RNA based individualized drug selection in breast cancer patients without patient-matched normal tissue.

Michael Forster, Adam Mark, Friederike Egberts, Elisa Rosati, Elke Rodriguez, Martin Stanulla, Dirk Bauerschlag, Christian Schem, Nicolai Maass, Anu Amallraja, Karla K Murphy, Bruce R Prouse, Raed A Sulaiman, Brandon M Young, Micaela Mathiak, Georg Hemmrich-Stanisak, David Ellinghaus, Stephan Weidinger, Philip Rosenstiel, Norbert Arnold, Brian Leyland-Jones, Casey B Williams, Andre Franke, Tobias Meißner
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While standard RNA expression tests stratify patients into risk groups, RNA-Seq can guide personalized drug selection based on expressed mutations, fusion genes, and differential expression (DE) between tumor and normal tissue. However, patient-matched normal tissue may be unavailable. Additionally, biological variability in normal tissue and technological biases may confound results. Therefore, we present normal expression reference data for two sequencing methods that are suitable for breast biopsies.We identified breast cancer related and drug related genes that are expressed uniformly across our normal samples. Large subsets of these genes are identical for formalin fixed paraffin embedded samples and fresh frozen samples. Adipocyte signatures were detected in frozen compared to formalin samples, prepared by surgeons and pathologists, respectively. Gene expression confounded by adipocytes was identified using fat tissue samples. Finally, immune repertoire statistics were obtained for healthy breast, tumor and fat tissues.Our reference data can be used with patient tumor samples that are asservated and sequenced with a matching aforementioned method. Coefficients of variation are given for normal gene expression. Thus, potential drug selection can be based on confidently overexpressed genes and immune repertoire statistics.Normal expression from formalin and frozen healthy breast tissue samples using Roche Kapa RiboErase (total RNA) (19 formalin, 9 frozen) and Illumina TruSeq RNA Access (targeted RNA-Seq, aka TruSeq RNA Exome) (11 formalin, 1 frozen), and fat tissue (6 frozen Access). Tumor DE using 10 formalin total RNA tumor samples and 1 frozen targeted RNA tumor sample.