biostudies-arrayexpressMarco MernbergerHomo sapienshttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-15265Do illuminate the vulnerabilities of CDK4/6i-resistant T47D breast cancer cells, we created CDK4/6i-resistant T47D cell clones through a gradual dose escalation protocol using the selective CDK4/6 inhibitors Ribociclib and Palbociclib. We then performed gene expression analysis targeting 2549 genes using the HTG Oncology Biomarker Panel (OBP) via HTG EdgeSeq.biostudies-arrayexpressGrowth Protocol - T47D cells lines were obtained from the American Tissue Collection Center (ATCC) and grown in high-glucose DMEM supplemented with 10% fetal bovine serum, 100U ml-1 and 100 µg ml-1 Streptomycin at 37°C with 5% CO2. All cell lines were regularly tested and found to be negative for mycoplasma contamination. The metabolic drugs and molecular inhibitors were obtained from Sigma-Aldrich unless indicated otherwise and used at the following concentrations: Ribociclib (MedChemExpress) 0,125 – 32 µM, and Palbociclib (MedChemExpress) 0,125 – 32 µM.Sample Treatment - CDK4/6i-resistant T47D cells were generated using a dose escalation approach. Starting concentrations were set at 100-fold lower than the IC50 values for the respective cell line-inhibitor combinations. Parental T47D cells were treated with Ribociclib at concentrations ranging from 9,1 nM to 1 µM, and Palbociclib at concentrations from 23,2 nM to 1 µM. Parental MCF-7 cells were treated with Ribociclib from 0,73 nM to 1 µM, and Palbociclib from 2,8 nM to 1 µM. The resistant cell clones were selected and continuously cultured with 1 µM of the respective CDK4/6i. For the experiments, the cells were cultured in absence of the CDK4/6-inhibitors.Sample Collection - 1 x 106 cells were plated and cultivated for 48 hours. For cell harvesting and counting, adherent cells were treated with Trypsin-EDTA solution (2x) for 5 minutes at 37°C. The cells were resuspended in DMEM (with 10% FBS) and centrifuged. After washing with PBS, viable cell count was determined, and the appropriate number of cells (either 4 million or 2 million) was collected in a 1.5 mL Eppendorf tube.Library Construction - Gene expression analysis was performed targeting 2549 genes using the HTG Oncology Biomarker Panel (OBP) and HTG’s extraction-free nuclease protection assay (HTG Molecular Diagnostics, Tucson, USA). Cells were lysed and diluted to 3000 cells/35 µL of lysis buffer. 35 µL of the lysate was processed in the HTG EdgeSeq processor.Sequencing - After library preparation and pooling, samples were sequenced for 75 cycles on a NextSeq 550 Dx sequencer (Illumina, San Diego, USA).OrganizationMINSEQE ScoreAssays and DataMAGE-TAB FilesSequence Alignment - Read alignment and transcript counting were performed using HTG EdgeSeq Parser v5.4 with the Bowtie 2 algorithm. Differential expression analysis of the obtained transcript counts was performed using edgeR (version 3.40.0)UnknownTranscriptomicsGenomicsProteomicsNextSeq 550RNA-seq of coding RNAHomo sapiensAnne-Sophie LitmeyerCarsten DenkertAlina StrohMarco MernbergerMarie WittUwe WagnerThomas WündischLuise von WichertNiklas GremkeThorsten StieweJulia Teply-SzymanskiMichael WanzelfalseHTG EdgeSeq analysis of parental and CDK4/6i-resistant (Ribociclib or Palbociclib) T47D breast cancer cell clonesDo illuminate the vulnerabilities of CDK4/6i-resistant T47D breast cancer cells, we created CDK4/6i-resistant T47D cell clones through a gradual dose escalation protocol using the selective CDK4/6 inhibitors Ribociclib and Palbociclib. We then performed gene expression analysis targeting 2549 genes using the HTG Oncology Biomarker Panel (OBP) via HTG EdgeSeq.2026-06-17T00:00:00Z2026-06-17T01:00:35.508Z2025-06-25T13:57:05.757ZE-MTAB-15265ERP174034EFO_0004170EFO_0003789EFO_0004917EFO_0005518EFO_0003738EFO_0003969EFO_0004184