The aim of our study was to set up a panel for targeted sequencing of chemoresistance genes and the main transcription factors driving their expression and to evaluate their predictive and prognostic value in breast cancer patients. Coding and regulatory regions of 509 genes, selected from PharmGKB and Phenopedia, were sequenced using massive parallel sequencing in blood DNA from 105 breast cancer patients in the testing phase. In total, 18,245 variants were identified of which 2565 were novel variants (without rs number in dbSNP build 150) in the testing phase. Variants with major allele frequency over 0.05 were further prioritized for validation phase based on a newly developed decision tree. Using emerging in silico tools and pharmacogenomic databases for functional predictions and associations with response to cytotoxic therapy or disease-free survival of patients, 55 putative variants were identified and used for validation in 805 patients with clinical follow up using KASPTM technology. In conclusion, associations of rs2227291, rs2293194, and rs4376673 (located in ATP7A, KCNAB1, and DFFB genes, respectively) with response to neoadjuvant cytotoxic therapy and rs1801160 in DPYD with disease-free survival of patients treated with cytotoxic drugs were validated and should be further functionally characterized.

• Klíčová slova
• breast cancer, chemoresistance, in silico prediction, next generation sequencing, pharmacogenomics,
• Publikační typ
• časopisecké články MeSH

BACKGROUND: This study addresses involvement of major 5-fluorouracil (5-FU) pathway genes in the prognosis of colorectal carcinoma patients. METHODS: Testing set and two validation sets comprising paired tumor and adjacent mucosa tissue samples from 151 patients were used for transcript profiling of 15 5-FU pathway genes by quantitative real-time PCR and DNA methylation profiling by high resolution melting analysis. Intratumoral molecular profiles were correlated with clinical data of patients. Protein levels of two most relevant candidate markers were assessed by immunoblotting. RESULTS: Downregulation of DPYD and upregulation of PPAT, UMPS, RRM2, and SLC29A1 transcripts were found in tumors compared to adjacent mucosa in testing and validation sets of patients. Low RRM2 transcript level significantly associated with poor response to the first-line palliative 5-FU-based chemotherapy in the testing set and with poor disease-free interval of patients in the validation set irrespective of 5-FU treatment. UPP2 was strongly methylated while its transcript absent in both tumors and adjacent mucosa. DPYS methylation level was significantly higher in tumor tissues compared to adjacent mucosa samples. Low intratumoral level of UPB1 methylation was prognostic for poor disease-free interval of the patients (P = 0.0002). The rest of the studied 5-FU genes were not methylated in tumors or adjacent mucosa. CONCLUSIONS: The observed overexpression of several 5-FU activating genes and DPYD downregulation deduce that chemotherapy naïve colorectal tumors share favorable gene expression profile for 5-FU therapy. Low RRM2 transcript and UPB1 methylation levels present separate poor prognosis factors for colorectal carcinoma patients and should be further investigated.

• Klíčová slova
• 5-fluorouracil, Colorectal carcinoma, Expression, Methylation, Prognosis,
• MeSH
• CpG ostrůvky MeSH
• fluoruracil farmakologie   MeSH
• kolorektální nádory genetika   metabolismus   mortalita   patologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• metylace DNA MeSH
• promotorové oblasti (genetika) MeSH
• protinádorové antimetabolity farmakologie   MeSH
• regulace genové exprese u nádorů účinky léků   MeSH
• ribonukleosiddifosfátreduktasa genetika   metabolismus   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• signální transdukce účinky léků   MeSH
• staging nádorů MeSH
• stanovení celkové genové exprese * MeSH
• stupeň nádoru MeSH
• transkriptom * MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fluoruracil MeSH
• protinádorové antimetabolity MeSH
• ribonucleotide reductase M2 MeSH Prohlížeč
• ribonukleosiddifosfátreduktasa MeSH