Name: Spheroids treated with and without Drug
Keywords: Spheroids

Dataset Information

Spheroids treated with and without Drug

ABSTRACT: MALDI MSI for monitoring of drug response in primary cancer spheroids

INSTRUMENT(S): ultraflex

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Primary Cell, Epithelial Cell

DISEASE(S): Malignant Neoplasm Of Ovary

SUBMITTER: Parul Mittal

LAB HEAD: Peter Hoffmann

PROVIDER: PXD013069 | Pride | 2019-09-03

REPOSITORIES: Pride

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Publications

Matrix Assisted Laser Desorption/Ionization Mass Spectrometry Imaging (MALDI MSI) for Monitoring of Drug Response in Primary Cancer Spheroids.

Mittal Parul P Price Zoe K ZK Lokman Noor A NA Ricciardelli Carmela C Oehler Martin K MK Klingler-Hoffmann Manuela M Hoffmann Peter P

Proteomics 20191015 21-22

Malignant ascites is a fluid, which builds up in the abdomen and contains cancer cells in the form of single cells or multicellular clusters called spheroids. Malignant ascites has been observed in patients suffering from ovarian, cervical, gastric, colorectal, pancreatic, endometrial, or primary liver cancer. The spheroids are believed to play a major role in chemo resistance and metastasis of the cancer. To ease the discomfort of patients, malignant ascites (MA) is often drained from the abdom ...[more]

PMID: 31474002

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Project description:It has been recognised for a long time that cell surface glycans plays a vital role in the biological process and their altered form can lead to cancer. However, the molecular details and regulatory mechanism between the glycans and cancer is yet not clear. Over the past decade mass spectrometry-based techniques has become a prominent method for analysing glycans especially N-glycan matrix assisted laser desorption/ionisation mass spectrometry imaging (MALDI MSI). It is a powerful technique that combines mass spectrometry with histology, enabling the visualisation and label free detection of N-linked glycans on a single tissue section. Here, we carried out N-glycan MALDI MSI on six endometrial cancer (EC) formalin fixed paraffin embedded (FFPE) tissue sections including the adjacent normal endometrium, and tissue microarrays (TMA) consisting of 8 EC patients with lymph node metastasis (LNM) and 20 without LNM. By doing that several m/z values were detected that can significantly distinguish normal endometrium from cancerous. Also, detected a m/z value that can discriminate the primary tumour with LNM from those without. Identification of those discriminative m/z values was performed using porous graphitized carbon liquid chromatography tandem mass spectrometry (PGC-LC-MS/MS). Overall, we observed higher abundance of oligomannose in tumour regions compared to normal with AUC ranges from 0.85-0.99. Whereas, complex N-glycans were detected in lower abundance with AUC ranges from 0.03-0.28. Comparison of N-glycans between the primary tumours with LNM and without LNM indicates reduced abundance of a complex core-fucosylated N-glycan in patients with metastasis relative to without. In summary, N-glycan MALDI MSI can be used to characterize the cancerous endometrium from the normal, also patients with LNM from those without. Identification of those discriminative m/z values was performed using porous graphitized carbon liquid chromatography tandem mass spectrometry (PGC-LC-MS/MS).

Project description:Spheroids, near spherical multicellular aggregates, are one of the most common types of threedimensional (3D) cell cultures. Despite decades of implementation of spheroid technology in various fields of life science and medical research, no minimal information (MI) guidelines are available to cope with heterogeneity and to stimulate transparency. To cope with this unmet need, we assembled an international consortium to develop the MISpheroID knowledgebase (https://www.mispheroid.org) and interrogation revealed heterogeneity and lack of transparency in published spheroid-related experiments. This steered us to empirically evaluate the impact of cell line, culture medium type, spheroid formation method and spheroid size on complementary spheroid metrics (RNA fingerprints, presence of cell death, ATP content, glucose to lactate conversion, secreted protein signatures, circularity, size and cancer therapy response). We measured media-induced transcriptional variation in lung cancer (A549), colorectal cancer (HCT116), ovarium cancer (SKOV3) and glioblastoma (U87MG) spheroids using RNA sequencing (RNA-seq). These spheroids were formed in ultra-low attachment plates and cultured in 6 different media types (DMEM high glucose, DMEM/F12, RPMI1640, DMEM low glucose, EMEM and MEM) for 5 (HCT116) or 7 (A549, SKOV3 and U87MG) days. RNA extraction was performed on 2 spheroids per condition using the miRNeasy micro kit (217084, Qiagen, Hilden, Germany). RNA-sequencing libraries were prepared from purified RNA using the QuantSeq 3' mRNA-Seq Library Prep Kit FWD for Illumina (Lexogen, Vienna, Austria) according to the manufacturer's instructions, using 27.5ng of RNA that was DNase treated using HL-dsDNA (Arcticzymes, TromsØ, Norway). The individual libraries were quantified by qPCR using the KAPA Library Quantification Kit (Roche, Pleasanton, CA, US) and equimolarly pooled. The pool concentration was measured with Qubit and 1.4pM with 1% PhiX was sequenced on a NextSeq 500 (Illumina, San Diego, CA, US) using a high-output 1x75 run. Reads were mapped to the human genome using Tophat and gene expression counts were generated using HTSeq. This data was used to perform Principal Component Analysis (PCA) and Gene Set Enrichment Analysis (GSEA).

Dataset Information

Spheroids treated with and without Drug

Publications

Matrix Assisted Laser Desorption/Ionization Mass Spectrometry Imaging (MALDI MSI) for Monitoring of Drug Response in Primary Cancer Spheroids.

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