Here we show that metabolomic profiling can be used to non-invasively identify novel biomarkers of AOM-induced colorectal neoplasia in mice. Altered metabolite levels were found in feces and plasma of tumor-bearing mice and in tumor tissue itself. In fact, a number of these metabolic changes overlapped across the 3 matrices.
Because colorectal tumors can shed cells or bleed, feces provide a potentially ideal matrix for identifying biomarkers of intestinal neoplasia. To this point, a large number of altered metabolites were detected in the feces of tumor-bearing mice. Ideally, a biomarker should reflect the burden of disease. We identified 43 fecal metabolites including heme that changed with disease progression (). The fact that levels of heme increased over time is consistent with enhanced gastrointestinal bleeding as tumor burden increased. In humans, the stool guaiac test detects heme and is used to non-invasively screen for colorectal neoplasia (6
). The current preclinical results suggest that a large number of metabolites in addition to heme change during colorectal carcinogenesis. If metabolic changes in addition to elevated heme levels can be detected in the feces of humans during colorectal carcinogenesis, it may be possible to develop improved metabolite-based non-invasive testing.
Plasma offers another potential matrix to identify biomarkers of colorectal carcinogenesis. Importantly, our study found a significant number of altered metabolites in the plasma of tumor-bearing mice, including many that changed in association with increased tumor burden (). A number of human studies have also utilized MS in an effort to identify blood-based signatures of CRC (15
). Consistent with prior reports in humans (15
), we found altered levels of urea, hippurate and 2-hydroxybutyrate in tumor-bearing mice (). We recognize that our ability to tightly control variables that can affect the metabolome including diet, age, gender and gut microbiota may result in different or additional changes than what has been found in humans (11
Metabolomic analysis can also provide new insights into the deregulation of molecular pathways in tumors. Of interest, serine (a product of phosphoglycerate dehydrogenase activity), sarcosine (an intermediate in glycine metabolism) and 2-HG (a product of glutamate/α-ketoglutarate metabolism) were increased in tumor tissue as compared with normal colorectal mucosa (). The observed expression changes in PHGDH, PSAT, PSPH and DMGDH, SARDH, GNMT within the tumor tissue can explain the increases in serine and sarcosine levels, respectively (Supplementary Table 1
). We also note that AOM-induced tumors contain β-catenin
mutations which may contribute to these metabolomic changes (31
The relevance of our findings is highlighted by several previous human findings. For example, increased serine levels have been reported in human colon cancer (13
). Moreover, a subset of human breast cancers appears to be dependent on the serine synthesis pathway (35
). Increased sarcosine levels have been observed in prostate cancer and may play a role in progression (20
). The finding of increased 2-HG levels is potentially significant given its association with leukemia and glioma in which mutant isocitrate dehydrogenase contributes to its increased production (28
). Future studies are warranted to elucidate the functional significance of the observed metabolic changes. Significantly, sarcosine was increased in both feces and tumor tissue, and 2-HG was increased in both plasma and tumor tissue, which may reflect tumor cell shedding. In this regard, changes found in feces and plasma occurred at an early stage of colorectal carcinogenesis ( and ), highlighting the potential utility of metabolomics for identifying biomarkers of early colorectal lesions. Based on the current findings, it will be of considerable interest to determine whether the serine-sarcosine or glutamate-2-HG pathways are altered in human colorectal tumors.
Future studies using this or related metabolomic approaches should be carried out in humans with the goal of both elucidating the metabolic changes that occur in colorectal neoplasia and identifying new biomarkers that have the potential to be useful for the early detection of colorectal neoplasia.