The aim of this study was to evaluate transcript levels of all 49 human ATP-binding cassette transporters (ABCs) in one of the most drug-resistant cancers, namely, the pancreatic ductal adenocarcinoma (PDAC). Association of ABCs levels with clinical-pathologic characteristics and KRAS mutation status was followed as well.Tumors and adjacent nonneoplastic tissues were obtained from 32 histologically verified PDAC patients. The transcript profile of ABCs was assessed using quantitative real-time polymerase chain reaction with a relative standard curve. KRAS mutations in exon 2 were assessed by high-resolution melting analysis and sequencing.Most ABCs were deregulated in PDAC and 10 ABCs were associated with clinical-pathologic characteristics. KRAS mutations did not change the global expression profile of ABCs.The expression of ABC transporters was significantly deregulated in PDAC tumors when compared to nonmalignant tissues. The observed up-regulation of ABCB4, ABCB11, ABCC1, ABCC3, ABCC5, ABCC10, and ABCG2 in tumors may contribute to the generally poor treatment response of PDAC. The up-regulation of ABCA1, ABCA7, and ABCG1 implicates a serious impairment of cellular cholesterol homeostasis in PDAC. On the other hand, the observed down-regulation of ABCA3, ABCC6, ABCC7, and ABCC8 suggests a possible role of stem cells in the development and progression of PDAC.
Panel of 509 pharmacogenes and oncogenes was selected for targeted next generation sequencing. Data mining in databases and functional impact software tools were used as an evidence for potential pathogenicity. Survey of several in silico methods revealed significant differences in resultant set of genes mostly due to number of tools used. Following the most stringent approach with fully concordant prediction in comparison to specific combination of tools and majority vote prediction, ten prioritized genes were statistically correlated with the clinical patient data. The highest burden of pathogenic variants correlating with clinical data was statistically significant in CFTR gene. Patients carrying these variants had significantly reduced disease-free-survival in comparison with patients without these variants.
Breast cancer is the most frequent cancer in the female population worldwide. The role of germline genetic variability in cytochromes P450 (CYP) in breast cancer prognosis and individualized therapy awaits detailed elucidation. In the present study, we used the next-generation sequencing to assess associations of germline variants in the coding and regulatory sequences of all human CYP genes with response of the patients to the neoadjuvant cytotoxic chemotherapy and disease-free survival (n = 105). A total of 22 prioritized variants associating with a response or survival in the above evaluation phase were then analyzed by allelic discrimination in the large confirmation set (n = 802). Associations of variants in CYP1B1, CYP4F12, CYP4X1, and TBXAS1 with the response to the neoadjuvant cytotoxic chemotherapy were replicated by the confirmation phase. However, just association of variant rs17102977 in CYP4X1 passed the correction for multiple testing and can be considered clinically and statistically validated. Replicated associations for variants in CYP4X1, CYP24A1, and CYP26B1 with disease-free survival of all patients or patients stratified to subgroups according to therapy type have not passed a false discovery rate test. Although statistically not confirmed by the present study, the role of CYP genes in breast cancer prognosis should not be ruled out. In conclusion, the present study brings replicated association of variant rs17102977 in CYP4X1 with the response of patients to the neoadjuvant cytotoxic chemotherapy and warrants further research of genetic variation CYPs in breast cancer.
Apoptosis plays a critical role in cancer cell survival and tumor development. We provide a hypothesis-generating screen for further research by exploring the expression profile and genetic variability of caspases (2, 3, 7, 8, 9, and 10) in breast carcinoma patients. This study addressed isoform-specific caspase transcript expression and genetic variability in regulatory sequences of caspases 2 and 9.Gene expression profiling was performed by quantitative real-time PCR in tumor and paired non-malignant tissues of two independent groups of patients. Genetic variability was determined by high resolution melting, allelic discrimination, and sequencing analysis in tumor and peripheral blood lymphocyte DNA of the patients.CASP3 A+B and S isoforms were over-expressed in tumors of both patient groups. The CASP9 transcript was down-regulated in tumors of both groups of patients and significantly associated with expression of hormonal receptors and with the presence of rs4645978-rs2020903-rs4646034 haplotype in the CASP9 gene. Patients with a low intratumoral CASP9A/B isoform expression ratio (predicted to shift equilibrium towards anti-apoptotic isoform) subsequently treated with adjuvant chemotherapy had a significantly shorter disease-free survival than those with the high ratio (p=0.04). Inheritance of CC genotype of rs2020903 in CASP9 was associated with progesterone receptor expression in tumors (p=0.003).Genetic variability in CASP9 and expression of its splicing variants present targets for further study.
Východiska: Kaspazy 8 a 9 (kodovane geny CASP8 a CASP9 ) jsou hlavnimi iniciacnimi kaspazami programovane buněcne smrti (apoptozy), jejiž deregulace může vest ke vzniku a progresi nadoroveho onemocněni, i k lekove rezistenci pacientek. Cilem prace bylo ověřit, zda existuje vztah mezi výskytem polymorfismů v CASP8 a CASP9 spojovaných s rizikem vzniku karcinomu prsu, hladinami transkriptu těchto genů (vcetně hladin alternativnich proti-apoptických variant) v nadorových tkanich karcinomu prsu a klinickým profilem pacientek. Material a metody: Polymorfismy byly stanoveny metodami Sangerova sekvenovani, pomoci high resolution melting analýzy (HRM) a alelicke diskriminace ve vzorcich DNA nadorových tkani a krevnich lymfocytů ziskaných od 60 pacientek. Hladiny celkoveho transkriptu CASP8, CASP9 a alternativnich variant CASP8L a CASP9B byly stanoveny ve vzorcich nadorove tkaně pomoci kvantitativni PCR v realnem case. Klinicky zajimave vztahy byly nasledně ověřeny na souboru vzorků DNA ziskane z periferni krve od 615 pacientek. Výsledky: Výskyt haplotypu rs4645978-rs2020903-rs4646034 v CASP9 významně asocioval s hladinou celkoveho transkriptu CASP9 v nadorech, expresi progesteronoveho receptoru, expresi ERBB2 a TNBC subtypem karcinomu prsu. Významne vztahy polymorfismu rs3834129 v CASP8 se stadiem onemocněni, polymorfismu rs6435074 s expresi estrogenniho receptoru, ERBB2 expresi a s gradem nadorů, a polymorfismu rs6723097 s expresi ERBB2 nebyly validacni studii potvrzeny. Nicmeně nosicky alely A v polymorfismu rs6723097 v CASP8 lecene hormonalni terapii vykazovaly delsi bezpřiznakove přeživani než nosicky genotypu CC. Zavěr: Soucasna studie odhalila dosud neznamý a potencialně ( in silico ) funkcni vztah haplotypu CASP9 , složeneho ze tři polymorfismů typu SNP, k molekularnimu i klinickemu fenotypu karcinomu prsu. Tento vztah naznacuje, za předpokladu, že bude ověřen nezavislými studiemi, možne využiti v prognostice onemocněni.
