Journal: Scientific Reports

Article Title: Phosphoserine aminotransferase 1 is associated to poor outcome on tamoxifen therapy in recurrent breast cancer

doi: 10.1038/s41598-017-02296-w

Figure Lengend Snippet: Schematic overview of experimental workflow. Panel A: a total of 379 FFPE tissues were captured on a tissue micro-array and analyzed by IHC. After filtering for ER positivity and hormonal naïve tumors, a total of 279 samples remained. Further filtering for missing data after IHC analysis yielded a panel of 261 tumors, on which survival analysis for the association of PSAT1 protein levels to TTP was performed. Parallel to this, PSAT1 mRNA expression was measured by RT-qPCR (n = 161) and Affymetrix chip (n = 155) approaches on frozen tumor specimens. These data were used for comparison between PSAT1 mRNA and protein levels (TMA and RT-qPCR; n = 56), correlation analysis (RT-qPCR and Affymetrix; n = 122), and pathway analysis (Affymetrix only; n = 155). Panel B shows tumor sample overlap between TMA, RT-qPCR and Affymetrix sets. Acronyms: ER: estrogen receptor; FFPE: formalin-fixed paraffin-embedded; IHC: immunohistochemistry; TMA: tissue microarray TTP: time to progression; RT-qPCR: quantitative reverse transcriptase polymerase chain reaction.

Article Snippet: Quantification of PSAT1 was performed using the TaqMan probe–based gene expression assay Hs00253548_m1 specific for splice variant beta (Applied Biosystems/Life Technologies, Warrington, WA, USA) as previously described , , For Affymetrix gene expression profiling, total RNA samples were cleaned and DNAse treated with the NucleoSpin RNA II kit according the manufacturers instruction (Machery-Nagel, Dueren, Germany) and shipped to ServiceXS (Leiden, The Netherlands) for downstream processing with the 3’IVT express kit and hybridization on the Human Genome (HG) U133 Plus 2.0 array (n = 20) and HG U133 Perfect Match.

Techniques: Microarray, Expressing, Quantitative RT-PCR, Formalin-fixed Paraffin-Embedded, Immunohistochemistry, Polymerase Chain Reaction

Journal: Scientific Reports

Article Title: Phosphoserine aminotransferase 1 is associated to poor outcome on tamoxifen therapy in recurrent breast cancer

doi: 10.1038/s41598-017-02296-w

Figure Lengend Snippet: PSAT1 expression and clinical relevance in the TMA and Affymetrix datasets. Breast carcinoma IHC stained tissues either displayed high or low PSAT1 protein levels. Two representative specimen having either high or low PSAT1 are presented (panel A). Kaplan-Meier analysis showed that high expression of PSAT1 protein was significantly associated to shorter TTP when compared to tumors with low PSAT1 levels (panel B). PSAT1 mRNA levels were assessed by Affymetrix GeneChip. Statistical analysis not only showed PSAT1 mRNA expression was enrihed in poor outcome patients (t test P = 0.014; panel C), but Kaplan Meier analysis showed that, also in this set, PSAT1 expression was significantly associated to shorter TTP (panel D).

Article Snippet: Quantification of PSAT1 was performed using the TaqMan probe–based gene expression assay Hs00253548_m1 specific for splice variant beta (Applied Biosystems/Life Technologies, Warrington, WA, USA) as previously described , , For Affymetrix gene expression profiling, total RNA samples were cleaned and DNAse treated with the NucleoSpin RNA II kit according the manufacturers instruction (Machery-Nagel, Dueren, Germany) and shipped to ServiceXS (Leiden, The Netherlands) for downstream processing with the 3’IVT express kit and hybridization on the Human Genome (HG) U133 Plus 2.0 array (n = 20) and HG U133 Perfect Match.

Techniques: Expressing, Staining

Journal: Scientific Reports

Article Title: Phosphoserine aminotransferase 1 is associated to poor outcome on tamoxifen therapy in recurrent breast cancer

doi: 10.1038/s41598-017-02296-w

Figure Lengend Snippet: Association of PSAT1 protein expression to clinical and histo-pathological characteristics.

Article Snippet: Quantification of PSAT1 was performed using the TaqMan probe–based gene expression assay Hs00253548_m1 specific for splice variant beta (Applied Biosystems/Life Technologies, Warrington, WA, USA) as previously described , , For Affymetrix gene expression profiling, total RNA samples were cleaned and DNAse treated with the NucleoSpin RNA II kit according the manufacturers instruction (Machery-Nagel, Dueren, Germany) and shipped to ServiceXS (Leiden, The Netherlands) for downstream processing with the 3’IVT express kit and hybridization on the Human Genome (HG) U133 Plus 2.0 array (n = 20) and HG U133 Perfect Match.

Techniques: Expressing

Journal: Scientific Reports

Article Title: Phosphoserine aminotransferase 1 is associated to poor outcome on tamoxifen therapy in recurrent breast cancer

doi: 10.1038/s41598-017-02296-w

Figure Lengend Snippet: Cox regression analysis for TTP of PSAT1 stained tumors.

Article Snippet: Quantification of PSAT1 was performed using the TaqMan probe–based gene expression assay Hs00253548_m1 specific for splice variant beta (Applied Biosystems/Life Technologies, Warrington, WA, USA) as previously described , , For Affymetrix gene expression profiling, total RNA samples were cleaned and DNAse treated with the NucleoSpin RNA II kit according the manufacturers instruction (Machery-Nagel, Dueren, Germany) and shipped to ServiceXS (Leiden, The Netherlands) for downstream processing with the 3’IVT express kit and hybridization on the Human Genome (HG) U133 Plus 2.0 array (n = 20) and HG U133 Perfect Match.

Techniques: Staining

Journal: Scientific Reports

Article Title: Phosphoserine aminotransferase 1 is associated to poor outcome on tamoxifen therapy in recurrent breast cancer

doi: 10.1038/s41598-017-02296-w

Figure Lengend Snippet: PSAT1 expression associated genes in the gene expression dataset. The 155 tumors in the Affymetrix cohort were stratified according to PSAT1 expression. All genes were annotated for KEGG terms and Global test was performed to assess which terms were associated to PSAT1 expression. Panel A and B display the top 2 KEGG pathways associated to PSAT1: Cytokine-cytokine receptor interaction ( A ) and Jak-STAT signaling pathway ( B ). Bar charts (left) represent enriched genes in each pathway, with red and green columns representing the association to high and low expression of PSAT1, respectively. Heatmaps of most significantly enriched genes in each pathway (enrichment statistic P < 0.01; genes are ordered based on decreasing average expression) in relation to PSAT1 expression are also shown (right).

Article Snippet: Quantification of PSAT1 was performed using the TaqMan probe–based gene expression assay Hs00253548_m1 specific for splice variant beta (Applied Biosystems/Life Technologies, Warrington, WA, USA) as previously described , , For Affymetrix gene expression profiling, total RNA samples were cleaned and DNAse treated with the NucleoSpin RNA II kit according the manufacturers instruction (Machery-Nagel, Dueren, Germany) and shipped to ServiceXS (Leiden, The Netherlands) for downstream processing with the 3’IVT express kit and hybridization on the Human Genome (HG) U133 Plus 2.0 array (n = 20) and HG U133 Perfect Match.

Techniques: Expressing