Multiplatform Urinary Metabolomics Profiling to Discriminate Cachectic from Non-Cachectic Colorectal Cancer Patients: Pilot Results from the ColoCare Study

Abstract

Cachexia is a multifactorial syndrome that is characterized by loss of skeletal muscle mass in cancer patients. The biological pathways involved remain poorly characterized. Here, we compare urinary metabolic profiles in newly diagnosed colorectal cancer patients (stage I-IV) from the ColoCare Study in Heidelberg, Germany. Patients were classified as cachectic (n = 16), pre-cachectic (n = 13), or non-cachectic (n = 23) based on standard criteria on weight loss over time at two time points. Urine samples were collected pre-surgery, and 6 and 12 months thereafter. Fat and muscle mass area were assessed utilizing computed tomography scans at the time of surgery. N = 152 compounds were detected using untargeted metabolomics with gas chromatography-mass spectrometry and n = 154 features with proton nuclear magnetic resonance spectroscopy. Thirty-four metabolites were overlapping across platforms. We calculated differences across groups and performed discriminant and overrepresentation enrichment analysis. We observed a trend for 32 compounds that were nominally significantly different across groups, although not statistically significant after adjustment for multiple testing. Nineteen compounds could be identified, including acetone, hydroquinone, and glycine. Comparing cachectic to non-cachectic patients, higher levels of metabolites such as acetone (Fold change (FC) = 3.17; p = 0.02) and arginine (FC = 0.33; p = 0.04) were observed. The two top pathways identified were glycerol phosphate shuttle metabolism and glycine and serine metabolism pathways. Larger subsequent studies are needed to replicate and validate these results.

Keywords: cancer cachexia; colorectal cancer; metabolomics; serial samples; urinary profiles.

Figure 1

Classification of patients into the three groups: Between baseline and 6 months, and between 6 months and 12 months.

Figure 2

Orthogonal projections to latent structures discriminant analysis. (A) Cachectic patients and non-cachectic patients. Urinary Metabolites only. (B) Cachectic and non-cachectic patients. Urinary metabolites and information on fat and muscle area from CT scans.

Figure 3

Orthogonal projections to latent structures discriminant analysis. (A) Pre-cachectic patients and non-cachectic patients. Urinary Metabolites only. (B) Pre-cachectic patients and non-cachectic patients. Urinary metabolites and information on fat and muscle area from CT scans.

Figure 4

Orthogonal projections to latent structures discriminant analysis. (A) Pre-cachectic patients and cachectic patients. Urinary Metabolites only. (B) Pre-cachectic patients and cachectic patients. Urinary metabolites and information on fat and muscle area from CT scans.

Figure 5

Summary plot for over-representation analyses (ORA). Overall urinary metabolites. For each of the listed pathways the respective metabolites identified are listed below: Glycine and serine metabolism: Glycine, L-arginine, L-methionine (3/53); glycerol phosphate shuttle: Hydroquinone (1/11); methionine metabolism: Glycine, L-methionine (2/43); ketone body metabolism: Acetone (1/13); arginine and proline metabolism: L-arginine (1/35); alanine metabolism: Glycine, L-arginine (2/53); spermidine and spermine biosynthesis: L-methionine (1/18); riboflavin metabolism: Hydroquinone (1/20); glutathione metabolism: Glycine (1/21); betaine metabolism: L-methionine (1/21); carnitine synthesis: Glycine (1/22); bile acid biosynthesis: Glycine, cholic acid (2/65); glycerolipid metabolism: Hydroquinone (1/25); tyrosine metabolism: p-Hydroxyphenylacetic acid, aminomalate (2/72); urea cycle: L-arginine (1/29); starch and sucrose metabolism: Sucrose (1/31); ammonia recycling: Glycine (1/32); beta-alanine metabolism: Uracil (1/34); aspartate metabolism: L-arginine (1/35); retinol metabolism: Glucoronide (1/37); galactose metabolism: Sucrose (1/38); porphyrin metabolism: Glycine (1/40); glutamate metabolism: Glycine (1/50); pyrimidine metabolism: Uracil (1/59); purine metabolism: Glycine (1/74).

Figure 6

Summary plot for over-representation analyses (ORA): Urinary metabolites that were different between cachectic and non-cachectic patients. For each of the listed pathways, the respective metabolites identified are listed below: Ketone body metabolism: Acetone (1/13); urea cycle: Arginine (1/29); aspartate metabolism: L-arginine (1/35); arginine and proline metabolism: L-arginine (1/35); glycine and serine metabolism: L-arginine (1/59); tyrosine metabolism: p-Hydroxyphenylacetic acid (1/72).

Figure 7

Summary plot for over-representation analyses (ORA). Metabolites that were different between pre-cachectic and non-cachectic patients. For each of the listed pathways, the respective metabolites identified are listed below: Glycine and serine metabolism: L-arginine and L-methionine (2/59); glycerol phosphate shuttle: Hydroquinone (1/11); ketone body metabolism: Acetone (1/13); spermidine and spermine biosynthesis: L-methionine (1/18); riboflavin metabolism: Hydroquinone (1/20); betaine metabolism: L-methionine (1/21); glycerolipid metabolism: Hydroquinone (1/25); urea cycle: L-arginine (1/29); beta-alanine metabolism: Uracil (1/34); aspartate metabolism: L-arginine (1/35); methionine metabolism: L-methionine (1/43); arginine and proline metabolism: L-arginine (1/53); pyrimidine metabolism: Uracil (1/59); bile acid biosynthesis: Cholic acid (1/65).

Figure 8

Summary plot for over-representation analyses (ORA). Metabolites that were different between pre-cachectic and cachectic patients. For each of the listed pathways, the respective metabolites identified are listed below: Glycerol phosphate shuttle: Hydroquinone (1/11); alanine metabolism: Glycine (1/17); riboflavin metabolism: Hydroquinone (1/20); glutathione metabolism: Glycine (1/21); carnitine synthesis: Glycine (1/22); glycerolipid metabolism: Hydroquinone (1/25); ammonia recycling: Glycine (1/32); porphyrin metabolism: Glycine (1/40); methionine metabolism: Glycine (1/43); glutamate metabolism: Glycine (1/49); arginine and proline metabolism: Glycine (1/53); glycine and serine metabolism: Glycine (1/59); bile acid biosynthesis: Glycine (1/65); tyrosine metabolism: Aminomalonate (1/72); purine metabolism: Glycine (1/74).