]. We report that organotypic slice cultures are suitable for automated histological analyses as well as whole transcriptome sequencing, thereby providing an adequate alternative with regard to individualized cancer research and therapy

Methods

Results

Conclusion