{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST002167","ANALYSIS_ID":"AN003550","VERSION":"1","CREATED_ON":"May 16, 2022, 7:14 am"},

"PROJECT":{"PROJECT_TITLE":"Remote solid cancers rewire hepatic nitrogen metabolism via host nicotinamide-N-methyltransferase","PROJECT_SUMMARY":"Cancers disrupt host homeostasis in various manners but the identity of host factors underlying such disruption remains largely unknown. Here we show that nicotinamide-N-methyltransferase (NNMT) is a novel host factor that mediates metabolic dysfunction in the livers of cancer-bearing mice. Multiple solid cancers distantly increase expression of Nnmt and its product 1-methylnicotinamide (MNAM) in the liver. Multi-omics analyses reveal suppression of the urea cycle accompanied by accumulation of amino acids, and enhancement of uracil biogenesis in the livers of cancer-bearing mice. Importantly, genetic deletion of Nnmt leads to alleviation of these metabolic abnormalities, and buffers cancer-dependent weight loss and reduction of the voluntary wheel-running activity. Our data also demonstrate that MNAM is capable of affecting urea cycle metabolites in the liver. These results suggest that cancers up-regulate the hepatic NNMT pathway to rewire liver metabolism towards uracil biogenesis rather than nitrogen disposal via the urea cycle, thereby disrupting host homeostasis.","INSTITUTE":"Tohoku University","LAST_NAME":"Kawaoka","FIRST_NAME":"Shinpei","ADDRESS":"4-1 Seiryo-cho, Sendai, Miyagi, 9808575, Japan","EMAIL":"kawaokashinpei@gmail.com","PHONE":"0227178568"},

"STUDY":{"STUDY_TITLE":"Remote solid cancers rewire hepatic nitrogen metabolism via host nicotinamide-N-methyltransferase (AML cells)","STUDY_SUMMARY":"Cancers disrupt host homeostasis in various manners but the identity of host factors underlying such disruption remains largely unknown. Here we show that nicotinamide-N-methyltransferase (NNMT) is a novel host factor that mediates metabolic dysfunction in the livers of cancer-bearing mice. Multiple solid cancers distantly increase expression of Nnmt and its product 1-methylnicotinamide (MNAM) in the liver. Multi-omics analyses reveal suppression of the urea cycle accompanied by accumulation of amino acids, and enhancement of uracil biogenesis in the livers of cancer-bearing mice. Importantly, genetic deletion of Nnmt leads to alleviation of these metabolic abnormalities, and buffers cancer-dependent weight loss and reduction of the voluntary wheel-running activity. Our data also demonstrate that MNAM is capable of affecting urea cycle metabolites in the liver. These results suggest that cancers up-regulate the hepatic NNMT pathway to rewire liver metabolism towards uracil biogenesis rather than nitrogen disposal via the urea cycle, thereby disrupting host homeostasis. Anionic polar metabolites (i.e., organic acids, sugar phosphates, nucleotides, etc.) were analyzed via IC/HR/MS/MS. Cationic polar metabolites (i.e., amino acids, bases, nucleosides, NAM, SAM, MNAM, SAH, me2PY, me4PY, etc) were analyzed via PFPP-LC/HR/MS/MS.","INSTITUTE":"Tohoku University","LAST_NAME":"Kawaoka","FIRST_NAME":"Shinpei","ADDRESS":"4-1 Seiryo-cho, Sendai, Miyagi, 9808575, Japan","EMAIL":"kawaokashinpei@gmail.com","PHONE":"0227178568"},

"SUBJECT":{"SUBJECT_TYPE":"Mammal","SUBJECT_SPECIES":"Mus musculus","TAXONOMY_ID":"10090"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"191015",
"Sample ID":"control_1",
"Factors":{"Culture conditions":"Control","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"1"}
},
{
"Subject ID":"191015",
"Sample ID":"control_2",
"Factors":{"Culture conditions":"Control","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"3"}
},
{
"Subject ID":"191015",
"Sample ID":"control_3",
"Factors":{"Culture conditions":"Control","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"5"}
},
{
"Subject ID":"191015",
"Sample ID":"control_4",
"Factors":{"Culture conditions":"Control","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"7"}
},
{
"Subject ID":"191015",
"Sample ID":"4T1_1",
"Factors":{"Culture conditions":"4T1-conditioned media","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"2"}
},
{
"Subject ID":"191015",
"Sample ID":"4T1_2",
"Factors":{"Culture conditions":"4T1-conditioned media","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"4"}
},
{
"Subject ID":"191015",
"Sample ID":"4T1_3",
"Factors":{"Culture conditions":"4T1-conditioned media","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"6"}
},
{
"Subject ID":"191015",
"Sample ID":"4T1_4",
"Factors":{"Culture conditions":"4T1-conditioned media","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"8"}
},
{
"Subject ID":"200401",
"Sample ID":"cont_1",
"Factors":{"Culture conditions":"Control","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"cell1"}
},
{
"Subject ID":"200401",
"Sample ID":"cont_2",
"Factors":{"Culture conditions":"Control","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"cell4"}
},
{
"Subject ID":"200401",
"Sample ID":"cont_3",
"Factors":{"Culture conditions":"Control","Treatment":"No treatment"},
"Additional sample data":{"RAW_FILE_NAME":"cell7"}
},
{
"Subject ID":"200401",
"Sample ID":"TNF20_1",
"Factors":{"Culture conditions":"Control","Treatment":"TNFalpha (20 ng/mL) treatment"},
"Additional sample data":{"RAW_FILE_NAME":"cell3"}
},
{
"Subject ID":"200401",
"Sample ID":"TNF20_2",
"Factors":{"Culture conditions":"Control","Treatment":"TNFalpha (20 ng/mL) treatment"},
"Additional sample data":{"RAW_FILE_NAME":"cell6"}
},
{
"Subject ID":"200401",
"Sample ID":"TNF20_3",
"Factors":{"Culture conditions":"Control","Treatment":"TNFalpha (20 ng/mL) treatment"},
"Additional sample data":{"RAW_FILE_NAME":"cell9"}
},
{
"Subject ID":"200401",
"Sample ID":"TNF200_1",
"Factors":{"Culture conditions":"Control","Treatment":"TNFalpha (200 ng/mL) treatment"},
"Additional sample data":{"RAW_FILE_NAME":"cell2"}
},
{
"Subject ID":"200401",
"Sample ID":"TNF200_2",
"Factors":{"Culture conditions":"Control","Treatment":"TNFalpha (200 ng/mL) treatment"},
"Additional sample data":{"RAW_FILE_NAME":"cell5"}
},
{
"Subject ID":"200401",
"Sample ID":"TNF200_3",
"Factors":{"Culture conditions":"Control","Treatment":"TNFalpha (200 ng/mL) treatment"},
"Additional sample data":{"RAW_FILE_NAME":"cell8"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"4T1 cells were cultured in 10 cm dishes for 48 hours and the culture supernatant was collected. The supernatant was stored as the 4T1-conditioned media at 4°C until use. AML cells per a well were cultured in a 6 well plate for 24 hours, and then the media was switched to the 4T1-conditioned media. After 24 hours, the treated AML12 cells were collected.","SAMPLE_TYPE":"AML cells"},

"TREATMENT":{"TREATMENT_SUMMARY":"AML cells per well were cultured in a 24 well plate for 24 hours. The media was then switched to the bovine-serum free media, and TNF alpha was added at the concentration of 20 ng/ml or 200 ng/ml (Roche). After 24 hours, the treated AML12 cells were collected."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"Metabolites were extracted from AML12 cells (less than 6 × 10E5 cells/well (6 well plate)) using the Bligh and Dyer’s method with some modifications. Briefly, each sample was mixed with 1 mL of cold methanol containing 10-camphorsulfonic acid (1.5 nmol) and piperazine-1,4-bis (2-ethanesulfonic acid) (PIPES, 1.5 nmol) as internal standards for mass spectrometry-based metabolomic analysis. The samples were vigorously mixed by vortexing for 1 min followed by 5 min of sonication. The extracts were then centrifuged at 16,000 × g for 5 min at 4 °C, and the resultant supernatant (400 uL) was collected. After mixing 400 uL of supernatant with 400 uL of chloroform and 320 uL of water, the aqueous and organic layers were separated by vortexing and subsequent centrifugation at 16,000 × g and 4 °C for 5 min. The aqueous (upper) layer (500 uL) was transferred into a clean tube. After the aqueous layer extracts were evaporated under vacuum, the dried extracts were stored at −80 °C until the analysis of hydrophilic metabolites. Prior to analysis, the dried aqueous layer was reconstituted in 50 uL of water."},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_SUMMARY":"Anionic polar metabolites (i.e., organic acids, sugar phosphates, nucleotides, etc.) were analyzed via IC/HRMS/MS.","CHROMATOGRAPHY_TYPE":"Other","INSTRUMENT_NAME":"Thermo Dionex ICS-5000+","COLUMN_NAME":"Dionex IonPac AS11-HC (2 um i.d. × 250 mm, 4 um particle size)"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS"},

"MS":{"INSTRUMENT_NAME":"Thermo Q Exactive Orbitrap","INSTRUMENT_TYPE":"Orbitrap","MS_TYPE":"ESI","ION_MODE":"NEGATIVE","MS_COMMENTS":"-"},

"MS_METABOLITE_DATA":{
"Units":"peak area",

"Data":[{"Metabolite":"10-Camphorsulfonic acid (IS)","cont_1":"5.E+08","cont_2":"4.E+08","cont_3":"4.E+08","TNF20_1":"5.E+08","TNF20_2":"4.E+08","TNF20_3":"3.E+08","TNF200_1":"5.E+08","TNF200_2":"4.E+08","TNF200_3":"4.E+08","control_1":"5.E+08","control_2":"5.E+08","control_3":"4.E+08","control_4":"6.E+08","4T1_1":"5.E+08","4T1_2":"6.E+08","4T1_3":"6.E+08","4T1_4":"4.E+08"},{"Metabolite":"PIPES (IS)","cont_1":"1.E+08","cont_2":"1.E+08","cont_3":"1.E+08","TNF20_1":"1.E+08","TNF20_2":"1.E+08","TNF20_3":"1.E+08","TNF200_1":"1.E+08","TNF200_2":"1.E+08","TNF200_3":"1.E+08","control_1":"8.E+07","control_2":"6.E+07","control_3":"7.E+07","control_4":"9.E+07","4T1_1":"8.E+07","4T1_2":"1.E+08","4T1_3":"1.E+08","4T1_4":"8.E+07"},{"Metabolite":"Fum","cont_1":"3.E+07","cont_2":"2.E+07","cont_3":"2.E+07","TNF20_1":"3.E+07","TNF20_2":"2.E+07","TNF20_3":"2.E+07","TNF200_1":"3.E+07","TNF200_2":"2.E+07","TNF200_3":"3.E+07","control_1":"4.E+07","control_2":"5.E+07","control_3":"3.E+07","control_4":"7.E+07","4T1_1":"2.E+07","4T1_2":"4.E+07","4T1_3":"4.E+07","4T1_4":"3.E+07"},{"Metabolite":"Uracil","cont_1":"0.E+00","cont_2":"0.E+00","cont_3":"0.E+00","TNF20_1":"0.E+00","TNF20_2":"0.E+00","TNF20_3":"0.E+00","TNF200_1":"0.E+00","TNF200_2":"0.E+00","TNF200_3":"0.E+00","control_1":"6.E+05","control_2":"6.E+05","control_3":"3.E+05","control_4":"8.E+05","4T1_1":"5.E+05","4T1_2":"2.E+06","4T1_3":"8.E+05","4T1_4":"7.E+05"}],

"Metabolites":[{"Metabolite":"10-Camphorsulfonic acid (IS)","cont_1":"5.E+08","cont_2":"4.E+08","cont_3":"4.E+08","TNF20_1":"5.E+08","TNF20_2":"4.E+08","TNF20_3":"3.E+08","TNF200_1":"5.E+08","TNF200_2":"4.E+08","TNF200_3":"4.E+08","control_1":"5.E+08","control_2":"5.E+08","control_3":"4.E+08","control_4":"6.E+08","4T1_1":"5.E+08","4T1_2":"6.E+08","4T1_3":"6.E+08","4T1_4":"4.E+08"},{"Metabolite":"PIPES (IS)","cont_1":"1.E+08","cont_2":"1.E+08","cont_3":"1.E+08","TNF20_1":"1.E+08","TNF20_2":"1.E+08","TNF20_3":"1.E+08","TNF200_1":"1.E+08","TNF200_2":"1.E+08","TNF200_3":"1.E+08","control_1":"8.E+07","control_2":"6.E+07","control_3":"7.E+07","control_4":"9.E+07","4T1_1":"8.E+07","4T1_2":"1.E+08","4T1_3":"1.E+08","4T1_4":"8.E+07"},{"Metabolite":"Fum","cont_1":"3.E+07","cont_2":"2.E+07","cont_3":"2.E+07","TNF20_1":"3.E+07","TNF20_2":"2.E+07","TNF20_3":"2.E+07","TNF200_1":"3.E+07","TNF200_2":"2.E+07","TNF200_3":"3.E+07","control_1":"4.E+07","control_2":"5.E+07","control_3":"3.E+07","control_4":"7.E+07","4T1_1":"2.E+07","4T1_2":"4.E+07","4T1_3":"4.E+07","4T1_4":"3.E+07"},{"Metabolite":"Uracil","cont_1":"0.E+00","cont_2":"0.E+00","cont_3":"0.E+00","TNF20_1":"0.E+00","TNF20_2":"0.E+00","TNF20_3":"0.E+00","TNF200_1":"0.E+00","TNF200_2":"0.E+00","TNF200_3":"0.E+00","control_1":"6.E+05","control_2":"6.E+05","control_3":"3.E+05","control_4":"8.E+05","4T1_1":"5.E+05","4T1_2":"2.E+06","4T1_3":"8.E+05","4T1_4":"7.E+05"}]
}

}