Metabolic Profiling in Formalin-Fixed and Paraffin-Embedded Prostate Cancer Tissues

Abstract

Metabolite profiling has significantly contributed to a deeper understanding of the biochemical metabolic networks and pathways in cancer cells. Metabolomics-based biomarker discovery would greatly benefit from the ability to interrogate retrospective annotated clinical specimens archived as formalin-fixed, paraffin-embedded (FFPE) material. Mass spectrometry–based metabolomic analysis was performed in matched frozen and FFPE human prostate cancers as well as isogenic prostate cancer cell lines. A total of 352 and 460 metabolites were profiled in human tissues and cell lines, respectively. Classes and physical–chemical characteristics of the metabolites preserved in FFPE material were characterized and related to their preservation or loss following fixation and embedding. Metabolite classes were differentially preserved in archival FFPE tissues, regardless of the age of the block, compared with matched frozen specimen, ranging from maximal preservation of fatty acids (78%) to loss of the majority of peptides and steroids. Generally, FFPE samples showed a decrease of metabolites with functional groups, such as carboxamide. As an adjunct technique, metabolic profiles were also obtained in situ from FFPE tissue sections where metabolites were extracted in a manner that preserves tissue architecture. Despite the fact that selected metabolites were not retained after processing, global metabolic profiles obtained from FFPE can be used to predict biologic states and study biologic pathways. These results pave the way for metabolomics-based biomarker discovery/validation utilizing retrospective and clinically annotated FFPE collections. Implications Metabolic profiles can be performed in archival tissue and may be used to complement other profiling methods such as gene expression for biomarker discovery or pathway analysis in the assessment of biologic states.

Reticular and erosive oral lichen planus have a distinct metabolomic profile: A preliminary study using gas chromatography‐mass spectrometry

“…The use of a single analytical platform could be associated with the detection of small differences in the metabolic profile of OLP clinical forms. Moreover, although metabolomics based on formalin‐fixed paraffin‐embedded tissue is a valuable tool to differentiate biological states, the process of formalin‐fixation with paraffin‐embedding causes a reduction in several classes of metabolites, such as peptides and steroids . Therefore, future metabolomics analyses of fresh frozen specimens are warranted to complement our findings.…”

Paraffin-Embedded Tissue as a Novel Matrix in Metabolomics Study: Optimization of Metabolite Extraction Method

“…The number of metabolites was similar to results obtained within presented study (157 compounds before filtration). However, the results presented by Cacciatore et al [16] were obtained with the use of two complementary techniques, whereas, in presented research, only one technique was applied. Moreover, authors did not distinguish how many compounds were detected by every applied technique.…”

“…However, some previously published studies covered the comparison of metabolic profiles obtained from FFPE tissues and fresh-frozen specimens. According to the literature, formalin fixation and paraffin embedding may cause loss of about 25% of detected metabolites when compared to fresh-frozen samples, especially such groups as amino acids, peptides, or carbohydrates [13,15,16]. Fortunately, the goal of our study was to analyze fatty compound synthesis and metabolism, which were found to be stable during FFPE sample preparation procedure.…”

“…As a result, 60 compounds were detected in FFPE samples. Recently, Cacciatore et al [16] performed untargeted metabolomic analysis, based on ultraperformance liquid chromatography-mass spectrometry (UPLC-MS) and GC-MS techniques, to compare two types of samples: fresh-frozen and FFPE cell lines of prostate cancer, and fresh-frozen, and FFPE prostate cancer tissues. As a result, 252 metabolites were detected in FFPE cell line specimens, while 140 metabolites were identified in FFPE prostate cancer tissues.…”

“…Consequently, tissue seems to be an adequate matrix when the aim is to investigate the mechanism of the disease [12]. In genomics or proteomics of cancer, formalin-fixed, paraffin-embedded (FFPE) tissues are commonly used samples, whereas they were rarely described in metabolomics research [13][14][15][16][17]. Preparation of FFPE samples includes three steps: formalin fixation, dehydratation, and paraffin embedding.…”