ʻO ka ʻike ʻokoʻa o nā nodules pulmonary i ʻike ʻia e ka computed tomography (CT) he mea paʻakikī ia i ka hana lapaʻau. Maanei, wehewehe mākou i ka metabolome honua o 480 mau laʻana serum, me nā kaohi olakino, nā nodules māmā benign, a me ka adenocarcinoma māmā stage I. Hōʻike nā Adenocarcinomas i nā ʻano metabolomic kū hoʻokahi, ʻoiai ʻo nā nodules benign a me nā kānaka olakino he like kiʻekiʻe i nā ʻano metabolomic. I ka hui ʻike (n = 306), ua ʻike ʻia kahi hui o 27 metabolites e hoʻokaʻawale ma waena o nā nodules benign a me nā malignant. ʻO ka AUC o ke kumu hoʻohālikelike i loko o ka hōʻoia kūloko (n = 104) a me ka hōʻoia waho (n = 111) mau hui he 0.915 a me 0.945, kēlā me kēia. Ua hōʻike ka loiloi ala i ka hoʻonui ʻia ʻana o nā metabolites glycolytic e pili ana me ka emi ʻana o ka tryptophan i loko o ka serum adenocarcinoma māmā i hoʻohālikelike ʻia me nā nodules benign a me nā kaohi olakino, a ua hōʻike ʻia e hoʻolaha ka uptake tryptophan i ka glycolysis i loko o nā cell cancer lung. Hōʻike kā mākou haʻawina i ka waiwai o nā biomarkers metabolite serum i ka loiloi ʻana i ka pilikia o nā nodules pulmonary i ʻike ʻia e CT.
He mea koʻikoʻi ka ʻike mua ʻana i ka maʻi kanesa no ka hoʻomaikaʻi ʻana i nā helu ola no nā poʻe maʻi kanesa. Ua hōʻike ʻia nā hopena mai ka US National Lung Cancer Screening Trial (NLST) a me ka European NELSON Study e hiki i ka nānā ʻana me ka tomography computed haʻahaʻa (LDCT) ke hōʻemi nui i ka make ʻana o ka maʻi kanesa māmā i nā hui pilikia kiʻekiʻe1,2,3. Mai ka hoʻohana nui ʻia ʻana o LDCT no ka nānā ʻana i ka maʻi kanesa māmā, ua hoʻomau ka piʻi ʻana o ka nui o nā ʻike radiographic incidental o nā nodules pulmonary asymptomatic 4. Ua wehewehe ʻia nā nodules pulmonary he mau opacities focal a hiki i ka 3 cm ke anawaena 5. Ke kū nei mākou i nā pilikia i ka loiloi ʻana i ka likelika o ka malignancy a me ka hana ʻana me ka nui o nā nodules pulmonary i ʻike ʻia ma ka LDCT. Hiki i nā palena o CT ke alakaʻi i nā hoʻokolohua hahai pinepine a me nā hopena wahaheʻe-positive, e alakaʻi ana i ke komo ʻana a me ka overtreatment pono ʻole6. No laila, aia kahi pono e hoʻomohala i nā biomarkers hilinaʻi a pono e ʻike pololei i ka maʻi kanesa māmā i nā pae mua a hoʻokaʻawale i ka hapa nui o nā nodules benign i ka ʻike mua ʻana 7.
ʻO ka loiloi molekala piha o ke koko (serum, plasma, peripheral blood mononuclear cells), me ka genomics, proteomics a i ʻole DNA methylation8,9,10, ua alakaʻi i ka hoihoi nui i ka ʻike ʻana i nā biomarkers diagnostic no ka maʻi kanesa māmā. I kēia manawa, ke ana nei nā ʻano metabolomics i nā huahana hopena cellular i hoʻopilikia ʻia e nā hana endogenous a me exogenous a no laila ua hoʻopili ʻia e wānana i ka hoʻomaka ʻana o ka maʻi a me ka hopena. ʻO ka Liquid chromatography-tandem mass spectrometry (LC-MS) kahi ʻano i hoʻohana nui ʻia no nā haʻawina metabolomics ma muli o kona ʻike kiʻekiʻe a me ka laulā dynamic nui, hiki ke uhi i nā metabolites me nā waiwai physicochemical like ʻole11,12,13. ʻOiai ua hoʻohana ʻia ka loiloi metabolomic honua o ka plasma/serum e ʻike i nā biomarkers e pili ana i ka maʻi kanesa māmā14,15,16,17 a me ka pono o ka mālama ʻana,18 serum metabolite classifiers e hoʻokaʻawale ma waena o nā nodules māmā benign a malignant e mau ana ke aʻo nui ʻia. -noiʻi nui.
ʻO Adenocarcinoma a me ka squamous cell carcinoma nā ʻano nui ʻelua o ka maʻi kanesa māmā ʻaʻole liʻiliʻi (NSCLC). Hōʻike nā hoʻokolohua CT screening like ʻole ʻo adenocarcinoma ke ʻano histological maʻamau o ka maʻi kanesa māmā1,19,20,21. Ma kēia haʻawina, ua hoʻohana mākou i ka ultra-performance liquid chromatography-high-resolution mass spectrometry (UPLC-HRMS) e hana i ka nānā ʻana o ka metabolomics ma ka huina o 695 mau laʻana serum, me nā kaohi olakino, nā nodules pulmonary benign, a me CT-i ʻike ʻia he ≤3 cm. Nānā no ka Stage I lung adenocarcinoma. Ua ʻike mākou i kahi panel o nā metabolites serum e hoʻokaʻawale i ka adenocarcinoma māmā mai nā nodules benign a me nā kaohi olakino. Ua hōʻike ka loiloi hoʻonui ala he mau hoʻololi maʻamau ka tryptophan abnormal a me ka metabolism glucose i ka adenocarcinoma māmā i hoʻohālikelike ʻia me nā nodules benign a me nā kaohi olakino. ʻO ka hope loa, ua hoʻokumu a hōʻoia mākou i kahi mea hoʻokaʻawale metabolic serum me ke kikoʻī kiʻekiʻe a me ka ʻike e hoʻokaʻawale ma waena o nā nodules pulmonary malignant a me benign i ʻike ʻia e LDCT, hiki ke kōkua i ka ʻike ʻokoʻa mua a me ka loiloi pilikia.
Ma ke aʻo ʻana o kēia manawa, ua hōʻiliʻili ʻia nā laʻana serum i kūlike i ke kāne a me ka makahiki mai 174 mau kaohi olakino, 292 mau maʻi me nā nodules pulmonary benign, a me 229 mau maʻi me ka adenocarcinoma māmā pae I. Hōʻike ʻia nā ʻano demographic o nā kumuhana 695 ma ka Papa Hoʻohui 1.
E like me ka mea i hōʻike ʻia ma ke Kiʻi 1a, ua hōʻiliʻili ʻia he 480 mau laʻana serum, me 174 mau mana olakino (HC), 170 mau nodules benign (BN), a me 136 mau laʻana adenocarcinoma māmā (LA) pae I, ma ke Kikowaena Kanesa o ke Kulanui ʻo Sun Yat-sen. ʻO ka hui ʻike no ka profiling metabolomic i hoʻopaʻa ʻole ʻia me ka hoʻohana ʻana i ka ultra-performance liquid chromatography-high-resolution mass spectrometry (UPLC-HRMS). E like me ka mea i hōʻike ʻia ma ke Kiʻi Hoʻohui 1, ua ʻike ʻia nā metabolites ʻokoʻa ma waena o LA a me HC, LA a me BN e hoʻokumu i kahi kumu hoʻohālike hoʻokaʻawale a ʻimi hou aku i ka nānā ʻana i ke ala ʻokoʻa. Ua hoʻokau ʻia nā laʻana 104 i hōʻiliʻili ʻia e ke Kikowaena Kanesa o ke Kulanui ʻo Sun Yat-sen a me 111 mau laʻana i hōʻiliʻili ʻia e ʻelua mau haukapila ʻē aʻe i ka hōʻoia kūloko a me waho.
a He heluna kanaka noiʻi i loko o ka hui ʻike i hana ʻia i ka loiloi metabolomics serum honua me ka hoʻohana ʻana i ka ultra-performance liquid chromatography-high-resolution mass spectrometry (UPLC-HRMS). b ʻO ka loiloi discriminant hapa liʻiliʻi loa (PLS-DA) o ka metabolome holoʻokoʻa o 480 mau laʻana serum mai ka hui noiʻi, me nā mana olakino (HC, n = 174), nā nodules benign (BN, n = 170), a me ke kahua I lung adenocarcinoma (Los Angeles, n = 136). +ESI, ʻano ionization electrospray maikaʻi, -ESI, ʻano ionization electrospray maikaʻi ʻole. c–e Hōʻike ʻia nā Metabolites me nā nui like ʻole i ʻelua mau hui i hāʻawi ʻia (hoʻāʻo kūlana Wilcoxon ʻelua-tailed, ka waiwai p i hoʻoponopono ʻia i ka helu ʻike wahaheʻe, FDR <0.05) i ka ʻulaʻula (hoʻololi pelu > 1.2) a me ka uliuli (hoʻololi pelu < 0.83). ) i hōʻike ʻia ma ke kiʻi lua pele. ʻO ka palapala wela o ka hui hierarchical e hōʻike ana i nā ʻokoʻa koʻikoʻi i ka helu o nā metabolites i hōʻike ʻia ma waena o LA a me BN. Hāʻawi ʻia ka ʻikepili kumu ma ke ʻano o nā faila ʻikepili kumu.
Ua kālailai ʻia ka metabolome serum holoʻokoʻa o 174 HC, 170 BN a me 136 LA i loko o ka hui ʻike me ka hoʻohana ʻana i ka loiloi UPLC-HRMS. Hōʻike mua mākou e hui pū ana nā laʻana o ka mana hoʻomalu maikaʻi (QC) ma ke kikowaena o kahi kumu hoʻohālike loiloi ʻāpana nui (PCA) i mālama ʻole ʻia, e hōʻoia ana i ke kūpaʻa o ka hana o kēia noiʻi (Kiʻi Hoʻohui 2).
E like me ka mea i hōʻike ʻia ma ka ʻāpana liʻiliʻi loa o nā squares-discriminant analysis (PLS-DA) ma ke Kiʻi 1 b, ua ʻike mākou he mau ʻokoʻa maopopo ma waena o LA a me BN, LA a me HC ma nā ʻano ionization electrospray maikaʻi (+ESI) a me nā ʻano maikaʻi ʻole (−ESI). Eia naʻe, ʻaʻohe ʻokoʻa koʻikoʻi i loaʻa ma waena o BN a me HC ma nā kūlana +ESI a me -ESI.
Ua loaʻa iā mākou he 382 mau hiʻohiʻona ʻokoʻa ma waena o LA a me HC, 231 mau hiʻohiʻona ʻokoʻa ma waena o LA a me BN, a me 95 mau hiʻohiʻona ʻokoʻa ma waena o BN a me HC (hoʻāʻo kūlana inoa ʻo Wilcoxon, FDR <0.05 a me ka loli he nui >1.2 a i ʻole <0.83) (Kiʻi .1c-e). . Ua hoʻopaʻa hou ʻia nā piko (ʻIkepili Hoʻohui 3) e kūʻē i kahi waihona ʻikepili (waihona waihona puke mzCloud/HMDB/Chemspider) e ka waiwai m/z, ka manawa paʻa a me ka ʻimi spectrum mass fragmentation (nā kikoʻī i wehewehe ʻia ma ka ʻāpana Methods) 22 . ʻO ka hope loa, ua ʻike ʻia he 33 a me 38 mau metabolites i hoʻopaʻa ʻia me nā ʻokoʻa koʻikoʻi i ka nui no LA versus BN (Kiʻi 1f a me ka Papa Hoʻohui 2) a me LA versus HC (Kiʻi Hoʻohui 3 a me ka Papa Hoʻohui 2), kēlā me kēia. I ka hoʻohālikelike ʻana, ʻo 3 wale nō metabolites me nā ʻokoʻa koʻikoʻi i ka nui i ʻike ʻia ma BN a me HC (Papa Hoʻohui 2), e kūlike ana me ka overlap ma waena o BN a me HC ma PLS-DA. Uhi kēia mau metabolites ʻokoʻa i kahi ākea o nā biochemicals (Kiʻi Hoʻohui 4). I ka hui pū ʻia, hōʻike kēia mau hopena i nā loli koʻikoʻi i ka serum metabolome e hōʻike ana i ka hoʻololi ʻino o ka maʻi kanesa māmā mua i hoʻohālikelike ʻia me nā nodules māmā benign a i ʻole nā mea olakino. I kēia manawa, ʻo ka like o ka serum metabolome o BN a me HC e hōʻike ana e hiki i nā nodules pulmonary benign ke kaʻana like i nā ʻano olaola he nui me nā poʻe olakino. No ka mea he mea maʻamau nā mutations gene epidermal growth factor receptor (EGFR) i ka adenocarcinoma māmā subtype 23, ua ʻimi mākou e hoʻoholo i ka hopena o nā mutations driver ma ka serum metabolome. A laila ua kālailai mākou i ka ʻikepili metabolomic holoʻokoʻa o 72 mau hihia me ke kūlana EGFR i ka hui adenocarcinoma māmā. ʻO ka mea hoihoi, ua loaʻa iā mākou nā ʻikepili hoʻohālikelike ma waena o nā maʻi mutant EGFR (n = 41) a me nā maʻi EGFR wild-type (n = 31) i ka loiloi PCA (Kiʻi Hoʻohui 5a). Eia nō naʻe, ua ʻike mākou i 7 mau metabolites nona ka nui i hoʻololi nui ʻia i nā poʻe maʻi me ka hoʻololi ʻana o ka EGFR i hoʻohālikelike ʻia me nā poʻe maʻi me ka EGFR wild-type (t test, p < 0.05 a me ka hoʻololi ʻana o ka fold > 1.2 a i ʻole < 0.83) (Kiʻi Hoʻohui 5b). ʻO ka hapa nui o kēia mau metabolites (5 mai loko mai o 7) he acylcarnitines, nona ke kuleana koʻikoʻi i nā ala oxidation fatty acid.
E like me ka mea i hōʻike ʻia ma ke kaʻina hana i hōʻike ʻia ma ke Kiʻi 2 a, ua loaʻa nā biomarkers no ka hoʻokaʻawale ʻana o ka nodule me ka hoʻohana ʻana i nā mea hoʻokele shrinkage liʻiliʻi loa a me ke koho ʻana ma muli o 33 mau metabolites ʻokoʻa i ʻike ʻia ma LA (n = 136) a me BN (n = 170). ʻO ka hui maikaʻi loa o nā loli (LASSO) - ke kumu hoʻohālike regression logistic binary. Ua hoʻohana ʻia ka cross-validation he ʻumi manawa e hoʻāʻo ai i ka hilinaʻi o ke kumu hoʻohālike. Hoʻoponopono ʻia ke koho loli a me ka regularization parameter e kahi hoʻopaʻi maximization likelihood me ka parameter λ24. Ua hana hou ʻia ka loiloi metabolomics honua ma ke kūʻokoʻa i loko o nā hui hōʻoia kūloko (n = 104) a me ka hōʻoia waho (n = 111) e hoʻāʻo i ka hana hoʻokaʻawale o ke kumu hoʻohālike discriminant. ʻO ka hopena, ua ʻike ʻia he 27 mau metabolites i loko o ka set ʻike ʻana ʻo ia ke kumu hoʻohālike discriminant maikaʻi loa me ka waiwai AUC awelika nui loa (Kiʻi 2b), ma waena o 9 i hoʻonui ʻia ka hana a me 18 i emi iho ka hana ma LA i hoʻohālikelike ʻia me BN (Kiʻi 2c).
Kaʻina hana no ke kūkulu ʻana i kahi mea hoʻokaʻawale nodule pulmonary, me ke koho ʻana i ka panela maikaʻi loa o nā metabolites serum i loko o ka set ʻike me ka hoʻohana ʻana i kahi kumu hoʻohālike regression logistic binary ma o ka ʻumi-fold cross-validation a me ka loiloi ʻana i ka hana wānana i nā set hōʻoia kūloko a me waho. b Nā helu helu hōʻoia cross o ke kumu hoʻohālike regression LASSO no ke koho biomarker metabolic. Hōʻike nā helu i hāʻawi ʻia ma luna nei i ka helu awelika o nā biomarkers i koho ʻia ma kahi λ i hāʻawi ʻia. Hōʻike ka laina kiko ʻulaʻula i ka waiwai AUC awelika ma ka lambda pili. Hōʻike nā kaha hewa hina i nā waiwai AUC liʻiliʻi a me ka nui loa. Hōʻike ka laina kiko i ke kumu hoʻohālike maikaʻi loa me nā biomarkers 27 i koho ʻia. AUC, wahi ma lalo o ke kaʻina hana hana mea hoʻokipa (ROC). c Nā loli pelu o 27 metabolites i koho ʻia i loko o ka hui LA i hoʻohālikelike ʻia me ka hui BN i loko o ka hui ʻike. Koluma ʻulaʻula - hoʻāla. He emi ka kolamu polū. d–f Nā kaʻina hana hana mea hoʻokipa (ROC) e hōʻike ana i ka mana o ke kumu hoʻohālike discriminant e pili ana i 27 mau hui metabolite i loko o nā set hōʻoia ʻike, kūloko, a me waho. Hāʻawi ʻia ka ʻikepili kumu ma ke ʻano o nā faila ʻikepili kumu.
Ua hana ʻia kekahi kumu hoʻohālike wānana ma muli o nā coefficients regression kaumaha o kēia mau metabolites 27 (Papa Hoʻohui 3). ʻO ka loiloi ROC ma muli o kēia mau metabolites 27 i loaʻa kahi waiwai ma lalo o ke piʻo (AUC) o 0.933, ʻo ka ʻike o ka hui ʻike he 0.868, a ʻo ka kikoʻī he 0.859 (Kiʻi 2d). I kēia manawa, ma waena o nā metabolites ʻokoʻa 38 i hōʻike ʻia ma waena o LA a me HC, ua loaʻa i kahi hui o 16 mau metabolites kahi AUC o 0.902 me ka ʻike o 0.801 a me ka kikoʻī o 0.856 i ka hoʻokaʻawale ʻana iā LA mai HC (Kiʻi Hoʻohui 6a-c). Ua hoʻohālikelike ʻia nā waiwai AUC ma muli o nā paepae loli pelu like ʻole no nā metabolites ʻokoʻa. Ua ʻike mākou ua hana maikaʻi loa ke kumu hoʻohālike hoʻokaʻawale i ka hoʻokaʻawale ʻana ma waena o LA a me BN (HC) i ka wā i hoʻonohonoho ʻia ai ka pae loli pelu i 1.2 e kūʻē i 1.5 a i ʻole 2.0 (Kiʻi Hoʻohui 7a,b). Ua hōʻoia hou ʻia ke kumu hoʻohālikelike, ma muli o 27 mau hui metabolite, i loko o nā cohorts kūloko a me waho. ʻO ka AUC he 0.915 (ʻike 0.867, kikoʻī 0.811) no ka hōʻoia kūloko a me 0.945 (ʻike 0.810, kikoʻī 0.979) no ka hōʻoia kūwaho (Kiʻi 2e, f). No ka loiloi ʻana i ka pono interlaboratory, ua kālailai ʻia he 40 mau laʻana mai ka cohort kūwaho i loko o kahi keʻena hoʻokolohua kūwaho e like me ka mea i wehewehe ʻia ma ka ʻāpana Methods. Ua loaʻa i ka pololei o ka hoʻohālikelike kahi AUC o 0.925 (Kiʻi Hoʻohui 8). No ka mea, ʻo ka lung squamous cell carcinoma (LUSC) ka lua o ka subtype maʻamau o ka maʻi kanesa māmā non-small cell (NSCLC) ma hope o ka lung adenocarcinoma (LUAD), ua hoʻāʻo pū mākou i ka pono kūpono o nā ʻikepili metabolic. BN a me 16 mau hihia o LUSC. ʻO ka AUC o ka hoʻokae ma waena o LUSC a me BN he 0.776 (Kiʻi Hoʻohui 9), e hōʻike ana i ka hiki ʻoi aku ka maikaʻi ʻole i hoʻohālikelike ʻia me ka hoʻokae ma waena o LUAD a me BN.
Ua hōʻike ʻia nā haʻawina e pili pono ana ka nui o nā nodules ma nā kiʻi CT me ka likelika o ka malignancy a ke noho nei he mea nui ia o ka mālama ʻana i ka nodule25,26,27. Ua hōʻike ʻia ka loiloi ʻana o ka ʻikepili mai ka cohort nui o ka haʻawina nānā NELSON ua like ka pilikia o ka malignancy i nā kumuhana me nā nodes <5 mm me nā kumuhana me ka ʻole o nā nodes 28. No laila, ʻo ka nui liʻiliʻi e pono ai ka nānā ʻana i ka CT maʻamau he 5 mm, e like me ka ʻōlelo a ka British Thoracic Society (BTS), a me 6 mm, e like me ka ʻōlelo a ka Fleischner Society 29. Eia nō naʻe, ʻo nā nodules i ʻoi aku ka nui ma mua o 6 mm a me nā hiʻohiʻona benign maopopo ʻole, i kapa ʻia ʻo indeterminate pulmonary nodules (IPN), he pilikia nui ia i ka loiloi a me ka hoʻokele ʻana i ka hana lapaʻau30,31. Ua nānā hou mākou inā ua hoʻopili ka nui o ka nodule i nā pūlima metabolomic me ka hoʻohana ʻana i nā laʻana i hui pū ʻia mai nā cohorts ʻike a me ka hōʻoia kūloko. Ke kālele nei i 27 mau biomarkers i hōʻoia ʻia, ua hoʻohālikelike mua mākou i nā ʻaoʻao PCA o HC a me BN sub-6 mm metabolomes. Ua ʻike mākou ua pili ka hapa nui o nā kiko ʻikepili no HC a me BN, e hōʻike ana ua like nā pae metabolite serum ma nā hui ʻelua (Kiʻi 3a). Ua mālama ʻia nā palapala ʻāina hiʻona ma nā ʻano nui like ʻole ma BN a me LA (Kiʻi 3b, c), ʻoiai ua ʻike ʻia kahi kaʻawale ma waena o nā nodules malignant a me benign ma ka pae 6-20 mm (Kiʻi 3d). Loaʻa i kēia cohort kahi AUC o 0.927, specificity o 0.868, a me ka sensitivity o 0.820 no ka wānana ʻana i ka malignancy o nā nodules e ana ana i 6 a 20 mm (Kiʻi 3e, f). Hōʻike kā mākou mau hopena hiki i ka classifier ke hopu i nā loli metabolic i hoʻokumu ʻia e ka hoʻololi malignant mua, me ka nānā ʻole i ka nui o ka nodule.
ad Hoʻohālikelike ʻia nā ʻikepili PCA ma waena o nā hui i kuhikuhi ʻia e pili ana i kahi mea hoʻokaʻawale metabolic o 27 metabolites. CC a me BN < 6 mm. b BN < 6 mm vs BN 6–20 mm. ma LA 6–20 mm e kūʻē iā LA 20–30 mm. g BN 6–20 mm a me LA 6–20 mm. GC, n = 174; BN < 6 mm, n = 153; BN 6–20 mm, n = 91; LA 6–20 mm, n = 89; LA 20–30 mm, n = 77. e Ke kiʻikuhi ʻano hana hoʻokipa (ROC) e hōʻike ana i ka hana hoʻohālike discriminant no nā nodules 6–20 mm. f Ua helu ʻia nā waiwai kūpono ma muli o ke kumu hoʻohālike regression logistic no nā nodules e ana ana i 6–20 mm. Hōʻike ka laina kiko hina i ka waiwai cutoff kūpono (0.455). Hōʻike nā helu ma luna i ka pakeneka o nā hihia i manaʻo ʻia no Los Angeles. E hoʻohana i kahi hoʻāʻo t a ka Haumāna ʻelua-tailed. PCA, ka loiloi ʻāpana nui. ʻĀpana AUC ma lalo o ke kūlou. Hāʻawi ʻia ka ʻikepili kumu ma ke ʻano o nā faila ʻikepili kumu.
Ua koho hou ʻia ʻehā mau laʻana (44-61 mau makahiki) me nā nui nodule pulmonary like (7-9 mm) e hōʻike i ka hana o ke kumu hoʻohālike wānana malignancy i manaʻo ʻia (Kiʻi 4a, b). I ka nānā mua ʻana, ua hōʻike ʻia ka Hihia 1 ma ke ʻano he nodule paʻa me ka calcification, kahi hiʻohiʻona e pili ana i ka benignity, ʻoiai ʻo ka Hihia 2 i hōʻike ʻia ma ke ʻano he nodule paʻa hapa ʻole me nā hiʻohiʻona benign maopopo ʻole. Ua hōʻike ʻekolu mau puni o nā scan CT hahai i noho paʻa kēia mau hihia ma luna o kahi manawa 4 mau makahiki a no laila ua manaʻo ʻia he mau nodules benign (Kiʻi 4a). Ke hoʻohālikelike ʻia me ka loiloi lapaʻau o nā scan CT serial, ua ʻike koke a pololei ka loiloi metabolite serum hoʻokahi me ke kumu hoʻohālike classifier o kēia manawa i kēia mau nodules benign e pili ana i nā palena probabilistic (Papa 1). Hōʻike ka Kiʻi 4b ma ka hihia 3 i kahi nodule me nā hōʻailona o ka retraction pleural, ka mea i pili pinepine ʻia me ka malignancy32. Ua hōʻike ʻia ka Hihia 4 ma ke ʻano he nodule paʻa hapa ʻole me ka ʻike ʻole o kahi kumu benign. Ua wānana ʻia kēia mau hihia a pau he ʻino e like me ke kumu hoʻohālike classifier (Papa 1). Ua hōʻike ʻia ka loiloi o ka adenocarcinoma māmā e ka hoʻokolokolo histopathological ma hope o ke ʻoki ʻana o ka māmā (Kiʻi 4b). No ka hoʻonohonoho hōʻoia waho, ua wānana pololei ka metabolic classifier i ʻelua mau hihia o nā nodules māmā indeterminate i ʻoi aku ka nui ma mua o 6 mm (Kiʻi Hoʻohui 10).
Nā kiʻi CT o ka puka makani axial o nā māmā o ʻelua mau hihia o nā nodules benign. I ka hihia 1, ua hōʻike ka CT scan ma hope o 4 mau makahiki i kahi nodule paʻa paʻa e ana ana i 7 mm me ka calcification ma ka lobe ʻākau haʻahaʻa. I ka hihia 2, ua hōʻike ka CT scan ma hope o 5 mau makahiki i kahi nodule paʻa, paʻa hapa me ke anawaena o 7 mm ma ka lobe ʻākau luna. b Nā kiʻi CT puka makani Axial o nā māmā a me nā haʻawina pathological e pili ana i ʻelua mau hihia o ka adenocarcinoma kahua I ma mua o ka ʻoki ʻana o ka māmā. Ua hōʻike ka hihia 3 i kahi nodule me ke anawaena o 8 mm ma ka lobe ʻākau luna me ka retraction pleural. Ua hōʻike ka hihia 4 i kahi nodule aniani honua paʻa hapa e ana ana i 9 mm ma ka lobe hema luna. ʻO ka hoʻoluʻu ʻana o Hematoxylin a me eosin (H&E) o ka ʻiʻo māmā i ʻoki ʻia (scale bar = 50 μm) e hōʻike ana i ke ʻano ulu acinar o ka adenocarcinoma māmā. Hōʻike nā pua i nā nodules i ʻike ʻia ma nā kiʻi CT. ʻO nā kiʻi H&E he mau kiʻi hōʻike o nā kahua microscopic he nui (>3) i nānā ʻia e ka pathologist.
I ka hui pū ʻia ʻana, hōʻike kā mākou mau hopena i ka waiwai kūpono o nā biomarkers metabolite serum i ka ʻike ʻokoʻa o nā nodules pulmonary, kahi mea e hoʻokū ai i nā pilikia i ka loiloi ʻana i ka nānā ʻana o CT.
Ma muli o kahi panela metabolite ʻokoʻa i hōʻoia ʻia, ua ʻimi mākou e ʻike i nā pilina olaola o nā loli metabolic nui. Ua ʻike ʻia ka loiloi hoʻonui ala KEGG e MetaboAnalyst i 6 mau ala i hoʻololi nui ʻia ma waena o nā hui ʻelua i hāʻawi ʻia (LA vs. HC a me LA vs. BN, i hoʻoponopono ʻia p ≤ 0.001, hopena > 0.01). Ua hōʻike ʻia kēia mau hoʻololi e nā haunaele i ka metabolism pyruvate, ka metabolism tryptophan, ka metabolism niacin a me ka nicotinamide, glycolysis, ka pōʻaiapuni TCA, a me ka metabolism purine (Kiʻi 5a). A laila ua hana hou mākou i nā metabolomics i kuhikuhi ʻia e hōʻoia i nā hoʻololi nui me ka hoʻohana ʻana i ka quantification absolute. Hoʻoholo ʻana i nā metabolites maʻamau i nā ala i hoʻololi pinepine ʻia e ka triple quadrupole mass spectrometry (QQQ) me ka hoʻohana ʻana i nā kūlana metabolite maoli. Ua hoʻokomo ʻia nā ʻano demographic o ka hāpana pahuhopu o ke aʻo ʻana i ka metabolomics i ka Papa Hoʻohui 4. E kūlike me kā mākou mau hopena metabolomics honua, ua hōʻoia ka loiloi quantitative ua hoʻonui ʻia ka hypoxanthine a me xanthine, pyruvate, a me lactate ma LA i hoʻohālikelike ʻia me BN a me HC (Kiʻi 5b, c, p <0.05). Eia naʻe, ʻaʻohe ʻokoʻa koʻikoʻi i loaʻa i kēia mau metabolites ma waena o BN a me HC.
ʻO ka loiloi hoʻonui alahele KEGG o nā metabolites ʻokoʻa koʻikoʻi i ka hui LA i hoʻohālikelike ʻia me nā hui BN a me HC. Ua hoʻohana ʻia kahi Globaltest ʻelua-tailed, a ua hoʻoponopono ʻia nā waiwai p me ka hoʻohana ʻana i ke ʻano Holm-Bonferroni (i hoʻoponopono ʻia ʻo p ≤ 0.001 a me ka nui o ka hopena > 0.01). b – d Nā kiʻi Violin e hōʻike ana i nā pae hypoxanthine, xanthine, lactate, pyruvate, a me tryptophan i loko o ka serum HC, BN, a me LA i hoʻoholo ʻia e LC-MS/MS (n = 70 no kēlā me kēia hui). Hōʻike nā laina kiko keʻokeʻo a me ka ʻeleʻele i ka median a me ka quartile, kēlā me kēia. e ʻO ka kiʻi Violin e hōʻike ana i ka hōʻike mRNA Log2TPM (transcripts no miliona) i hoʻonohonoho ʻia o SLC7A5 a me QPRT i ka adenocarcinoma māmā (n = 513) i hoʻohālikelike ʻia me ka ʻiʻo māmā maʻamau (n = 59) ma ka ʻikepili LUAD-TCGA. Hōʻike ka pahu keʻokeʻo i ka laulā interquartile, ʻo ka laina ʻeleʻele ākea ma ke kikowaena e hōʻike ana i ka median, a ʻo ka laina ʻeleʻele kū pololei e hoʻolōʻihi ana mai ka pahu e hōʻike ana i ka 95% interval hilinaʻi (CI). f ʻO ka Pearson correlation plot o SLC7A5 a me ka GAPDH expression i loko o ka lung adenocarcinoma (n = 513) a me ka maʻamau o ka lung tissue (n = 59) ma ka dataset TCGA. Hōʻike ka ʻāpana hina i ka 95% CI. r, Pearson correlation coefficient. g Nā pae tryptophan cellular i hoʻonohonoho ʻia i loko o nā cell A549 i hoʻololi ʻia me ka nonspecific shRNA control (NC) a me shSLC7A5 (Sh1, Sh2) i hoʻoholo ʻia e LC-MS/MS. Hōʻike ʻia ka loiloi helu o ʻelima mau laʻana kūʻokoʻa biologically i kēlā me kēia hui. h Nā pae cellular o NADt (NAD holoʻokoʻa, me NAD+ a me NADH) i loko o nā cell A549 (NC) a me SLC7A5 knockdown A549 cell (Sh1, Sh2). Hōʻike ʻia ka loiloi helu o ʻekolu mau laʻana kūʻokoʻa biologically i kēlā me kēia hui. Ua ana ʻia ka hana glycolytic o nā pūnaewele A549 ma mua a ma hope o ka hoʻokuʻu ʻana o SLC7A5 e ka extracellular acidification rate (ECAR) (n = 4 mau laʻana kūʻokoʻa biologically no kēlā me kēia hui). 2-DG,2-deoxy-D-glucose. Ua hoʻohana ʻia ka hoʻāʻo t ʻelua-tailed Student ma (b–h). Ma (g–i), hōʻike nā kaha hewa i ka awelika ± SD, ua hana ʻia kēlā me kēia hoʻokolohua ʻekolu manawa kūʻokoʻa a ua like nā hopena. Hāʻawi ʻia ka ʻikepili kumu ma ke ʻano o nā faila ʻikepili kumu.
Ke noʻonoʻo nei i ka hopena koʻikoʻi o ka metabolism tryptophan i hoʻololi ʻia i loko o ka hui LA, ua loiloi pū mākou i nā pae tryptophan serum i loko o nā hui HC, BN, a me LA me ka hoʻohana ʻana iā QQQ. Ua ʻike mākou ua hoʻemi ʻia ka tryptophan serum ma LA i hoʻohālikelike ʻia me HC a i ʻole BN (p <0.001, Kiʻi 5d), kahi mea e kūlike me nā ʻike mua e haʻahaʻa ana nā pae tryptophan e kahe ana i nā poʻe maʻi me ka maʻi kanesa māmā ma mua o nā kaohi olakino mai ka hui kaohi33,34,35. Ua ʻike ʻia kekahi noiʻi e hoʻohana ana i ka PET/CT tracer 11C-methyl-L-tryptophan ua hoʻonui nui ʻia ka manawa paʻa hōʻailona tryptophan i loko o ka ʻiʻo kanesa māmā i hoʻohālikelike ʻia me nā ʻeha benign a i ʻole nā ʻiʻo maʻamau36. Ke kuhi nei mākou e hōʻike paha ka emi ʻana o ka tryptophan i loko o ka serum LA i ka lawe ʻana o ka tryptophan ikaika e nā cell kanesa māmā.
Ua ʻike ʻia hoʻi ʻo ka huahana hope loa o ke ala kynurenine o ka tryptophan catabolism ʻo NAD+37,38, he substrate koʻikoʻi ia no ka hopena o ka glyceraldehyde-3-phosphate me 1,3-bisphosphoglycerate i ka glycolysis39. ʻOiai ua kālele nā haʻawina mua i ke kuleana o ka tryptophan catabolism i ka hoʻoponopono ʻana i ka pale ʻana, ua ʻimi mākou e wehewehe i ka pilina ma waena o ka dysregulation tryptophan a me nā ala glycolytic i ʻike ʻia i ke aʻo ʻana o kēia manawa. ʻIke ʻia ka ʻohana transporter solute 7 lālā 5 (SLC7A5) he tryptophan transporter43,44,45. ʻO ka Quinolinic acid phosphoribosyltransferase (QPRT) kahi enzyme i loaʻa ma lalo o ke ala kynurenine e hoʻololi i ka quinolinic acid i NAMN46. Ua hōʻike ka nānā ʻana o ka ʻikepili LUAD TCGA ua hoʻonui nui ʻia ʻo SLC7A5 a me QPRT i loko o ka ʻiʻo tumor i hoʻohālikelike ʻia me ka ʻiʻo maʻamau (Fig. 5e). Ua ʻike ʻia kēia piʻi ʻana ma nā pae I a me II a me nā pae III a me IV o ka adenocarcinoma māmā (Kiʻi Hoʻohui 11), e hōʻike ana i nā haunaele mua i ka metabolism tryptophan e pili ana me ka tumorigenesis.
Eia kekahi, ua hōʻike ka ʻikepili LUAD-TCGA i kahi pilina maikaʻi ma waena o ka hōʻike ʻana o SLC7A5 a me GAPDH mRNA i nā laʻana maʻi kanesa (r = 0.45, p = 1.55E-26, Kiʻi 5f). I ka hoʻohālikelike ʻana, ʻaʻohe pilina koʻikoʻi i loaʻa ma waena o ia mau pūlima gene i loko o ka ʻiʻo māmā maʻamau (r = 0.25, p = 0.06, Kiʻi 5f). ʻO ka hoʻohaʻahaʻa ʻana o SLC7A5 (Kiʻi Hoʻohui 12) i loko o nā pūnaewele A549 ua hōʻemi nui i nā pae tryptophan cellular a me NAD(H) (Kiʻi 5g,h), e hopena ana i ka hana glycolytic attenuated e like me ke ana ʻia e ka extracellular acidification rate (ECAR) (Kiʻi 1). 5i). No laila, ma muli o nā loli metabolic i loko o ke serum a me ka ʻike in vitro, ke kuhi nei mākou e hiki i ka metabolism tryptophan ke hana iā NAD+ ma o ke ala kynurenine a pāʻani i kahi kuleana koʻikoʻi i ka hoʻolaha ʻana i ka glycolysis i ka maʻi kanesa māmā.
Ua hōʻike ʻia nā haʻawina e hiki i ka nui o nā nodules pulmonary indeterminate i ʻike ʻia e LDCT ke alakaʻi i ka pono no nā hoʻāʻo hou e like me PET-CT, biopsy māmā, a me ka mālama nui ʻana ma muli o kahi hōʻailona wahaheʻe o ka malignancy.31 E like me ka mea i hōʻike ʻia ma ke Kiʻi 6, ua ʻike kā mākou haʻawina i kahi panela o nā metabolites serum me ka waiwai diagnostic hiki ke hoʻomaikaʻi i ka stratification pilikia a me ka hoʻokele ʻana ma hope o nā nodules pulmonary i ʻike ʻia e CT.
Loiloi ʻia nā nodules pulmonary me ka hoʻohana ʻana i ka tomography computed haʻahaʻa (LDCT) me nā hiʻohiʻona kiʻi e hōʻike ana i nā kumu maikaʻi ʻole a maikaʻi ʻole paha. ʻO ka hopena maopopo ʻole o nā nodules hiki ke alakaʻi i nā kipa hahai pinepine, nā hana pono ʻole, a me ka mālama nui ʻana. ʻO ka hoʻokomo ʻana o nā mea hoʻokaʻawale metabolic serum me ka waiwai diagnostic e hoʻomaikaʻi paha i ka loiloi pilikia a me ka hoʻokele ʻana o nā nodules pulmonary ma hope. PET positron emission tomography.
Hōʻike ka ʻikepili mai ka noiʻi US NLST a me ka noiʻi ʻEulopa NELSON e hiki i ka nānā ʻana i nā hui pilikia kiʻekiʻe me ka tomography computed haʻahaʻa (LDCT) ke hōʻemi i ka make ʻana o ka maʻi kanesa māmā1,3. Eia nō naʻe, ʻo ka loiloi pilikia a me ka hoʻokele lapaʻau ma hope o ka nui o nā nodules pulmonary i ʻike ʻia e LDCT ke paʻakikī loa. ʻO ka pahuhopu nui ka hoʻomaikaʻi ʻana i ka hoʻokaʻawale pololei ʻana o nā protocols LDCT e kū nei ma o ka hoʻokomo ʻana i nā biomarkers hilinaʻi.
Ua ʻike ʻia kekahi mau biomarkers molekala, e like me nā metabolites koko, ma ka hoʻohālikelike ʻana i ka maʻi kanesa māmā me nā kaohi olakino15,17. Ma ke aʻo ʻana o kēia manawa, ua kālele mākou i ka hoʻopili ʻana o ka loiloi metabolomics serum e hoʻokaʻawale ma waena o nā nodules pulmonary benign a malignant i ʻike ʻia e LDCT. Ua hoʻohālikelike mākou i ka metabolome serum honua o ka kaohi olakino (HC), nā nodules māmā benign (BN), a me nā laʻana adenocarcinoma māmā (LA) pae I me ka hoʻohana ʻana i ka loiloi UPLC-HRMS. Ua ʻike mākou he like nā ʻano metabolic o HC a me BN, ʻoiai ua hōʻike ʻo LA i nā loli koʻikoʻi i hoʻohālikelike ʻia me HC a me BN. Ua ʻike mākou i ʻelua mau pūʻulu o nā metabolites serum e hoʻokaʻawale iā LA mai HC a me BN.
ʻO ke ʻano hana ʻike LDCT o kēia manawa no nā nodules benign a malignant e pili nui ana i ka nui, ka nui, ke ʻano a me ka ulu ʻana o nā nodules i ka hala ʻana o ka manawa30. Ua hōʻike ʻia nā haʻawina mua e pili pono ana ka nui o nā nodules i ka hiki ke loaʻa i ka maʻi kanesa māmā. ʻOiai i nā poʻe maʻi kiʻekiʻe, ʻo ka pilikia o ka malignancy i nā nodes <6 mm he <1%. ʻO ka pilikia o ka malignancy no nā nodules e ana ana i 6 a 20 mm mai 8% a 64%30. No laila, ke paipai nei ka Fleischner Society i kahi anawaena cutoff o 6 mm no ka hahai ʻana i ka CT maʻamau.29 Eia nō naʻe, ʻaʻole i hana pono ʻia ka loiloi pilikia a me ka hoʻokele ʻana i nā nodules pulmonary indeterminate (IPN) ʻoi aku ka nui ma mua o 6 mm 31. ʻO ka hoʻokele ʻana o kēia manawa i ka maʻi puʻuwai hānau e pili pinepine ʻia ma ke kali makaʻala me ka nānā pinepine ʻana i ka CT.
Ma muli o ka metabolome i hōʻoia ʻia, ua hōʻike mākou no ka manawa mua i ka hui pū ʻana o nā pūlima metabolomic ma waena o nā kānaka olakino a me nā nodules benign <6 mm. Kūlike ka like ʻana o ke olaola me nā ʻike CT mua e like me ka haʻahaʻa o ka pilikia o ka malignancy no nā nodules <6 mm e like me nā kumuhana me ka ʻole o nā nodes.30 Pono e hoʻomaopopo ʻia e hōʻike pū ana kā mākou mau hopena he like nui nā nodules benign <6 mm a me ≥6 mm i nā ʻano metabolomic, e hōʻike ana he kūlike ka wehewehe hana o ka benign etiology me ka nānā ʻole i ka nui o ka nodule. No laila, hiki i nā panela metabolite serum diagnostic hou ke hāʻawi i kahi assay hoʻokahi ma ke ʻano he hoʻāʻo rule-out ke ʻike mua ʻia nā nodules ma CT a hiki ke hōʻemi i ka nānā ʻana i ka serial. I ka manawa like, ua hoʻokaʻawale ka panela like o nā biomarkers metabolic i nā nodules malignant ≥6 mm ka nui mai nā nodules benign a hāʻawi i nā wānana pololei no nā IPN o ka nui like a me nā hiʻohiʻona morphological ambiguous ma nā kiʻi CT. Ua hana maikaʻi kēia mea hoʻokaʻawale metabolism serum i ka wānana ʻana i ka malignancy o nā nodules ≥6 mm me ka AUC o 0.927. I ka hui pū ʻia ʻana, hōʻike kā mākou mau hopena e hiki i nā pūlima metabolomic serum kū hoʻokahi ke hōʻike pono i nā loli metabolic mua i hoʻokumu ʻia e ka tumor a loaʻa ka waiwai kūpono e like me nā wānana pilikia, kūʻokoʻa i ka nui o ka nodule.
ʻO ka mea nui, ʻo ka adenocarcinoma māmā (LUAD) a me ka squamous cell carcinoma (LUSC) nā ʻano nui o ka maʻi kanesa māmā ʻaʻole liʻiliʻi (NSCLC). No ka mea ua pili pono ʻo LUSC me ka hoʻohana ʻana i ka paka47 a ʻo LUAD ka histology maʻamau o nā nodules māmā i ʻike ʻia ma ka nānā ʻana o CT48, ua kūkulu kūikawā ʻia kā mākou kumu hoʻohālike classifier no nā laʻana adenocarcinoma pae I. Ua kālele pū ʻo Wang a me nā hoa hana ma LUAD a ua ʻike i ʻeiwa mau pūlima lipid e hoʻohana ana i ka lipidomics e hoʻokaʻawale i ka maʻi kanesa māmā mua mai nā poʻe olakino17. Ua hoʻāʻo mākou i ke kumu hoʻohālike classifier o kēia manawa ma 16 mau hihia o ka pae I LUSC a me 74 mau nodules benign a ua ʻike i ka pololei o ka wānana LUSC haʻahaʻa (AUC 0.776), e hōʻike ana he mau pūlima metabolomic ko LUAD a me LUSC. ʻOiaʻiʻo, ua hōʻike ʻia ʻo LUAD a me LUSC he ʻokoʻa i ka etiology, ke kumu olaola a me nā aberrations genetic49. No laila, pono e hoʻokomo ʻia nā ʻano histology ʻē aʻe i nā kumu hoʻomaʻamaʻa no ka ʻike ʻana i ka maʻi kanesa māmā ma muli o ka heluna kanaka i nā papahana nānā.
Maanei, ua ʻike mākou i nā ala ʻeono i hoʻololi pinepine ʻia i ka adenocarcinoma māmā i hoʻohālikelike ʻia me nā kaohi olakino a me nā nodules benign. ʻO Xanthine a me hypoxanthine nā metabolites maʻamau o ke ala metabolic purine. E kūlike me kā mākou mau hopena, ua hoʻonui nui ʻia nā mea waena e pili ana me ka metabolism purine i loko o ka serum a i ʻole nā ʻiʻo o nā maʻi me ka adenocarcinoma māmā i hoʻohālikelike ʻia me nā kaohi olakino a i ʻole nā maʻi ma ke kahua preinvasive15,50. ʻO nā pae serum xanthine a me hypoxanthine kiʻekiʻe e hōʻike paha i ka anabolism e pono ai e nā cell cancer proliferating wikiwiki. ʻO ka dysregulation o ka metabolism glucose kahi hōʻailona kaulana o ka metabolism cancer51. Maanei, ua ʻike mākou i kahi piʻi nui o ka pyruvate a me ka lactate i ka hui LA i hoʻohālikelike ʻia me ka hui HC a me BN, kahi e kūlike me nā hōʻike mua o nā ʻano ʻino o ke ala glycolytic i nā ʻano serum metabolome o nā maʻi maʻi kanesa māmā ʻaʻole liʻiliʻi (NSCLC) a me nā kaohi olakino. kūlike nā hopena52,53.
ʻO ka mea nui, ua ʻike mākou i kahi pilina inverse ma waena o ka pyruvate a me ka metabolism tryptophan i loko o ke serum o nā adenocarcinomas māmā. Ua hoʻemi ʻia nā pae tryptophan serum i ka hui LA i hoʻohālikelike ʻia me ka hui HC a i ʻole BN. ʻO ka mea hoihoi, ua ʻike ʻia kahi noiʻi nui ma mua e hoʻohana ana i kahi cohort prospective e pili ana nā pae haʻahaʻa o ka tryptophan e kahe ana me ka hoʻonui ʻia o ka pilikia o ka maʻi kanesa māmā 54. ʻO Tryptophan kahi waikawa amino koʻikoʻi a mākou e loaʻa ai mai ka meaʻai. Hoʻoholo mākou e hōʻike ana ka pau ʻana o ka tryptophan serum i ka adenocarcinoma māmā i ka pau wikiwiki o kēia metabolite. Ua ʻike maopopo ʻia ʻo ka huahana hope o ka tryptophan catabolism ma o ke ala kynurenine ke kumu o ka de novo NAD+ synthesis. No ka mea, ua hana nui ʻia ʻo NAD+ ma o ke ala salvage, ʻo ke koʻikoʻi o NAD+ i ka metabolism tryptophan i ke olakino a me nā maʻi e hoʻoholo ʻia46. Ua hōʻike kā mākou loiloi o ka waihona ʻikepili TCGA ua hoʻonui nui ʻia ka hōʻike ʻana o ka tryptophan transporter solute transporter 7A5 (SLC7A5) i ka adenocarcinoma māmā i hoʻohālikelike ʻia me nā kaohi maʻamau a ua pili pono me ka hōʻike ʻana o ka enzyme glycolytic GAPDH. Ua kālele nui nā haʻawina mua i ke kuleana o ka tryptophan catabolism i ka hoʻopau ʻana i ka pane pale antitumor40,41,42. Maanei mākou e hōʻike nei i ka pale ʻana o ka lawe ʻana o ka tryptophan ma o ka hoʻokuʻi ʻana o SLC7A5 i loko o nā cell cancer lungs e hopena i ka emi ʻana o nā pae NAD cellular a me ka hoʻēmi ʻana o ka hana glycolytic. I ka hōʻuluʻulu manaʻo, hāʻawi kā mākou haʻawina i kahi kumu olaola no nā loli i ka metabolism serum e pili ana me ka hoʻololi ʻino o ka adenocarcinoma māmā.
ʻO nā hoʻololi EGFR nā hoʻololi hoʻokele maʻamau i nā poʻe maʻi me NSCLC. I kā mākou noiʻi, ua ʻike mākou he like nā ʻano metabolomic o nā poʻe maʻi me ka EGFR wild-type (n = 41), ʻoiai ua ʻike mākou i ka emi ʻana o nā pae serum o kekahi mau poʻe maʻi EGFR mutant i nā poʻe maʻi acylcarnitine. ʻO ka hana i hoʻokumu ʻia o nā acylcarnitines ka lawe ʻana i nā hui acyl mai ka cytoplasm i loko o ka matrix mitochondrial, e alakaʻi ana i ka oxidation o nā waikawa momona e hana i ka ikehu 55. E like me kā mākou mau ʻike, ua ʻike pū kekahi noiʻi hou i nā ʻano metabolome like ma waena o nā ʻōpū EGFR mutant a me EGFR wild-type ma ka nānā ʻana i ka metabolome honua o 102 mau laʻana ʻiʻo adenocarcinoma māmā 50. ʻO ka mea hoihoi, ua loaʻa pū ka ʻike acylcarnitine i loko o ka hui EGFR mutant. No laila, inā paha e hōʻike ana nā loli i nā pae acylcarnitine i nā loli metabolic i hoʻokomo ʻia e EGFR a me nā ala molekala ma lalo e pono ai ke aʻo hou ʻia.
I ka hopena, ua hoʻokumu kā mākou noiʻi i kahi mea hoʻokaʻawale metabolic serum no ka ʻike ʻokoʻa o nā nodules pulmonary a hāpai i kahi kaʻina hana e hiki ai ke hoʻomaikaʻi i ka loiloi pilikia a hoʻomaʻamaʻa i ka hoʻokele lapaʻau e pili ana i ka nānā ʻana o ka CT scan.
Ua ʻāpono ʻia kēia haʻawina e ke Kōmike Hoʻopono o ka Halemai Kanesa o ke Kulanui ʻo Sun Yat-sen, ka Halemai Pili Mua o ke Kulanui ʻo Sun Yat-sen, a me ke Kōmike Hoʻopono o ka Halemai Kanesa o ke Kulanui ʻo Zhengzhou. Ma nā hui hōʻoia ʻike a me loko, ua hōʻiliʻili ʻia he 174 sera mai nā kānaka olakino a me 244 sera mai nā nodules benign mai nā kānaka e hana ana i nā hoʻokolohua lapaʻau makahiki ma ka ʻOihana Hoʻomalu a me ka Kāohi ʻana i ke Kanesa, ke Kikowaena Kanesa o ke Kulanui ʻo Sun Yat-sen, a me 166 nodules benign. serum. Ua hōʻiliʻili ʻia nā adenocarcinomas māmā Stage I mai ke Kikowaena Kanesa o ke Kulanui ʻo Sun Yat-sen. Ma ka hui hōʻoia waho, aia he 48 mau hihia o nā nodules benign, 39 mau hihia o ka adenocarcinoma māmā stage I mai ka Halemai Hoʻopono Mua o ke Kulanui ʻo Sun Yat-sen, a me 24 mau hihia o ka adenocarcinoma māmā stage I mai ka Halemai Kanesa ʻo Zhengzhou. Ua hōʻiliʻili pū ke Kikowaena Kanesa o ke Kulanui ʻo Sun Yat-sen i 16 mau hihia o ka maʻi kanesa māmā squamous cell stage I e hoʻāʻo i ka hiki ke diagnostic o ka mea hoʻokaʻawale metabolic i hoʻokumu ʻia (hōʻike ʻia nā ʻano o ka mea maʻi ma ka Papa Hoʻohui 5). Ua hōʻiliʻili ʻia nā laʻana mai ka hui ʻike a me ka hui hōʻoia kūloko ma waena o Ianuali 2018 a me Mei 2020. Ua hōʻiliʻili ʻia nā laʻana no ka hui hōʻoia kūwaho ma waena o ʻAukake 2021 a me ʻOkakopa 2022. No ka hōʻemi ʻana i ka manaʻo hoʻokae kāne, ua hāʻawi ʻia ka nui like o nā hihia kāne a me nā wahine i kēlā me kēia hui. Hui ʻIke a me ka Hui Loiloi Kūloko. Ua hoʻoholo ʻia ke kāne a me ke wahine o ka mea komo ma muli o ka hōʻike ponoʻī. Ua loaʻa ka ʻae ʻia mai nā mea komo āpau a ʻaʻohe uku i hāʻawi ʻia. ʻO nā kumuhana me nā nodules benign ʻo ia ka poʻe me ka helu CT scan paʻa ma 2 a 5 mau makahiki i ka manawa o ka nānā ʻana, koe wale nō 1 hihia mai ka laʻana hōʻoia kūwaho, i hōʻiliʻili ʻia ma mua o ke ʻoki ʻana a ʻike ʻia e ka histopathology. Me ka ʻokoʻa o ka bronchitis mau. Ua hōʻiliʻili ʻia nā hihia adenocarcinoma māmā ma mua o ka resection māmā a ua hōʻoia ʻia e ka maʻi pathological. Ua hōʻiliʻili ʻia nā laʻana koko hoʻokē ʻai i loko o nā ʻōmole hoʻokaʻawale serum me ka ʻole o nā anticoagulants. Ua hoʻopaʻa ʻia nā laʻana koko no 1 hola ma ka mahana o ka lumi a laila centrifuged ma 2851 × g no 10 mau minuke ma 4°C e hōʻiliʻili i ka supernatant serum. Ua hoʻopaʻahau ʻia nā aliquots serum ma -80°C a hiki i ka unuhi ʻana o ka metabolite. Ua hōʻiliʻili ke Keʻena o ka Pale ʻana i ke Kanesa a me ka Hoʻokolokolo Lapaʻau o ke Kikowaena Kanesa o ke Kulanui ʻo Sun Yat-sen i kahi waihona serum mai 100 mau mea hāʻawi olakino, me ka nui like o nā kāne a me nā wahine i piha i ka makahiki 40 a 55. Ua hui pū ʻia nā nui like o kēlā me kēia hāpana mea hāʻawi, ua hoʻokaʻawale ʻia ka waihona hopena a mālama ʻia ma -80°C. Ua hoʻohana ʻia ka hui serum ma ke ʻano he mea kuhikuhi no ka kaohi maikaʻi a me ka hoʻohālikelike ʻikepili.
Ua hoʻoheheʻe ʻia ka serum kuhikuhi a me nā laʻana hoʻāʻo a ua unuhi ʻia nā metabolites me ka hoʻohana ʻana i kahi ʻano unuhi hui (MTBE/methanol/wai) 56. I ka pōkole, ua hui pū ʻia he 50 μl o ka serum me 225 μl o ka methanol anuanu a me 750 μl o ka methyl tert-butyl ether anuanu (MTBE). E hoʻoulu i ka hui ʻana a hoʻomoʻa ma luna o ka hau no 1 hola. A laila ua hui pū ʻia nā laʻana a ua hui pū ʻia me 188 μl o ka wai MS-grade e loaʻa ana nā kūlana kūloko (13C-lactate, 13C3-pyruvate, 13C-methionine, a me 13C6-isoleucine, i kūʻai ʻia mai Cambridge Isotope Laboratories). A laila ua centrifuged ʻia ka hui ʻana ma 15,000 × g no 10 min ma 4 °C, a ua hoʻoili ʻia ke kahua haʻahaʻa i loko o ʻelua mau paipu (125 μL kēlā me kēia) no ka loiloi LC-MS ma nā ʻano maikaʻi a maikaʻi ʻole. ʻO ka mea hope loa, ua hoʻomaloʻo ʻia ka hāpana i loko o kahi mea hoʻonui vacuum wikiwiki.
Ua hoʻohui hou ʻia nā metabolites maloʻo i loko o 120 μl o 80% acetonitrile, vortexed no 5 min, a centrifuged ma 15,000 × g no 10 min ma 4°C. Ua hoʻoili ʻia nā supernatants i loko o nā vials aniani amber me nā microinserts no nā haʻawina metabolomics. ʻO ka nānā ʻana i ka metabolomics i hoʻolālā ʻole ʻia ma kahi kahua ultra-performance liquid chromatography-high-resolution mass spectrometry (UPLC-HRMS). Ua hoʻokaʻawale ʻia nā metabolites me ka hoʻohana ʻana i kahi ʻōnaehana Dionex Ultimate 3000 UPLC a me kahi kolamu ACQUITY BEH Amide (2.1 × 100 mm, 1.7 μm, Waters). Ma ke ʻano ion maikaʻi, ʻo nā pae mobile he 95% (A) a me 50% acetonitrile (B), kēlā me kēia me 10 mmol/L ammonium acetate a me 0.1% formic acid. Ma ke ʻano maikaʻi ʻole, aia nā pae mobile A a me B i loko o 95% a me 50% acetonitrile, kēlā me kēia, aia nā pae ʻelua i loko o 10 mmol/L ammonium acetate, pH = 9. Penei ka papahana gradient: 0–0.5 min, 2% B; 0.5–12 min, 2–50% B; 12–14 min, 50–98% B; 14–16 min, 98% B; 16–16.1. min, 98 –2% B; 16.1–20 min, 2% B. Ua mālama ʻia ke kolamu ma 40°C a me ka laʻana ma 10°C i loko o ka autosampler. ʻO ka nui o ke kahe ʻana he 0.3 ml/min, ʻo ka nui o ka injection he 3 μl. Ua hoʻohana ʻia kahi spectrometer mass Q-Exactive Orbitrap (Thermo Fisher Scientific) me kahi kumu electrospray ionization (ESI) ma ke ʻano scan piha a ua hui pū ʻia me ke ʻano nānā ʻana ddMS2 e hōʻiliʻili i nā nui o ka ʻikepili. Ua hoʻonohonoho ʻia nā palena MS penei: voltage spray +3.8 kV/- 3.2 kV, mahana capillary 320°C, kinoea pale 40 arb, kinoea kōkua 10 arb, mahana mea hoʻomehana probe 350°C, laulā scanning 70–1050 m/ h, hoʻonā. 70 000. Ua loaʻa nā ʻikepili me ka hoʻohana ʻana iā Xcalibur 4.1 (Thermo Fisher Scientific).
No ka loiloi ʻana i ka maikaʻi o ka ʻikepili, ua hana ʻia nā laʻana kaohi maikaʻi (QC) i hui pū ʻia ma ka wehe ʻana i 10 μL aliquots o ka supernatant mai kēlā me kēia laʻana. Ua kālailai ʻia ʻeono mau inikini laʻana kaohi maikaʻi i ka hoʻomaka ʻana o ke kaʻina loiloi e loiloi i ke kūpaʻa o ka ʻōnaehana UPLC-MS. A laila hoʻokomo pinepine ʻia nā laʻana kaohi maikaʻi i loko o ka pūʻulu. Ua kālailai ʻia nā pūʻulu 11 a pau o nā laʻana serum i loko o kēia haʻawina e LC-MS. Ua hoʻohana ʻia nā aliquots o kahi hui serum pool mai 100 mau mea hāʻawi olakino ma ke ʻano he mea kuhikuhi i nā pūʻulu e nānā i ke kaʻina hana unuhi a hoʻoponopono no nā hopena batch-to-batch. Ua hana ʻia ka loiloi metabolomics i kuhikuhi ʻole ʻia o ka cohort discovery, ka cohort hōʻoia kūloko, a me ka cohort hōʻoia waho ma ke Kikowaena Metabolomics o ke Kulanui ʻo Sun Yat-sen. Ua kālailai pū ka hale hana waho o ke Kulanui ʻo Guangdong o Technology Analysis and Testing Center i 40 mau laʻana mai ka cohort waho e hoʻāʻo i ka hana o ke kumu hoʻohālike classifier.
Ma hope o ka unuhi ʻana a me ka hoʻohui hou ʻana, ua ana ʻia ka nui piha o nā metabolites serum me ka hoʻohana ʻana i ka ultra-high performance liquid chromatography-tandem mass spectrometry (Agilent 6495 triple quadrupole) me kahi kumu electrospray ionization (ESI) ma ke ʻano multiple reaction monitoring (MRM). Ua hoʻohana ʻia kahi kolamu ACQUITY BEH Amide (2.1 × 100 mm, 1.7 μm, Waters) e hoʻokaʻawale i nā metabolites. ʻO ka pae mobile he 90% (A) a me 5% acetonitrile (B) me 10 mmol/L ammonium acetate a me 0.1% ammonia solution. Penei ka papahana gradient: 0–1.5 min, 0% B; 1.5–6.5 min, 0–15% B; 6.5–8 min, 15% B; 8–8.5 min, 15%–0% B; 8.5–11.5 min, 0%B. Ua mālama ʻia ke kolamu ma 40 °C a me ka hāpana ma 10 °C i loko o ka autosampler. ʻO ka nui o ke kahe ʻana he 0.3 mL/min a ʻo ka nui o ka injection he 1 μL. Ua hoʻonohonoho ʻia nā palena MS penei: ka voltage capillary ±3.5 kV, ke kaomi nebulizer 35 psi, ke kahe ʻana o ke kinoea sheath 12 L/min, ka mahana o ke kinoea sheath 350°C, ka mahana o ke kinoea maloʻo 250°C, a me ke kahe ʻana o ke kinoea maloʻo 14 l/min. ʻO nā hoʻololi MRM o ka tryptophan, pyruvate, lactate, hypoxanthine a me ka xanthine he 205.0–187.9, 87.0–43.4, 89.0–43.3, 135.0–92.3 a me 151.0–107.9 pakahi. Ua hōʻiliʻili ʻia nā ʻikepili me ka hoʻohana ʻana iā Mass Hunter B.07.00 (Agilent Technologies). No nā laʻana serum, ua helu ʻia ka tryptophan, pyruvate, lactate, hypoxanthine, a me ka xanthine me ka hoʻohana ʻana i nā piʻo calibration o nā hopena hui maʻamau. No nā laʻana cell, ua hoʻohālikelike ʻia ka nui o ka tryptophan i ke kūlana kūloko a me ka nui o ka protein cell.
Ua hana ʻia ka unuhi ʻana o ka piko (m/z a me ka manawa paʻa (RT)) me ka hoʻohana ʻana i ka Compound Discovery 3.1 a me TraceFinder 4.0 (Thermo Fisher Scientific). No ka hoʻopau ʻana i nā ʻokoʻa hiki ke loaʻa ma waena o nā ʻāpana, ua puʻunaue ʻia kēlā me kēia piko hiʻohiʻona o ka hāpana hoʻāʻo e ka piko hiʻohiʻona o ka mea kuhikuhi mai ka ʻāpana like e loaʻa ai ka nui pili. Hōʻike ʻia nā ʻokoʻa maʻamau pili o nā kūlana kūloko ma mua a ma hope o ka hoʻohālikelike ʻana ma ka Papa Hoʻohui 6. Ua wehewehe ʻia nā ʻokoʻa ma waena o nā hui ʻelua e ka helu ʻike wahaheʻe (FDR<0.05, hoʻāʻo kūlana i kau inoa ʻia ʻo Wilcoxon) a me ka loli pelu (>1.2 a i ʻole <0.83). Hōʻike ʻia ka ʻikepili MS maka o nā hiʻohiʻona i unuhi ʻia a me ka ʻikepili MS i hoʻoponopono ʻia i ka serum kuhikuhi ma ka ʻIkepili Hoʻohui 1 a me ka ʻIkepili Hoʻohui 2, kēlā me kēia. Ua hana ʻia ka annotation piko ma muli o ʻehā mau pae i wehewehe ʻia o ka ʻike ʻana, me nā metabolites i ʻike ʻia, nā hui i hōʻike ʻia, nā papa hui i hōʻike ʻia, a me nā hui i ʻike ʻole ʻia 22. Ma muli o nā ʻimi waihona ʻikepili ma Compound Discovery 3.1 (mzCloud, HMDB, Chemspider), ua koho hope ʻia nā hui olaola me nā kūlana i hōʻoia ʻia e MS/MS a i ʻole nā annotations kūlike pololei ma mzCloud (helu > 85) a i ʻole Chemspider ma ke ʻano he waena ma waena o ka metabolome ʻokoʻa. Hoʻokomo ʻia nā annotations Peak no kēlā me kēia hiʻohiʻona i ka ʻIkepili Hoʻohui 3. Ua hoʻohana ʻia ʻo MetaboAnalyst 5.0 no ka loiloi univariate o ka nui o ka metabolite i hoʻonohonoho ʻia. Ua loiloi pū ʻo MetaboAnalyst 5.0 i ka loiloi hoʻonui ala KEGG ma muli o nā metabolites ʻokoʻa koʻikoʻi. Ua kālailai ʻia ka loiloi ʻāpana nui (PCA) a me ka loiloi discriminant hapa liʻiliʻi loa (PLS-DA) me ka hoʻohana ʻana i ka pūʻolo polokalamu ropls (v.1.26.4) me ka normalization stack a me ka autoscaling. Ua hoʻokumu ʻia ke kumu hoʻohālike biomarker metabolite kūpono no ka wānana ʻana i ka malignancy nodule me ka hoʻohana ʻana i ka regression logistic binary me ka liʻiliʻi loa o ka shrinkage absolute a me ka mea koho (LASSO, pūʻolo R v.4.1-3). Ua hōʻike ʻia ka hana o ke kumu hoʻohālikelike i nā seti ʻike a me ka hōʻoia ma o ka helu ʻana iā AUC ma muli o ka loiloi ROC e like me ka pūʻolo pROC (v.1.18.0.). Ua loaʻa ka palena kūpono kūpono ma muli o ka helu helu Youden kiʻekiʻe loa o ke kumu hoʻohālike (sensitivity + specificity - 1). ʻO nā laʻana me nā waiwai i emi a ʻoi aku paha ma mua o ka paepae e wānana ʻia he mau nodules benign a me ka adenocarcinoma māmā, kēlā me kēia.
Ua ulu ʻia nā pūnaewele A549 (#CCL-185, American Type Culture Collection) i loko o ka medium F-12K me 10% FBS. Ua hoʻokomo ʻia nā moʻo RNA hairpin pōkole (shRNA) e kuhikuhi ana iā SLC7A5 a me kahi kaohi nontargeting (NC) i loko o ka vector lentiviral pLKO.1-puro. Penei nā moʻo antisense o shSLC7A5: Sh1 (5′-GGAGAAACCTGAACAGTT-3′), Sh2 (5′-GCCGTGGACTTCGGGAACTAT-3′). Ua kūʻai ʻia nā antibodies iā SLC7A5 (#5347) a me tubulin (#2148) mai Cell Signaling Technology. Ua hoʻohana ʻia nā antibodies iā SLC7A5 a me tubulin ma kahi dilution o 1:1000 no ka nānā ʻana o Western blot.
Ke ana nei ka Seahorse XF Glycolytic Stress Test i nā pae extracellular acidification (ECAR). I ka hoʻāʻo ʻana, ua lawelawe ʻia ka glucose, oligomycin A, a me 2-DG ma ke ʻano he hoʻāʻo e hoʻāʻo i ka mana glycolytic cellular e like me ke ana ʻia e ECAR.
Ua hoʻopili ʻia nā pūnaewele A549 i hoʻololi ʻia me ka mana hoʻokele ʻole (NC) a me shSLC7A5 (Sh1, Sh2) i ka pō i loko o nā kīʻaha 10 cm ke anawaena. Ua unuhi ʻia nā metabolites cell me 1 ml o ka methanol wai anuanu 80%. Ua ʻoki ʻia nā pūnaewele i loko o ka hopena methanol, hōʻiliʻili ʻia i loko o kahi paipu hou, a centrifuged ma 15,000 × g no 15 min ma 4°C. E hōʻiliʻili i 800 µl o ka supernatant a maloʻo me ka hoʻohana ʻana i kahi vacuum concentrator wikiwiki. A laila ua kālailai ʻia nā pellets metabolite maloʻo no nā pae tryptophan me ka hoʻohana ʻana iā LC-MS/MS e like me ka mea i wehewehe ʻia ma luna. Ua ana ʻia nā pae NAD(H) cellular i loko o nā pūnaewele A549 (NC a me shSLC7A5) me ka hoʻohana ʻana i kahi pahu colorimetric NAD+/NADH quantitative (#K337, BioVision) e like me nā kuhikuhi a ka mea hana. Ua ana ʻia nā pae protein no kēlā me kēia laʻana e hoʻomaʻamaʻa i ka nui o nā metabolites.
ʻAʻohe ʻano helu i hoʻohana ʻia e hoʻoholo mua i ka nui o ka laʻana. Ua manaʻo ʻia nā haʻawina metabolomics ma mua i kuhikuhi ʻia i ka ʻike ʻana o ka biomarker15,18 he mau pae hoʻohālikelike no ka hoʻoholo ʻana i ka nui, a i hoʻohālikelike ʻia me kēia mau hōʻike, ua lawa kā mākou laʻana. ʻAʻohe laʻana i hoʻokaʻawale ʻia mai ka hui noiʻi. Ua hoʻokaʻawale ʻia nā laʻana serum i kahi hui ʻike (306 mau hihia, 74.6%) a me kahi hui hōʻoia kūloko (104 mau hihia, 25.4%) no nā haʻawina metabolomics i kuhikuhi ʻole ʻia. Ua koho maʻamau mākou i 70 mau hihia mai kēlā me kēia hui mai ka hoʻonohonoho ʻike no nā haʻawina metabolomics i kuhikuhi ʻia. Ua makapō nā mea noiʻi i ka hoʻokaʻawale ʻana o ka hui i ka wā o ka hōʻiliʻili ʻikepili LC-MS a me ka nānā ʻana. Ua wehewehe ʻia nā loiloi helu o ka ʻikepili metabolomics a me nā hoʻokolohua cell ma nā ʻāpana Results, Figure Legends, a me Methods. Ua hana ʻia ka helu ʻana o ka tryptophan cellular, NADT, a me ka hana glycolytic ʻekolu manawa kūʻokoʻa me nā hopena like.
No ka ʻike hou aku e pili ana i ka hoʻolālā haʻawina, e ʻike i ka Natural Portfolio Report Abstract e pili ana me kēia ʻatikala.
Hōʻike ʻia ka ʻikepili MS maka o nā hiʻohiʻona i unuhi ʻia a me ka ʻikepili MS maʻamau o ka serum kuhikuhi ma ka ʻIkepili Hoʻohui 1 a me ka ʻIkepili Hoʻohui 2, kēlā me kēia. Hōʻike ʻia nā hōʻike kiʻekiʻe no nā hiʻohiʻona ʻokoʻa ma ka ʻIkepili Hoʻohui 3. Hiki ke hoʻoiho ʻia ka ʻikepili LUAD TCGA mai https://portal.gdc.cancer.gov/. Hāʻawi ʻia ka ʻikepili hoʻokomo no ke kaha kiʻi ʻana i ka pakuhi ma ka ʻikepili kumu. Hāʻawi ʻia ka ʻikepili kumu no kēia ʻatikala.
ʻO ka National Lung Screening Study Group, etc. Ka hoʻemi ʻana i ka make ʻana o ka maʻi kanesa māmā me ka tomography computed haʻahaʻa. ʻEnelani ʻĀkau. J. Med. 365, 395–409 (2011).
ʻO Kramer, BS, Berg, KD, Aberle, DR a me Prophet, PC Nānā ʻana i ka maʻi ʻaʻai māmā me ka hoʻohana ʻana i ka CT helical haʻahaʻa: nā hopena mai ka National Lung Screening Study (NLST). J. Med. Screen 18, 109–111 (2011).
ʻO De Koning, HJ, et al. Ke hōʻemi nei i ka make ʻana o ka maʻi kanesa māmā me ka nānā ʻana o ka CT volumetric i kahi hoʻokolohua randomized. ʻEnelani ʻĀkau. J. Med. 382, 503–513 (2020).
Ka manawa hoʻouna: Sep-18-2023