Identification of hypoxanthine as a urine marker for non-Hodgkin lymphoma by low-mass-ion profiling.
BACKGROUND
Non-Hodgkin lymphoma (NHL) is a hematologic malignancy for which good diagnostic markers are lacking. Despite continued improvement in our understanding of NHL, efforts to identify diagnostic markers have yielded dismal results. Here, we translated low-mass-ion information in urine samples from patients with NHL into a diagnostic marker.


METHODS
To minimize experimental error, we tested variable parameters before MALDI-TOF analysis of low-mass ions in urine. Urine from 30 controls and 30 NHL patients was analyzed as a training set for NHL prediction. All individual peak areas were normalized to total area up to 1000 m/z. The training set analysis was repeated four times. Low-mass peaks that were not affected by changes in experimental conditions were collected using MarkerView software. Human Metabolome Database (HMDB) searches and ESI LC-MS/MS analyses were used to identify low-mass ions that exhibited differential patterns in control and NHL urines. Identified low-mass ions were validated in a blinded fashion in 95 controls and 66 NHL urines to determine their ability to discriminate NHL patients from controls.


RESULTS
The 30 highest-ranking low-mass-ion peaks were selected from the 60-urine training set, and three low-mass-ion peaks with high intensity were selected for identification. Of these, a 137.08-m/z ion showed lower mass-peak intensity in urines of NHL patients, a result that was validated in a 161-urine blind validation set (95 controls and 66 NHL urines). The 130.08-m/z ion was identified from HMDB searches and ESI LC-MS/MS analyses as hypoxanthine (HX). The HX concentration in urines of NHL patients was significantly decreased (P < 0.001) and was correlated with the mass-peak area of the 137.08-m/z ion. At an HX concentration cutoff of 17.4 microM, sensitivity and specificity were 79.2% and 78.4%, respectively.


CONCLUSIONS
The present study represents a good example of low-mass-ion profiling in the setting of disease screening using urine. This technique can be a powerful non-invasive diagnostic tool with high sensitivity and specificity for NHL screening. Furthermore, HX identified in the study may be a useful single urine marker for NHL screening.
Yoo
Byong Chul
BC
Colorectal Cancer Branch, Division of Translational and Clinical Research I, Research Institute, National Cancer Center, Goyang-si, Republic of Korea.
Kong
Sun-Young
SY
Jang
Sang-Geun
SG
Kim
Kyung-Hee
KH
Ahn
Sun-A
SA
Park
Weon-Seo
WS
Park
Sohee
S
Yun
Tak
T
Eom
Hyeon-Seok
HS
D000368:Aged; D016022:Case-Control Studies; D002624:Chemistry, Clinical / Q000379:methods; D005260:Female; D006801:Humans; D019271:Hypoxanthine / Q000652:urine*; D008228:Lymphoma, Non-Hodgkin / Q000175:diagnosis* / Q000652:urine*; D008297:Male; D055432:Metabolomics / Q000379:methods; D008875:Middle Aged; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
D019271:Hypoxanthine