Project description:Although formalin fixed paraffin embedded (FFPE) tissue is a major biological source in cancer research, it is challenging to work with due to macromolecular fragmentation and nucleic acid crosslinking. Therefore, it is important to characterise the quality of data that can be obtained from FFPE samples. We have compared three independent platforms (next generation sequencing, microarray and NanoString) for profiling microRNAs (miRNAs) using clinical FFPE samples from hepatoblastoma (HB) patients. The number of detected miRNAs ranged from 228 to 345 (median = 294) using the next generation sequencing platform, whereas 79 to 125 (median = 112) miRNAs were identified using microarrays in three HB samples, including technical replicates. NanoString identified 299 to 372 miRNAs in two samples. Between the platforms, we observed high reproducibility and significant levels of shared detection. However, for commonly detected miRNAs, a strong correlation between platforms was not observed. Analysis of 10 additional HB samples with NanoString identified significantly overlapping miRNA expression profiles, and an alternative pattern was identified in a poorly differentiated HB with an aggressive phenotype. This investigation serves as a roadmap for future studies investigating miRNA expression in clinical FFPE samples, and as a guideline for the selection of an appropriate platform.
Project description:This work highlights similarities and differences between three platforms (next-generation sequencing, microarray and NanoString) for detecting miRNAs and compares their strengths and weaknesses.
Project description:This work highlights similarities and differences between three platforms (next-generation sequencing, microarray and NanoString) for detecting miRNAs and compares their strengths and weaknesses.
Project description:This work highlights similarities and differences between three platforms (next-generation sequencing, microarray and NanoString) for detecting miRNAs and compares their strengths and weaknesses.
Project description:This work highlights similarities and differences between three platforms (next-generation sequencing, microarray and NanoString) for detecting miRNAs and compares their strengths and weaknesses. miRNA expression profiles were determined in Hepatoblastoma FFPE samples using a NanoString platform.
Project description:This work highlights similarities and differences between three platforms (next-generation sequencing, microarray and NanoString) for detecting miRNAs and compares their strengths and weaknesses. miRNA expression profiles were determined in 6 Hepatoblastoma FFPE samples using a next-generation sequencing platform.
Project description:This work highlights similarities and differences between three platforms (next-generation sequencing, microarray and NanoString) for detecting miRNAs and compares their strengths and weaknesses. miRNA expression profiles were determined in 6 Hepatoblastoma FFPE samples using a microarray platform.
Project description:Current criteria for identifying cancer patients suitable for immunotherapy with immune checkpoint blockers (ICBs) are subjective and prone to misinterpretation, as they mainly rely on the visual assessment of CD274 (best known as PD-L1) expression levels by immunohistochemistry (IHC). To address this issue, we developed a RNA sequencing (RNAseq)-based approach that specifically measures the abundance of immune transcripts in formalin-fixed paraffin embedded (FFPE) specimens. Besides exhibiting superior sensitivity as compared to whole transcriptome RNAseq, our assay requires little starting material, implying that it is compatible with RNA degradation normally caused by formalin. Here, we demonstrate that a targeted RNAseq panel reliably profiles mRNA expression levels in FFPE samples from a cohort of ovarian carcinoma patients. The expression profile of immune transcripts as measured by targeted RNAseq in FFPE versus freshly frozen (FF) samples from the same tumor was highly concordant, in spite of the RNA quality issues associated with formalin fixation. Moreover, the results of targeted RNAseq on FFPE specimens exhibited a robust correlation with mRNA expression levels as measured on the same samples by quantitative RT-PCR, as well as with protein abundance as determined by IHC. These findings demonstrate that RNAseq profiling on archival FFPE tissues can be used reliably in studies assessing the efficacy of cancer immunotherapy.