Combined Transcription-metabolism to Explore the Role of Tryptophan Metabolic Pathway in Trastuzumab Resistance in HER2-positive Breast Cancer

OBJECTIVE  To explore metabolomic and transcriptomic differences in the tryptophan pathway between trastuzumab-sensitive and trastuzumab-resistant cells in human epidermal growth factor receptor 2(HER2) positive breast cancer.

METHODS  Real-timequantitative polymerase chain reaction(qRT-PCR) and targeted metabolomics basing on ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) were used to detect levels of metabolic enzyme genes(AFMID, KYNU, AADAT, KYAT3, KYAT1, HAAO, KMO, CAT) and metabolites(quinolinic acid, picolinic acid, cinnabarinic acid, 3-hydroxykynurenine, xanthurenic acid, kynurenic acid, 3-hydroxyanthranilic acid, nicotinamide, tryptophan, kynurenine) in trastuzumab-sensitive cells and trastuzumab-resistant cells. Orthogonal partial least-squares discrimination analysis(OPLS-DA) was used to perform pattern recognition analysis of metabolic enzyme genes and metabolites. The differential metabolic enzyme genes and metabolites were screened by the criteria of P<0.05 and fold change(FC)>1.5 or FC<0.67. And the receiver operating characteristic curve(ROC) analysis was used to access the classification ability of differential metabolites and differential metabolic enzyme genes.

RESULTS The OPLS-DA models based on the expression levels of metabolic enzyme genes and metabolites could distinguish trastuzumab-sensitive cells and trastuzumab-resistant cells. Comparing to trastuzumab-sensitive cells, expression levels of HAAO were up-regulated in trastuzumab-resistant cells(P<0.05), the levels of KMO and AFMID were down-regulated in trastuzumab-resistant cells (P<0.05), the levels of 3-hydroxyanthranilic acid, kynurenine, kynurenic acid and cinnabarinic acid were up-regulated in trastuzumab-resistant cells(P<0.05) and levels of nicotinamide were down-regulated in trastuzumab-resistant cells(P<0.05). The ROC analysis revealed that the area under curve of kynurenine, kynurenic acid, vincristine, nicotinamide and KMO were >0.8.

CONCLUSION Abnormal expression of the tryptophan metabolic pathway metabolizing enzyme gene KMO and metabolites(kynurenine, kynurenic acid, nicotinamide, and cinnabarinic acid) in trastuzumab-resistant and sensitive cells of HER2-positive breast cancers is expected to contribute to the study of tryptophan metabolic profiles of trastuzumab-resistant cells of HER2-positive breast cancers.