Mass Spectrometry Imaging Reveals Molecular Changes Associated with Aging in Pancreatic Cancer
Introduction
Aging is one of the most important risk factors for pancreatic ductal adenocarcinoma (PDAC) with the typical median age of onset in the 70s at the time of diagnosis. However, a subset of patients experiences disease onset much earlier, especially among women, without any known genetic predisposition. The increasing recognition of age-associated pancreatic cancer incidence necessitates understanding diverse molecular changes associated with variations in age of onset and gender. Mass spectrometry imaging allows for detection of hundreds of biomolecules from tissue sections without target-specific reagents and multiple classes of analytes can be analyzed from the same section using sequential imaging. Here, we seek to interrogate molecular differences between early and typical onset pancreatic cancer as well as any gender-specific differences.
Methods
FFPE tissue sections were dewaxed with xylene before being coated with DAN matrix using an HTX M5 Sprayer. Metabolite images were collected in negative ion mode using a Bruker timsTOF fleX MALDI QTOF mass spectrometer. After image collection, matrix was removed and PNGase F digestion was performed on the same section before coating with CHCA matrix and N-glycan image collection. Subsequent to N-glycan imaging, matrix was removed and tryptic digestion was performed on the same section before coating with CHCA and peptide image collection. All images were collected at 20 μm resolution. Finally, the imaged sections were H&E stained for histological evaluation. Image visualization was performed using SCiLS Lab and metabolite and glycan identification were carried out using MetaboScape.
Results
In this study we sought to understand molecular differences between early onset PDAC (< 55 year) versus typical onset (>75 year) as well as any gender differences. Sequential imaging of the same tissue section allows us to maximize data coverage and perform direct co-registration of different classes of molecules to each other. Preliminary analysis focused on imaging tumor samples from two early onset and two elderly patients, one male and one female in each age group. Metabolite imaging revealed higher abundance of taurine and palmitic acid in samples from elderly patients, while more guanine was detected in samples from early onset patients. Cytosine and uracil were slightly more abundant in male patients than female patients with higher abundance in the early onset patient. Glycan imaging showed more abundant signal for Hex5dHex1HexNAc4 and an unknown glycan at m/z 2056.743 in tumor tissues from female patients. Hex5HexNAc 2 was most abundant in the sample from the elderly female patient. Finally, tryptic digest imaging was performed on the sections. From these data, we found an unknown peptide at m/z 958.572 to be more abundant in the elderly patients, as was actin. An unidentified peptide at m/z 1240.267 was almost exclusively detected in the early onset male patient. Histone H2A displayed similar abundance across all samples, while an unidentified peptide at m/z 1487.674 was more prevalent in the samples from elderly patients. Work is ongoing to replicate these results in additional samples and to identify more of the molecules detected in these imaging studies. We also plan to integrate these MSI results with other spatial-omics technologies, including spatial transcriptomics. The results gleaned from this work will aid in our understanding of how the aging process impacts dynamic changes of multi-analytes in pancreatic cancer.
Conclusion
Molecular differences in metabolites, glycans, and peptides were observed between early onset versus elderly as well as male versus female PDAC tumors.